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Dear Editor Neonatal suppurative parotitis (NSP) is rare; fewer than 90 cases were reported before 1970[1]. A study by Spiegel et al identified only 32 cases in the English literature between 1970 and 2004[2]. Another study by Ismail et al identified 12 new cases in the English literature since 2004[3]. We are reporting another case. Most cases were male (77%) and one third of them were preterm infants[2,3]. Staphylococcus aureus was the most frequent offending organism. Ascending infection from the oral cavity through Stensen’s duct or, less commonly, hematogenous seeding of the parotid gland have both been implicated in the development of NSP[4]. Several risk factors for the development of NSP are recognized: dehydration, prematurity, excessive oropharyngeal suction, prolonged gavaging, ductal obstruction and immune suppression[2,3]. Typically, infants present in the second week of life with parotid gland swelling, fever and erythema in the overlying skin. Purulent material can drain spontaneously or be expressed from the Stensen’s duct.

rematurity, excessive oropharyngeal suction, prolonged gavaging, ductal obstruction and immune suppression[2,3]. Typically, infants present in the second week of life with parotid gland swelling, fever and erythema in the overlying skin. Purulent material can drain spontaneously or be expressed from the Stensen’s duct. A 19-day-old female infant presented with a 3-day history of fever, irritability, and right-sided periauricular swelling [Fig. 1]. She was born at 37 weeks + 3days gestation via normal vaginal delivery. Pregnancy was uneventful. Birth weight was 3200 g. On admission the baby was irritable. Her weight was 3450 g, and rectal temperature 38°C. A diffuse, tender, unilateral swelling in the right parotid region was observed. It was warm to touch and fluctuated. The rest of the physical examination was unremarkable. The mother reported no history suggestive of mastitis or recent infection. Fig. 1 19-day-old female infant with right-sided periauricular swelling Initial laboratory tests were as follows: white blood cell count 18600/mm3 with 53.4% neutrophils and 41% lymphocytes, hemoglobin 9.5 g/dl and erythrocyte sedimentation rate (ESR) 42 mm/h. Ultrasound examination of the mass demonstrated a cystic mass with septation and debris in the right parotid. Percutaneous aspiration of fluctuant area of the parotid gland was done and sample sent for culture. After obtaining blood and cerebrospinal fluid for culture empiric therapy with amikacin, vancomycin and ceftazidim was initiated. Surgical drainage was done 12 hours later.

ith septation and debris in the right parotid. Percutaneous aspiration of fluctuant area of the parotid gland was done and sample sent for culture. After obtaining blood and cerebrospinal fluid for culture empiric therapy with amikacin, vancomycin and ceftazidim was initiated. Surgical drainage was done 12 hours later. After four days of therapy, the swelling was not regressed. Culture of parotid abscess showed Staphylococcus aureus growth. With regard to antibiogram, clindamycin and meropenem was started and vancomycin and ceftazidim discontinued. Thereon blood culture, cerebrospinal fluid analysis and culture were normal. Daily drainage was not needed and after 10 days of therapy she was discharged. To our knowledge, acute bacterial infection of parotid gland in the neonatal age has not been reported yet from Iran. Our patient illustrates the typical manifestation of an acute suppurative parotitis.

After four days of therapy, the swelling was not regressed. Culture of parotid abscess showed Staphylococcus aureus growth. With regard to antibiogram, clindamycin and meropenem was started and vancomycin and ceftazidim discontinued. Thereon blood culture, cerebrospinal fluid analysis and culture were normal. Daily drainage was not needed and after 10 days of therapy she was discharged. To our knowledge, acute bacterial infection of parotid gland in the neonatal age has not been reported yet from Iran. Our patient illustrates the typical manifestation of an acute suppurative parotitis. Our patient was female unlike the most of reported patients[2,3]. She had unilateral parotid gland involvement, like the most of cases[2,3]. One third of the patients were prematures[2,3]. With regard to the average worldwide prematurity rate of 9.6%[5], it should be considered as a major risk factor for the infection. The patient was febrile on admission, however fever was reported in fewer than half of the patients[3]. As in our case, most patients had peripheral WBC count more than 15000/mm3[2,3]. Her ESR was 42mm/h which was elevated in only 20% of the patients[2]. S. aureus was the most frequently isolated pathogen in cultures of pus [2,3] same as in our case. However other Gram-positive organisms, Gram-negative organisms and anaerobic species were also isolated[2,6]. In our case blood culture was negative, like the most of cases[2,3]. She was product of a normal vaginal delivery. The causative agents are thought to be derived from the patients’ mouth flora. The newborns acquire their first microflora of the mouth, ear and skin from the mother’s birth canal during normal vaginal delivery[7]. She was breastfed, like the most of the patients with NSP[2] and raises the possibility of insufficient breast-feeding as a responsible factor for dehydration in these patients. Ultrasound is a useful device for diagnosis, and excludes other predisposing factors like Stenson’s duct abnormality, sialolith, and parotid gland neoplasm. The shortest effective duration reported in treating NSP due to S. aureus and in the absence of septicemia was 7 days[6]. The prognosis seems to be excellent, however as complications salivary fistula, facial palsy, mediastinitis, septicemia and meningitis were reported[1]. No deaths were reported in the patients studied after 1970[1].

A late preterm male baby with a birth weight of 2.5 kg was born to primi mother. Baby cried immediately after birth with an Apgar score of 8/8/9. Baby was antenatally diagnosed as a case of omphalocele with umbilical cord cyst. There was no history of any drug intake or any other chronic illness in the mother. Baby was delivered by LSCS. On physical examination infant had an omphalocele sac measuring about 14×10 cm with intestine as its content and umbilical cord cyst (Fig 1). X-Ray showed intestine as content of the sac (Fig. 2). Transillumination test was positive. Baby had webbing of neck (Fig. 3) and no other malformations. Fig. 1 Omphalocele sac 14×10 cm with intestine as its content and umbilical cord cyst Fig. 2 X-ray showing intestine as the content of the sac Infant was further evaluated with ECHO and Neurosonogram which were normal. Baby was operated on day 2 of life and intestines were reduced. Baby was gradually started on feeding and post-operative course was uneventful. Karyotypic analysis of the infant was normal. The infant was discharged successfully and is now in follow up. Omphalocele also known as exomphalos is a midline defect characterized by the evisceration of abdominal contents covered by a protective sac. The wall of the sac is formed by (inside to outside) peritoneum, Wharton’s jelly and amnion[1]. The defect is most commonly located at the base of the umbilical stalk in the midline. In omphalocele, the bowel does not return to the abdominal cavity between 10-12th week of gestation[2]. Fig. 3 Short webbed neck of the infant

Omphalocele also known as exomphalos is a midline defect characterized by the evisceration of abdominal contents covered by a protective sac. The wall of the sac is formed by (inside to outside) peritoneum, Wharton’s jelly and amnion[1]. The defect is most commonly located at the base of the umbilical stalk in the midline. In omphalocele, the bowel does not return to the abdominal cavity between 10-12th week of gestation[2]. Fig. 3 Short webbed neck of the infant The incidence of omphalocele ranges from 1.5 and 3 per 10,000 births[3,4]. Associated anomalies in omphalocele range from 50% -70% and are most important determinants in prognosis[5]. The treatment involves stabilization and surgical reduction of the sac content. The syndromes associated with omphalocele include[6]: • Otopalatodigital syndrome type II, • Melnick-needles syndrome, • Rieger syndrome, • Meckel syndrome, • Shprintzen-Goldberg omphalocele syndrome, • Lethal omphalocele-cleft palate syndrome, • Cerebro-costo-mandibular syndrome, • Fetal valproate syndrome, • Marshall-Smith syndrome, • Fryns syndrome, •Donnai-barrow syndrome, • Charge syndrome, • Goltz syndrome, • Carpenter syndrome, • Toriello-Carey syndrome, • Cornelia de Lange syndrome, • Sprengel anomaly, • Kennerknecht syndrome. What should be done? • Perinatal identification of omphalocele should be evaluated for omphalocele-related disorders and familial inheritance and a thorough genetic counseling should be done promptly for the parents. • Omphalocele associated malformations should be searched as there are very high chances of having them and help in prognostication of the parents.

• Perinatal identification of omphalocele should be evaluated for omphalocele-related disorders and familial inheritance and a thorough genetic counseling should be done promptly for the parents. • Omphalocele associated malformations should be searched as there are very high chances of having them and help in prognostication of the parents. • Karyotypic study should be done to rule out chromosomal disorders associated with omphalocele.

On 26th of April 2012, the “Autonomous Province of Trento” has brought into effect the Law n.31/2006 “Regulations for Diagnostic Post Mortem Investigation in Victims of the Sudden Infant Death Syndrome (SIDS) and Unexpected Fetal Death”, establishing an agreement with the “Lino Rossi” Research Center of the University of Milan. The Law states that all infants suddenly died within the first year of life, suspected of SIDS, and all fetuses died after the 25th week of gestation without any apparent cause, must undergo an anatomopathological examination[1]. We report the results until now obtained, focusing on the neuropathology of reflexogenic sudden perinatal and infant death. The study included 9 Sudden Intrauterine Unexpected Fetal Death Syndrome (SIUDS) (aged 19-40 gestational weeks) and 3 SIDS (aged from 20 hours to 6 months), occurring in the Autonomous Province of Trento (North Italy) in a 2-year period (2011-2012). For each case, all available information about pregnancy, fetal development and delivery and, in cases of infant death, about the environmental and familiar situation where the death occurred, besides information related to the potential risk factors (such as maternal smoking, maternal obesity, feeding, position the baby was last left in), were collected and categorised during post-mortem family interviews. The information sheets were recorded in a dedicated data bank, according to the art. 3 of the Law 31/2006, predisposed by the Autonomous Province of Trento and including two subsections: one for fetal loss and another for infant deaths. After removal two fresh samples of cerebral cortex (0.5-1 cm3 thick) for the genetic and toxicological investigations, respectively, the brainstem and the cerebellum, where the main vital centers are located (cardiorespiratory, arousal, upper digestive tract, etc.), were processed and embedded in paraffin. Transverse serial sections were made at intervals of 30 μm. For each level, six to seven 5 μm sections were obtained, two of which were routinely stained for histological examination using alternately hematoxylin-eosin and Klüver Barrera stains.

al, upper digestive tract, etc.), were processed and embedded in paraffin. Transverse serial sections were made at intervals of 30 μm. For each level, six to seven 5 μm sections were obtained, two of which were routinely stained for histological examination using alternately hematoxylin-eosin and Klüver Barrera stains. The other two sections were employed for immunohistochemical detection of the neuronal nuclear antigen 3 (NeuN). The remaining sections were saved for further investigations and stained as deemed necessary. The routine histological evaluation of the brainstem was focused on the locus coeruleus, parafacial/facial complex, superior olivary complex, retrotrapezoid nucleus, superior olivary nucleus, parabrachial/Kölliker-Fuse complex in the pons/mesencephalon; on the hypoglossus, the dorsal motor vagal, the tractus solitarius, the ambiguus, the pre-Bötzinger, the inferior olivary and the arcuate nuclei in the medulla oblongata and the intermediolateral nucleus in the spinal cord. We can summarize the results as follows: 1) Both unexplained fetal and infant deaths share common congenital anomalies of the central autonomic nervous system, so indicating that SIUDS should not be regarded as distinct from SIDS. 2) Noteworthy in the brainstem of the unexplained fetal death is the frequent association of the hypoplasia of the facial/parafacial complex with the hypoplasia of the raphe system. In particular, the hypodevelopment of the parafacial nucleus, consisting of “pre-inspiratory” neurons with the main function of hierarchical modulation of the breathing circuitry, is a very frequent finding in unexplained stillbirths. The raphe neurons in fact, known as major producers of serotonin and responsible for the serotonergic transmission during intrauterine life, play a trophic role in the neuronal development of fetal brains. A dysfunction in serotonergic transmission during intrauterine life could affect all neuronal structures checking vital functions, thus preventing the fetal life. 3) The NeuN protein is not expressed in both SIUDS and SIDS cases (Fig. 1). These congenital anomalies are likely to disrupt the vital actions by reflex mechanism, centered within bulbospinal structures in the domain of baro-(mechano)-chemoreflexes. These respond to variations in blood pressure and chemical breathing-depended components of blood and cerebrospinal fluid (e.g. pO2, pCO2, pH).

). These congenital anomalies are likely to disrupt the vital actions by reflex mechanism, centered within bulbospinal structures in the domain of baro-(mechano)-chemoreflexes. These respond to variations in blood pressure and chemical breathing-depended components of blood and cerebrospinal fluid (e.g. pO2, pCO2, pH). Fig.1 Left. NeuN-immunonegative neurons in a pontine histological section of a SIDS victim dead at 2 months. Right. NeuN_immunopositivity in an equivalent section of a control case (late fetal death -38 gestational weeks) for comparison - NeuN immunostaining. Magnification: 40×

). These congenital anomalies are likely to disrupt the vital actions by reflex mechanism, centered within bulbospinal structures in the domain of baro-(mechano)-chemoreflexes. These respond to variations in blood pressure and chemical breathing-depended components of blood and cerebrospinal fluid (e.g. pO2, pCO2, pH). Fig.1 Left. NeuN-immunonegative neurons in a pontine histological section of a SIDS victim dead at 2 months. Right. NeuN_immunopositivity in an equivalent section of a control case (late fetal death -38 gestational weeks) for comparison - NeuN immunostaining. Magnification: 40× The pathological involvement of the vagal-glossopharingeal neuronal circuitry explains some lethal reflexogenic mechanism, quoted in the literature as “drive”, “death-feigning” and/or “Ondina’s course” reflexes. Moreover, parahypoglossal glossopharingeal brainstem damage can promote suffocation by tongue hypotonia during supine sleep, especially if an abnormal reticular formation 4 impairs respiration and/or gastro-esophageal reflux goes into trachea. Overall the results obtained in the 12 cases of sudden death reported here, in agreement with those highlighted in our previous study[2] performed on a very large survey of perinatal deaths (140 SIDS, 95 SIUDS and 78 controls), show the common congenital origin of SIUDS and SIDS and support the statement of the National Institute of Child Health and Human Development: “SIDS is a developmental disorder. It takes its origin from fetal development”. In the world, only Italy has a national law that orders “Regulations for Diagnostic Post Mortem Investigation in Victims of the SIDS and Unexpected Fetal Death”. This law can be an example for the other nations in the world, and we believe it should be known. The present paper is the first result of the application of the law. The province of Trento (North Italy) applies this law because it is an autonomous province. The use of NeuN immunohistochemistry is another new important marker in the study of this pathology.

There is extensive evidence for short-term and long-term health benefits of breastfeeding for mothers and babies[1]. However, for some women coping with common breastfeeding problems in early postpartum is a physically and emotionally exhausting task, and they may harbor doubts about the continuation of breastfeeding[2]. A study conducted in Iran has reported that difficulties such as sore nipples or the mother's perception of having insufficient milk were the reasons for the exclusive breastfeeding discontinuation[3]. The mother’s perception and reaction to breastfeeding difficulties are affected by multiple factors, such as psychological and physical health, socio-demographic characteristics, quality of marital relationship and living condition[4]. Quality of life (QOL) is a broad ranging concept that includes all the mentioned aspects[5]. Therefore, the present study aims to investigate the relationships between QOL and breastfeeding difficulties in a sample of mothers from northeast Iran.

characteristics, quality of marital relationship and living condition[4]. Quality of life (QOL) is a broad ranging concept that includes all the mentioned aspects[5]. Therefore, the present study aims to investigate the relationships between QOL and breastfeeding difficulties in a sample of mothers from northeast Iran. 358 women who attended urban health centers, agreed to participate in the study. The inclusion criterion was gestational age of at least 28 weeks. The participants completed the World Health Organization Quality of Life - brief version (WHOQOL-BREF) in the third trimester of pregnancy. It contains 24 questions divided between 4 domains: Physical, Psychological, Social Relationships and Environment[5]. The validity and reliability of the Iranian version of WHOQOL-BREF have been supported in a previous study[6]. Women completed the Breastfeeding Experience Scale (BES) at 4 weeks postpartum. The first 18 questions of the BES rate the severity of common breastfeeding difficulties using a 5‐point Likert scale. Content validity and internal consistency of this scale (alpha coefficient 0.76) has been supported in a previous study[7]. In our study, the alpha coefficient was 0.82.

le (BES) at 4 weeks postpartum. The first 18 questions of the BES rate the severity of common breastfeeding difficulties using a 5‐point Likert scale. Content validity and internal consistency of this scale (alpha coefficient 0.76) has been supported in a previous study[7]. In our study, the alpha coefficient was 0.82. Mean age of women was 26.17 and 58.7% of them were primigravida. The mean total score of the breastfeeding difficulties questionnaire was 31.4±8.5. Common difficulties experienced by women were baby nursing too frequently (81.6%), difficulty in combining housekeeping and breastfeeding (69%), feeling very tired or fatigued (59%), and worry about having enough milk (52%). The correlation coefficients between breastfeeding difficulties score and the score of physical, mental, social, environmental, and global score of QOL were -0.217 (P<0.001), -0.172 (P=0.001), -0.157 (P=0.004), -0.154 (P=0.004), and -0.168 (P=0.002) respectively. Multiple regression analysis controlling for the effects of confounder variables showed that the global score of QOL was a predictor of breastfeeding difficulties (B= -0.21, CI [-0.20, -0.07]). The variables remained in the model explained 22.3% of the variance in breastfeeding difficulties (F=10.1, P=0.002). We explored differences of breastfeeding difficulties means according to different levels of satisfaction revealed in the independent Q1 of the WHOQOL-BREF, which asks about an individual’s overall perception of QOL. The mean breastfeeding difficulties score (SD) according to each level of satisfaction was as follows: ‘poor’ or ‘neither poor nor good’, 32.8 (8.5); ‘good’, 31.8 (8.5); and ‘’very good, 29.6 (8.5). There was a significant difference of breastfeeding difficulties scores according to 3 levels of perception of quality of life (F=3.15, P=0.04).

difficulties score (SD) according to each level of satisfaction was as follows: ‘poor’ or ‘neither poor nor good’, 32.8 (8.5); ‘good’, 31.8 (8.5); and ‘’very good, 29.6 (8.5). There was a significant difference of breastfeeding difficulties scores according to 3 levels of perception of quality of life (F=3.15, P=0.04). Our results indicate that there is a weak and negative correlation between quality of life scores and breastfeeding difficulties scores. However, prenatal QOL was related independently to breastfeeding difficulties. Mothers with poor QOL are more likely to experience breastfeeding difficulties in early postpartum. These findings are comparable with a Brazilian research, which found a correlation between breastfeeding self-efficacy and maternal QOL[8]. In addition, we found that women who perceived their QOL as ‘good’ or ‘very good’ had lower breastfeeding difficulties scores than mothers who perceived their QOL as ‘not bad, not good’. QOL was independently related to breastfeeding difficulties. Mothers with poor QOL are more likely to experience breastfeeding difficulties in early postpartum. To promote EBF, mothers with low QOL should be supported during the early postpartum.

Eruption of deciduous teeth, their exfoliation followed by eruption of permanent dentition is an orderly sequential and age specific event[1]. Significant deviations from accepted norms of eruption time are often observed. Most parents are anxious about delayed tooth eruption, as it is considered to be an important milestone during child’s development. Delayed tooth eruption might be the primary or sole manifestation of local or systemic pathology[2]. The systemic conditions like malnut-rition, rickets, endocrinopathies, chemotherapy, cerebral palsy and low birth weight can lead to delayed tooth eruption. Local factors delaying eruption may include presence of mucosal barriers, supernumerary teeth, odontogenic and non odontogenic tumors, local infections, injury and ankylosis of deciduous teeth.

Eruption of deciduous teeth, their exfoliation followed by eruption of permanent dentition is an orderly sequential and age specific event[1]. Significant deviations from accepted norms of eruption time are often observed. Most parents are anxious about delayed tooth eruption, as it is considered to be an important milestone during child’s development. Delayed tooth eruption might be the primary or sole manifestation of local or systemic pathology[2]. The systemic conditions like malnut-rition, rickets, endocrinopathies, chemotherapy, cerebral palsy and low birth weight can lead to delayed tooth eruption. Local factors delaying eruption may include presence of mucosal barriers, supernumerary teeth, odontogenic and non odontogenic tumors, local infections, injury and ankylosis of deciduous teeth. A 12 year old boy referred with a complaint of missing right central incisor tooth. Examination of his upper jaw revealed that the central incisor tooth was unerupted and the deciduous counterpart was retained with mesial drifting of the lateral tooth creating an anesthetic appearance (Fig. 1). Radiographic examination showed presence of multiple circular and irregularly shaped radiopaque structures in anterior maxilla, overlapping the crown of unerupted permanent tooth (Fig. 2). Based on the diagnosis of complex odontoma, the area was surgically explored and the tumorous masses were removed. The gross examination and histological evaluation confirmed the diagnosis of complex odontoma. Patient is presently under observation as parents opted to wait and orthodontic extrusion is planned if no physiologic eruption took place in 6 months post operative period. Odontomas are mixed odontogenic tumors, since they are composed of both epithelial and mesenchymal tissues. However, biologically regarded as hamartomas rather than neoplasms[3]. Most of the cases of odontomas are often undetected because they are clinically asymptomatic and nonaggressive. They are usually identified on routine radiographic examinations or during evaluation of delayed tooth eruption as in this case.

tissues. However, biologically regarded as hamartomas rather than neoplasms[3]. Most of the cases of odontomas are often undetected because they are clinically asymptomatic and nonaggressive. They are usually identified on routine radiographic examinations or during evaluation of delayed tooth eruption as in this case. A thorough visual, manual and radiographic examination should be performed for all pediatric patients who present with clinical evidence of delayed eruption, missing or displaced tooth. Fig. 1 Intra oral view showing unerupted permanent incisor and drifting of adjacent permanent lateral incisor Fig. 2 Radiograph showing presence of multiple radiopaque masses and retained permanent incisor Early identification and removal of odontomas help us to: Adopt less complex and invasive treatment Ensure better prognosis Avoid displacement or devitalization of adjacent tooth After removal of the obstacle from the path of eruption, an impacted tooth either erupts spontaneously if it has conserved its eruptive force or orthodontic force is required to bring the tooth in normal position[4].

Proliferation culture medium was prepared, which consisted of DMEM culture medium (GIBCO), 10% fetal bovine serum (FBS, GIBCO), penicillin (Shanghai new pioneer pharmaceuticals company) 100U/ml, streptomycin (Northern China pharmaceuticals company) 100µg/ml. PMCs were plated into 6 well culture-plates at the density of 1×107 cells/ml with volume of 2 ml per well, cultured for 3 days in a humidified atmosphere (37 °C, 5% CO2). The differentiation culture of peripheral mononuclear cells (PMC) [ 6 ] : After proliferation culture of peripheral mononuclear cells for 3 days, the culture medium was eliminated, and sterilized PBS was added, then PMCs were gently repeatedly beaten by pipette so that the unattached cells could be eliminated. 2 wells were again continually added with proliferation culture medium as negative controls, and the other 4 wells were added with changed culture medium, namely differentiation culture medium, which consisted of DMEM/F12 (GIBCO), 10% fetal bovine serum (GIBCO), penicillin (Shanghai new pioneer pharmaceuticals company) 100U/ml, streptomycin (Northern China Pharmaceuticals Co) 100µg/ml, all trans retinoid acid (Sigma Co) 0.5 µM, nerve growth factor (Sigma Co) 100ng/ml, insulin (Bo-Yun Biotech) 25 µg/ml, transferring (Bo-Yun Biotech) 100µg/ml, heparin (Wanbang Pharmaceuticals Co) 2 µg/ml, progesterone (Shanghai Pharmaceutical Co Ltd) 60 nM. PMCs were cultured in a humidified atmosphere (37°C, 5% CO2), and half volume of culture medium in each well was renewed with the same medium with the original medium in the well (Proliferation culture medium or differentiation culture medium) every 3 days. The cell shape and growth were daily observed under inverted phase contrast microscope. On the 10th day of differentiation culture, the immunohistochemistry was carried out to identify the differentiated cells.

Introduction Kawasaki disease (KD) is an acute febrile illness of childhood which presents as a systemic disease. While many organs are affected, involvement of cardiovascular system during early and late stages of the disease is considered as the leading cause of acquired heart disease in children in developed countries[1,2]. With more recognition of KD, it seems that the reported incidence is also increasing in some developing countries, even more than attacks of acute rheumatic fever in this age group[3-5]. While several decades have elapsed since Dr. Tomisaku Kawasaki described this disease for the first time, the cause is still unknown. Furthermore, epidemiological studies have revealed different incidence among different ethnic groups, and so the presence of certain human leukocyte antigens has been considered as a possible inherited predisposition for development of the disease[6-13]. It is postulated that exposure of susceptible person to the triggering pathogen may cause release of certain inflammatory mediators which can provoke vasculitis as the main pathological process with resultant clinical manifestations[14]. In this study we determined the distribution of human leukocyte B antigens among 90 Iranian patients with diagnosis of KD according to American Heart Association criteria[15]. We used the PCR-SSP technique as described in literature for antigen typing[16-19]. Subjects and Methods

While several decades have elapsed since Dr. Tomisaku Kawasaki described this disease for the first time, the cause is still unknown. Furthermore, epidemiological studies have revealed different incidence among different ethnic groups, and so the presence of certain human leukocyte antigens has been considered as a possible inherited predisposition for development of the disease[6-13]. It is postulated that exposure of susceptible person to the triggering pathogen may cause release of certain inflammatory mediators which can provoke vasculitis as the main pathological process with resultant clinical manifestations[14]. In this study we determined the distribution of human leukocyte B antigens among 90 Iranian patients with diagnosis of KD according to American Heart Association criteria[15]. We used the PCR-SSP technique as described in literature for antigen typing[16-19]. Subjects and Methods The study population: The study population consisted of 90 Iranian patients with final diagnosis of KD at the time of hospital discharge between 1991 and 2008 who were under follow up of pediatric immunology and pediatric cardiology services of Shiraz University of Medical Sciences, Shiraz, Iran. One hundred and nineteen patients were found to fulfill classic criteria of Kawasaki disease according to American Heart Association, among which 90 patients included in our study. Exclusion criteria were all patients presenting less than 4 criteria of Kawasaki. All of the patients were residents of south Iran. Informed consent for participation was obtained from the parents or their guardians. HLA-B distribution of 89 healthy Iranians was used as control[20]. The protocol of the study was approved by Ethical Committee of Shiraz University.

ng less than 4 criteria of Kawasaki. All of the patients were residents of south Iran. Informed consent for participation was obtained from the parents or their guardians. HLA-B distribution of 89 healthy Iranians was used as control[20]. The protocol of the study was approved by Ethical Committee of Shiraz University. Method: Two samples of one milliliter of blood was obtained from each subject and preserved in -20⁰C for Antigen typing by PCR. DNA extraction: Total DNA was extracted from 200 μL of whole blood by column based method using amplification DNA blood mini Kit (Qiagnene, England). Determination of DNA concentration: DNA concentration was measured by absorbance of the samples at 260 nm and 280 nm versus distilled water as blank. Optical density ratio between 260 nm/280 nm of all samples was above 1.8. The following formula was used to quantify DNA concentration. DNA (μg/ml)=Optical density 260 nm×dilution factor×50. Finally DNA concentration was adjusted to 50-100 μg/ml with distilled water and stored at -200C for analysis.

Determination of DNA concentration: DNA concentration was measured by absorbance of the samples at 260 nm and 280 nm versus distilled water as blank. Optical density ratio between 260 nm/280 nm of all samples was above 1.8. The following formula was used to quantify DNA concentration. DNA (μg/ml)=Optical density 260 nm×dilution factor×50. Finally DNA concentration was adjusted to 50-100 μg/ml with distilled water and stored at -200C for analysis. Antigen-B typing : Typing was performed by method of PCR-SSP, using BAG HISTO type low resolution kit (BAG Health Care GmbH, Germany). Genomic DNA (50-100 μg/ml) was amplified by PCR–SSP method with 48 pairs of sequence specific primers. Internal positive control was included in each reaction. A total of 48 reactions were done for each sample. This method is based on the fact that primer extension and hence successful PCR relies on an exact match at the 3-end of both primers. So, only if the primers entirely match the target sequence, amplification is obtained (Fig.1). For the PCR procedure the following components were included in 10 μL of reaction mixture: 1 × Polymerase chain reaction buffer (includes dNTps1, MgC12), Taq DNA polymerase, internal and sequence specific primers, template DNA. Amplification of PCR-SSP was carried out by using Eppendorf Mastercycler.

obtained (Fig.1). For the PCR procedure the following components were included in 10 μL of reaction mixture: 1 × Polymerase chain reaction buffer (includes dNTps1, MgC12), Taq DNA polymerase, internal and sequence specific primers, template DNA. Amplification of PCR-SSP was carried out by using Eppendorf Mastercycler. Gel electrophoresis : PCR products (48 for each sample) were subjected to 3% Agarose gel electrophoresis at 100-150 volt for 30-60 minutes. Then after termination of the run, the gels were stained in ethidium bromide solution for 30 minutes, and under ultraviolet illumination (220-310 nm) PCR products were visualized (Fig. 2). Documentation and interpretation: For interpretation of the results, specific tables and evaluation diagram were used. Fig. 1 Principle of PCR–SSP Fig. 2 Gel electrophoresis picture of PCR products Only bands with correct size were considered as positive reaction. Internal positive control bands were also observed in all reactions. For further reassurance, score evaluation software (BAG Health Care, Germany) was used for analysis of the results. The control group included 89 healthy Iranians who were residents of Shiraz and selected from a population pool on whom a genetic study was done by Farjadian et al in the area[20]. Data were analyzed by Pearson chi-square test and Fisher's exact test according to the frequency specific type antigen in the patients. SPSS version 15 was used for statistical analysis and a P value less than 0.05 considered statistically significant.

n whom a genetic study was done by Farjadian et al in the area[20]. Data were analyzed by Pearson chi-square test and Fisher's exact test according to the frequency specific type antigen in the patients. SPSS version 15 was used for statistical analysis and a P value less than 0.05 considered statistically significant. Findings The frequency of human leukocyte antigen type B of the patients and the control subjects is presented in Table 1. Comparison between frequency of 30 types of B* antigens among patients and control group revealed only 7 (3.9%) patients with positive antigen-B40* type, while 18 (10%) subjects of control group had this antigen with a statistically significant difference (95%CI: 0.13-0.84, RR=1.15 and P= 0.02). The frequency of the other 29 types of human leukocyte antigen type B had no statistically significant difference between the patients and control group (Table 2). Discussion While most of the early and late manifestations of KD are controlled with the use of intravenous immunoglobulin and acetylsalicylic acid, the etiology and possible predisposing factors are still undetermined. For development of KD, different etiological factors such as bacteria, viruses, mycoplasma, superantigens or even rug shampoo have been accused[1]. Table 1 Prevalence of complete Kawasaki criteria in the patients Parameter Frequency Sex: male/female 1:4 Lymph node 74% Conjunctivitis 94% Oral mucosa involvement 90% Extremity involvement 74% Rash 78% Cardiac involvement including coronary artery aneurism 26%

he original medium in the well (Proliferation culture medium or differentiation culture medium) every 3 days. The cell shape and growth were daily observed under inverted phase contrast microscope. On the 10th day of differentiation culture, the immunohistochemistry was carried out to identify the differentiated cells. Immunohistochemistry identification: Cells in each well were fixed with 4% paraform for 25 min, washed thoroughly with PBS 5 times, permeabilized with 1g/L Triton for 15 min, again washed thoroughly with PBS 5 times, rinsed with 3% H2O2 for 10 min, washed with PBS 5 times. Table 1 The clinical data of patients with neonatal hypoxic ischemic encephalopathy (HIE) Group (No) Controls (n=10) Mild HIE (n=10) Moderate HIE (n=10) Serious HIE With good Improveme nt (n=10) Serious HIE with poor improvement (n=10) F (X2) P. value Time after birth 11.8 (7.4) 8.3 (6.5) 13.8 (6.9) 13.2 (6.5) 9.8 (7.5) 1.12 0.4 Gestation weeks 38.6 (1.1) 39.1 (1.6) 39.1 (1.6) 39.1 (1.5) 39.1 (1.6) 0.20 0.9 Birth weight 333.3 (464.6) 3112.4 (362.7) 3233.2 (491.9) 3186.3 (536.0) 3241.1 (485.3) 0.29 0.9 Sex Male Female 6 4 5 5 6 4 7 3 4 6 2.17 0.8 Delivery Cesarean

Discussion While most of the early and late manifestations of KD are controlled with the use of intravenous immunoglobulin and acetylsalicylic acid, the etiology and possible predisposing factors are still undetermined. For development of KD, different etiological factors such as bacteria, viruses, mycoplasma, superantigens or even rug shampoo have been accused[1]. Table 1 Prevalence of complete Kawasaki criteria in the patients Parameter Frequency Sex: male/female 1:4 Lymph node 74% Conjunctivitis 94% Oral mucosa involvement 90% Extremity involvement 74% Rash 78% Cardiac involvement including coronary artery aneurism 26% Coronary artery aneurism 0.06% Table 2 Prevalence and percentage of human leukocyte antigen type B in the patients and controls Human Leukocyte Agntigen -B Patients (90) n % Controls (89) n % P value *07 9 5 4 2.2 0.2 *08 5 2.8 11 6.2 0.1 *13 6 3.3 2 1.1 0.3 *14 4 2.2 4 2.2 1.0 *15 3 1.7 3 1.7 1.0 *18 9 5 16 9 0.1 *27 2 1.1 4 2.2 0.4 *35 43 23.9 35 19.7 0.3 *.38 4 2.2 - 0 0.1 *39 3 1.7 1 0.6 0.6 *40 7 3.9 18 10.1 0.02 *41 7 3.9 1 0.6 0.07 *42 - 0 2 1.1 0.2 *44 8 4.4 5 2.8 0.4 *45 1 0.6 3 1.7 0.4 *49 2 1.1 1 0.6 1.0 *50 7 3.9 2 1.1 0.2 *51 17 9.4 21 11.8 0.5 *52 8 4.4 7 3.9 1.0 *53 5 2.8 13 7.3 0.05 *55 6 3.3 7 3.9 0.8 *56 1 0.6 1 0.6 1.0 *57 4 2.2 3 1.7 1.0 *58 7 3.9 9 5.1 0.6 *73 3 1.7 1 0.6 0.3 *81 - 0 4 2.2 0.06 *37 5 2.7 0 0 0.06 *54 2 1.1 0 0 0.5 *36 1 0.6 0 0 1.0

*40 7 3.9 18 10.1 0.02 *41 7 3.9 1 0.6 0.07 *42 - 0 2 1.1 0.2 *44 8 4.4 5 2.8 0.4 *45 1 0.6 3 1.7 0.4 *49 2 1.1 1 0.6 1.0 *50 7 3.9 2 1.1 0.2 *51 17 9.4 21 11.8 0.5 *52 8 4.4 7 3.9 1.0 *53 5 2.8 13 7.3 0.05 *55 6 3.3 7 3.9 0.8 *56 1 0.6 1 0.6 1.0 *57 4 2.2 3 1.7 1.0 *58 7 3.9 9 5.1 0.6 *73 3 1.7 1 0.6 0.3 *81 - 0 4 2.2 0.06 *37 5 2.7 0 0 0.06 *54 2 1.1 0 0 0.5 *36 1 0.6 0 0 1.0 *47 1 0.6 0 0 1.0 Rare presentation of KD in infants younger than 4 months may suggest protective role of maternal antibodies[14]. Having higher incidence among those with Asian background, especially 5000-6000 new cases annually in Japan, and the greatest occurrence in twins, suggest the role of a genetic predisposition[10]. This genetic background may predispose the patient to a super-antigen driven response with a selective expression of immune system and then the release of certain mediators of inflammation. The production of various inflammatory cytokines and elevated level of matrix metalloproteinases may mediate the vascular endothelial damages[14,21]. Accordingly, the presence of certain human leukocyte antigens may increase susceptibility of the patients to KD or may protect them against the disease or its complications. Kato et al demonstrated increased presence of human leukocyte antigen BW22 in patients with KD in comparison with control group (P<0.0005)[6]. Krensky et al in their report from Boston hospital showed increased prevalence of BW51 antigen among Kawasaki patients (P<0.002)[7]. While Kaslow et al in their study reported that the combination of different human leukocyte antigens may have a role. Chang et al from China, Hong Kong, did not show any association between presence of human leukocyte antigens, including type B and KD[8-9].

valence of BW51 antigen among Kawasaki patients (P<0.002)[7]. While Kaslow et al in their study reported that the combination of different human leukocyte antigens may have a role. Chang et al from China, Hong Kong, did not show any association between presence of human leukocyte antigens, including type B and KD[8-9]. Recently Maggioli et al reported the possible role of HLA class III region and susceptibility to KD[22].

valence of BW51 antigen among Kawasaki patients (P<0.002)[7]. While Kaslow et al in their study reported that the combination of different human leukocyte antigens may have a role. Chang et al from China, Hong Kong, did not show any association between presence of human leukocyte antigens, including type B and KD[8-9]. Recently Maggioli et al reported the possible role of HLA class III region and susceptibility to KD[22]. Also with respect to development of coronary artery aneurysms, polymorphism of transmembrane region of human leukocyte antigen genes and presence of certain alleles with increased susceptibility or a possible protective role for coronary complication in KD has been suggested[23-25]. So far studies from around the world have shown varied and inconsistent association between KD and single or summation of alleles (halotype)[26]. However, the possibility of a protective role for human leukocyte antigen in KD has not been reported as far as we are aware. For certain infectious diseases Cresio Alves et al have reviewed the association between human leukocyte antigen genes and increased susceptibility or protection against certain infections and their complication[27]. In their report, among Chilean patients with Chagas disease the incidence of cardiomyopathy was lower in those patients with antigen type B40*, however a report from Venezulea did not confirm this finding. Also for leprosy a protective role for type-A and type-B summations (halotype) has been suggested. The prevalence of antigen-B40* among our control group was about 10.1%. This antigen also has been the most frequent allele among Mexican, Korean, Chinese from Hainan province and Omanis[9,12,28,29]. Considering the higher frequency of antigen type-B40+ allele among our control subjects, may assume the possibility of a protective role for this antigen in our area. However, more studies are needed to find out the role of B-40* antigen and KD among mentioned ethnic groups with higher frequency of this antigen.

8,29]. Considering the higher frequency of antigen type-B40+ allele among our control subjects, may assume the possibility of a protective role for this antigen in our area. However, more studies are needed to find out the role of B-40* antigen and KD among mentioned ethnic groups with higher frequency of this antigen. Limitation of study: It is shown that many genetic characters can be transferred in halotype pattern, i.e. summation of multiple genes. The major limitation of our study is that we checked only the type-B antigens. Certainly further studies are needed to determine the possibility of a protective role for certain single gene or summation of genes for KD in our area. Conclusion The presence of higher frequency of allele type-B40* in the control group may represent a protective role for this antigen with resultant decreased susceptibility to KD in our area. Presence or lack of certain human leukocyte antigens has been assumed for expression of certain genes which can induce production of mediators of inflammatory response in this disease. Acknowledgment This work was financially supported by Vice Chancellor for Research of Shiraz University of Medical Sciences (Grant Number 86-3719). We would like to thank the families of Kawasaki patients who kindly contributed to this work. We are also thankful to Mrs. Akbarzadeh for her secretarial assistance. Conflict of Interest: None Authors’ Contribution Gh. Ajami: conceived and designed the study and prepared the manuscript.

Acknowledgment This work was financially supported by Vice Chancellor for Research of Shiraz University of Medical Sciences (Grant Number 86-3719). We would like to thank the families of Kawasaki patients who kindly contributed to this work. We are also thankful to Mrs. Akbarzadeh for her secretarial assistance. Conflict of Interest: None Authors’ Contribution Gh. Ajami: conceived and designed the study and prepared the manuscript. Kh. Aflaki: acquisition of Data, initially analyzed, interpreted the data and contributed in writing of initial manuscript. S. Alyasin: performed the literature search and assisted in first draft of manuscript writing. B. Gharesi-fard: acquisition of Data and HLA analysis. M. Borzouee: and contributed in the revision of the manuscript. H. Amoozgar: modified the analysis and contributed in the revision of the manuscript. All authors read and approved the final manuscript.

Introduction Germ cell tumors (GCT) are classified as malignant or benign tumors[1]. They consist of a broad variety of neoplasms that all originate from primordial germ cells. Although all of them have the same origin; but are different in pathological behavior and clinical presentation. These types of tumors are not common in children and comprise only 4 percent of childhood cancer[1]. The most common types of GCT are: teratomas, germinomas, endodermal sinus tumor or yolk sac tumors (YSTs)[2], choriocarcinoma and embryonal carcinoma. YSTs are the most prevalent malignant tumors of the gonads in children[1]. Despite their misleading name which appears to be in association with their anatomic place in the gonadal glands, most of them are extragonadal[3]. Our current knowledge regarding GCT in Iran is incomplete and since our center is a referral center we decided to design a study in order to gather demographic information of germ cell tumors and their treatment outcomes. Subjects and Methods Study population: All patients with the pathological diagnosis germ cell tumor (GCT) who were treated in Mofid Children’s Hospital during January 1999 to January 2009 were included. Data regarding age, sex, tumor site, bio-chemical assay, pathology, treatment and outcomes were gathered. Patient Treatment and Follow up: All patients underwent surgery in our center. Patient’s follow-up was carried out either in person in the clinic or by phone.

Study population: All patients with the pathological diagnosis germ cell tumor (GCT) who were treated in Mofid Children’s Hospital during January 1999 to January 2009 were included. Data regarding age, sex, tumor site, bio-chemical assay, pathology, treatment and outcomes were gathered. Patient Treatment and Follow up: All patients underwent surgery in our center. Patient’s follow-up was carried out either in person in the clinic or by phone. Statistical analysis: It is a retrospective descriptive study. For qualitative variables we computed frequency and percentage and for quantitative variables, mean and standard deviation. Survival analysis was performed using Kaplan-Meier. All the statistical analyses were performed by SPSS version16.0. Findings Forty four patients entered our study, of which 32 (72.7%) were girls and 12 (27.3%) boys; with a median age of 23 months. The pathological types are shown in Table 1. We had both gonadal and extra gonadal tumors in our patients with extra gonadal tumors being the more prevalent (72.72%). The anatomic distributions of the tumors are shown in Table2. The most prevalent stage at the time of diagnosis was stage 1 and in 5 cases the staging was unknown. Staging of the tumor at the time of diagnosis is summarized in Table 3. AFP and BHCG was measured in all patients. Table 1 Differentiation of pathology type Pathologic type Frequency (%) Mature teratoma (benign) 18 (40.9) Immature teratoma 6 (13.6) Yolk sac tumor 14 (31.8) Mixed tumors 3 (6.8) Malignant teratoma 1 (2.3) Dysgerminoma 1 (2.3) Choriocarcinoma 1 (2.3)

Findings Forty four patients entered our study, of which 32 (72.7%) were girls and 12 (27.3%) boys; with a median age of 23 months. The pathological types are shown in Table 1. We had both gonadal and extra gonadal tumors in our patients with extra gonadal tumors being the more prevalent (72.72%). The anatomic distributions of the tumors are shown in Table2. The most prevalent stage at the time of diagnosis was stage 1 and in 5 cases the staging was unknown. Staging of the tumor at the time of diagnosis is summarized in Table 3. AFP and BHCG was measured in all patients. Table 1 Differentiation of pathology type Pathologic type Frequency (%) Mature teratoma (benign) 18 (40.9) Immature teratoma 6 (13.6) Yolk sac tumor 14 (31.8) Mixed tumors 3 (6.8) Malignant teratoma 1 (2.3) Dysgerminoma 1 (2.3) Choriocarcinoma 1 (2.3) Total 44 (100) AFP was elevated in 14 cases of yolk sac tumor and one patient of immature teratoma. Also BHCG was high in one case of choriocarcinoma. Those patients who had very high level of AFP (3 times more than the upper limit of normal) died. All patients were treated surgically: total resection in 41 (93.2%) cases, partial resection in 1 (2.3%) case and open biopsy in 2 (4.5%) cases. Fifteen (34.1%) patients received chemotherapy. Follow up results in our patients demonstrated that we had 31 (77.5%) cases with complete remission and 9 (22.5%) deaths; 4 patients were lost to follow up. The median survival in our study was 16 months (IQR 4-49 months). Table 2 Gonadal and extra-gonadal tumor locations Sex/ Site Gonadal Extra Gonadal Total

Total 44 (100) AFP was elevated in 14 cases of yolk sac tumor and one patient of immature teratoma. Also BHCG was high in one case of choriocarcinoma. Those patients who had very high level of AFP (3 times more than the upper limit of normal) died. All patients were treated surgically: total resection in 41 (93.2%) cases, partial resection in 1 (2.3%) case and open biopsy in 2 (4.5%) cases. Fifteen (34.1%) patients received chemotherapy. Follow up results in our patients demonstrated that we had 31 (77.5%) cases with complete remission and 9 (22.5%) deaths; 4 patients were lost to follow up. The median survival in our study was 16 months (IQR 4-49 months). Table 2 Gonadal and extra-gonadal tumor locations Sex/ Site Gonadal Extra Gonadal Total Female 9 (Ovary) 15 (Sacrococcygeal) 5 (Abdomen and Retroperitoneal) 2 (Mediastinum) 1 (Head and Neck) 32 (72.72%) Male 3 (Testis) 6 (Sacrococcygeal) 2 (Abdomen and Retroperitoneal) 1 (Head and Neck) 12 (27.27%) Total 12 (27.27%) 32 (72.72%) 44 (100%) Table 3 Staging of the tumor at the time of diagnosis Stage Frequency (%) 1 23 (52.3) 2 3 (6.8) 3 2 (4.5) 4 11 (25) Total 39 (86.1) The Kaplan-Meier survival plot of the patients is shown in Fig 1. The highest mortality rate was found in patients with yolk sac tumors (8 of 13 cases). One other case of mortality was found in a patient with immature teratoma (1 of 6 cases).

Total 12 (27.27%) 32 (72.72%) 44 (100%) Table 3 Staging of the tumor at the time of diagnosis Stage Frequency (%) 1 23 (52.3) 2 3 (6.8) 3 2 (4.5) 4 11 (25) Total 39 (86.1) The Kaplan-Meier survival plot of the patients is shown in Fig 1. The highest mortality rate was found in patients with yolk sac tumors (8 of 13 cases). One other case of mortality was found in a patient with immature teratoma (1 of 6 cases). Discussion GCTs are a rare and diverse group of heterogeneous tumors that include both benign and malignant histologies[4]. Since they are rare, conducting a retrospective research seemed the best policy for gathering information on these tumors in our country and paving the way for future studies. Pathologic type: In our study there were a total of 44 patients, and the most common pathologic type was mature teratoma (41%), in a review by Zachary Horton et al teratomas are considered to be the most prevalent type of germ cell tumors in children[5]. In a study by Billmire and colleagues, which was carried out on 131 children with primary ovarian tumor the most common histology was reported to be teratoma + EST (endodermal sinus tumor ) and teratoma +other malignant elements[6]. Billmire et al. in another study on 26 patients with malignant retroperitoneal and abdominal germ cell tumors reported that the most common pathologic type was pure yolk sac tumor[7].

n tumor the most common histology was reported to be teratoma + EST (endodermal sinus tumor ) and teratoma +other malignant elements[6]. Billmire et al. in another study on 26 patients with malignant retroperitoneal and abdominal germ cell tumors reported that the most common pathologic type was pure yolk sac tumor[7]. Location: We had 32 cases of extragonadal and 12 cases of gonadal involvement. In Marina et al[8], study within 73 patients with extra cranial immature teratomas, there were 51 gonadal involvements and 22 extra gonadal cases. In Brodeur GM et al[9] study with the total number of57 patients, gonadal involvement was seen in 30 cases and extragonadal involvement in 27 cases. Fig. 1 Kaplan–Meier survival plot of the 40 patients after surgery for pediatric germ cell tumors Stage at diagnosis: In our study the most common stage at diagnosis was stage one (53%), and in Billmire et al. study[1] with a total number of 26 cases they had 3 stage I to II, 5 stage III, and 17 stage IV. In another study by Billmire et al[6], on 131 patients with primary ovarian tumor the most common stage was III (58 cases) followed by stage I (41 cases).

ommon stage at diagnosis was stage one (53%), and in Billmire et al. study[1] with a total number of 26 cases they had 3 stage I to II, 5 stage III, and 17 stage IV. In another study by Billmire et al[6], on 131 patients with primary ovarian tumor the most common stage was III (58 cases) followed by stage I (41 cases). Serum marker: Mann et al[10] evaluated 126 patients aged 0 to younger than 16 years with malignant GCT, serum AFP measured in 123 patients was elevated in 115 patients, whereas HCG was high in 19 of 77 cases. In our study only in 14 cases of yolk sac tumor and one patient of immature teratoma, AFP was elevated, and also BHCG was increased in one case of choriocarcinoma. Those patients in whom AFP level was very high (3 times more than upper limit of normal) died. As stated in the International Germ Cell Consensus Classification[11] the degree of elevation of alpha-fetoprotein, human gonadotropin, and lactic dehydrogenase in adults is a risk factor for poor prognosis but this is not true in pediatric GCTs and only degree of elevation of AFP is associated with a poor prognosis in the pediatric population. This is consistent with the finding of our study.

gree of elevation of alpha-fetoprotein, human gonadotropin, and lactic dehydrogenase in adults is a risk factor for poor prognosis but this is not true in pediatric GCTs and only degree of elevation of AFP is associated with a poor prognosis in the pediatric population. This is consistent with the finding of our study. Survival: The 5-year relative survival rate in Smith et al[12] was 83.9%, and varied by histologic subtype, race, stage of disease, and age at diagnosis. The estimated 5-year survival rate in Marina et al[13] is 100% for patients with stage I disease, 87% for stage II, 72% stage III, and 56% for stage IV. Six-year survival rate was 95% in stage I, 94% in stage II, 98% in stage III, and 93% in stage IV, in Billmire et al. study[6]. In Mann et al study[14] the 5-year survival rate in all 184 patients was 93.2%. In our study follow up results demonstrated complete remission in 31 (77.5%), patients and 9 (22.5%) patients died. Conclusion It seems that patients with extra-gonadal GCT and high AFP levels have a worse prognosis and lower survival rate; but a more powerful study including more patients from different pediatric surgery centers in Iran should be carried out in order to reach a solid conclusion. Combination of surgery and chemotherapy can lead to a better prognosis. Acknowledgment This study was financially supported by the office of the Vice-chancellor for Clinical Research of the Tehran University of Medical Sciences. Conflict of Interest: None Authors’ Contribution A.A Khaleghnejad: Concept, design, data analysis and surgery.

Conclusion It seems that patients with extra-gonadal GCT and high AFP levels have a worse prognosis and lower survival rate; but a more powerful study including more patients from different pediatric surgery centers in Iran should be carried out in order to reach a solid conclusion. Combination of surgery and chemotherapy can lead to a better prognosis. Acknowledgment This study was financially supported by the office of the Vice-chancellor for Clinical Research of the Tehran University of Medical Sciences. Conflict of Interest: None Authors’ Contribution A.A Khaleghnejad: Concept, design, data analysis and surgery. B.A. Mirshemirani: Critical revision of manuscript, surgery and funds. C.M. Rouzrokh: Acquisition of data, manuscript preparation and surgery. D.L. Mohajerzadeh: Data collection and analysis, and surgery. E.N, Khaleghnejad: Data collection and analysis. F.Sh, Hasas-yeganeh: Data collection and analysis All authors approved the final version of the manuscript

Introduction Neonatal hypoxic ischemic encephalopathy (HIE) is one of the most common causes of death and long-term neurological impairment in full-term neonates worldwide. Up to now, there is no specific effective treatment for the severe HIE. The potential use of stem cells to reduce brain damage is a possibility[1]. It has been found that asphyxia influences proliferation and differentiation of brain neural stem cells in newborn animal models[2-4], and that peripheral blood stem cells play an important role in repairing brain damage[5]. But It has not been reported yet whether asphyxia influences peripheral blood stem cells differentiating into neural cells, and whether with the progress of the disease there is a change of peripheral blood stem cells differentiating into neural cells in newborns with HIE. This research will focus on these questions.

. But It has not been reported yet whether asphyxia influences peripheral blood stem cells differentiating into neural cells, and whether with the progress of the disease there is a change of peripheral blood stem cells differentiating into neural cells in newborns with HIE. This research will focus on these questions. Subjects and Methods Fifty term newborn infants from the pediatric department of Shanghai Tongji hospital affiliated to Tongji University, Shanghai, China, were enrolled into the study from March 2007 to March 2010. 72 hours after birth, the severity of HIE was evaluated according to The Sarnat Grading Scale of HIE, and 20 serious cases, 10 moderate cases, and 10 mild cases were identified. All the patients met the HIE diagnostic criteria. 10 severe cases improved obviously 14 days after birth, which were defined as no consciousness, no seizure, no abnormal muscle tone, no brain stem symptom, whereas 10 severe cases poorly improved, which were defined as having one or more of the above manifestations 14 days after birth or died. 2 cases died within 1 week, and 6 cases died after 1 week. 10 newborn infants with wet lung acted as controls, presenting with breath difficulty, but oxygen saturation was normal. The study protocol was reviewed and approved by the local institutional ethics committee and written informed consent was obtained from all guardians prior to the study Blood sample collection:

Subjects and Methods Fifty term newborn infants from the pediatric department of Shanghai Tongji hospital affiliated to Tongji University, Shanghai, China, were enrolled into the study from March 2007 to March 2010. 72 hours after birth, the severity of HIE was evaluated according to The Sarnat Grading Scale of HIE, and 20 serious cases, 10 moderate cases, and 10 mild cases were identified. All the patients met the HIE diagnostic criteria. 10 severe cases improved obviously 14 days after birth, which were defined as no consciousness, no seizure, no abnormal muscle tone, no brain stem symptom, whereas 10 severe cases poorly improved, which were defined as having one or more of the above manifestations 14 days after birth or died. 2 cases died within 1 week, and 6 cases died after 1 week. 10 newborn infants with wet lung acted as controls, presenting with breath difficulty, but oxygen saturation was normal. The study protocol was reviewed and approved by the local institutional ethics committee and written informed consent was obtained from all guardians prior to the study Blood sample collection: 5 ml peripheral blood was collected from all newborn infants within 24 hours after birth. Again 5 ml blood was collected from severe HIE patients on 7th day after birth if the patients were still alive. The mononuclear cells were separated and collected with Ficoll separating medium (GIBCO ) 1500r/min×5min, re-suspended in proliferation culture medium (see below), which were applied in the following process. The proliferation culture of peripheral mononuclear cells (PMCs) [ 6 ] :

5 ml peripheral blood was collected from all newborn infants within 24 hours after birth. Again 5 ml blood was collected from severe HIE patients on 7th day after birth if the patients were still alive. The mononuclear cells were separated and collected with Ficoll separating medium (GIBCO ) 1500r/min×5min, re-suspended in proliferation culture medium (see below), which were applied in the following process. The proliferation culture of peripheral mononuclear cells (PMCs) [ 6 ] : Proliferation culture medium was prepared, which consisted of DMEM culture medium (GIBCO), 10% fetal bovine serum (FBS, GIBCO), penicillin (Shanghai new pioneer pharmaceuticals company) 100U/ml, streptomycin (Northern China pharmaceuticals company) 100µg/ml. PMCs were plated into 6 well culture-plates at the density of 1×107 cells/ml with volume of 2 ml per well, cultured for 3 days in a humidified atmosphere (37 °C, 5% CO2). The differentiation culture of peripheral mononuclear cells (PMC) [ 6 ] :

improvement (n=10) F (X2) P. value Time after birth 11.8 (7.4) 8.3 (6.5) 13.8 (6.9) 13.2 (6.5) 9.8 (7.5) 1.12 0.4 Gestation weeks 38.6 (1.1) 39.1 (1.6) 39.1 (1.6) 39.1 (1.5) 39.1 (1.6) 0.20 0.9 Birth weight 333.3 (464.6) 3112.4 (362.7) 3233.2 (491.9) 3186.3 (536.0) 3241.1 (485.3) 0.29 0.9 Sex Male Female 6 4 5 5 6 4 7 3 4 6 2.17 0.8 Delivery Cesarean vaginal 7 3 6 4 5 5 4 6 3 7 3.94 0.5 Agent A (Normal horse serum) from immunohistochemistry kit was dropped into culture wells (Hangzhou Baitong Biotechnology Co Ltd), incubated and protected from light for 45 min to block antigens. 5μg/ml anti-Nestin antibody (R&D systems) was added, incubated and protected from light at 4℃ overnight. Each cell was rinsed and washed with PBS for 5 min×5 times, and agent B (biotin labeled second antibody) from immunohistochemistry kit was added, incubated and protected from light at room temperature for 60 min, rinsed and washed with PBS for 5min×5 times. Agent C (horseradish peroxydase complex) was added and incubated at room temperature for 15 min, washed with PBS 5 for 5 min×5 times. DAB was added into each well to react for 3 min, and then immediately washed with double distilled water thoroughly. Immunohistochemistry positive cells in each well further identified by cell shape, which had at least two long cell processes, were counted under microscope, and the average number of positive cells in 4 wells (added with differentiation culture medium) was taken to be the positive cells number. Statistical methods:

vaginal 7 3 6 4 5 5 4 6 3 7 3.94 0.5 Agent A (Normal horse serum) from immunohistochemistry kit was dropped into culture wells (Hangzhou Baitong Biotechnology Co Ltd), incubated and protected from light for 45 min to block antigens. 5μg/ml anti-Nestin antibody (R&D systems) was added, incubated and protected from light at 4℃ overnight. Each cell was rinsed and washed with PBS for 5 min×5 times, and agent B (biotin labeled second antibody) from immunohistochemistry kit was added, incubated and protected from light at room temperature for 60 min, rinsed and washed with PBS for 5min×5 times. Agent C (horseradish peroxydase complex) was added and incubated at room temperature for 15 min, washed with PBS 5 for 5 min×5 times. DAB was added into each well to react for 3 min, and then immediately washed with double distilled water thoroughly. Immunohistochemistry positive cells in each well further identified by cell shape, which had at least two long cell processes, were counted under microscope, and the average number of positive cells in 4 wells (added with differentiation culture medium) was taken to be the positive cells number. Statistical methods: SPSS13.0 for windows was adopted to analyze the data. The difference of Neuron marker nestin positive cells, time after birth, gestational weeks, birth weight among HIE newborns with different severity within 24 hr was analyzed by ANOVA, The difference of sex and delivery mode among HIE newborns with different severity was analyzed by chi-square test. The difference of Neuron marker nestin positive cells in the severe HIE newborns between different improvement groups on 7th day was analyzed by t test.

ferent severity within 24 hr was analyzed by ANOVA, The difference of sex and delivery mode among HIE newborns with different severity was analyzed by chi-square test. The difference of Neuron marker nestin positive cells in the severe HIE newborns between different improvement groups on 7th day was analyzed by t test. Findings The patients’ clinical data was as follows (Table 1). There was no difference among groups as to sex, delivery, time after birth, gestational weeks and birth weight. All severe HIE patients received tracheal intubation and mechanical ventilation. After 10 days of cell culture in vitro, nestin positive cells with long cell processes could be identified among PMCs (Fig. 1). The number of cells with neuron marker nestin had no difference among the mild HIE group, the moderate HIE group, the severe HIE with good improvement group, the severe HIE with poor improvement group and the controls (Table 2). The number of cells with neuron marker nestin differentiated from PMCs in the severe HIE with good improvement on 7th day was obviously higher than that within 24 hours after birth, and also higher than the number of cells with neuron marker nestin differentiated from PMCs in the severe HIE with poor improvement on 7th day. Table 2 Induced neuron marker nestin positive cells in HIE newborns within 24 hr after birth Group Control (n=10) Mild (n=10) Moderate (n=10) Severe with good improvement (n=10) Severe with poor improvement (n=10) F P. value Nestin Positive cells 71.13 (7.19) 70.46 (6.66) 73.34 (6.46) 68.99 (7.85) 71.43 (6.88) 0.51 0.7 HIE: hypoxic ischemic encephalopathy

Table 2 Induced neuron marker nestin positive cells in HIE newborns within 24 hr after birth Group Control (n=10) Mild (n=10) Moderate (n=10) Severe with good improvement (n=10) Severe with poor improvement (n=10) F P. value Nestin Positive cells 71.13 (7.19) 70.46 (6.66) 73.34 (6.46) 68.99 (7.85) 71.43 (6.88) 0.51 0.7 HIE: hypoxic ischemic encephalopathy Fig. 1 PMCs induced to differentiate into nestin positive cells Note:“ * ” indicates nestin positive cells, DAB stain. Control: A (×200, <24h), B (×400, <24h), C (×200, 7d), D (×400, 7d); mild HIE: E (×200, <24h), F (×400, <24h), G (×200, 7d), H (×400, 7d) ; moderate HIE: I (×200, <24h), J (×400, <24h), K (×200, 7d), L (×400, 7d); Severe HIE with poor improvemnet: M (×200, <24h), N (×400, <24h), O (×200,7d), P (×400, 7d); Severe HIE with poor improvement: Q (×200, <24h), R (×400, <24h), S (×200, 7d), T (×400,7d); Negative control: U (×200, <24h), V (×400, <24h), W (×200, 7d), X (×400,7d) The number of cells with neuron marker nestin in the severe HIE with poor improvement on 7th day was not different from that within 24 hours after birth (Table 3). Discussion HIE is one of the main causes of death and long- term neurological impairment in full-term neonates. There is no specific effective treatment for the severe HIE. Many investigations have identified endogenous neural stem cells were mobilized when HIE took place, namely theneural stem cells in brain take part in HIE repair[1], and there are also investigations revealing that peripheral blood stem cells have close relation to the damage repair of HIE brain[5,7-11]

the severe HIE. Many investigations have identified endogenous neural stem cells were mobilized when HIE took place, namely theneural stem cells in brain take part in HIE repair[1], and there are also investigations revealing that peripheral blood stem cells have close relation to the damage repair of HIE brain[5,7-11] Umbilical mononuclear cells consist of multipotent stem cells[12] and mesenchymal stem cells[13] with neuron differentiation potential. Umbilical blood is a part of the neonatal peripheral blood, so it is likely that the neonatal peripheral blood cells possess similar character of the umbilical blood cells, namely consist of stem cells with neuron differentiation potential. Table 3 Induced neuron marker nestin positive cells in the severe HIE newborns with different improvement on 7th day after birth Group/Time 24 hr 7 d t P. value Severe HIE with good improvement (n=10) 68.99 (7.85) 94.50 (15.57) 4.66 <0.001 Severe HIE with poor improvement (n=8 * ) 72.60 (7.27) 69.48 (5.32) 1.46 0.2 t 1.001 4.621 P. value 0.332 0.000 * 8 cases: 2 cases died within 1 week, and the data on 7th day was absent, so the 2 cases were deleted when the data within 24 hrs after birth was analyzed only among the alive patients on 7th day

Severe HIE with good improvement (n=10) 68.99 (7.85) 94.50 (15.57) 4.66 <0.001 Severe HIE with poor improvement (n=8 * ) 72.60 (7.27) 69.48 (5.32) 1.46 0.2 t 1.001 4.621 P. value 0.332 0.000 * 8 cases: 2 cases died within 1 week, and the data on 7th day was absent, so the 2 cases were deleted when the data within 24 hrs after birth was analyzed only among the alive patients on 7th day Our data confirmed that PMCs from newborns and newborns with HIE could differentiate into cells with neural marker namely possessing neuron cells differentiation potential. Human peripheral CD14+ monocytes[14] and CD133+ stem cells in umbilical blood[15] can differentiate into cells with neural markers. It needs to be further confirmed which PMCs gradients in HIE newborn patients can differentiate into neuron cells. The influence of asphyxia on neonatal peripheral blood stem cells differentiating into neuron cells has not yet been reported by now. Ischemia and anoxia have definite influence on neural stem cells in brain[2-4,16,17]. We could not find the influence of asphyxia on the ability of neonatal PMCs differentiating into neural cells, which may suggest the different influence of asphyxia on central neural stem cells and peripheral blood stem cells with neuron differentiation potential.

ite influence on neural stem cells in brain[2-4,16,17]. We could not find the influence of asphyxia on the ability of neonatal PMCs differentiating into neural cells, which may suggest the different influence of asphyxia on central neural stem cells and peripheral blood stem cells with neuron differentiation potential. We found the ability of PMCs from severe HIE with good improvement on 7th day after birth to neural cells obviously increase, which may suggest these patients predominate at promoting neural cells differentiation, and may also suggest peripheral blood stem cells play a possible role in the HIE repair. The mechanism of peripheral blood stem cells taking part in central neural cells repair is unclear, which may relate to directly entering central nervous system to differentiate into neural cells to replace the injured brain tissues or to produce neurotrophin to promote the proliferation and differentiaton of brain internal neural stem cells[18-27]. The different differentiation ability of PMCs with the progress of HIE has close relationship with the prognosis. If the differentiation ability of PMCs can be identified as early as possible by improving the culture methods to shorten the culture time, it may be valuable to judge the prognosis.

ural stem cells[18-27]. The different differentiation ability of PMCs with the progress of HIE has close relationship with the prognosis. If the differentiation ability of PMCs can be identified as early as possible by improving the culture methods to shorten the culture time, it may be valuable to judge the prognosis. The ability of peripheral mononuclear cells differentiating into neural cells in term infants with good improvement suffering from severe HIE was enhanced, which may suggest possible relationship between the brain repair and the peripheral stem cells. The mechanism of PMCs taking part in the protection of central nervous system injury suffered from asphyxia need to be further investigated. Conclusion The ability of peripheral mononuclear cells differentiating into neural cells in term infants with good improvement suffering from severe hypoxic ischemic encephalopathy was enhanced, which may suggest possible relationship between the brain repair and the peripheral stem cells. Acknowledgment This work was supported by Shanghai Municipal Health Bureau Scientific Research Projects (No 2009139) and The key neonatal laboratory of the Ministry of Health Open Projects (No 200801). Conflict of Interest: None Authors’ Contribution Concept / Design: G. Xiaohui Acquisition of Data: W. Dong; Z. Yuwen Data Analysis / Interpretation: W. Dong; Z. Yuwen Manuscript Preparation: W. Dong Critical Revision of the Manuscript: G. Xiaohui Funds Collection: W. Dong All authors approved final version of the manuscript.

Nonalcoholic fatty liver disease (NAFLD) has become the most common cause of chronic liver disease in the pediatric population in the last few decades[1,2]. In Western countries, the prevalence of NAFLD is estimated up to 10% of children. In Iran fatty liver was diagnosed by ultrasound in 7.1% of children aged 7-18 years[1]. Increased prevalence of this form of liver disease runs parallel with the dramatic rise in childhood obesity and diabetes mellitus over the past 2 decades. Most of children with NAFLD are low active and obese, and suffer from metabolic impairments (including increased baseline waist circumference, hypertension and high insulin level) and dyslipidemia[3]. There was a strong relationship between NAFLD and the abnormal metabolic variables in children[4]. Higher age, higher fasting insulin, abnormality in total cholesterol, LDL cholesterol and triglycerides is linked with higher risk of NAFLD[1]. Pediatric NAFLD is not a benign condition, and it can be a predictive of cardiovascular disease and diabetes mellitus in adulthood and some times it can progress to severe liver diseases[4].

gher fasting insulin, abnormality in total cholesterol, LDL cholesterol and triglycerides is linked with higher risk of NAFLD[1]. Pediatric NAFLD is not a benign condition, and it can be a predictive of cardiovascular disease and diabetes mellitus in adulthood and some times it can progress to severe liver diseases[4]. The mechanisms leading to NAFLD in children is the same as in adults and include genetic background, epigenetics and environmental factors (i.e. high caloric intake, daily consumption of junk food and soft drinks, low level of physical activity, and beyond average weight) all of which concur during development and progression of the disease[5]. Obesity is the circle between the risk factors showing that 70% of NAFLD in pediatric group is associated with obesity[6]. Faghih et al found a tremendous trend toward over-weight and obesity with a 3-fold increase of obesity among adolescent girl students in Ahvaz between 1997 and 2006. The data showed that there was a significant relation between BMI and food habits including number of meals and missing the breakfast. The authors recommend, regarding the harmful ramifications of obesity, to encourage healthy eating patterns and increased physical activity among adolescent girls[7].

een 1997 and 2006. The data showed that there was a significant relation between BMI and food habits including number of meals and missing the breakfast. The authors recommend, regarding the harmful ramifications of obesity, to encourage healthy eating patterns and increased physical activity among adolescent girls[7]. Diagnosis is established by liver biopsy. Laboratory tests and radiographic findings provide clues to the potential presence of fatty liver disease. Liver enzymes elevation in childhood is a cardio-metabolic risk factor and an additional component of the metabolic syndrome[8]. In all children with abnormal level of liver enzymes, evaluation of NAFLD should be done. To screen children susceptible to NAFLD, it will be very useful to assess waist circumference, fasting blood sugar, fasting insulin and serum lipid profile in regular intervals.

n additional component of the metabolic syndrome[8]. In all children with abnormal level of liver enzymes, evaluation of NAFLD should be done. To screen children susceptible to NAFLD, it will be very useful to assess waist circumference, fasting blood sugar, fasting insulin and serum lipid profile in regular intervals. Nutrition plays essential role in development of NAFLD and it can regress the fatty accumulations in the liver. Lifestyle modification, including slow and steady weight loss, improved dietary habits, and increased daily, aerobic physical activity, remains the first-line approach in treating pediatric fatty liver disease[4]. The ministries of health and education have agreed in 2007 about the instructions for healthy eating in schools. Healthy nutrition bases or markets inside the schools should be established for providing, maintaining and promotion of physical, mental and social activities of the students. The main goals are providing healthy food and avoiding unhealthy food supply for students. Healthy food means providing body’s nutritional needs with safe, adequate, varied and balanced foods. It is a common knowledge that healthy food contains low salt and low-fat with less than 10% of trans-fatty oil. Soft drinks should be eliminated from student’s regimen, mineral water and fresh fruits can replace it.

Introduction High risk pregnancy refers to pregnancy accompanied by factors which increase the risk of neonatal mortality and morbidity. Based on statistics 10-20% of pregnancies are reported as high risk pregnancies[1]. Neonatal state of health has a considerable effect on future health and life. Since neonate`s immune system and other organs in preterm neonate are not developed completely, they are at risk of many threats resulting neonatal admission in neonatal intensive care units (NICUs) for a short or long time in the first month of life[2].

te of health has a considerable effect on future health and life. Since neonate`s immune system and other organs in preterm neonate are not developed completely, they are at risk of many threats resulting neonatal admission in neonatal intensive care units (NICUs) for a short or long time in the first month of life[2]. Previous studies showed that preterm and late preterm babies had an increased risk ratio of both acute and long term morbidities and such complications affect the length of NICU stay compared with term infants[3,4]. Premature rapture of membranes (PROM) is another major clinical complication that is often related to high rates of neonatal morbidity and mortality[5]. Some studies have also indicated increased rates of morbidity and mortality in late preterm infants of women with gestational hypertension or preeclampsia. More neonatal intensive care unit admissions, hypoglycemia, respiratory distress, and re-hospitalization were seen in these children[6]. Maternal gestational diabetes mellitus (GDM) is another clinical complication associated with increased prenatal morbidity. Neonatal hyperglycaemia-related events, such as hypoglycemia, respiratory distress syndrome (RDS), hyperbilirubinemia, congenital anomaly, large for gestational age (LGA), primary cesarean section, polyhydramnios, preterm delivery, admission to NICU >24 h, are more frequent in mothers with GDM. NICU admission is reported in 29% of GDM and 40% in type 2 diabetes mellitus (DM) pregnancies[7-9]. Longer NICU stay was seen in pregnancies complicated by concurrent existence of hypertension and diabetes[10].

an section, polyhydramnios, preterm delivery, admission to NICU >24 h, are more frequent in mothers with GDM. NICU admission is reported in 29% of GDM and 40% in type 2 diabetes mellitus (DM) pregnancies[7-9]. Longer NICU stay was seen in pregnancies complicated by concurrent existence of hypertension and diabetes[10]. The aim of this study was to identify the influence of pregnancy complications on the period of neonatal length of stay in NICU. The provision of neonatal intensive care unit for complicated newborns is a great burden on the health care system. In the USA the cost of preterm babies care has been estimated about $8 billion annually[11]. Although there have been many studies looking at those factors resulting in newborn’s admission in NICU, but there is little information regarding the effects of maternal complications on duration of neonatal hospitalization. Such investigation would be beneficial for health organizations to access plans and proper strategies to decrease high risk pregnancies and consequently neonatal NICU hospitalization period. As a result, these strategies not only would be effective in preventing poor antenatal outcome, but also decrease the health system cost significantly.

ld be beneficial for health organizations to access plans and proper strategies to decrease high risk pregnancies and consequently neonatal NICU hospitalization period. As a result, these strategies not only would be effective in preventing poor antenatal outcome, but also decrease the health system cost significantly. Subjects and Methods A cross-sectional-descriptive analytical study was carried out in the NICU ward of Akbarabadi Hospital in Tehran, during 6 months in 2011. All data were gathered from neonates and their mothers’ medical records. Of 526 NICU admitted children, 37 newborns were excluded due to transfer from other centers (lack of data). The target population consisted of 489 newborns, admitted in the NICU for at least one day. Neonatal gestational age, sex, newborns’ problems, the length of neonatal stay at NICU (days) were recorded in a check list. At the same time we assessed maternal obstetric medical records and gathered data for mothers' complications (PROM, preeclampsia, urinary tract infection (UTI), GDM, vaginal bleeding, addiction). Finally we evaluated statistically the effects of these complications on neonates' admission period in NICU. The software package SPSS version 19 was used to perform the statistical analysis. The t-test, Chi square, regression and ANOVAs analysis were applied where applicable. The level of significance was considered P<0.05.

y we evaluated statistically the effects of these complications on neonates' admission period in NICU. The software package SPSS version 19 was used to perform the statistical analysis. The t-test, Chi square, regression and ANOVAs analysis were applied where applicable. The level of significance was considered P<0.05. Patients’ data were handled confidentially and as no intervention was performed in our study, we did not ask for patients consent. Ethics approval for the study was obtained from the institutional review board of Tehran University of Medical Sciences. Findings Thirty seven of 526 neonatal medical records were excluded. Of the remaining 489 babies hospitalized at NICU for 1 to 54 days (mean 7.9 days), 281 were males, and 28.42% born preterm. 308 newborns weighed <2500 g at birth and 170 neonates had a birth weight between 2500 and 4000 gr. 80.5% were singletons. Of 489 mothers, 150 had PROM, 4.1% GDM, 1.84% UTI, 13.91% preeclampsia, 12.1% vaginal bleeding, 10.4% needed assisted reproductive techniques (ART), and in 1.6% drug abuse was recorded. Some demographic neonatal and maternal characteristics are shown in Table 1. Among neonates admitted to NICU, 322 cases (65.8%) had RDS, 20 cases (1.4%) showed seizures, 5 (1%) had sepsis and 1 (0.2%) had NEC. Twenty-two percent of admitted neonates died mostly (9%) due to RDS. Table1 Demographic characteristics of admitted neonates and their mothers in NICU Variable Frequency (%) n = 489 Gender male 281 (57.5) female 208 (42.5) Gestational age (week) <37 132 (26.9) >37 357 (73.1)

Of 489 mothers, 150 had PROM, 4.1% GDM, 1.84% UTI, 13.91% preeclampsia, 12.1% vaginal bleeding, 10.4% needed assisted reproductive techniques (ART), and in 1.6% drug abuse was recorded. Some demographic neonatal and maternal characteristics are shown in Table 1. Among neonates admitted to NICU, 322 cases (65.8%) had RDS, 20 cases (1.4%) showed seizures, 5 (1%) had sepsis and 1 (0.2%) had NEC. Twenty-two percent of admitted neonates died mostly (9%) due to RDS. Table1 Demographic characteristics of admitted neonates and their mothers in NICU Variable Frequency (%) n = 489 Gender male 281 (57.5) female 208 (42.5) Gestational age (week) <37 132 (26.9) >37 357 (73.1) Birth weight (gram) <2500 308 (63.0) 2500-4000 170 (34.8) >4000 11 (2.2) Expired newborns in NICU days 108 (22.1) Complications Twin 74 (14.9) Triplet 22 (4.5) PROM 150 (30.7) Gestational diabetes mellitus 20 (4.1) Urinary tract infection 9 (1.8) Preeclampsia 68 (13.9) Drug addiction 8 (1.1) Vaginal bleeding 59 (12.1) Assisted reproductive techniques 51 (10.4) Oligohydramnios 28 (5.7) NICU: neonatal intensive care units; PROM: Premature rapture of membranes

) Triplet 22 (4.5) PROM 150 (30.7) Gestational diabetes mellitus 20 (4.1) Urinary tract infection 9 (1.8) Preeclampsia 68 (13.9) Drug addiction 8 (1.1) Vaginal bleeding 59 (12.1) Assisted reproductive techniques 51 (10.4) Oligohydramnios 28 (5.7) NICU: neonatal intensive care units; PROM: Premature rapture of membranes There was a significant relation between duration of neonatal NICU length of stay and maternal PROM (P=0.001), preeclampsia (P=0.01), UTI (P=0.02), multiple gestation (P=0.03), and oligohydramnios (P=0.003) (Table 2). A positive correlation between neonatal complications and length of stay in NICU (P<0.001) was noticeable. The longest and shortest periods of NICU hospitalization belonged to neonates with prematurity and necrotizing enterocolitis (9.02 vs 5.10 days). Moreover, the highest mean NICU admission period were seen in premature neonates with both RDS and neonatal seizure symptoms (38 days). We also found a positive correlation between numbers of gestation and length of NICU stay (P=0.03). Length of stay at NICU for singletons was 7.53 while in twin and triple gestations it was 9.52 and 9.54 days, respectively. Mortality rate in newborns whose mothers’ pregnancy was complicated by DM, vaginal bleeding and pregnancies following ART were significantly more than others (P=0.02, P=0.001, P=0.04, respectively). Table 2 Comparison of length of stay in NICU of neonates of mothers with and without complications Groups Days in NICU (mean) P value Premature Rapture of Membranes Yes 10.8 0.001 No 6.09 Gestational Diabetes Mellitus Yes 8.45 >0.05 No 7.92

Mortality rate in newborns whose mothers’ pregnancy was complicated by DM, vaginal bleeding and pregnancies following ART were significantly more than others (P=0.02, P=0.001, P=0.04, respectively). Table 2 Comparison of length of stay in NICU of neonates of mothers with and without complications Groups Days in NICU (mean) P value Premature Rapture of Membranes Yes 10.8 0.001 No 6.09 Gestational Diabetes Mellitus Yes 8.45 >0.05 No 7.92 Urinary Tract Infection Yes 5.22 0.02 No 7.99 Preeclampsia Yes 10.82 0.01 No 7.47 Drug addiction Yes 7.62 >0.05 No 7.94 Vaginal bleeding Yes 7.42 >0.05 No 8.01 Assisted Reproductive Techniques Yes 7.87 >0.05 No 8.52 Oligohydramnios Yes 11.78 0.003 No 7.70 Discussion Neonatal mortality and morbidity is common following complicated pregnancies. This study examined whether high risk pregnancies could influence neonatal outcome and length of NICU hospitalization.

Vaginal bleeding Yes 7.42 >0.05 No 8.01 Assisted Reproductive Techniques Yes 7.87 >0.05 No 8.52 Oligohydramnios Yes 11.78 0.003 No 7.70 Discussion Neonatal mortality and morbidity is common following complicated pregnancies. This study examined whether high risk pregnancies could influence neonatal outcome and length of NICU hospitalization. We found that maternal PROM, preeclampsia, oligohydramnios, UTI, and multiple gestations had increased duration of NICU stay. Previous studies have shown that PROM and preterm birth after PROM are associated with small-for-gestational age and low birth weight newborns. In addition the high rate of cesarean sections, chorioamnionits, fetal distress and placental accidents were seen more frequently in these groups[5]. PROM accounts for 25-40% of all preterm deliveries that increase the risk of neonatal morbidity by 75%. In addition, improvement in survival may be associated with adverse long term sequels needing more treatment and NICU hospitalization[11]. There is no doubt that pregnancies complicated with hypertension have higher rates of neonatal morbidity than normotensive pregnancies. Many studies have indicated that mothers with preeclampsia have increased rates of small for gestational age (SGA) infants[3]. Preeclampsia increases the rate of induction at each gestational week and cesarean section, both of them resulting in SGA infants’ birth with higher incidence of complications like RDS. Some forms of IUGR etiologically are linked to preeclampsia and its placental dysfunction. Hypertensive disorders in pregnancy also raise the incidence of NICU admission at 35, 36, and 37 weeks of gestation and longer neonatal stay. The gestational week of delivery rather than severity of complication had greater role on NICU admission and total stay. Even IUGR neonates from preeclamptic mothers were at a higher risk of NICU stay 7 days or more than unexplained IUGR neonates[6,12].

ion at 35, 36, and 37 weeks of gestation and longer neonatal stay. The gestational week of delivery rather than severity of complication had greater role on NICU admission and total stay. Even IUGR neonates from preeclamptic mothers were at a higher risk of NICU stay 7 days or more than unexplained IUGR neonates[6,12]. Oligohydramnios, systemic or regional infection (such as urinary tract infection) and multi fetal pregnancy are predisposing factors for preterm birth. Both lower gestational age and birth weight have adverse roles on morbidity following preterm birth. In developed countries, preterm birth is responsible for 75% of neonatal morbidity including neurodevelop-mental complication, pulmonary disease, and visual problem. Moreover singletons survive better in comparison with twins. The incidence of intraventricular hemorrhage (IVH) and RDS are higher in preterm twins than in singletons[11]. We also found that the multiple gestation affects length of NICU stay. In recent decades multiple births have increased due to increased employing of assisted reproductive techniques. Multiple pregnancies increase the rate of stillbirth, early and late neonatal mortality and morbidity. The main cause is related to prematurity, LBW and intra partum complications. In a tertiary referral center, 90% of triplets with gestational age <32 weeks were admitted in neonatal unit. Cerebral palsy in triplets and twins occurs 47 and 8 times more compared to singletons. Triplets and high multiple gestations stayed longer in neonatal unites (51 days) than twins (40 days)[13,14].

mplications. In a tertiary referral center, 90% of triplets with gestational age <32 weeks were admitted in neonatal unit. Cerebral palsy in triplets and twins occurs 47 and 8 times more compared to singletons. Triplets and high multiple gestations stayed longer in neonatal unites (51 days) than twins (40 days)[13,14]. We found that the highest mean length of NICU stay was seen in neonates with RDS and neonatal seizure, also correlation between neonatal mortality and RDS was noticeable. Our results were consistent with other studies. An investigation has demonstrated higher morbidity rate in preterm neonates at NICUs. Moreover RDS is the main common cause of neonatal mortality. Of infants born at 30-34 weeks, 28% had an acute lung disorder[15]. Late preterm birth (34-36 weeks) with higher risk of RDS accounts for 71.7% NICU admissions in USA[16]. Mortality rate and the risk of RDS are high among 26-29 weeks SGA neonates. The effect of SGA and prematurity may influence events resulting in increasing mortality and neonatal morbidity rates. Fetal hypoxemia, nutrient restriction and altered endocrine milieu could be affecting factors[17]. In addition, prevalence of neonatal seizures is around 1-3 per 1000 live births and more frequent in preterm population. Seizures increase neurological and developmental morbidity. In preterm babies 40% of seizures result from hypoxic-ischemic events. In a cohort study, 28% of neonatal seizure survivors had poor outcome like hypoxic-ischemic encephalopathy, focal infarction, subarachnoid hemorrhage, metabolic disorder, meningitis, or congenital brain malformation. Seizure with long adverse outcome increases neonatal need for prolonged medical treatment and clinical attention[18].

udy, 28% of neonatal seizure survivors had poor outcome like hypoxic-ischemic encephalopathy, focal infarction, subarachnoid hemorrhage, metabolic disorder, meningitis, or congenital brain malformation. Seizure with long adverse outcome increases neonatal need for prolonged medical treatment and clinical attention[18]. Finally our analysis revealed that mortality rate in newborns whose mothers pregnancy was complicated by GDM, vaginal bleeding, and ART was higher than in others. We speculated that all these conditions could be predisposing factors for preterm births, SGA and VLBW infants resulting rise in neonatal mortality rate. Low birth weight was common in our study population. Our results were compatible with other reported studies; neonatal mortality rate was higher in mothers with vaginal bleeding and third trimester complications[19,12]. In an investigation carried out in Nigeria, among 87 neonatal deaths; SGA and LBW infants had greatest portion (21.7% and 20.1% respectively)[20]. Furthermore studies showed that 85% of transferred embryos by ART could not produce a live birth[21]. ART also increases obstetric and perinatal adverse outcomes like perinatal death, chromosomal abnormalities, low birth weight, preterm labor, GDM, placental accidents and preeclampsia[22]. Limitation: In our study we considered only a few obstetrical complications like preeclampsia, vaginal bleeding, GDM, oligohydramnios, and preterm birth. We recommend to survey the effect of other direct and indirect obstetric morbidities in future studies.

Finally our analysis revealed that mortality rate in newborns whose mothers pregnancy was complicated by GDM, vaginal bleeding, and ART was higher than in others. We speculated that all these conditions could be predisposing factors for preterm births, SGA and VLBW infants resulting rise in neonatal mortality rate. Low birth weight was common in our study population. Our results were compatible with other reported studies; neonatal mortality rate was higher in mothers with vaginal bleeding and third trimester complications[19,12]. In an investigation carried out in Nigeria, among 87 neonatal deaths; SGA and LBW infants had greatest portion (21.7% and 20.1% respectively)[20]. Furthermore studies showed that 85% of transferred embryos by ART could not produce a live birth[21]. ART also increases obstetric and perinatal adverse outcomes like perinatal death, chromosomal abnormalities, low birth weight, preterm labor, GDM, placental accidents and preeclampsia[22]. Limitation: In our study we considered only a few obstetrical complications like preeclampsia, vaginal bleeding, GDM, oligohydramnios, and preterm birth. We recommend to survey the effect of other direct and indirect obstetric morbidities in future studies. Conclusion Maternal complication can influence neonatal outcome significantly. By increasing the maternal health level and prenatal care services, we can improve neonatal outcome and decrease length of stay in NICU. Acknowledgment This was taken from the medical student thesis by N. Afrasiabi with ID 4980. Conflict of Interest: None

Conclusion Maternal complication can influence neonatal outcome significantly. By increasing the maternal health level and prenatal care services, we can improve neonatal outcome and decrease length of stay in NICU. Acknowledgment This was taken from the medical student thesis by N. Afrasiabi with ID 4980. Conflict of Interest: None Authors’ Contribution P. Mohagheghi and M. Kalani: Concept, design, acquisition of data N. Afrasiabi and Z. Farahani: Data analysis and interpretation, literature, drafting of the manuscript. Gh. Mohades: Critical revision of the manuscript. All authors approved final version of the manuscript.

Introduction Febrile convulsion (FC) is defined as a type of seizure accompanied by fever which is seen in children aged 6 months to 5 years with no history of electrolyte imbalance and infection in central nervous system[1]. Although seizures are generally benign in character, care and attention and is appropriate, as occasionally seizures may have properties of recurrence and as there is a risk of the seizures becoming epileptic[1]. The pathogenesis of febrile convulsion is not as yet fully understood. Attempts have been made to explain why fever experienced by all children, develops into convulsion in only some of them. Studies have been conducted on children with febrile convulsion related to interferon-alpha[2], prolactin[3], low levels of cortisol[3], anemia[4] and zinc deficiency[5], but the role of all of these in the pathogenesis of FC remains controversial. It has been suggested that oxidative damage in neurological diseases and changes in the antioxidant defense system which result in increased lipid peroxidation may play a role in the pathogenesis of epilepsy and febrile convulsion[6]. Selenium, an important antioxidant for humans, has been studied in childhood epilepsy and particularly in resistant epilepsy and its levels were found to be low compared to those of normal children. Thus, it has been proposed that selenium may become a part of epilepsy treatment in future[7].

and febrile convulsion[6]. Selenium, an important antioxidant for humans, has been studied in childhood epilepsy and particularly in resistant epilepsy and its levels were found to be low compared to those of normal children. Thus, it has been proposed that selenium may become a part of epilepsy treatment in future[7]. Platelets play a fundamental role in thrombosis and homeostasis. However, recent studies have revealed that platelets play an even greater role in infection and inflammation[8]. Activated platelets cause an increase in the size of platelets by expressing inflammatory factors such as chemokines and cytokines. In other words, increased mean platelet volume is an indicator that thrombocytes have been activated[9]. Thrombocyte size measured as mean platelet volume (MPV), thrombocyte aggregation, thromboxane A2, thrombocyte factor 4 and thromboglobulin expression are good indicators of specific activities of thrombocytes[9]. The changes in MPV have been studied in several diseases[10,11]. This study aimed to evaluate the changes of the mean platelet volume values and serum selenium levels which follow a convulsion in patients with simple febrile convulsion.

Platelets play a fundamental role in thrombosis and homeostasis. However, recent studies have revealed that platelets play an even greater role in infection and inflammation[8]. Activated platelets cause an increase in the size of platelets by expressing inflammatory factors such as chemokines and cytokines. In other words, increased mean platelet volume is an indicator that thrombocytes have been activated[9]. Thrombocyte size measured as mean platelet volume (MPV), thrombocyte aggregation, thromboxane A2, thrombocyte factor 4 and thromboglobulin expression are good indicators of specific activities of thrombocytes[9]. The changes in MPV have been studied in several diseases[10,11]. This study aimed to evaluate the changes of the mean platelet volume values and serum selenium levels which follow a convulsion in patients with simple febrile convulsion. Subjects and Methods The study comprised 42 patients aged between 9 months and 60 months who presented at the pediatric clinic and the pediatric emergency clinic between 1 January 2012 and 1 April 2013 having undergone a generalized tonic clonic convulsion lasting less than 15 minutes and not recurring within 24 hours and who were then diagnosed with simple febrile convulsion. The control group comprised 30 healthy children aged 8 months to 60 months who were brought to the general pediatrics clinic for vaccination and/or a routine health check and who had no history of convulsions, epilepsy or neurological impairment. Informed consent was obtained from the parents of all the participants. Approval for the study was granted by the Local Ethics Committee. After taking a detailed anamnesis, the health status of all the children was determined from physical examination.

had no history of convulsions, epilepsy or neurological impairment. Informed consent was obtained from the parents of all the participants. Approval for the study was granted by the Local Ethics Committee. After taking a detailed anamnesis, the health status of all the children was determined from physical examination. Those with metabolic disease, hematologic disorders, chronic disease, a history of asphyxia and afebrile convulsion, neurological sequelae, neurological findings following a convulsion and those with a diagnosis of degenerative central nervous system and demyelinisation disease were excluded from the study. Blood samples were taken 2 hours after the febrile convulsion. For complete blood count an automatic blood count device (Abbott Celldyn 3500 Ill, USA) was employed in all febrile convulsion patients and all healthy control group children. The blood samples were centrifuged at 3500 rpm for 10 minutes, and then the formed elements were discarded with the tube. Part of the serum samples was stored at -80ºC. The remaining serum samples were tested for electrolytes, kidney and liver function (Abbott Aeroset, Abbott Diagnostics, Abbott Park, IL, USA) on the same day and on the study day, serum selenium levels were measured with the hidrure generation method on an atomic absorption spectometry device from the serum samples stored at -80ºC.

serum samples were tested for electrolytes, kidney and liver function (Abbott Aeroset, Abbott Diagnostics, Abbott Park, IL, USA) on the same day and on the study day, serum selenium levels were measured with the hidrure generation method on an atomic absorption spectometry device from the serum samples stored at -80ºC. Data were analyzed using SPSS (Statistical Package for the Social Sciences, version 11.5 for Windows, SPSS® Inc, Chicago, IL). Distribution of parametric variables was assessed with one-sample Kolmogorov–Smirnov test and all parametric variables were found to be normally distributed. The results were presented as mean±standard deviation. Demographic data was performed using chi-square test. Independent samples t test was used. Binary logistic regression analysis was performed to find independent predictors of patients with simple febrile convulsions. Receiver operating curve (ROC) analysis was performed to assess the value of selenium levels to detect the cutoff value of the risk of febrile convulsion.  A two-tailed P value of less than 0.05 was considered statistically significant. Findings The 42 patients with simple febrile convulsion included in the study consisted of 30 (71.4%) males and 12 (28.6%) females with a mean age of 2.6±1.2 years. The healthy control group comprised 16 (53.3%) males and 14 (46.7%) females with a mean age of2.4±1.2 years. No statistically significant difference was found between the two groups in terms of age or gender (P>0.05) (Table 1).

study consisted of 30 (71.4%) males and 12 (28.6%) females with a mean age of 2.6±1.2 years. The healthy control group comprised 16 (53.3%) males and 14 (46.7%) females with a mean age of2.4±1.2 years. No statistically significant difference was found between the two groups in terms of age or gender (P>0.05) (Table 1). Table 1 Selenium, MPV, platelet count and demographic data of the patient group and the control group Variable Patient group (n=42) Mean (SD) Control group (n=30) Mean (SD) P. value Gender, (Male/Female) 30/12 16/14 0.09 Age (years) 2.7 (1.5) 2.5 (1.2) 0.5 Selenyum ( μg/L ) 44.1 (13.4) 53.5 (10.9) 0.002 MPV (/fL) 6.4 (1.1) 5.8 (0.5) 0.002 Platelet (/μL) 353.4 (105.3) 407.2 (69.4) 0.01 Student t test for Independent Samples was used. SD: Standard deviation; MPV: mean platelet volume Both viral and bacterial infection findings of upper respiratory tract (tonsillitis, otitis) existed in all patients. When the mean selenium levels of the febrile convulsion patients were compared with those of the control group, they were found to be statistically significantly lower (P=0.002) (Table 1). While the MPV values were found to be statistically significantly higher compared to those of the control group (P=0.002), total platelets counts were found to be significantly lower (P<0.01) (Table 1).

When the mean selenium levels of the febrile convulsion patients were compared with those of the control group, they were found to be statistically significantly lower (P=0.002) (Table 1). While the MPV values were found to be statistically significantly higher compared to those of the control group (P=0.002), total platelets counts were found to be significantly lower (P<0.01) (Table 1). Binary logistic regression analysis revealed that decreased selenium level was an independent factor (B=-0.063, SE=0.022, Wald=7.99, P=0.005). In ROC-curve analysis, selenium levels below 49.05 mg/ml showed 73.3% sensitivity and 66.7% specificity for the risk of developing simple febrile seizure [area under the curve of 0.290 (95% Confidence Interval: 0.17-0.41)] (Fig. 1). Discussion To the best of our knowledge, the present study is the first in literature to investigate the relationship between Selenium, MPV, platelets and febrile seizures; and gave intriguing results: (i) Se and MPV values were found to be lower, (ii) platelet counts were found to be higher than control group, (iii) decreased selenium levels were independent predictors for simple febrile convulsions, and (iv) analysis of ROC for sensitivity and specificity revealed that the selenium levels below 49.05 mg/ml showed 73.3% sensitivity and 66.7% specificity for the risk of developing simple febrile seizure.

r than control group, (iii) decreased selenium levels were independent predictors for simple febrile convulsions, and (iv) analysis of ROC for sensitivity and specificity revealed that the selenium levels below 49.05 mg/ml showed 73.3% sensitivity and 66.7% specificity for the risk of developing simple febrile seizure. Although febrile convulsion is one of the most frequently occurring forms of convulsion seen in childhood, the pathogenesis is not yet fully understood. In recent years, studies have been conducted on the oxidative damage, changes in antioxidant enzymes and lipid peroxidation in neurological diseases which accompany epilepsy and convulsions. Fig. 1 Graph demonstrating receiver operating curve (ROC) analysis of selenium levels with area under the curve of 0.290 There are data suggesting that the occurrence of active oxygen metabolites and decreased activity of the antioxidative defense mechanisms can increase the risk of seizures[12,13]. It is thought that oxidative stress may play a role as a mechanism in the etiology of neuronal generation which is triggered by convulsion[14].

There are data suggesting that the occurrence of active oxygen metabolites and decreased activity of the antioxidative defense mechanisms can increase the risk of seizures[12,13]. It is thought that oxidative stress may play a role as a mechanism in the etiology of neuronal generation which is triggered by convulsion[14]. In previous studies it has been understood that there exists a cause and effect relationship between oxidative stress, reactive oxygen species production and epileptic seizures. Due to the protective effect against oxidative damage and the life-extending effect on neuronal cells of selenium and selenoproteins, when the serum selenium levels of epileptic and healthy children have been compared, those of children having suffered seizures have been found to be lower[7]. In studies which have compared mean selenium levels of febrile convulsion patients with those of a healthy control group, the mean selenium levels of febrile convulsion patients have been reported to be significantly low[15]. In our study, the mean serum selenium levels of the febrile convulsion patients were found to be significantly lower compared to those of the control group. Therefore, it can be said that free radicals are both the cause and result of epileptic seizures, as the serum selenium level may fall associated with the use of selenium as an antioxidant to neutralize oxidative damage which has been created as a result of oxygen radicals produced in the nervous system[7] just as in mitochondria which occur all over the body from oxidative phosphorylation.

t of epileptic seizures, as the serum selenium level may fall associated with the use of selenium as an antioxidant to neutralize oxidative damage which has been created as a result of oxygen radicals produced in the nervous system[7] just as in mitochondria which occur all over the body from oxidative phosphorylation. In events of tissue oxygenation and nutritional impairment and in cases such as diabetes mellitus[16], myocardial infarct[17], ischemic shock[18], cigarette smoking[19], renal artery stenosis[20] and hypertrophic dilated cardiomyopathy[21], thrombocyte volume has been reported to increase. MPV values have been reported to fall in diseases such as inflammatory bowel disease[22], pneumonia[11], Kawasaki disease [23] and acute phase Familial Mediterranean Fever [10]. In a study by Ozaydin et al MPV values of simple febrile convulsion patients were found to be higher than those of complex febrile convulsion patients[24].

ve been reported to fall in diseases such as inflammatory bowel disease[22], pneumonia[11], Kawasaki disease [23] and acute phase Familial Mediterranean Fever [10]. In a study by Ozaydin et al MPV values of simple febrile convulsion patients were found to be higher than those of complex febrile convulsion patients[24]. In the current study, the MPV values of the febrile convulsion patients were compared with the MPV values of the control group and the MPV values of the febrile convulsion patients were found to be significantly higher than those of the control group. The mean thrombocyte count was significantly lower compared to that of the control group. This may be caused by a reduced number of thrombocytes in circulation resulting from thrombocytes of smaller diameter having been used first. To compensate for the decreased number of platelets, the bone marrow rapidly produces thrombocytes. The mean size of these new thrombocytes is greater and this may be a reason for the increase in MPV. Thrombocyte volume is a parameter for defining thrombocyte function and thrombocytes of greater volume are hemostatically active.

ate for the decreased number of platelets, the bone marrow rapidly produces thrombocytes. The mean size of these new thrombocytes is greater and this may be a reason for the increase in MPV. Thrombocyte volume is a parameter for defining thrombocyte function and thrombocytes of greater volume are hemostatically active. In thrombocytopenic patients with associated thrombocyte consumption or thrombocyte loss, higher MPV values are seen and this increase is thought to arise from thrombocyte consumption [25]. Another reason, is that thrombocytes as immune cells like polymorph nuclear leukocytes, play a significant role in the phagocytosis and chemotaxis of micro-organisms such as viruses, bacteria and parasites and by expression of some inflammatory cytokines, these functions have been shown to be restored[26]. The increased size of platelets may be due to activated platelets expressing inflammatory factors such as chemokines and cytokines[27]. As thrombocyte function and volume are correlated, they contain more granules and in metabolic and enzymatic terms are more active[28], so MPV may be a simple inflammatory marker for inflammation[29,30].

of platelets may be due to activated platelets expressing inflammatory factors such as chemokines and cytokines[27]. As thrombocyte function and volume are correlated, they contain more granules and in metabolic and enzymatic terms are more active[28], so MPV may be a simple inflammatory marker for inflammation[29,30]. Conclusion While serum selenium levels and thrombocyte count were found to be statistically significantly low in FC cases compared to the control group, the MPV values were found to be high. It is thought that there may be a reduction in selenium associated with its consumption as an antioxidant to neutralize the increased oxidative damage in febrile convulsions. It can be said that this drop in selenium level causes the onset of a seizure. In the event of acute inflammation and infection which create an immune response, increased MPV, which is associated with structural changes in thrombocytes, even showing an anti-microbial effect, shows infection activity causing simple febrile convulsions. Conflict of Interest: None Authors’ Contribution Concept / Design: M. Abuhandan; A. Solmaz; S. Geter; C. Kaya; B. Koca Acquisition of Data: M. Abuhandan; S. Geter (A,B, D, G); Cemil Kaya (A, C, D, G); B. Guzel (B, D, G); B. Koca Data Analysis / Interpretation: M. Abuhandan; A. Solmaz; C. Kaya; I. Yetkin Manuscript Preparation: M. Abuhandan; S. Geter; C. Kaya; B. Guzel; I. Yetkin; B. Koca Critical Revision of the Manuscript: M. Abuhandan; A. Solmaz; I. Yetkin; B. Koca All authors approved the final version of the paper.

Introduction Obesity is an important factor threatening the health all over the world. It is known that obesity increases cardiac diseases by increasing tendency to atherosclerosis. Cardiac diseases in the adulthood are originated from childhood period, and atherosclerosis has a long asymptomatic period in children. It has been shown that intra-abdominal adiposity has a correlation with insulin resistance and metabolic syndrome (MS), and it was a risk marker for cardiovascular diseases[1]. In adult studies, it has been shown that epicardial adipose tissue is the marker of visceral adiposity, and it is a risk factor for cardiac diseases[2]. Correlation between body mass index (BMI) and epicardial adipose tissue thickness has been confirmed[3,4]. It is also reported that epicardial adipose tissue thickness was correlated with visceral adiposity in children, but not a marker for MS[5]. In this present study, it was aimed to define whether there was any relationship between measurements of epicardial adipose tissue thickness (easily measured and a non-invasive technique) and MS as well as insulin resistance. Subjects and Methods

It has been shown that intra-abdominal adiposity has a correlation with insulin resistance and metabolic syndrome (MS), and it was a risk marker for cardiovascular diseases[1]. In adult studies, it has been shown that epicardial adipose tissue is the marker of visceral adiposity, and it is a risk factor for cardiac diseases[2]. Correlation between body mass index (BMI) and epicardial adipose tissue thickness has been confirmed[3,4]. It is also reported that epicardial adipose tissue thickness was correlated with visceral adiposity in children, but not a marker for MS[5]. In this present study, it was aimed to define whether there was any relationship between measurements of epicardial adipose tissue thickness (easily measured and a non-invasive technique) and MS as well as insulin resistance. Subjects and Methods Study participants: The study was performed on patients, who applied to outpatient clinics of pediatric endocrinology and pediatric cardiology divisions. According to age and gender, patients with BMI over 95 percentile were accepted as obese[6]. Patients with three or more of parameters like abdominal obesity, hypertension, hyperglycemia, high triglyceride and low high density lipoprotein levels were defined as MS[7]. Auxological data of participants like height, weight, BMI, and waist circumference were recorded. Standard deviation score (SDS) of height, weight, and BMI were calculated[8]. Age and gender matched patients, who applied to outpatient clinic of pediatric cardiology with chest pain or murmur, and in whom pathology was not defined, were taken into the control group. All control cases were healthy. Subjects with systemic diseases, syndromic diseases, and chronic diseases were excluded from the study.

d gender matched patients, who applied to outpatient clinic of pediatric cardiology with chest pain or murmur, and in whom pathology was not defined, were taken into the control group. All control cases were healthy. Subjects with systemic diseases, syndromic diseases, and chronic diseases were excluded from the study. Blood samples: Fasting glucose, fasting insulin, alanine transaminase, plasma triglycerides, total cholesterol, low density lipoprotein, high density lipoprotein, high sensitive C-reactive protein levels of cases were measured in the blood samples. The homeostasis model assessment-estimated insulin resistance (HOMA-IR) index (glucose×insulin/405) was used for the detection of insulin resistance. Cases with HOMA-IR value above 3.16 were accepted as cases with insulin resistance[9]. Measurement of lipid profiles and high sensitive C-reactive protein: Plasma triglycerides, total cholesterol, low density lipoprotein, and high density lipoprotein were measured by an automated chemistry analyzer (Aeroset, Abbott, USA) using Abbott commercial kits. Serum high sensitive C-reactive protein level was measured using an available commercial kit (Roche).

e C-reactive protein: Plasma triglycerides, total cholesterol, low density lipoprotein, and high density lipoprotein were measured by an automated chemistry analyzer (Aeroset, Abbott, USA) using Abbott commercial kits. Serum high sensitive C-reactive protein level was measured using an available commercial kit (Roche). Echocardiographic measurements: Echocardiographic examination was performed using a General Electric Medical Systems, USA Vivid S6 device with 4-MHz phase transducer. Examinations have been made in left lateral position on standard parasternal long axis and apical four chamber views. An electrocardiogram was simultaneously recorded in all subjects. Interventricular septum thickness was measured from the two dimensional targeted M-mode echocardiographic tracings in the parasternal long axis. The left ventricular mass was calculated by the formula (LVM=[0.8×{1.04x (IVSd+LVEDd+ LVPWd)3–LVEDd3}]+0.6)[10]. Epicardial adipose tissue was measured in two-dimensional echocardiography as an echo-free space over the pericardial layers and its thickness was measured on the free wall of the right ventricle, perpendicular to the wall, from parasternal long-axis view at end-diastole for three cardiac cycles (Fig. 1)[11].

0.6)[10]. Epicardial adipose tissue was measured in two-dimensional echocardiography as an echo-free space over the pericardial layers and its thickness was measured on the free wall of the right ventricle, perpendicular to the wall, from parasternal long-axis view at end-diastole for three cardiac cycles (Fig. 1)[11]. Statistical Analysis: Data were analyzed using SPSS (Statistical Package for the Social Sciences, version 11.5 for Windows, SPSS® Inc, Chicago, IL). Distribution of parametric variables was assessed with one-sample Kolmogorov–Smirnov test and all parametric variables were not distributed normally. The results were presented as mean±standard deviation. Continuous variables were compared with independent sample t-test or the Mann-Whitney U test for two groups. Kruskal-Wallis test was used to determine for differences between three groups. A two-tailed P value less than 0.05 was considered statistically significant. Fig. 1 Echocardiographic measurement of epicardial adipose tissue (EAT)(plus signs indicate epicardial area) Findings In the study, 94 cases (50 females and 44 males) were enrolled. Of them, 30 were in obesity with MS (15F and 15M); 33 were in obesity without MS (16F and 17M); and 31 (19F and 12M) were in control groups. Mean ages in obesity with MS, obesity without MS and control groups were 12.97±1.80 years, 12.93±1.79 years, and 13.08±1.78 years, respectively. Table 1 Age, gender and auxological data of cases Obesity with MS (n=30) Obesity without MS (n=33) Controls (n=31) P. value Gender 15F/15M 16F/17M 19F/12M 0.752 Age (years) 12.97(1.80) 12.93 (1.79) 13.08 (1.78) 0.640

Findings In the study, 94 cases (50 females and 44 males) were enrolled. Of them, 30 were in obesity with MS (15F and 15M); 33 were in obesity without MS (16F and 17M); and 31 (19F and 12M) were in control groups. Mean ages in obesity with MS, obesity without MS and control groups were 12.97±1.80 years, 12.93±1.79 years, and 13.08±1.78 years, respectively. Table 1 Age, gender and auxological data of cases Obesity with MS (n=30) Obesity without MS (n=33) Controls (n=31) P. value Gender 15F/15M 16F/17M 19F/12M 0.752 Age (years) 12.97(1.80) 12.93 (1.79) 13.08 (1.78) 0.640 Height (cm) 156.60(9.72) 156.52 (9.47) 155.06 (9.20) 0.324 Height SDS 0.19(0.99) 0.10 (-1.10) -0.80 (0.91) 0.212 Weight (kg) 72.51(18.84) 72.04 (17.28) 44.63 (8.79) <0.001a,b Weight SDS 2.14(0.58) 2.12(0.67) -0.07 (0.91) <0.001a,b Body mass index (%) 29.08(4.83) 28.97 (4.03) 18.67 (4.17) <0.001a,b Body mass index SDS 2.42(0.31) 2.33 (0.33) 0.12 (0.81) <0.001a,b Waist circumference (cm) 91.06(12.04) 90.1 (10.43) 66.5 (8.99) <0.001a,b MS; Metabolic syndrome, SDS; Standard deviation score, P Kruskal Wallis test, a Difference between obesity with MS and control groups, b Difference between obesity without MS and control groups c Difference between obesity groups

Body mass index SDS 2.42(0.31) 2.33 (0.33) 0.12 (0.81) <0.001a,b Waist circumference (cm) 91.06(12.04) 90.1 (10.43) 66.5 (8.99) <0.001a,b MS; Metabolic syndrome, SDS; Standard deviation score, P Kruskal Wallis test, a Difference between obesity with MS and control groups, b Difference between obesity without MS and control groups c Difference between obesity groups There was no difference in mean age, gender, height and height SDS between the groups. Mean weight, weight SDS, BMI, BMI SDS, and waist circumference were markedly higher in obesity groups when compared with controls (P<0.001, Table 1). While there was no difference in mean fasting glucose levels between the groups, fasting glucose and HOMA-IR values were higher in obesity groups when compared with control group. Mean fasting insulin and HOMA-IR value were statistically significantly higher in obesity with MS group when compared with the obesity without MS group (P<0.001). Mean alanine transaminase, high sensitive C-reactive protein level, interventricular septum thickness and left ventricular mass were statistically higher in obesity groups when compared with the controls. Mean epicardial adipose tissue thickness in obesity with MS, obesity without MS, and control groups were 0.64±0.23 cm, 0.60±0.20 cm, and 0.27±0.12 cm, respectively (P<0.001). There was no significant difference in epicardial adipose tissue thickness between obesity groups (Fig 2).

roups when compared with the controls. Mean epicardial adipose tissue thickness in obesity with MS, obesity without MS, and control groups were 0.64±0.23 cm, 0.60±0.20 cm, and 0.27±0.12 cm, respectively (P<0.001). There was no significant difference in epicardial adipose tissue thickness between obesity groups (Fig 2). Laboratory and cardiac evaluation are shown in Table 2. When all cases were evaluated, strong positive correlations were detected between epicardial adipose tissue thickness, and weight, weight SDS, BMI, BMI SDS, waist circumference, interventricular septum thickness, and left ventricular mass (P<0.01, Table 3). No correlation was detected in epicardial adipose tissue thickness and fasting glucose, insulin, alanine transaminase, high sensitive C-reactive protein, interventricular septum thickness, and left ventricular mass between cases obesity with and without MS (P>0.05). Fig. 2 Changes in epicardial adipose tissue thickness according to groups Table 2 Laboratory and echocardiographic data of case groups Variable Obesity with MS (n=30) Obesity without MS (n=33) Controls (n=31) P. value Fasting glucose (mg/dl) 94,58 (12.87) 90.41 (8.10) 89.64 (4.85) 0.151 Fasting insulin (mU/ml) 25.36 (22.22) 10.76 (5.21) 6.68 (3.35) <0.001a,b,c HOMA-IR 6.69 (7.50) 2.53 (1.36) 1.53 (0.75) <0.001a,b,c Alanine transaminase (IU/ml) 37.91 (22.73) 23.83 (11.36) 14.25 (5.88) <0.001a,b Total Cholesterol (mg/dl) 185.42 (40.48) 176.18 (71.04) 150.65 (16.79) 0.048a Low-density lipoprotein (mg/dl) 100.64 (33.33) 109.11 (69.43) 90.25 (18.45) 0.354 High-density lipoprotein (mg/dl) 37.10 (5.52) 45.37 (9.74) 43.58 (9.43) 0.010a,c

HOMA-IR 6.69 (7.50) 2.53 (1.36) 1.53 (0.75) <0.001a,b,c Alanine transaminase (IU/ml) 37.91 (22.73) 23.83 (11.36) 14.25 (5.88) <0.001a,b Total Cholesterol (mg/dl) 185.42 (40.48) 176.18 (71.04) 150.65 (16.79) 0.048a Low-density lipoprotein (mg/dl) 100.64 (33.33) 109.11 (69.43) 90.25 (18.45) 0.354 High-density lipoprotein (mg/dl) 37.10 (5.52) 45.37 (9.74) 43.58 (9.43) 0.010a,c Triglycerids (mg/dl) 204.68 (71.46) 104.80 (41.29) 92.13 (37.86) <0.001a,c High sensitive CRP (mg/l) 0.57 (0.37) 0.54 (0.61) 0.19 (0.26) <0.001a,b Epicardial adipose tissue thickness (cm) 0.64 (0.23) 0.60 (0.20) 0.27 (0.12) <0.001a,b IVS (mm) 9.03 (1.73) 8.84 (1.46) 7.48 (1.12) <0.001a,b Left ventricular mass (g) 135.50 (62.75) 135.87 (43.31) 96.20 (32.01) 0.002a,b MS; Metabolic syndrome, SDS; Standard deviation score, CRP; C-reactive protein, IVS; Diastolic interventricular septum thickness, HOMA-IR: Homeostasis model assessment-estimated insulin resistance P Kruskal Wallis test, a Difference between obesity with MS and control groups, b Difference between obesity without MS and control groups c Difference between obesity groups When cases with obesity were evaluated according to presence of insulin resistance (when HOMA-IR threshold value was defined as 3.16), waist circumference was markedly increased in the group with insulin resistance (P<0.05), whereas no difference was defined in weight SDS, BMI SDS, epicardial adipose tissue thickness, alanine transaminase, high sensitive C-reactive protein, interventricular septum thickness and left ventricular mass (Table 4).

3.16), waist circumference was markedly increased in the group with insulin resistance (P<0.05), whereas no difference was defined in weight SDS, BMI SDS, epicardial adipose tissue thickness, alanine transaminase, high sensitive C-reactive protein, interventricular septum thickness and left ventricular mass (Table 4). Discussion Intra-abdominal and epicardial adipose tissues are originated from brown adipose tissue during embryogenesis, and differentiated into white adipose tissue[12]. Adipose tissue is an active organ. Marked inflammatory response is observed if visceral adipose tissue has been increased. Studies indicated that epicardial adipose tissue thickness had inflammatory activity potential as the other adipose tissue storages, as well as more proinflammatory cytokines like IL-1b, IL-6, and TNF alpha than the subcutaneous adipose tissue[13]. In our study, high sensitive C-reactive protein levels, the marker of inflammatory response, were increased markedly in obesity and MS groups. However, there was no correlation between increased high sensitive C-reactive protein and increased epicardial adipose tissue thickness. Unfavorable impacts of abdominal adiposity on cardiovascular system have not only been observed in adults, but in children and adolescents as well. It has been reported in studies that waist circumference values indicating abdominal adiposity were more significant than BMI values[14]. Table 3 Correlation between epicardial adiposity, left ventricle parameters and auxological data in all cases Variable Epicardial adipose tissue thickness, r IVSD, r LVM, r

Discussion Intra-abdominal and epicardial adipose tissues are originated from brown adipose tissue during embryogenesis, and differentiated into white adipose tissue[12]. Adipose tissue is an active organ. Marked inflammatory response is observed if visceral adipose tissue has been increased. Studies indicated that epicardial adipose tissue thickness had inflammatory activity potential as the other adipose tissue storages, as well as more proinflammatory cytokines like IL-1b, IL-6, and TNF alpha than the subcutaneous adipose tissue[13]. In our study, high sensitive C-reactive protein levels, the marker of inflammatory response, were increased markedly in obesity and MS groups. However, there was no correlation between increased high sensitive C-reactive protein and increased epicardial adipose tissue thickness. Unfavorable impacts of abdominal adiposity on cardiovascular system have not only been observed in adults, but in children and adolescents as well. It has been reported in studies that waist circumference values indicating abdominal adiposity were more significant than BMI values[14]. Table 3 Correlation between epicardial adiposity, left ventricle parameters and auxological data in all cases Variable Epicardial adipose tissue thickness, r IVSD, r LVM, r Weight (kg) 0.504* 0.597* 0.710* Weight SDS 0.593* 0.507* 0.548* Body mass index (%) 0.605* 0.493* 0.507* Body mass index SDS 0.627* 0.465* 0.459* Waist circumference (cm) 0.585* 0.581* 0.594* Epicardial adipose tissue thickness (cm) 0.377* 0.351*

Table 3 Correlation between epicardial adiposity, left ventricle parameters and auxological data in all cases Variable Epicardial adipose tissue thickness, r IVSD, r LVM, r Weight (kg) 0.504* 0.597* 0.710* Weight SDS 0.593* 0.507* 0.548* Body mass index (%) 0.605* 0.493* 0.507* Body mass index SDS 0.627* 0.465* 0.459* Waist circumference (cm) 0.585* 0.581* 0.594* Epicardial adipose tissue thickness (cm) 0.377* 0.351* IVS (mm) 0.802* * P<0.01; IVS; Diastolic interventricular septal thickness, LVM; Left ventricular mass; SDS; Standard deviation score Table 4 Evaluation of all cases with obesity and metabolic syndrome according to presence of insulin resistance Variable IR (+) (n=26) IR (-) (n=37) P. value * Weight SDS 2.01 (0.58) 1.76 (0.64) 0.1 Body mass index SDS 2.04 (0.33) 1.91 (0.31) 0.1 Waist circumference (cm) 94.4 (11.7) 88.3 (10.15) 0.04 Epicardial adipose tissue thickness (cm) 0.61 (0.22) 0.63 (0.21) 0.7 Alanine transaminase (IU/ml) 36.05 (24.71) 24.86 (10.99) 0.08 High sensitive C-reactive protein (mg/l) 0.56 (0.34) 0.56 (0.38) 1 Diastolic interventricular septum thickness (mm) 8.96 (1.66) 8.91 (1.55) 0.9 Left ventricular mass (g) 139.47 (60.74) 133.03 (50.81) 0.6 * P. value Mann Whitney U test; IR; Insulin resistance (Homeostasis model assessment- insulin resistance >3.16), SDS; Standard deviation score

High sensitive C-reactive protein (mg/l) 0.56 (0.34) 0.56 (0.38) 1 Diastolic interventricular septum thickness (mm) 8.96 (1.66) 8.91 (1.55) 0.9 Left ventricular mass (g) 139.47 (60.74) 133.03 (50.81) 0.6 * P. value Mann Whitney U test; IR; Insulin resistance (Homeostasis model assessment- insulin resistance >3.16), SDS; Standard deviation score Similarly, a strong correlation was detected between epicardial adipose tissue thickness and waist circumference[15]. In our study, a strong correlation was detected between epicardial adipose tissue thickness and weight as well as waist circumference. Epicardial adipose tissue, which is a part of visceral adiposity, accumulates all over the heart especially around coronary arteries. Framingham heart study supported that pericardial adiposity increased the risk of coronary artery disease, and it is related to myocardial infarction[16]. It was defined that mean epicardial adipose tissue thickness was higher in patients with unstable ischemic cardiac disease[17]. Moreover, the correlation between epicardial adipose tissue thickness and cardiac arrhythmia was shown[18]. Optimal cut-off for epicardial adipose tissue thickness associated obesity was found 0.36 cm (90% sensitivity and 87% specificity)[19]. Also, epicardial fat cut-off point for insulin resistance was shown 0.41 cm with 90% sensitivity and 61% specificity[4]. In another study, Okyay et al[20] showed a cutoff point of 0.435 cm determined MS with 61.7% sensitivity and 79.2% specificity. We did not study any epicardial fat cut-off point for obesity or MS.

]. Also, epicardial fat cut-off point for insulin resistance was shown 0.41 cm with 90% sensitivity and 61% specificity[4]. In another study, Okyay et al[20] showed a cutoff point of 0.435 cm determined MS with 61.7% sensitivity and 79.2% specificity. We did not study any epicardial fat cut-off point for obesity or MS. MS, which progresses with insulin resistance, is an important factor for atherosclerosis and cardiac disease. Low high density lipoprotein, one of MS criteria, indicates that there is decrease in an important antioxidant system, and a tendency for atherosclerosis[21]. The correlation between epicardial adipose tissue thickness and obesity, and MS was studied especially in adults. In a study, a strong correlation was defined between epicardial adipose tissue thickness, pericoronary adipose tissue thickness and MS, it was reported that they might be markers for MS[22]. Mazur et al[5] evaluated 52 obese children with the mean age of 11.6 years, and they emphasized that there was no difference in epicardial adipose tissue thickness between obese cases with and without MS. We found that there was no difference in epicardial adipose tissue thickness between MS and obesity groups.

MS[22]. Mazur et al[5] evaluated 52 obese children with the mean age of 11.6 years, and they emphasized that there was no difference in epicardial adipose tissue thickness between obese cases with and without MS. We found that there was no difference in epicardial adipose tissue thickness between MS and obesity groups. Manco et al[23] evaluated epicardial adipose tissues of 30 obese children with mean age of 11.2 years by the magnetic resonance imaging technique. They reported that epicardial adipose tissue thickness was statistically significantly increased in cases with insulin resistance (HOMA-IR>2.5) in the group, and this condition caused cardiovascular risk. In our study, HOMA-IR threshold value was accepted as 3.16, which was more significant in children; no correlation was detected with epicardial adipose tissue thickness. Abacı et al[4] reported that there was no correlation between epicardial adipose tissue thickness and insulin resistance. We found no positive correlation between epicardial adipose tissue thickness and fasting glucose, insulin level, and HOMA-IR in obesity groups. Moreover, epicardial adipose tissue thickness was not statistically different in IR group. In another study performed on 25 obese subjects and 24 controls with mean age of 13.0 years, marked correlation was reported between epicardial adipose tissue thickness and triglycerides, uric acid, and alanine transaminase[24]. In our study, no correlation was found between epicardial adipose tissue thickness and alanine transaminase, triglyceride in MS and obesity groups.

ontrols with mean age of 13.0 years, marked correlation was reported between epicardial adipose tissue thickness and triglycerides, uric acid, and alanine transaminase[24]. In our study, no correlation was found between epicardial adipose tissue thickness and alanine transaminase, triglyceride in MS and obesity groups. The correlation between obesity and left cardiac function disorder was defined in adults, and left ventricular diastolic filling defect abnormalities were also shown[25]. In adults, it was shown that increased left ventricular mass related to obesity, left ventricle hypertrophy, and diastolic function disorders were recovered after weight loss[26]. In another study, it was demonstrated that systolic and diastolic functions of both ventricles were decreased in adult obese patients[27]. It was reported that obesity caused cardiac function disorders starting from the childhood, and left ventricle hypertrophy[28,29]. Ozdemir et al[3] found correlation between epicardial adipose tissue thickness and left ventricular mass, left atrium diameter in obese cases. Atabek et al[30] reported that left ventricular mass in MS group was markedly increased than in the group of obese children and adolescents with the mean age of 11.9 years, so it might be a marker for MS. In our study, positive correlation was detected between epicardial adipose tissue thickness and interventricular septum thickness, left ventricular mass in obese cases with and without MS. However, these two parameters were not significant in indicating MS; it did not differ in insulin resistant group.

a marker for MS. In our study, positive correlation was detected between epicardial adipose tissue thickness and interventricular septum thickness, left ventricular mass in obese cases with and without MS. However, these two parameters were not significant in indicating MS; it did not differ in insulin resistant group. Conclusion Epicardial adipose tissue thickness is correlated with BMI, and waist circumference also in children. It is detected that epicardial adipose tissue thickness was not significant in indicating MS and insulin resistance. However, interventricular septum thickness and left ventricular mass, which are markedly increased in obese patients with and without MS, have indicated that obesity affects cardiac functions starting from the childhood. Acknowledgment This study was presented as a poster in 9th Winter Pediatric Meeting in Uludag, 17-20 March, 2013. Conflict of Interest: None Authors’ Contribution Concept / Design: E. Eren, B. Koca Acquisition of Data: E. Eren, B. Koca, M. Ture, B. Guzel Data Analysis / Interpretation: E. Eren, B. Koca, M. Ture, B. Guzel Manuscript Preparation: E. Eren, B. Koca Critical Revision of the Manuscript: E. Eren, B. Koca All authors approved final version of the paper.

Introduction Retinoblastoma, a malignant intraocular tumor, is commonly detected by parents seeing general signs of this disease[1,2]. In spite of being a rare childhood cancer, it is a common primary ocular tumor in children[3,4]. the incidence rate of retinoblastoma is approximately 1 in 15000 and 1 in 16600 live births in the United States and northern Europe respectively [5,6]. This malignancy is more frequent in Latin America, Africa and India[7,8]. During past 80 years, the curability rate has been dramatically increased to more than 90%[9,10]. The five years overall survival rate in retinoblastoma is greater than 90%[11-13]. Early diagnosis shortly after the detection of symptoms can improve survival rate[14]. The delay in clinical diagnosis depends on the medical culture of the country and awareness of the parents[14]. The estimated time interval between the first symptoms and clinical diagnosis can be a landmark of risk factor for altering survival rate in retinoblastoma, as the late diagnosis results in poor prognosis[14,15]. In order to investigate the consequences of delayed diagnosis in treatment of children with retinoblastoma, we conducted this study in MAHAK Pediatric Cancer Treatment and Research Center (MPCTRC), a referral childhood malignancy center in Tehran, Iran.

During past 80 years, the curability rate has been dramatically increased to more than 90%[9,10]. The five years overall survival rate in retinoblastoma is greater than 90%[11-13]. Early diagnosis shortly after the detection of symptoms can improve survival rate[14]. The delay in clinical diagnosis depends on the medical culture of the country and awareness of the parents[14]. The estimated time interval between the first symptoms and clinical diagnosis can be a landmark of risk factor for altering survival rate in retinoblastoma, as the late diagnosis results in poor prognosis[14,15]. In order to investigate the consequences of delayed diagnosis in treatment of children with retinoblastoma, we conducted this study in MAHAK Pediatric Cancer Treatment and Research Center (MPCTRC), a referral childhood malignancy center in Tehran, Iran. Subjects and Methods We undertook a retrospective review of all children with retinoblastoma, who were admitted to MPCTRC from April 2007 to Dec 2011. Only patients with clinically confirmed tumor by pediatric ophthalmologist were included in the study. Information for each patient including sex, age at diagnosis, the clinical manifestations, date of first symptoms, date of diagnosis, date of initiation of therapy, laterality and stage of the tumor was recorded. MPCTRC, being a large and well-known referral childhood malignancy center in the capital, receives children with retinoblastoma referred for chemotherapy from the whole country.

Subjects and Methods We undertook a retrospective review of all children with retinoblastoma, who were admitted to MPCTRC from April 2007 to Dec 2011. Only patients with clinically confirmed tumor by pediatric ophthalmologist were included in the study. Information for each patient including sex, age at diagnosis, the clinical manifestations, date of first symptoms, date of diagnosis, date of initiation of therapy, laterality and stage of the tumor was recorded. MPCTRC, being a large and well-known referral childhood malignancy center in the capital, receives children with retinoblastoma referred for chemotherapy from the whole country. Based on the clinical examinations and according to International Classification of Retinoblastoma (ICRB), groupings were applied as A to E for intraocular retinoblastoma[16,17]. Data was analyzed by SPSS version 19. Comparison of mean values was performed using the non-parametric Mann-Whitney test for unpaired samples, and chi-squared for parametric variables. All P-values were two-sided and considered statistically significant when <0.05.

Based on the clinical examinations and according to International Classification of Retinoblastoma (ICRB), groupings were applied as A to E for intraocular retinoblastoma[16,17]. Data was analyzed by SPSS version 19. Comparison of mean values was performed using the non-parametric Mann-Whitney test for unpaired samples, and chi-squared for parametric variables. All P-values were two-sided and considered statistically significant when <0.05. Findings The inclusion criteria were met by 157 patients (n=91, 58% boys). The mean age was 1.21±0.11 years (median 1.0 years; interquartile range 0-2 years; range: 0 to 6 years). The mean interval between the first detection of symptoms and the clinical diagnosis was 3.4±0.53 months (median 1 month; interquartile range 5.7–88.7 days; range 0-36.5 months). A delay in the diagnosis of less than 5 months was found for 105 (66.9%) children, and 9 (5.7%) patients were diagnosed with retinoblastoma longer than 15 months after the first symptoms were recognized.

the clinical diagnosis was 3.4±0.53 months (median 1 month; interquartile range 5.7–88.7 days; range 0-36.5 months). A delay in the diagnosis of less than 5 months was found for 105 (66.9%) children, and 9 (5.7%) patients were diagnosed with retinoblastoma longer than 15 months after the first symptoms were recognized. In 93 (59.2%) children, the tumor was unilateral. Intraocular tumor was found in 141 (89.8%) patients. Classification of D in children with bilateral tumor formed the largest group (n=40 eyes; 38.5%), followed by E (n=27 eyes; 26%), B (n=16 eyes; 15.4%), C (n=12 eyes; 11.5%) and finally by the A group (n=9 eyes; 8.6%). Children with unilateral tumor showed class D as the largest group (n=37 eyes; 45.1%), followed by E (n=28 eyes; 34.1%), C (n=9 eyes; 11.1%), B (n=7 eyes; 8.5%) and finally by the A group (n=1 eye; 1.2%). Chi-squared test showed the significant relation (P=0.05) between tumor grouping and delayed diagnosis in enrolled patients (median delay time: group A 1.2±0.7 months; group B 0.2±0.1 months; group C 1.5±0.6 months; group D 3.6±0.9 months; group E: 4.8 ±1.06 months).

s; 8.5%) and finally by the A group (n=1 eye; 1.2%). Chi-squared test showed the significant relation (P=0.05) between tumor grouping and delayed diagnosis in enrolled patients (median delay time: group A 1.2±0.7 months; group B 0.2±0.1 months; group C 1.5±0.6 months; group D 3.6±0.9 months; group E: 4.8 ±1.06 months). Categorizing the whole enrolled patients by age into subgroups of 0-2 years, 2-4 years and more than 4 years showed that the diagnostic delay was significantly (P<0.05) the longest in the elder aged group of more than 4 year olds (7.6±3.8 months; median 11.5 days; interquartile range 4.5 days to 18.7 months; range 0-36.5 months) compared with the youngest age group 0-2 years (3.5±0.6 months; median 1.0 month; interquartile range 6.5 to 94 days; range 0-29.3 months ) and the middle-aged group 2-4 years (2.1±0.53 months; median 1.0 month; interquartile range 5 to 65 days; range 0-14.13 months) (Fig. 1). The most frequent symptoms were leukocoria in 106 (67.51%) children with a mean delay in the diagnosis of 3.4±0.58 months (median 1 month; interquartile range 7 to 88 days; range 0-29.3 months) (Fig. 2). Strabismus was the leading symptom for 33 (25.4%) children, in whom the mean diagnostic delay was 2.9±0.7 months (median 31 days; interquartile range 10 to 121 days; range 0-18 months). As shown in Fig 2, presence of poor vision and eye inflammation predicted a late diagnosis of the tumor. Fig. 1 Age groups and delayed diagnosis

The most frequent symptoms were leukocoria in 106 (67.51%) children with a mean delay in the diagnosis of 3.4±0.58 months (median 1 month; interquartile range 7 to 88 days; range 0-29.3 months) (Fig. 2). Strabismus was the leading symptom for 33 (25.4%) children, in whom the mean diagnostic delay was 2.9±0.7 months (median 31 days; interquartile range 10 to 121 days; range 0-18 months). As shown in Fig 2, presence of poor vision and eye inflammation predicted a late diagnosis of the tumor. Fig. 1 Age groups and delayed diagnosis The most frequent symptoms were leukocoria in 106 (67.51%) children with a mean delay in the diagnosis of 3.4±0.58 months (median 1 month; interquartile range 7 to 88 days; range 0-29.3 months) (Fig. 2). Strabismus was the leading symptom for 33 (25.4%) children, in whom the mean diagnostic delay was 2.9±0.7 months (median 31 days; interquartile range 10 to 121 days; range 0-18 months). As shown in Fig 2, presence of poor vision and eye inflammation predicted a late diagnosis of the tumor. Taking the whole study population, Fig. 3 shows the association of the length of diagnostic delay and clinical finding of the tumor. It can be seen that extra ocular location of bilateral retinoblastoma was found in children with a diagnostic delay of 10 to 15 months, while extra ocular location with unilateral retinoblastoma was distinguished with a diagnostic delay of more than 15 months.

ngth of diagnostic delay and clinical finding of the tumor. It can be seen that extra ocular location of bilateral retinoblastoma was found in children with a diagnostic delay of 10 to 15 months, while extra ocular location with unilateral retinoblastoma was distinguished with a diagnostic delay of more than 15 months. Stratifying the whole study population into patients with unilateral retinoblastoma (n=93, 59.2%) and patients with bilateral retinoblastoma (n=64, 40.8%) revealed that age was significantly (P=o.oo1) lower in the subgroup with bilateral retinoblastomas than in the subgroup with unilateral retinoblastomas (0.6±0.12 years; median 0.1 year; interquartile range 0-1 year; range 0-5 years vs 1.6±0.15 years; median 1.5 years; interquartile range 0-3 years; range 0-6 years). The delay in the diagnosis of the tumor did not vary significantly between the subgroup with unilateral retinoblastomas (mean 3.9±0.87 months; median 1 month; interquartile range 7.2-91.7 days; range 0-36.5 months), and the subgroup with bilateral retinoblastoma (mean 2.6±0.62 months; median 27days; interquartile range 1.7-61 days; range 0-24.3 months). Fig. 2 Presenting symptoms of retinoblastoma Fig. 3 The clinical patterns of retinblastoma The delay in the diagnosis of the tumor varied significantly (P=0.006) between the subgroup with intraocular retinoblastomas (mean 2.9±0.52 months; median 1 month; interquartile range 6-61 days; range 0-36.5 months), and the subgroup with extra ocular retinoblastoma (mean 8.7±2.9 months; median 3 months; interquartile range 5 days-14.2 months; range 0-24.3 months).

significantly (P=0.006) between the subgroup with intraocular retinoblastomas (mean 2.9±0.52 months; median 1 month; interquartile range 6-61 days; range 0-36.5 months), and the subgroup with extra ocular retinoblastoma (mean 8.7±2.9 months; median 3 months; interquartile range 5 days-14.2 months; range 0-24.3 months). Between 2007 and 2011, retinoblastoma was one of the most frequent solid tumors admitted to MPCTRC, accounting for 9.6% of pediatric malignancies. Retinoblastoma represents 6.1% of all cancers in children <5 years of age in the United States[6]. In our study, the mean age of patients was 1.21±0.11 years (59.2% unilateral), was consistent with the literature. Approximately 80% of patients are diagnosed when <4 years old[14]; 60% of cases represent as unilateral and the remaining bilateral[1,2]. Retrospective analysis of 157 cases of retinoblastoma between age and laterality pattern at the time of diagnosis revealed that bilateral retinoblastoma presents at an earlier age than unilateral retinoblastoma. This finding supports the Knudson "two-hit" hypothesis of retino-blastoma genetics[16]. The key point in the early diagnosis of retinoblastoma is referring to pediatricians who can detect ocular disorders, which parents do not perceive most times. Distinguishing the signs and symptoms, funduscopic screening and initiation of therapy as soon as possible are of paramount importance[14].

Retrospective analysis of 157 cases of retinoblastoma between age and laterality pattern at the time of diagnosis revealed that bilateral retinoblastoma presents at an earlier age than unilateral retinoblastoma. This finding supports the Knudson "two-hit" hypothesis of retino-blastoma genetics[16]. The key point in the early diagnosis of retinoblastoma is referring to pediatricians who can detect ocular disorders, which parents do not perceive most times. Distinguishing the signs and symptoms, funduscopic screening and initiation of therapy as soon as possible are of paramount importance[14]. Ophthalmic disorders can present as strabismus and leukocoria[14]. Studies conducted in USA, UK, Brazil and China state that leukocoria is the most frequent symptom[3,14]. This was found in 67.51% of cases in our series, followed by strabismus (25.4%) and eye inflammation (7%). Literature reviews demonstrated that 50-60% of cases were diagnosed by leukocoria, 25% by strabismus and 6-10% by inflammation[14]. Bai and coworkers found the most frequent symptoms were leukocoria and poor vision, in addition to presence of strabismus related to late diagnosis of the tumor[3].

eye inflammation (7%). Literature reviews demonstrated that 50-60% of cases were diagnosed by leukocoria, 25% by strabismus and 6-10% by inflammation[14]. Bai and coworkers found the most frequent symptoms were leukocoria and poor vision, in addition to presence of strabismus related to late diagnosis of the tumor[3]. The mean delay in the diagnosis of retinoblastomas in Iranian children treated in the MPCTRC was 3.4±0.53 months. Duration of symptoms for longer than 10 months in enrolled patients was a prognostic factor for poor outcome. Rodriguez-Gallindo by reviewing cases of retinoblastoma at Brazil in 1995 demonstrated that 68% of patients showed a delay in diagnosis shorter than six months[7]. Bai and Rodrigues by different screening showed that delay diagnosis in their cases was 4.1 and 8.3 months respectively in China (2011) and Brazil (2004) [3,14]. In our study, the risk of delayed diagnosis was the same in both groups of patients with strabismus (2.9±0.7 months) and leukocoria (3.4±0.58 months), while Rodrigues et al found that cases with strabismus showed higher risk of delayed diagnosis (8.8 months) than cases with leukocoria (5.6 months) [14]. The diagnostic delay was significantly longer in the age group > 4 years old than other age groups (P<0.05). While Bai et al showed this relation in the age group of 2-4 years.

The mean delay in the diagnosis of retinoblastomas in Iranian children treated in the MPCTRC was 3.4±0.53 months. Duration of symptoms for longer than 10 months in enrolled patients was a prognostic factor for poor outcome. Rodriguez-Gallindo by reviewing cases of retinoblastoma at Brazil in 1995 demonstrated that 68% of patients showed a delay in diagnosis shorter than six months[7]. Bai and Rodrigues by different screening showed that delay diagnosis in their cases was 4.1 and 8.3 months respectively in China (2011) and Brazil (2004) [3,14]. In our study, the risk of delayed diagnosis was the same in both groups of patients with strabismus (2.9±0.7 months) and leukocoria (3.4±0.58 months), while Rodrigues et al found that cases with strabismus showed higher risk of delayed diagnosis (8.8 months) than cases with leukocoria (5.6 months) [14]. The diagnostic delay was significantly longer in the age group > 4 years old than other age groups (P<0.05). While Bai et al showed this relation in the age group of 2-4 years. There was a predominance of patients with intraocular tumor, which shows shorter duration of symptoms compared to those with advanced extra ocular tumors. Rodrigues et al reported improvement of prognosis in patients with intraocular retinoblastomas[14]. This improvement can be due to focusing on early recognition of retinoblastoma by public awareness. In our study, the patients with extra-ocular tumor exhibited signs for over 10 months, and both parents and ophthalmologists failed to make an early diagnosis. Patients with low educational level living in the rural and countryside showed advanced disease. Thus, the influence of educational awareness on parents and health services can lead to overcome delayed diagnoses.

xhibited signs for over 10 months, and both parents and ophthalmologists failed to make an early diagnosis. Patients with low educational level living in the rural and countryside showed advanced disease. Thus, the influence of educational awareness on parents and health services can lead to overcome delayed diagnoses. Conclusion According to these facts, for an infant/child with positive family history of retinoblastoma, early referring to ophthalmologist and pediatric oncologist is recommended. In addition, educational programs should be offered to bring into public awareness symptoms of retinoblastoma such as leukocoria, strabismus and eye inflammation to result in early diagnosis. Early diagnosis of retinoblastoma can be the critical risk factor in managing the patients as delay in diagnosis accounts for highly advanced tumor and poor prognosis. Acknowledgment The authors thank ophthalmologists who cared for enrolled patients. Research department thanks staff of medical archive in MPCTRC. Conflict of Interest: None Authors’ Contribution Concept / Design: M. Mehrvar, M. Alebouyeh, M. Tashvighi Data selection, Clinical examination and treatment of patients: M. Faranoush, A.A. Hedayati Asl, A. Alebouyeh, M. Tashvighi Data collection; R. Zangooei Data analysis; E. Abadi, N. Mehrvar Manuscript Preparation: N. Mehrvar, M.Faranoush Critical Revision of the Manuscript: A. Mehrvar, M. Alebouyeh All authors approved the final version of the paper.

Introduction Cerebral palsy (CP) is defined as a clinical syndrome characterized by a persistent disorder of posture or movement due to a non-progressive disorder of the immature brain[1]. The prevalence of CP is 2 to 2.5 per 1,000 live births[2] and its incidence may be increasing secondary to improved care in neonatal intensive care units and improved survival of low birth-weight infants[3]. Most children with CP will have spasticity as the main motor disorder and it can be classified either according to which body areas is affected: hemiplegia, diplegia, tetraplegia, or the movement disorder type: spastic, athetoid, ataxic and hypotonic cerebral palsy[2,3,5]. Spasticity is a major challenge for rehabilitation of children with cerebral palsy. Spasticity can prevent or hamper function, cause pain, disturb sleep, cause unnecessary complications and present major difficulties for care workers[6]. The paper is based on literature searches in PubMed, ISI Web of Science and Google Scholar using the key phrases «management of spasticity and cerebral palsy», with the emphasis on clinical studies. Our assessments also rest on our own clinical experience and research at Baqiyatallah Hospital. Literature Review: There are epidemiological, clinical and review studies about management of spasticity in children with cerebral palsy. Definitions of Spasticity

The paper is based on literature searches in PubMed, ISI Web of Science and Google Scholar using the key phrases «management of spasticity and cerebral palsy», with the emphasis on clinical studies. Our assessments also rest on our own clinical experience and research at Baqiyatallah Hospital. Literature Review: There are epidemiological, clinical and review studies about management of spasticity in children with cerebral palsy. Definitions of Spasticity Most physicians and therapists working with children with cerebral palsy probably feel that they can recognize spasticity when they see or feel it[1]. Spasticity is defined as a velocity dependent increased resistance to passive muscle stretch, or alternatively as inappropriate involuntary muscle activity associated with upper motor neuron paralysis[7,8]. Spasticity can result in functional problems with daily living activities (ADL) such as gait, feeding, washing, toileting and dressing[9]. Over time, spasticity may also cause problems, such as muscle pain or spasms, trouble moving in bed, difficulty with transfers, poor seating position, impaired ability to stand and walk, dystonic posturing muscle, contracture leading to joint deformity, bony deformation, joint subluxation or dislocation and diminished functional independence. Contractures occur when there is loss of joint motion due to structural changes in the muscles, ligaments and tendons surrounding the joint. Shortening and stiffness of the soft tissues make the joint resistant to stretching and prevent normal movement[4,5,10-12]. However, spasticity is a benefit for children with cerebral palsy. Increased tone may be useful for the child. It helps to keep the legs straight, thereby supporting the child’s weight against gravity. The child with increased tone in trunk extensors may stand and take a few steps. Spasticity may help preserve muscle bulk and bone density (Table 1)[11]. The extent and type of spasticity can fluctuate widely according to position of head and limbs, fatigue, stress and mood of children. One limb may have one pattern of spasticity whilst another may have a different pattern[6].

and take a few steps. Spasticity may help preserve muscle bulk and bone density (Table 1)[11]. The extent and type of spasticity can fluctuate widely according to position of head and limbs, fatigue, stress and mood of children. One limb may have one pattern of spasticity whilst another may have a different pattern[6]. Table 1 Adverse and beneficial effects of Spasticity Effects of spasticity Negative effects Abnormal posture Difficulty in hygiene and dressing Difficulty in movements Difficulty in sitting and transfers Inhibits muscle growth Joint subluxation or dislocation Leads to contractures Masks contraction in the antagonist Muscle Pain Pressure sores Shortening and stiffness of the soft tissues Positive effects Extensor tone in the limbs help standing Preserve bone density Preserve muscle bulk Causes of Spasticity

Difficulty in movements Difficulty in sitting and transfers Inhibits muscle growth Joint subluxation or dislocation Leads to contractures Masks contraction in the antagonist Muscle Pain Pressure sores Shortening and stiffness of the soft tissues Positive effects Extensor tone in the limbs help standing Preserve bone density Preserve muscle bulk Causes of Spasticity Spasticity in children can result from any disease process that affects the upper motor neuron within the central nervous system. Injury to the upper motor neuron decreases cortical input to the descending reticulospinal and corticospinal tracts, which causes weakness, loss of motor control, and reduction in the number of voluntarily active motor units. The reduction of these descending tracts removes the normal inhibition of the reflex arcs within the grey matter of the spinal cord, leading to a hyperactive reflex arc and spasticity[13]. While in certain cases there is no identifiable cause, typical causes include problems in intrauterine development (e.g. exposure to radiation, infection), asphyxia before birth, hypoxia of the brain, birth trauma during labor and delivery, and complications in the prenatal period or during childhood. Infections in the mother, low birth weight (less than 2.0 Kg) is a risk factor for CP. Also, between 40 and 50% of all children who develop CP were born prematurely. Premature infants are vulnerable, in part because their organs are not fully developed, increasing the risk of hypoxic injury to the brain that may manifest as cerebral palsy[14]. Measuring Spasticity

Spasticity in children can result from any disease process that affects the upper motor neuron within the central nervous system. Injury to the upper motor neuron decreases cortical input to the descending reticulospinal and corticospinal tracts, which causes weakness, loss of motor control, and reduction in the number of voluntarily active motor units. The reduction of these descending tracts removes the normal inhibition of the reflex arcs within the grey matter of the spinal cord, leading to a hyperactive reflex arc and spasticity[13]. While in certain cases there is no identifiable cause, typical causes include problems in intrauterine development (e.g. exposure to radiation, infection), asphyxia before birth, hypoxia of the brain, birth trauma during labor and delivery, and complications in the prenatal period or during childhood. Infections in the mother, low birth weight (less than 2.0 Kg) is a risk factor for CP. Also, between 40 and 50% of all children who develop CP were born prematurely. Premature infants are vulnerable, in part because their organs are not fully developed, increasing the risk of hypoxic injury to the brain that may manifest as cerebral palsy[14]. Measuring Spasticity The diagnosis of spasticity in children with CP requires a complete physical examination, with ancillary testing as needed. The physical examination should focus on motor power, muscle tone, active and passive range of motion of joints, sensation, deep tendon reflexes, station (pelvic and leg alignment while standing, if there is a possibility), presence of upper and lower limbs deformity, spinal alignment[13]. Mechanical instruments and electrophysiological techniques can also be used to assess spasticity. Mechanical instruments measuring the resistance of the muscle to passive stretch and electrophysiological measures showing the hyper excitability of the stretch reflex are used only for research purposes[15]. One of most important tests in rehabilitation for physical examination of spasticity is the Ashworth scale (Table 2). Always test the patient while he or she is in a relaxed supine position.

h and electrophysiological measures showing the hyper excitability of the stretch reflex are used only for research purposes[15]. One of most important tests in rehabilitation for physical examination of spasticity is the Ashworth scale (Table 2). Always test the patient while he or she is in a relaxed supine position. Table 2 Ashworth Scale of Muscle Tone Ashworth Scale Degree of Muscle Tone 1 No increase in tone 2 Slight increase in tone, “catch” when limb is moved 3 Marked increase in tone, passive movements difficult 4 Considerable increase in tone, passive movements difficult 5 Affected part is rigid in flexion or extension Passively move the joint rapidly and repeatedly through the available range of motion and grade the resistance using the definitions[8,12,16]. Individual assessment, prefer- ably with the aid of video clips from before and after treatment, may be useful for assessing effectiveness. One important parameter will always be whether the aims of the treatment were fulfilled.

ough the available range of motion and grade the resistance using the definitions[8,12,16]. Individual assessment, prefer- ably with the aid of video clips from before and after treatment, may be useful for assessing effectiveness. One important parameter will always be whether the aims of the treatment were fulfilled. Management of spasticity is a major challenge to treatment team. Various forms of therapy are available to people living with cerebral palsy as well as caregivers and parents caring for someone with this disability. They can all be useful at all stages of this disability and are vital in a CP person's ability to function and live more effectively[17]. There is no standardized approach to spasticity management of cerebral palsy. But adequate assessment of the specific impairments causing disability is necessary for appropriate interventions to be instituted[18]. The treatment strategy depends on the degree of functional failure caused by the spasticity and its location. In general, treatment options for management of spasticity in children with cerebral palsy include oral medications, physical and occupational therapy, splinting and casting, chemodenervation with botulinum toxin or phenol, selective dorsal rhizotomy, intrathecal baclofen, and orthopedic surgery[4-6,8,10,11,17,18]. Oral Medications Oral medications are a systemic, rather than focal, treatment for spasticity in children with cerebral palsy. Oral medications commonly used in children are baclofen, diazepam, clonazepam, dantrolene and tizanidine[19]. Botulinum Toxin

Management of spasticity is a major challenge to treatment team. Various forms of therapy are available to people living with cerebral palsy as well as caregivers and parents caring for someone with this disability. They can all be useful at all stages of this disability and are vital in a CP person's ability to function and live more effectively[17]. There is no standardized approach to spasticity management of cerebral palsy. But adequate assessment of the specific impairments causing disability is necessary for appropriate interventions to be instituted[18]. The treatment strategy depends on the degree of functional failure caused by the spasticity and its location. In general, treatment options for management of spasticity in children with cerebral palsy include oral medications, physical and occupational therapy, splinting and casting, chemodenervation with botulinum toxin or phenol, selective dorsal rhizotomy, intrathecal baclofen, and orthopedic surgery[4-6,8,10,11,17,18]. Oral Medications Oral medications are a systemic, rather than focal, treatment for spasticity in children with cerebral palsy. Oral medications commonly used in children are baclofen, diazepam, clonazepam, dantrolene and tizanidine[19]. Botulinum Toxin Botulinum toxin (BT) injection is now an established first-line treatment for focal spasticity[10,12,20-22]. Botulinum toxin type A produces dose-related weakness of skeletal muscle by impairing the release of acetylcholine at the neuromuscular junction. This partially interrupts muscle contraction making the muscle temporarily weaker[20-22]. Muscles commonly treated with BT include the gastrocnemius-soleus complex, hamstrings[10,12], hip adductors and flexor synergy muscles of the upper extremity[21,22]. Intramuscular injections can be localized by surface landmarks, electromyography stimulation, and/or ultrasound[20,22]. Following injection, muscle relaxation is evident within 48 to 72 hours and persists for a period of 3 to 6 months[23]. Botox injection can help improve a child’s ability to walk or use hands and allow for a better fitting orthotics by reducing spasticity. Therapists can take advantage of the time when an overly powerful muscle is weakened to work on strengthening the muscle on the opposite side of the joint (antagonist). Sometimes, casting of the involved extremity is done after the injection to increase the stretch of the tight muscle[10,12,20-22].

reducing spasticity. Therapists can take advantage of the time when an overly powerful muscle is weakened to work on strengthening the muscle on the opposite side of the joint (antagonist). Sometimes, casting of the involved extremity is done after the injection to increase the stretch of the tight muscle[10,12,20-22]. Intrathecal Baclofen Intrathecal baclofen (ITB) was approved for the treatment of spasticity of cerebral origin in 1996. ITB is a surgically implanted system used to control spasticity by infusing baclofen directly into the spinal canal and around the spinal cord[24]. Baclofen inhibits spasticity by blocking excitatory neurotransmitters in the spinal dorsal horn. ITB maximizes the dose delivered to spinal receptors and minimizes the side effects associated with oral baclofen[25]. Selective dorsal rhizotomy Selective dorsal rhizotomy (SDR) derives from late 19th century procedures for spasticity. SDR is a neurosurgical procedure that involves partial sensory deafferentation at the levels of L1 through S2 nerve rootlets[26]. After a series of tone management with rehabilitation punctuated with botulinum toxin injections, the child would probably be around 4 to 5 years old and SDR can be considered. A suitable candidate for selective dorsal rhizotomy is typified by 1) spasticity is still a problem 2) good strength of lower limbs and trunk muscles 3) able to stand straight with good alignment 4) intellectually good enough for carrying out training[27]. Splinting, Casting and Orthoses

Selective dorsal rhizotomy (SDR) derives from late 19th century procedures for spasticity. SDR is a neurosurgical procedure that involves partial sensory deafferentation at the levels of L1 through S2 nerve rootlets[26]. After a series of tone management with rehabilitation punctuated with botulinum toxin injections, the child would probably be around 4 to 5 years old and SDR can be considered. A suitable candidate for selective dorsal rhizotomy is typified by 1) spasticity is still a problem 2) good strength of lower limbs and trunk muscles 3) able to stand straight with good alignment 4) intellectually good enough for carrying out training[27]. Splinting, Casting and Orthoses Casts, splints, and orthoses are all devices that are designed to keep the body in a certain position. These devices are used to prevent or correct deformities in the spastic limb and/or to help children with cerebral palsy overcome activity limitations, such as difficulties with standing and walking[28,29] and serial casting can improve the range of movement in a joint that is already contracted[6]. Serial casting is an intervention practice that is becoming more commonly used in occupational therapy practice, in addition to other treatment modalities/protocols for children with cerebral palsy to manage spasticity and related contractures[30]. Serial casting is based on the premise that shortened muscles maintain the plasticity for lengthening. Providing a prolonged stretch offers biomechanical benefits and inhibits spasticity. But there is a difference between inhibitive casting and serial casting. in inhibitive casting only a single static cast is used and the purpose is to reduce tone rather than lengthen muscle, thereby improving function[31]. The most common type of orthosis is the ankle-foot orthosis (AFO). AFOs are typically designed to limit unwanted ankle movements, specifically ankle plantar flexion (foot pointed toward the ground) (Fig. 1). AFOs can be fixed (to block ankle movement) or articulating (to allow for some movement at the ankle)[32]. Preventing plantar flexion through the use of AFOs has been found to improve walking efficiency in children with spastic diplegic cerebral palsy[33] and in children with hemiplegic cerebral palsy. When AFO use is compared to barefoot walking, the children's walking patterns are better when wearing AFOs[34]. For children with cerebral palsy who tend to walk on their toes, AFOs have been shown to improve their ability to move from sit to stand. However, children with cerebral palsy who are able to stand on a flat foot did not benefit from AFOs for moving from sit to stand as the AFOs tended to slow them down[35]. AFOs have also been shown to affect how much energy children with cerebral palsy use to walk.

wn to improve their ability to move from sit to stand. However, children with cerebral palsy who are able to stand on a flat foot did not benefit from AFOs for moving from sit to stand as the AFOs tended to slow them down[35]. AFOs have also been shown to affect how much energy children with cerebral palsy use to walk. One study found that children with spastic diplegic cerebral palsy had lower oxygen needs during walking when they wore hinged AFOs[36]. Fig. 1 Ankle-foot orthosis (AFO) Orthopedic Surgery Orthopedic surgery is no option for managing spasticity. Instead, it is used to help correct the secondary problems that occur with growth in the face of spastic muscles and poor motion control. Those problems include muscle shortening, joints contractures and bony deformities[37]. Occupational Therapy and Physical Therapy

Orthopedic surgery is no option for managing spasticity. Instead, it is used to help correct the secondary problems that occur with growth in the face of spastic muscles and poor motion control. Those problems include muscle shortening, joints contractures and bony deformities[37]. Occupational Therapy and Physical Therapy Occupational therapy (OT) and physical therapy (PT) are a fundamental part of spasticity management. Muscle overactivity produces muscle shortening and muscle shortening increases spindle sensitivity. Muscle contracture and stretch sensitive muscle overactivity are intertwined. Therefore rehabilitation and physical treatments aimed at lengthening the overactive muscles are fundamental. Address both shortening and overactivity[38]. There are a number of different dynamic Occupational and Physical therapy approaches, including the Bobath technique[4,5], Sensory integration therapy[5], poprioceptive neuromuscular facilitation[39] and the Brunnstrom technique[40]. Consider applying various techniques such as ice (cold), heat, positioning, stretching exercises and use of orthotic devices for these purposes. Cold inhibits spastic muscles, but the effect is short-lived, perhaps outlasting the application of the cold by about half an hour[41]. Paradoxically, heat is also used for relaxation of a spastic muscle[42]. Position the child to stretch the spastic muscles and decrease the sensitivity of the stretch reflex and the brain stem reflexes that trigger spasticity. Also, the therapists should teach these positions to the family so that the child lies and sits this way most of the time at home[43]. Massage and stretching muscles may prevent contractures and promote muscle growth[44-46]. Spasticity decreases with slow and continuous stretching[47]. This effect lasts from 30 minutes to 2 hours. Use stretching exercises before bracing and serial casting to obtain the necessary joint position[44,46,47]. Also, Orthoses are generally used in conjunction with occupational therapy and physical therapy with the aims of increasing muscle length (through providing a prolonged stretch), breaking up mass patterns of movement and improving biomechanics and stability[48-51]. Muscle relaxation after stretching exercises lasts for a short period of time. For longer duration the stretch on the muscle should be maintained for several hours every day. This is possible with the use of rigid splints or serial casting[17,49,52].

patterns of movement and improving biomechanics and stability[48-51]. Muscle relaxation after stretching exercises lasts for a short period of time. For longer duration the stretch on the muscle should be maintained for several hours every day. This is possible with the use of rigid splints or serial casting[17,49,52]. Conclusion The management of spasticity following a cerebral palsy is complex and is a major challenge to treatment team. Initial management should focus on the elimination of externally exacerbating causes. If the spasticity interferes with function, causes pain, and produces deformity, then clear treatment goals should be established. There is not a standardized approach. The treatment needs to be evidence-based and depends on the degree of functional failure caused by the spasticity and its location. This management often requires a variety of different approaches including oral medications, but botulinum toxin, intrathecal baclofen, occupational and physical therapy and often surgical interventions such as selective dorsal rhizotomy and orthopedic surgery. Conflict of Interest: None

Dear editor, In Iran, according to WHO, the prevalence of G6PD enzyme deficiency is 10-14.9%[1]. An epidemiological study showed in the Fars province (southern Iran) 12% of males and 1.8% of females are G6PD deficient[2]. Initial neutrophil bactericidal activity depends on oxygen free radical production by the NADPH oxidase. In G6PD deficient state, decreasing the production of neutrophil NADPH has been reported. Few studies so far on the G6PD deficiency and increased chance of infection was carried out[3]. Regarding the higher probability of septicemia in neonates with G6PD deficiency, several studies were done[4,5]. Some studies showed it may be due to lower level of the G6PD enzyme activity in white blood cells[3]. In contrast other studies revealed that G6PD activity is higher in premature infants with gestational age of 29-32 weeks than in term neonates and this does not interfere with diagnosis of G6PD deficiency. In a prospective study from April 2007 till April 2009, in teaching hospitals of Shiraz University of Medical Sciences 150996 neonates were screened for G6PD enzyme activity in the first week of life. From these, 660 neonates have been admitted and screened for sepsis based on the clinical features in neonatal intensive care units. Parental written informed consent form was obtained from all patients for inclusion in the study. The Shiraz University of Medical Sciences Ethics Committee approved the study.

k of life. From these, 660 neonates have been admitted and screened for sepsis based on the clinical features in neonatal intensive care units. Parental written informed consent form was obtained from all patients for inclusion in the study. The Shiraz University of Medical Sciences Ethics Committee approved the study. Neonatal sepsis was defined as a clinical syndrome characterized by signs and symptoms of infection with or without accompanying bacteremia in the first month of life. Patients were divided into three groups: 1) Non-sepsis cases based on clinical symptoms and paraclinical data of following days were excluded. 2) Suspected sepsis was diagnosed according to clinical symptoms and laboratory abnormalities including 5000≤WBC≥25000, C- reactive protein (CRP) >6 mg/lit, immature neutrophils/total neutrophils >0.2, and negative blood cultures. 3) Definite sepsis diagnosis was established with positive blood cultures. We enrolled second and third groups in the study. For all neonates in whom sepsis was suspected, screening tests including complete blood count, CRP, and micro erythrocyte sedimentation rate (ESR), blood culture, urine culture, cerebrospinal fluid (CSF) analysis and culture were performed. G6PD activity was measured by qualitative method, fluorescent spot test, using kits (Kymia Pajohan, Iran) with 99% specificity and sensitivity. The prevalence of G6PD deficiency in male and female were 10932 (14.09%) and 8650 (11.77%) Table 1 The prevalence of G6PD deficiency and sepsis among 150996 newborns in southern Iran Total Male Female Newborns 150996 77536 (51.34%) 73460 (48.66%)

For all neonates in whom sepsis was suspected, screening tests including complete blood count, CRP, and micro erythrocyte sedimentation rate (ESR), blood culture, urine culture, cerebrospinal fluid (CSF) analysis and culture were performed. G6PD activity was measured by qualitative method, fluorescent spot test, using kits (Kymia Pajohan, Iran) with 99% specificity and sensitivity. The prevalence of G6PD deficiency in male and female were 10932 (14.09%) and 8650 (11.77%) Table 1 The prevalence of G6PD deficiency and sepsis among 150996 newborns in southern Iran Total Male Female Newborns 150996 77536 (51.34%) 73460 (48.66%) G6PD deficiency 19582 (12.9%) 10932 (14.09%) 8650 (11.77%) Sepsis 110 (0.07%) 68 (0.04%) 42 (0.02%) G6PD deficiency among newborns with sepsis 15 (13.6%) 10 (9%) 5 (4.5%) respectively. The overall incidence of G6PD enzyme deficiency was 12.96%. Of the total neonates 660 neonates suspected to have sepsis were referred to neonatal intensive care units, but finally after exclusion of other causes such as metabolic disorders and hypoxic ischemic encephalopathy, 110 patients (68 males and 42 females) were admitted and screened with impression of sepsis. The G6PD enzyme deficiency was proved in 15 (13.6%) of them. There was no statistically significant difference between male and female gender in this group (P=0.78). Table 1 shows the pre-valence of G6PD deficiency and sepsis among 150996 newborns in southern Iran. Few studies on the G6PD deficiency and increased chance of infection were so far carried out. Several reasons for this relationship are considered.

ficant difference between male and female gender in this group (P=0.78). Table 1 shows the pre-valence of G6PD deficiency and sepsis among 150996 newborns in southern Iran. Few studies on the G6PD deficiency and increased chance of infection were so far carried out. Several reasons for this relationship are considered. Our study showed that G6PD deficiency does not increase the chance of neonatal septicemia. Because of different variety of G6PD deficiency in different populations more studies by larger study groups is recommended for evaluation of relationship between G6PD deficiency and bacterial infection and sepsis.

Introduction Periodic fever syndromes are a group of diseases characterized by episodes of fever with healthy intervals between febrile episodes. These disorders are self-limited and improve spontaneously without specific therapy. Episodes of fever are usually associated with presence of inflammation in different parts of the body such as peritoneum, pleural spaces, testis, etc. with elevated serum acute phase reactants level. These diseases should be differentiated from infections, malignancies and other autoimmune disorders[1]. Existence of family history of periodic fever and prolonged illnesses are diagnostic clues of these disorders. At least eight hereditary periodic fever syndromes have been known so far which are common in fever with cutaneous manifestations, and musculoskeletal involvement[2], each of them having specific characteristics. Fever and symptoms occur due to the increment of inflammation which is caused by mutation in genes. Most of these disorders result from gene mutations that cause defect in production of some proteins which have important role in control of inflammation and apoptosis. Usually there are symptoms of the disease for a long time prior to the definite diagnosis. Mostly fever attacks start during childhood, but sometimes they may start in adolescence or in adulthood. Clinical examinations are helpful for diagnosis which is confirmed by specific genetic testing[3].

At least eight hereditary periodic fever syndromes have been known so far which are common in fever with cutaneous manifestations, and musculoskeletal involvement[2], each of them having specific characteristics. Fever and symptoms occur due to the increment of inflammation which is caused by mutation in genes. Most of these disorders result from gene mutations that cause defect in production of some proteins which have important role in control of inflammation and apoptosis. Usually there are symptoms of the disease for a long time prior to the definite diagnosis. Mostly fever attacks start during childhood, but sometimes they may start in adolescence or in adulthood. Clinical examinations are helpful for diagnosis which is confirmed by specific genetic testing[3]. After a brief review of pathogenesis, we present a new approach to the periodic fever in Iranian patients based on epidemiology of infectious and auto-inflammatory disorders in Iran. In addition, we will explain the most important clinical and laboratory data of different types of periodic fever syndromes based on our experience. Definition and Clinical Manifestations

After a brief review of pathogenesis, we present a new approach to the periodic fever in Iranian patients based on epidemiology of infectious and auto-inflammatory disorders in Iran. In addition, we will explain the most important clinical and laboratory data of different types of periodic fever syndromes based on our experience. Definition and Clinical Manifestations Periodic fever disorders are a group of recurrent and episodic disorders with fever as a main complaint. Arthritis or arthralgia, abdominal or chest pain due to serositis, skin rash, and oral ulcers are other common presentations of periodic fever syndromes, although these symptoms can be found in infectious diseases with prolonged or recurrent fever[4,5]. Headache, seizure, aseptic meningitis, development delay as well as adenopathy, organomegaly and cardiac involvement are uncommon findings in these disorders. Renal involvement is uncommon but in some types, renal failure due to amyloidosis may occur. Periodic fever is named when there are 3 episodes of fever in at least 3-6 months with 7 days asymptomatic period between each episode[5]. Rarely, periodic fever syndrome without fever has been reported[6,7]. In this situation, one of the other symptoms is repeated periodically. For this reason, we believe that periodic fever should be considered as a diagnosis in a patient, who shows similar problem repeatedly. Some periodic fever syndromes occur in regular intervals especially periodic fever, aphthous stomatitis, pharyngitis, cervical adenitis (PFAPA) and cyclic neutropenia, however, Familial Mediterranean Fever (FMF) and hyperimmunoglobulin D (hyper IgD) generally have regular patterns. On the other hand, auto-inflammatory syndromes in infancy such as chronic infantile neurologic cutaneous and articular (CINCA) syndrome, Muckle-Wells syndrome, familial cold urticaria and tumor necrosis factor receptor–associated periodic syndrome (TRAPS) usually have irregular patterns[8,9].

lly have regular patterns. On the other hand, auto-inflammatory syndromes in infancy such as chronic infantile neurologic cutaneous and articular (CINCA) syndrome, Muckle-Wells syndrome, familial cold urticaria and tumor necrosis factor receptor–associated periodic syndrome (TRAPS) usually have irregular patterns[8,9]. Pathogenesis Periodic fever syndrome disorders are categorized under the term monogenic auto-inflammatory syndromes. The characteristic of these disorders is episodic attacks of systemic inflammations without presence of infection or auto-antibodies[2,10,11]. The pathogenesis of these disorders is disregulation of inflammation control due to mutations of genes coding for some proteins with regulation role[11]. It has been believed that dysregulation of innate immune response and abnormalities in activity of interleukin1 and pro-inflammatory cytokines may contribute to fever production and systemic inflammation[2,12]. Innate immune cells such as macrophages, neutrophils, and monocytes are involved[12]. Unlike adaptive immunity, innate immunity is programmed genetically[13] and most of these diseases are caused by mutation in genes which make proteins participant in inflammatory response[14]. It is thought that mutate proteins lead to increased or prolonged secretion of pro-inflammatory cytokines[15]. Activation of caspase-1 and the release of IL-1β are the end point of pathophysiologic cycle of these disorders[16]. Treatment with anti-IL1 drugs is mandatory[15]. Approach to Periodic Fever

Periodic fever syndrome disorders are categorized under the term monogenic auto-inflammatory syndromes. The characteristic of these disorders is episodic attacks of systemic inflammations without presence of infection or auto-antibodies[2,10,11]. The pathogenesis of these disorders is disregulation of inflammation control due to mutations of genes coding for some proteins with regulation role[11]. It has been believed that dysregulation of innate immune response and abnormalities in activity of interleukin1 and pro-inflammatory cytokines may contribute to fever production and systemic inflammation[2,12]. Innate immune cells such as macrophages, neutrophils, and monocytes are involved[12]. Unlike adaptive immunity, innate immunity is programmed genetically[13] and most of these diseases are caused by mutation in genes which make proteins participant in inflammatory response[14]. It is thought that mutate proteins lead to increased or prolonged secretion of pro-inflammatory cytokines[15]. Activation of caspase-1 and the release of IL-1β are the end point of pathophysiologic cycle of these disorders[16]. Treatment with anti-IL1 drugs is mandatory[15]. Approach to Periodic Fever In a patient with periodic fever, acquired infection with chronic and periodic nature should be ruled out. It depends on epidemiology of infectious diseases. In Iran and other Middle East countries, brucellosis, Malaria, infectious mononucleosis and borelliosis should be considered in differential diagnosis of periodic fever disorders especially with fever and arthritic manifestation[17-19]. Malaria and borrelliosis can be ruled out by evaluation of peripheral blood smear and brucellosis and Lyme disease can be excluded by serological studies or blood culture. In children, urinary tract infection may present as a periodic disorder, so urine analysis and culture are mandatory in a child with periodic symptoms[4]. Some malignancies such as leukemia and tumoral lesions should be excluded in patients with periodic syndromes and weight loss in any age and weight gain disorder or malaise in children[9,20]. For this reason, abdominal ultrasound and chest X-ray are necessary and bone marrow aspiration in selected patients is recommended. Table 1 shows paraclinical evaluation in suspected patients with periodic fever syndromes.

iodic syndromes and weight loss in any age and weight gain disorder or malaise in children[9,20]. For this reason, abdominal ultrasound and chest X-ray are necessary and bone marrow aspiration in selected patients is recommended. Table 1 shows paraclinical evaluation in suspected patients with periodic fever syndromes. After ruling out infection and malignancies, auto-inflammatory disorders should be considered. However, physician needs to reevaluate each patient with periodic disorder for non-inflammatory syndromes. In Iran after excluding infections, malignancy and cyclic neutropenia, FMF and PFAPA are the most common periodic fever disorders. Similar to other countries, hyper IgD, CINCA syndrome, TRAPS and other auto-inflammatory syndromes are rare causes of periodic fever disorders in our system registry of periodic fever in Iran. Fig. 1 and Fig. 2 show clinical approach to periodic fever in Iranian patients in Periodic Fever Clinic in Children’s Medical Center, based on the epidemiology of periodic fever in this country. Familial Mediterranean Fever (FMF) FMF or recurrent hereditary polyserositis is the most common disease among hereditary periodic fever syndromes, and it is an autosomal recessive disorder[21]. Algorithm 1 First step to approach to Periodic Fever CBC: Complete Blood Cells; ESR: Erythrocyte Sedimentation Rate; CRP: C-Reactive Protein Algorithm 2 Second step to approach to Periodic Fever * It can be as a coincidence of FMF and other periodic fever disorders (such as PFAPA)

FMF or recurrent hereditary polyserositis is the most common disease among hereditary periodic fever syndromes, and it is an autosomal recessive disorder[21]. Algorithm 1 First step to approach to Periodic Fever CBC: Complete Blood Cells; ESR: Erythrocyte Sedimentation Rate; CRP: C-Reactive Protein Algorithm 2 Second step to approach to Periodic Fever * It can be as a coincidence of FMF and other periodic fever disorders (such as PFAPA) FMF: Familial Mediterranean Fever ; PFAPA: Periodic Fever, Aphthus stomatitis, Pharyngitis, Cervical Adenitis; U/A: Urinaly sis; BUN; Blood Urea Nitrogen; Cr: Creatinine; ESR: Erythrocyte Sedimentation Rate; CRP: C - Reactive Protein Algorithm 2 (continue) Second step to approach to Periodic Fever IgD: Hyperimmunoglobulin D; TRAPS: Tumor Necrosis Factor Receptor–Associated Periodic Syndrome; CINCA: Chronic Infantile Neurologic Cutaneous a nd Articular Syndrome Table 1 Laboratory tests for patients with periodic fever Study Indication Time CBC, ESR, CRP All patients At least 2 times in febrile and afebrile period Chest X-ray All patients In febrile period Abdominal ultrasound All patients In febrile period Serum Igs All patients No difference Wright, Coombs-Wright In endemic region At least once in febrile period EBV serology All patients with sore throat and lymphadenopathy In febrile period Peripheral blood smear In endemic region At least once in febrile period Blood culture All patients At least once in febrile period Throat culture All patients with sore throat In febrile period Urine analysis and culture All children At least once in febrile period

EBV serology All patients with sore throat and lymphadenopathy In febrile period Peripheral blood smear In endemic region At least once in febrile period Blood culture All patients At least once in febrile period Throat culture All patients with sore throat In febrile period Urine analysis and culture All children At least once in febrile period Stool exam All children At least once, no difference Bone marrow aspiration In selected patients with cytopenia, bone pain, weight loss, abdominal and/or mediastinal adenopathy No difference Liver function tests All patients At least once in febrile period FANA All children At least once, no difference *CBC: Complete Blood Cells; ESR: Erythrocyte Sedimentation Rate; CRP: C-Reactive Protein; Ig: Immunoglobulin; EBV: Epstein Barr Virus; FANA: Fluorescent Antinuclear Antibody Test Epidemiology: This disease is seen most frequently in people from Mediterranean area such as Jews, Arabs, Turks and Armenians, but it is sometimes reported from all over the world. Prevalence of FMF is 1:200 to 1:1000[8,22]. Although FMF is a autosomal receive disorders, positive family history is present in less than 50% of patients[9,23,24]. This disease is slightly more common in women perhaps due to the influence of sex hormones. Sometimes fever attacks disappear in pregnancy and return after delivery. Rarely infertility may occur due to defect in ovulation and adhesion in pelvic peritoneum. Some complications of FMF such as renal amyloidosis are more common in males[21].

ightly more common in women perhaps due to the influence of sex hormones. Sometimes fever attacks disappear in pregnancy and return after delivery. Rarely infertility may occur due to defect in ovulation and adhesion in pelvic peritoneum. Some complications of FMF such as renal amyloidosis are more common in males[21]. Pathophysiology: The disease is characterized by periodic fever that occasionally occurs at regular intervals, usually with neutrophil induced serositis. The responsible gene for the disease is MEFV that is located on short arm of chromosome 16[2]. This gene produces a protein called Pyrin or Marenostrin that is found in neutrophils and plays an important role in reduction of inflammation[25]. Mutation in this gene leads to defection in production of these proteins and onset of inflammatory cycle. In addition to gene mutation, environmental factors are important and in many patients there is a trigger factor such as emotional stress, infection, extreme physical exercise, fatigue, trauma and menses[12,21,26,27]. Clinical Manifestations: Familial Mediterranean fever usually begins in early childhood and 60-90% of patients are younger than 20 years[12,21]. The mean age of patients at onset of symptoms is 2.5 years. Mean age of patients at the time of diagnosis is 4 years. Very rarely it may occur after 30 years of age[28].

Pathophysiology: The disease is characterized by periodic fever that occasionally occurs at regular intervals, usually with neutrophil induced serositis. The responsible gene for the disease is MEFV that is located on short arm of chromosome 16[2]. This gene produces a protein called Pyrin or Marenostrin that is found in neutrophils and plays an important role in reduction of inflammation[25]. Mutation in this gene leads to defection in production of these proteins and onset of inflammatory cycle. In addition to gene mutation, environmental factors are important and in many patients there is a trigger factor such as emotional stress, infection, extreme physical exercise, fatigue, trauma and menses[12,21,26,27]. Clinical Manifestations: Familial Mediterranean fever usually begins in early childhood and 60-90% of patients are younger than 20 years[12,21]. The mean age of patients at onset of symptoms is 2.5 years. Mean age of patients at the time of diagnosis is 4 years. Very rarely it may occur after 30 years of age[28]. Classical symptoms of FMF are periods of fever and abdominal pain. Frequency of these periods is variable from once in a week to several months. Fever is the most common symptom and it is seen in almost all cases[8,12,23]. The fever is usually short-term and usually takes 1 to 3 days and it resolves without treatment[8,12]. Recurrent oral aphthae may occur without correlation with fever attacks. Fever may initially be the only symptom for many years without another sign in children[8,12,25].

m and it is seen in almost all cases[8,12,23]. The fever is usually short-term and usually takes 1 to 3 days and it resolves without treatment[8,12]. Recurrent oral aphthae may occur without correlation with fever attacks. Fever may initially be the only symptom for many years without another sign in children[8,12,25]. Abdominal pain is the second most common symptom that is seen in more than 80% of patients[9,23,29]. It is usually generalized and has an acute onset, is usually associated with vomiting and diarrhea and sometimes with guarding and rebound tenderness that mimicks acute appendicitis but prophylactic appendectomy for prevention of misdiagnosis is not recommended[8,21,25,30]. Sometimes this pain leads to laparotomy before the definite diagnosis[30]. Rarely a condition called chronic abdominal disease results from recurrent inflammation leading to peritoneal adhesion and subclinical inflammation between fever attacks[21]. Serositis in different cavities like abdomen (peritonitis), chest (pleuritis and pericarditis), joints (arthritis and sinovitis) is another clinical presentation of FMF[9,12,31]. Another common symptom is febrile chest attack that occurs in about 30% and it characterized with painful breathing[8,21,23,29]. It is usually unilateral and increases with inspiration. On physical examination decrease of breath sounds may be present and in chest radiography minimal pleural effusion or pleural thickening may be observed[25]. Patients with homozygous mutation of M694V experience more episodes of pleural attacks[21].

[8,21,23,29]. It is usually unilateral and increases with inspiration. On physical examination decrease of breath sounds may be present and in chest radiography minimal pleural effusion or pleural thickening may be observed[25]. Patients with homozygous mutation of M694V experience more episodes of pleural attacks[21]. Arthritis is seen in 70% of patients and is usually monoarthritis, nondestructive and occurs in large joints of lower extremities such as ankle, knee and hip[25,32]. The involved joint is red, tender and swollen resembling septic arthritis. Synovial fluid is inflammatory in which neutrophil count is elevated but microbial culture is negative[32]. Arthritis disappears with nonsteroidal anti-inflammatory drugs (NSAIDs). The period of this arthritis is longer than other manifestations of FMF and rarely takes more than one month. Arthritis in adults may be destructive and lead to joint replacement[25]. Muscular pain is observed in 10% of patients with FMF, and is one of the debilitating manifestations that sometimes last several weeks in patients treated with NSAIDs, and may be associated with abdominal pain without peritonitis[21]. Acute phase reactants and sedimentation rate are higher in myalgia than in other manifestations of FMF. Muscular enzymes and muscle biopsy are normal[2]. Myalgia responds well to treatment with corticosteroids, but not to colchicine. It must be differentiated from colchicine-induced myopathy[8,21].

out peritonitis[21]. Acute phase reactants and sedimentation rate are higher in myalgia than in other manifestations of FMF. Muscular enzymes and muscle biopsy are normal[2]. Myalgia responds well to treatment with corticosteroids, but not to colchicine. It must be differentiated from colchicine-induced myopathy[8,21]. Erysipelas like erythema (ELE) is found in less than 30% of patients[23,27,29,33,34]. This rash is sometimes associated with arthritis and is usually observed in lower extremities, the skin of which is red, warm and tender, sometimes being difficult to differentiate from cellulitis. Pathologic feature of these lesions is inflammation in superficial dermis and perivascular infiltration without vasculitis. Direct immunofluorescence shows C3 deposits in the wall of small vessels[8]. Incidence of Henoch-Schoenlein purpura (HSP) and polyarteritis nodosa (PAN) are more prevalent in patients with FMF and the age of onset is younger than in other people[8,14]. Renal amyloidosis is the worst complication of FMF that is seen in untreated patients. Persistent inflammation causes amyloid deposition on kidneys and can lead to nephrotic syndrome and renal impairment[14]. This condition progresses in several years. We discuss about risk factors of amyloidosis later in prognosis section. Scarcely, renal insufficiency may be the initial presentation of the FMF (phenotype II)[35]. Sometimes transient microscopic hematuria is one of the findings[8]. Treatment with colchicine may prevent progression to amyloidosis[36,37].

veral years. We discuss about risk factors of amyloidosis later in prognosis section. Scarcely, renal insufficiency may be the initial presentation of the FMF (phenotype II)[35]. Sometimes transient microscopic hematuria is one of the findings[8]. Treatment with colchicine may prevent progression to amyloidosis[36,37]. Other less common manifestations of FMF are acute scrotum (involvement of tunica vaginalis), acute pericarditis, thyroiditis, meningitis and splenomegaly[8,9,27,38,39]. Laboratory findings: There are no specific laboratory findings during and between febrile attacks in FMF. Leukocytosis, ESR, C-reactive protein and fibrinogen are usually elevated during febrile episodes[4,21,23,31]. Serum amyloid A (SAA) is an acute-phase reactant and a good marker for diagnosis of FMF between febrile attacks or in patients without febrile attacks, although the specificity of SAA is low[40]. In addition, SAA can be used in amyloidosis suspicion. The risk for amyloidosis increases when SAA is elevated[40]. It also can be used for adjustment of colchicine dose[40]. There is a high correlation between SAA and CRP level. So when SAA is not available, CRP can be used for amyloidosis between febrile attacks[40].

In addition, SAA can be used in amyloidosis suspicion. The risk for amyloidosis increases when SAA is elevated[40]. It also can be used for adjustment of colchicine dose[40]. There is a high correlation between SAA and CRP level. So when SAA is not available, CRP can be used for amyloidosis between febrile attacks[40]. Diagnosis and differential diagnosis: Diagnosis of FMF is based on clinical manifestation and exclusion of other diseases, but definite diagnosis needs genetically confirmation and finding of mutation in MEFV gene. Irritable bowel syndrome, recurrent infections, functional abdominal pain, and other periodic fever including: hyper-IgD immunoglobulinemia (HIDS), familial Hibernian fever and Marshall's syndrome should be excluded before diagnosis of FMF[8,26]. For many years diagnosis of FMF was based on clinical criteria and exclusion of other differential diseases. However as we mentioned previously, nowadays confirmation of FMF is based on genetic study and mutation in MEFV gene.

Diagnosis and differential diagnosis: Diagnosis of FMF is based on clinical manifestation and exclusion of other diseases, but definite diagnosis needs genetically confirmation and finding of mutation in MEFV gene. Irritable bowel syndrome, recurrent infections, functional abdominal pain, and other periodic fever including: hyper-IgD immunoglobulinemia (HIDS), familial Hibernian fever and Marshall's syndrome should be excluded before diagnosis of FMF[8,26]. For many years diagnosis of FMF was based on clinical criteria and exclusion of other differential diseases. However as we mentioned previously, nowadays confirmation of FMF is based on genetic study and mutation in MEFV gene. There are some clinical criteria to suggest the diagnosis of FMF. Tel Hashomor criteria are most often applied in adults[41] but its specificity in children is low[23,29]. The criteria are classified into two groups of major and minor criteria. Major criteria include: (1) recurrent fever plus serositis, (2) secondary amyloid A amyloidosis, and (3) response to treatment with colchicine. Minor criteria include: (1) recurrent fever, (2) erysipelas-like erythema (ELE) and (3) familial history of FMF. The presence of two major criteria or one major criterion plus two minor ones leads to definite diagnosis[29,41]. On the other hand, presence of a major criterion plus a minor criterion suggests probable diagnosis[21].

ria include: (1) recurrent fever, (2) erysipelas-like erythema (ELE) and (3) familial history of FMF. The presence of two major criteria or one major criterion plus two minor ones leads to definite diagnosis[29,41]. On the other hand, presence of a major criterion plus a minor criterion suggests probable diagnosis[21]. The clinical criteria which are introduced by Yalcinkaya et al are as follows: fever (axillary temperature >38oC , 6-72 hours of duration, ≥3 attacks) abdominal pain (6-72 hours of duration, ≥3 attacks), Chest pain (6-72 hours of duration, ≥3 attacks), arthritis (6-72 hours of duration, ≥3 attacks, oligoarthritis), and family history of FMF[23]. Genetic study provides the facility to confirm FMF in difficult cases with atypical signs, late onset disease and negative family history of FMF and wider indications for molecular approach may result in more frequent diagnosis of FMF[42]. Nowadays, more than 80 mutations have been reported in FMF patients, the majority being located in exon 10[26,43,44]. Five common mutations are M694V, M680I, V726A, M694I, and E148Q that account for 85% of patient suffering from FMF[4,24,26,33,34]. Onset of the disease in patients with homozygous mutation occurs at earlier age[29]. M694V is the most popular mutation and patients with this mutation are more likely to suffer from the severe form of FMF and higher prevalence of arthritis and progress to amyloidosis[44].

of patient suffering from FMF[4,24,26,33,34]. Onset of the disease in patients with homozygous mutation occurs at earlier age[29]. M694V is the most popular mutation and patients with this mutation are more likely to suffer from the severe form of FMF and higher prevalence of arthritis and progress to amyloidosis[44]. Treatment: The drug of choice for FMF is colchicine that is also used for prevention. It is recommended to use this drug for a long time. A minority group of patients do not respond to colchicine may be due to non-adherence[21,27]. Starting dose is 1mg/day and it may be increased to 1.5 to 2 mg/day until remission is achieved[8]. Based on weight or body surface area, the colchicine dose is 0.03±0.02 mg/kg/day and 1.16±0.45 mg/m2/day, respectively[8]. In children younger than 5 years, higher doses of colchicine, 0.07 mg/kg/day or 1.9 mg/m2/day, may be required[8]. The most common side effects of colchicine are diarrhea and nausea which are seen rarely. Fetal malformation from colchicine has not been reported[8]. In colchicine resistance cases interferon-alpha may be helpful[26,28]. In patients with incomplete remission with colchicine, or in whom amyloid A level is high despite treatment with colchicine, IL1 blockade (Anakinra) may be effective[12,16,31]. Other drugs such as NSAIDs and corticosteroids have limited indication for treatment of FMF. Myalgia responds to treatment with corticosteroids, but not colchicines. NSAIDs are used for treatment of arthralgia[21]. Febrile attacks do not respond to and are not preventable by NSAIDs or steroids.

be effective[12,16,31]. Other drugs such as NSAIDs and corticosteroids have limited indication for treatment of FMF. Myalgia responds to treatment with corticosteroids, but not colchicines. NSAIDs are used for treatment of arthralgia[21]. Febrile attacks do not respond to and are not preventable by NSAIDs or steroids. Severity Scoring: The severity of FMF can be predicted by Pras’s severity-scale scores (Table 2)[45]. This scoring can be used for establishing a treatment and follow-up strategy. In this system, mild severity is defined with a score of 2-5 points, moderate severity with a score of 6-10, and severe disease has a score more than 10. New FMF severity score was suggested by Mor et al with more sensitivity and specificity (>92%) (Table 3)[46]. Patients with severe FMF should be treated with high dose colchicine and should be followed closely. Prognosis: Generally, prognosis of FMF is related to subsequent amyloidosis. Before initiating colchicine therapy, the incidence of amyloidosis was near 50%[12], but it has decreased to less than 30%. The effect of several factors for development of amyloidosis has been assessed in different studies. Some authors believed there is an association between severity of FMF and its prognosis[46]. All factors evaluated for disease severity can play a role in FMF prognosis[46]. Although some investigators believed that the severity of FMF does not play a major role in development of amyloidosis in FMF[32]. Patients Table 2 Pras’s severity-scale scores for Familial Mediterranean fever[45] Variable Score Age of onset Younger than 5 years 5-10 years

Prognosis: Generally, prognosis of FMF is related to subsequent amyloidosis. Before initiating colchicine therapy, the incidence of amyloidosis was near 50%[12], but it has decreased to less than 30%. The effect of several factors for development of amyloidosis has been assessed in different studies. Some authors believed there is an association between severity of FMF and its prognosis[46]. All factors evaluated for disease severity can play a role in FMF prognosis[46]. Although some investigators believed that the severity of FMF does not play a major role in development of amyloidosis in FMF[32]. Patients Table 2 Pras’s severity-scale scores for Familial Mediterranean fever[45] Variable Score Age of onset Younger than 5 years 5-10 years 10-20 years 20 years or older 3 points 2 points 1 point 0 point Frequency of attacks (number per month) More than 2 1-2 Less than 1 3 points 2 points 1 point Colchicine dosage to control attacks (tablets per day) More than 4 (no response) 4 3 2 4 points 3 points 2 points 1 point Arthritis Protracted Acute 3 points 2 points Erysipelas like erythema Present 2 points Amyloidosis Present Phenotype II 3 points 4 points with low compliance to colchicine therapy are in risk for renal amyloidosis [8,21,36,37]. There is an association between low compliance for treatment and continuous inflammation (number and duration of fever attacks). Genetic predisposition is one of the other important factors in development of amyloidosis[36]. Mutation of M694V has a correlation with amyloidosis even in patients without fever[44,47].

37]. There is an association between low compliance for treatment and continuous inflammation (number and duration of fever attacks). Genetic predisposition is one of the other important factors in development of amyloidosis[36]. Mutation of M694V has a correlation with amyloidosis even in patients without fever[44,47]. Other factors which can decrease the risk of amyloidosis include: earlier treatment and continuous therapy with colchicine[37]. It seems that environmental factors such as geography play a role in development of amyloidosis[12]. Amyloidosis is more common in North Africans Jews, Armenians, and Turks with M694V homozygous and positive family history of amyloidosis. This complication is less seen in Iraqi Jews, American Armenians, Arabs and Iranians[7,27]. Sometimes serum amyloid A level during the attack-free periods is elevated, and high level of amyloid A level may be the first symptom of renal amyloidosis, so monitoring serum amyloid A level may be helpful to predict the progress of renal amyloidosis[40]. Late onset disease may have a milder clinical presentation and has a better prognosis[26]. Amyloidosis rarely occurs in adults with late onset FMF[28,40].

Sometimes serum amyloid A level during the attack-free periods is elevated, and high level of amyloid A level may be the first symptom of renal amyloidosis, so monitoring serum amyloid A level may be helpful to predict the progress of renal amyloidosis[40]. Late onset disease may have a milder clinical presentation and has a better prognosis[26]. Amyloidosis rarely occurs in adults with late onset FMF[28,40]. Familial Mediterranean fever in Iran: People with different ethnicities live in Iran and consanguineous marriage is common between Iranian people. These leads to more frequent diagnosis of genetic disorders like FMF in this country. The disease is seen sporadically in different parts of Iran[7], however, it seems that Iranian Azeri Turks are more prone to FMF[33]. Demographic and clinical presentations of patients with FMF in Iran are comparable with those of other populations (Table 4). Consanguineous marriages (<40%) and disease occurrence in siblings (33%) are common in Iranian patients[33,34]. Table 3 Second set of criteria for Familial Mediterranean fever severity score[46] Criteria 1. > 1 site in a single attack (In at least 25% of attacks) 2. > 2 sites in the course of the disease 3. > 2 mg/day colchicine to achieve remission 4. > 2 pleuritic attacks during the course of the disease 5. > 2 Erysipelas-like erythema attacks during the course of the disease 6. Age of onset <10 years. Severe disease >3 criteria; intermediate disease 2 criteria; mild disease < 1 criterion Table 4 Comparison of clinical characteristics of Familial Mediterranean fever in Iran with other countries

1. > 1 site in a single attack (In at least 25% of attacks) 2. > 2 sites in the course of the disease 3. > 2 mg/day colchicine to achieve remission 4. > 2 pleuritic attacks during the course of the disease 5. > 2 Erysipelas-like erythema attacks during the course of the disease 6. Age of onset <10 years. Severe disease >3 criteria; intermediate disease 2 criteria; mild disease < 1 criterion Table 4 Comparison of clinical characteristics of Familial Mediterranean fever in Iran with other countries Country [Reference No] Mean age at the onset of symptoms Frequency of clinical findings In >70% of patients In >20% and <70% of patients In <20% of patients Prevalence of amyloidosis The most prevalent mutations Iran/ Tehran [ 33 ] 11.4 (1 month- 28 years) Fever, Abdominal pain Chest pain Erysipelas-like erythema 5.6% M694V, M680I Iran/ Tabriz [ 34 ] 18.5 (2-66 years) Abdominal pain, Fever Arthritis and chest pain Erysipelas-like erythema 4% M694V, V726A Iran/ Tehran [ 7 ] 49.2 mo (2 mo-15 yrs) Fever, Abdominal pain Joint pain and chest pain Erysipelas-like erythema and bone pain 0 M694V, V726A Iran/ Semnan [ 28 ] 29 ±7.8 Fever, Abdominal pain Chest pain, Scrotal pain, Headache Arthritis and Erysipelas-like erythema Not reported Not studed Japan [ 48 ] -- Fever Abdominal pain, chest pain, Arthritis Erysipelas-like erythema 3.7% M694I/ E148Q Turky [ 49 ] Range: 1-12 Fever, Abdominal pain, Arthritis Chest pain Vvasculitis rash 3.5% M694V, M680I Arabs/Egypt [ 50 , 51 ] 6.9±2.8 Fever, Abdominal pain, Arthralgia, Myalgia and Operation Arthritis, chest pain Not reported V726A/M694V

Japan [ 48 ] -- Fever Abdominal pain, chest pain, Arthritis Erysipelas-like erythema 3.7% M694I/ E148Q Turky [ 49 ] Range: 1-12 Fever, Abdominal pain, Arthritis Chest pain Vvasculitis rash 3.5% M694V, M680I Arabs/Egypt [ 50 , 51 ] 6.9±2.8 Fever, Abdominal pain, Arthralgia, Myalgia and Operation Arthritis, chest pain Not reported V726A/M694V Armenia [ 24 ] 10.8±8.4 Fever, Abdominal pain, Chest pain, Arthritis -- -- 21.2% M694V, M680I Fever and abdominal pain are the most common clinical findings occurring in more than 85% of patients while erysipelas-like erythema is reported in less than 10%[7,33,34]. Molecular approach for diagnosis of FMF is rapidly expanding in Iran and some referral laboratories for genetic study such as the laboratory of Children’s Medical Center in Tehran. In Iranian people, M694V is the most common mutation[33,34]. Other common mutations in Iranians are M680I, V726A, E148Q and M694I. Recently, in a study on a group of Iranian patients in our center, we found 2 rare mutations in 2 patients in exon 10 in codons[52]. Most Iranian patients have good (60%-100%) response to colchicine [7,28,33,34]. Amyloidosis has been reported in 0-5.6% of Iranian FMF patients[7,33.34]. Periodic fever, Aphthous stomatitis, Pharyngitis, Cervical adenitis (PFAPA) Marshall's syndrome or Periodic fever, aphthous stomatitis, pharyngitis, cervical adenitis which stands for PFAPA is a sporadic syndrome characterized by recurrent febrile disease with symptoms in head and neck[4,8,53]. Clinical Pearls in FMF It is an autosomal recessive disorder and the most common disease among HPFS.

Periodic fever, Aphthous stomatitis, Pharyngitis, Cervical adenitis (PFAPA) Marshall's syndrome or Periodic fever, aphthous stomatitis, pharyngitis, cervical adenitis which stands for PFAPA is a sporadic syndrome characterized by recurrent febrile disease with symptoms in head and neck[4,8,53]. Clinical Pearls in FMF It is an autosomal recessive disorder and the most common disease among HPFS. The MEFV Gene which is responsible for the disease is located on short arm of chromosome 16. It usually begins in early childhood. Common clinical manifestations are: recurrent fever, abdominal pain, arthritis and erysipelas-like erythema. Renal amyloidosis is the worst complication of FMF. Definite diagnosis of FMF is based on finding of the mutation in MEFV gene. The main drug for treatment of FMF is colchicine. Table 5 Clinical criteria of Periodic Fever, Aphthus stomatitis, Pharyngitis, Cervical Adenitis Regulatory recurring fevers with an early age of onset (<5 years of age) Symptoms in the absence of upper respiratory tract infection with at least one of the following clinical signs: a) aphthous stomatitis b) cervical lymphadenitis c) pharyngitis Exclusion of cyclic neutropenia, completely asymptomatic interval between episodes, normal growth and development. Epidemiology: The age of onset is before 5 years[4,9,31,53] and this disease is slightly more common in males[4,31].

Symptoms in the absence of upper respiratory tract infection with at least one of the following clinical signs: a) aphthous stomatitis b) cervical lymphadenitis c) pharyngitis Exclusion of cyclic neutropenia, completely asymptomatic interval between episodes, normal growth and development. Epidemiology: The age of onset is before 5 years[4,9,31,53] and this disease is slightly more common in males[4,31]. Etiology and Pathophysiology: The etiology of the illness is still unknown, but it is thought that impairment in regulation of cytokine production may be involved in pathogenesis of PFAPA[4,8,31]. Several cytokines such as IFN, TNF, and IL-6 are elevated in febrile episodes[8]. It seems that an abnormal immune response to an antigen in oral cavity or tonsils is responsible for symptoms. Pathologic findings in tonsils are nothing but just nonspecific chronic inflammation[54]. Clinical Manifestations: PFAPA usually occurs in regular intervals and episodes of fever every 2 to 12 weeks, but when the child gets older the intervals of the disease increase[9,31,53]. Patients feel well between episodes of the illness. The onset of the disease is along with malaise and then an abrupt fever which may rise up to 40oC. Exudative tonsillitis is seen in most patients, and aphtous stomatitis is seen in 70% of patients. Cervical adenitis presents in 88-100% of patients. Chills, sweating, headache, and myalgia are common[8,53]. Sometimes mild hepatospleno-megaly and arthralgia may be seen[8]. Symptoms of the illness usually last for 3-4 days and resolve slowly[4,31] without sequels[9].

and aphtous stomatitis is seen in 70% of patients. Cervical adenitis presents in 88-100% of patients. Chills, sweating, headache, and myalgia are common[8,53]. Sometimes mild hepatospleno-megaly and arthralgia may be seen[8]. Symptoms of the illness usually last for 3-4 days and resolve slowly[4,31] without sequels[9]. Laboratory findings: Laboratory findings are mild leukocytosis, elevated ESR and CRP[8,9,31], and high level of serum IgD in 66% of patients and elevated IgE level in 50% of patients[8]. Radiologic findings are normal in all patients[8]. Diagnosis and differential diagnosis: The diagnosis is made clinically with exclusion of other diseases and with significant response to single dose of corticosteroids[53]. In 1999 “Thomas et al” ,diagnostic criteria were determined in order to facilitate diagnosis of PFAPA (Table 5)[53]. PFAPA should be differentiated from other periodic febrile illnesses such as recurrent tonsillitis, hyperglobulinemia D syndrome, cyclic neutron-penia and infectious diseases[8,12,53].

rticosteroids[53]. In 1999 “Thomas et al” ,diagnostic criteria were determined in order to facilitate diagnosis of PFAPA (Table 5)[53]. PFAPA should be differentiated from other periodic febrile illnesses such as recurrent tonsillitis, hyperglobulinemia D syndrome, cyclic neutron-penia and infectious diseases[8,12,53]. Treatment: Treatment with antibiotics and NSAIDs is ineffective and PFAPA dramatically responds to a single dose of prednisolone 2mg/kg or 0.3mg/kg of bethamethasone[8]. They are the first line therapy but the treatment may be associated with decreased intervals of the disease[9] and prednisolone cannot prevent subsequent episodes[53]. Fever subsides within 2 to 4 hours but other symptoms such as aphthous stomatitis disappear slower than the fever[8]. Cimetidine is used for prevention of episodes in some centers[9]. Recently it is proven that tonsillectomy with or without adenoidectomy resolves symptoms of the disease in more than 90% of cases and improves symptoms in 4.6% of cases[53,54] but not all experts do recommend it.

titis disappear slower than the fever[8]. Cimetidine is used for prevention of episodes in some centers[9]. Recently it is proven that tonsillectomy with or without adenoidectomy resolves symptoms of the disease in more than 90% of cases and improves symptoms in 4.6% of cases[53,54] but not all experts do recommend it. Prognosis: Prognosis of PFAPA is very excellent. Affected children have normal development and growth[8]. No morbidity or mortality has been reported up to now in PFAPA patients. Unlike other periodic fevers, amyloidosis or chronic organ involvement is not a complication of PFAPA, but there is a case report on IgA nephropathy in a child after 5 years periodic fever and PFAPA. Exacerbation of hematuria has been reported in this case during fever attacks. Treatment with methyl prednisolone pulse therapy and immunosuppressives improved urine findings, but successful treatment for control of PFAPA episodes and IgA nephropathy was tonsil-ectomy[55]. Clinical Pearls in Periodic Fever, Aphthous stomatitis, Pharyngitis, Cervical Adenitis PAFAP is a periodic fever with unknown pathophysiology. It usually affects children younger than 5 years old. Prognosis is good without any prolonged complications.

Prognosis: Prognosis of PFAPA is very excellent. Affected children have normal development and growth[8]. No morbidity or mortality has been reported up to now in PFAPA patients. Unlike other periodic fevers, amyloidosis or chronic organ involvement is not a complication of PFAPA, but there is a case report on IgA nephropathy in a child after 5 years periodic fever and PFAPA. Exacerbation of hematuria has been reported in this case during fever attacks. Treatment with methyl prednisolone pulse therapy and immunosuppressives improved urine findings, but successful treatment for control of PFAPA episodes and IgA nephropathy was tonsil-ectomy[55]. Clinical Pearls in Periodic Fever, Aphthous stomatitis, Pharyngitis, Cervical Adenitis PAFAP is a periodic fever with unknown pathophysiology. It usually affects children younger than 5 years old. Prognosis is good without any prolonged complications. PFAPA in Iran: Although there is no report on characteristics of PFAPA in Iranian patients, it seems not to be rare in Iran. An unpublished report in our system registry shows PFAPA diagnosis in nearly 20% of patients with periodic fever. In a recent report from Iran, 30% of patients with PFAPA had MEFV gene mutations, although these mutations do not have any effect on presentation and course of PFAPA[56].

s, it seems not to be rare in Iran. An unpublished report in our system registry shows PFAPA diagnosis in nearly 20% of patients with periodic fever. In a recent report from Iran, 30% of patients with PFAPA had MEFV gene mutations, although these mutations do not have any effect on presentation and course of PFAPA[56]. Other periodic fever syndromes and auto- inflammatory disorders include TRAPS, hyper IgD sydromes and cryopyrin (CINCA or NOMID, Muckle-Wells syndrome, familial cold urticaria), PAPA (pyogenic arthritis, pyoderma gangraenosum, cystic acne), CRMO (chronic recurrent multifocal osteomyelitis), DIRA (deficiency of the interleukin-1–receptor antagonist) and Majeed syndrome will be discussed in part II of this paper. Conflict of Interest: None

Introduction China has the largest floating population in the world. The floating population refers to the large and increasing number of migrants without local household registration status. Most of these migrants came from the rural areas of the central and western parts of China. They face daunting problems particularly with access to health care, adequate housing, employment opportunities. The children of floating population can not fully enjoy the social medical services, and that should bring a bad impact on their health. According to a systematic analysis reported in Lancet, of the estimated 8.795 million deaths in children younger than 5 years worldwide in 2008, infectious diseases caused 68%, with the largest percentages due to pneumonia[1]. According to the concept of recurrent respiratory tract infections (RRIs), a child with upper respiratory infection at least 6 times or lower respiratory infection at least 2 times per year was defined as a child with recurrent respiratory infection[2]. Diagnosis of rhinitis, nasopharyngitis, oropharyngitis, tonsillitis or laryngitis, Otitis media was evaluated as upper respiratory tract infections. Tracheitis, bronchitis, bropnchiolitis and pneumonia were assessed as low respiratory tract infections.

ear was defined as a child with recurrent respiratory infection[2]. Diagnosis of rhinitis, nasopharyngitis, oropharyngitis, tonsillitis or laryngitis, Otitis media was evaluated as upper respiratory tract infections. Tracheitis, bronchitis, bropnchiolitis and pneumonia were assessed as low respiratory tract infections. Some literatures reported the associated factors for recurrent respiratory infections, such as socioeconomic status, family characteristics, air pollution, smoking, maternal age, breast feeding, micronutrient Condition of children et al[3-10]. But often only a few risk factors have been studied at a time. The present study aimed to identify and compare risk factors for recurrent respiratory infections in preschool children between resident and floating population in Yiwu, China; and to give suggestions for the health care provider to prevent recurrent respiratory infections according to the key findings for resident and floating population children respectively. Subjects and Methods Study Area and Population

Some literatures reported the associated factors for recurrent respiratory infections, such as socioeconomic status, family characteristics, air pollution, smoking, maternal age, breast feeding, micronutrient Condition of children et al[3-10]. But often only a few risk factors have been studied at a time. The present study aimed to identify and compare risk factors for recurrent respiratory infections in preschool children between resident and floating population in Yiwu, China; and to give suggestions for the health care provider to prevent recurrent respiratory infections according to the key findings for resident and floating population children respectively. Subjects and Methods Study Area and Population Yiwu, located in the central part of Zhejiang Province, covers an area of 1,105 sq km with 6 towns and 7 sub-districts and has 740,000 resident populations and 1,430,000 floating population. The study was conducted in Yiwu from June 2009 to September 2010. According to the principle of random sampling, we recruited 844 participants，resident population 639 (75.7%) and floating population 205 (24.3%) who were preschool children. They were sampling among nursery schools registered in the bureau of education in Yiwu. The sample size was calculated according the prevalence rate and the feasibility of the investigation. The sampling procedure was as follows. The registered children were coded and then selected by the method of random number which generated by Microsoft Office Excel with the function RAND. Children with congenital disease, hereditary disease and tumor as well as having a history of surgery were excluded from this study. Children using immunosuppressive agent or hormonal drugs were also excluded. In this paper, floating children were defined as preschool children who had urban registered permanent residence of Yiwu and lived in Yiwu for three and more months.

d tumor as well as having a history of surgery were excluded from this study. Children using immunosuppressive agent or hormonal drugs were also excluded. In this paper, floating children were defined as preschool children who had urban registered permanent residence of Yiwu and lived in Yiwu for three and more months. Diagnosis procedure for RRIs Diagnosis was made by a physician according to their hospital records. Based on the clinical concept and management of recurrent respiratory tract infections in children of 3-5 years old in 2008[2], a child with upper respiratory infection at least 6 times or lower respiratory infection at least 2 times per year was defined as a patient with recurrent respiratory infection. The interval between every two infections should be at least 7 days. If upper respiratory infections were less than 6 times annually, lower respiratory infections could be added to meet the diagnostic criteria. Data Collection

Diagnosis was made by a physician according to their hospital records. Based on the clinical concept and management of recurrent respiratory tract infections in children of 3-5 years old in 2008[2], a child with upper respiratory infection at least 6 times or lower respiratory infection at least 2 times per year was defined as a patient with recurrent respiratory infection. The interval between every two infections should be at least 7 days. If upper respiratory infections were less than 6 times annually, lower respiratory infections could be added to meet the diagnostic criteria. Data Collection In the present study, when questioning whether participants had experienced infections, diagnosis was required by a physician according to hospital records. In order to explore the influence factors in preschool children with recurrent respiratory infections, structured questionnaires were addressed the mother or guardians. The questionnaire was designed according to the refereces[3,4]. The first part of the questionnaire included demographic details, such as age, gander, weight and height. The second part consisted of questions about the associated factors of RRIs, including the following item categories: birth characteristics, socioeconomic factors, fostering factors,housing conditions.

ereces[3,4]. The first part of the questionnaire included demographic details, such as age, gander, weight and height. The second part consisted of questions about the associated factors of RRIs, including the following item categories: birth characteristics, socioeconomic factors, fostering factors,housing conditions. A medical doctor and a registered nurse who was familiar with anthropometric measurements (weight, height) of children were trained for 2 days before the questionnaire investigation. The investigation was performed between June 2009 and September 2010. The serum of children was collected after questionnaire investigation. All the guardians who took part in the study gave their informed consent for participation. Trace element and Immunoglobulin in serum of the children were measured by atomic absorption method and immunological turbidity kit (Beijing Pu MB5) respectively. The laboratory results were also recorded. Statistical Methods The database was established by Epidata3.0 and analyzed by SPSS16.0 software. Chi square test was used to compare the distribution of some health related factors in preschool children between resident and floating population. If p value was less than 0.05, then statistical significance was considered.

A medical doctor and a registered nurse who was familiar with anthropometric measurements (weight, height) of children were trained for 2 days before the questionnaire investigation. The investigation was performed between June 2009 and September 2010. The serum of children was collected after questionnaire investigation. All the guardians who took part in the study gave their informed consent for participation. Trace element and Immunoglobulin in serum of the children were measured by atomic absorption method and immunological turbidity kit (Beijing Pu MB5) respectively. The laboratory results were also recorded. Statistical Methods The database was established by Epidata3.0 and analyzed by SPSS16.0 software. Chi square test was used to compare the distribution of some health related factors in preschool children between resident and floating population. If p value was less than 0.05, then statistical significance was considered. Risk factor analyses were performed in resident and floating population respectively. Risk factor analyses were carried out using Chi-square test. Based on the results of Chi-square test; a multivariate regression model was constructed by Binary logistic regression. The SPSS16.0 software was used for the Logistic Model, Forward likelihood ratio was chosen as a method of analysis. For inclusion in the model P=0.05 was used, whereas P=0.10 was chosen for exclusion from the model. In all the analyses, the confidence level was 95%. Findings Study Population and Influence Factor Analyses

Risk factor analyses were performed in resident and floating population respectively. Risk factor analyses were carried out using Chi-square test. Based on the results of Chi-square test; a multivariate regression model was constructed by Binary logistic regression. The SPSS16.0 software was used for the Logistic Model, Forward likelihood ratio was chosen as a method of analysis. For inclusion in the model P=0.05 was used, whereas P=0.10 was chosen for exclusion from the model. In all the analyses, the confidence level was 95%. Findings Study Population and Influence Factor Analyses For resident and floating population respectively, the height of preschool children were 104.610.6cm and 103.69.9cm with no significant difference (t=1.154，P=0.2); the weight were 17.44.5kg and 16.7 3.7kg with significant difference (t=2.190，P=0.03). Childbearing age were 27.54.0 years and 26.83.9 years respectively with significant difference (t=0.744, P=0.04); the birth weight were 3.4±0.6kg and 3.3±0.6kg with no significant difference (t=-0.744, P=0.5).

erence (t=1.154，P=0.2); the weight were 17.44.5kg and 16.7 3.7kg with significant difference (t=2.190，P=0.03). Childbearing age were 27.54.0 years and 26.83.9 years respectively with significant difference (t=0.744, P=0.04); the birth weight were 3.4±0.6kg and 3.3±0.6kg with no significant difference (t=-0.744, P=0.5). The prevalence rate for recurrent respiratory infections in resident populations and floating population children were 25.2% and 23.9% respectively with significant difference(t=2.190, P=0.03). Tables 1, 2 present the comparison of related social factors, fostering factors and living environment factors between resident populations and floating population preschool children in Yiwu, China. When the gender, parents occupation, monthly income were similar, education (father and mother) were significantly different. There were significant differences between resident populations and floating population preschool children on a number of fostering and living environment factors (delivery, vaccination, sleeping habit, taking care). Risk Factors in Resident Preschool Children with Recurrent Respiratory Infections The height of children were 104.811.0cm and 103.99.5cm in children with RRIs and control respectively, and there was no significant difference between the two groups (t=0.949, P=0.343). The weight were 17.54.6kg and 17.14.2kg in the two groups, and there was no significant difference between the two groups(t=0.958, P=0.338). Table1 Comparison of related social factors in preschool children between resident and floating population in Yiwu, China, 2009-2010 Characteristics

The height of children were 104.811.0cm and 103.99.5cm in children with RRIs and control respectively, and there was no significant difference between the two groups (t=0.949, P=0.343). The weight were 17.54.6kg and 17.14.2kg in the two groups, and there was no significant difference between the two groups(t=0.958, P=0.338). Table1 Comparison of related social factors in preschool children between resident and floating population in Yiwu, China, 2009-2010 Characteristics Resident population Floating population χ2 P. value Gender Male 373 (58.4) 114 (55.6) 0.379 0.5 Female 266 (41.6) 91 (44.4) Occupation Brain work 324 (50.7) 96 (46.8) 1.367 0.5 Physical force work 88 (13.8) 34 (16.6) Both 227 (35.5) 75 (6.6) Monthly income(RMB) <1000 28 (4.4) 12 (5.9) 3.255 0.4 >=1000 170 (26.6) 46 (22.4) >=3000 228 (35.7) 68 (33.2) >=6000 213 (33.3) 79 (38.5) Education, father Junior school 134 (21.0) 57 (27.8) 16.673 <0.001 Senior school 176 (27.5) 76 (37.1) College 329 (51.5) 72 (35.1) Education, mother Junior school 139 (21.8) 82 (40.0) 32.665 <0.001 Senior school 199 (31.1) 65 (31.7) College 301 (47.1) 58 (28.3) Table 2 Comparison of related social factors in preschool children between resident and floating population in Yiwu, China, 2009-2010 Characteristics Resident population Floating population χ 2 P Gender Male 373 58.4% 114 55.6% 0.379 0.538 Female 266 41.6% 91 44.4% Occupation Brain work 324 50.7% 96 46.8% 1.367 0.505 Physical force work 88 13.8% 34 16.6% Both 227 35.5% 75 36.6% Monthly income(RMB) <1000 28 4.4% 12 5.9% 3.255 0.354 >=1000 170 26.6% 46 22.4% >=3000 228 35.7% 68 33.2% >=6000 213 33.3% 79 38.5%

population χ 2 P Gender Male 373 58.4% 114 55.6% 0.379 0.538 Female 266 41.6% 91 44.4% Occupation Brain work 324 50.7% 96 46.8% 1.367 0.505 Physical force work 88 13.8% 34 16.6% Both 227 35.5% 75 36.6% Monthly income(RMB) <1000 28 4.4% 12 5.9% 3.255 0.354 >=1000 170 26.6% 46 22.4% >=3000 228 35.7% 68 33.2% >=6000 213 33.3% 79 38.5% Education, father Junior school 134 21.0% 57 27.8% 16.673 <0.001 Senior school 176 27.5% 76 37.1% College 329 51.5% 72 35.1% Education, mother Junior school 139 21.8% 82 40.0% 32,665 <0.001 Senior school 199 31.1% 65 31.7% College 301 47.1% 58 28.3% Childbearing age were 27.73.9 years and 27.04.2 years in children with RRIs and control respectively, and there was significant difference between the two groups (t=1.969, P=0.049). The birth weight were 3.30.5kgand 3.40.7kg in the two groups, and there was no significant difference between the two groups (t=-1.459, P=0.145). Tables 3, 4 present the results of the univariate risk factor analyses in resident populations preschool children. A number of factors were significantly associated with recurrent respiratory infections (delivery, asthma, rickets, food preference, snack, drinking water, sleeping habit, trip mode, someone smoking in the same room).

present the results of the univariate risk factor analyses in resident populations preschool children. A number of factors were significantly associated with recurrent respiratory infections (delivery, asthma, rickets, food preference, snack, drinking water, sleeping habit, trip mode, someone smoking in the same room). The variables were chosen as those with a p value of less than 0.10 in the univariate analyses. The multivariate equation model consisted of the following variables: childbearing age (OR=0.93, 95%CI: 0.88-0.99, P=0.01), asthma (OR=3.20, 95%CI: 1.60-6.45, P=0.001), rickets (OR=5.30, 95%CI: 1.99-14.09, P=0.001), food preference (OR=1.57, 95%CI: 1.02-2.43, P=0.04), snack (OR=1.50, 95%CI: 1.07-2.07, P=0.02). There was no significant difference between children with RRIs and control on trace element and immunoglobulin in serum (P >0.05) (Table 5). Risk Factors in Floating Preschool Children with Recurrent respiratory Infections The height of children were 102.79.3cm and 105.810.6cm in children with RRIs and control respectively, and there was no significant difference between the two groups (t=-1.945, P=0.05). The weight were 16.23.3kg and 17.84.2kg in the two groups, and there was significant difference between the two groups (t=-2.648, P=0.009). Childbearing age were 27.13.9years and 26.03.8 years in children with RRIs and control respectively, and there was no significant difference between the two groups (t=1.718, P=0.09). The birth weight were 3.30.6kg and 3.40.6kg in the two groups, and there was no significant difference between the two groups (t=-1.078, P=0.3).

aring age were 27.13.9years and 26.03.8 years in children with RRIs and control respectively, and there was no significant difference between the two groups (t=1.718, P=0.09). The birth weight were 3.30.6kg and 3.40.6kg in the two groups, and there was no significant difference between the two groups (t=-1.078, P=0.3). Tables 3, 4 present the results of the univariate risk factor analyses in floating population preschool children rural child. A number of factors were significantly associated with recurrent respiratory infections (infant feeding, rickets, and snake). The variables were chosen as those with a P value of less than 0.10 in the univariate analyses. The multivariate equation model consisted of the following variables: infant feeding (OR=2.24, 95%CI: 1.31-3.81, P=0.003), snack (OR=2.06, 95%CI: 1.07-3.98, P=0.03), trip mode (OR=2.77, 95%CI: 1.11-6.94, P =0.03). There were significant differences between children with RRIs and control on ferrum and alkaline phosphatase level when detecting trace element and immunoglobulin in serum. Preschool children with recurrent respiratory infection have lower level of ferrum and high level of alkaline phosphatase (P <0.05) (Table 5). Table 3 Risk factors for recurrent respiratory infections in preschool children between resident and floating population in Yiwu, China, 2009-2010 Characteristics Resident population Floating population Children with RRIs (%) Control (%) P. value Children with RRIs (%) Control (%) P. value Age(year) <=3 24 (14.9) 90 (18.8) 0.09 11 (22.4) 36 (23.7) 0.2 <=4 57 (35.4) 120 (25.1) 11 (22.4) 45 (29.6) <=5 38 (23.6) 126 (26.4) 9 (18.4) 38 (25.0)

Table 3 Risk factors for recurrent respiratory infections in preschool children between resident and floating population in Yiwu, China, 2009-2010 Characteristics Resident population Floating population Children with RRIs (%) Control (%) P. value Children with RRIs (%) Control (%) P. value Age(year) <=3 24 (14.9) 90 (18.8) 0.09 11 (22.4) 36 (23.7) 0.2 <=4 57 (35.4) 120 (25.1) 11 (22.4) 45 (29.6) <=5 38 (23.6) 126 (26.4) 9 (18.4) 38 (25.0) >5 42 (26.1) 142 (29.7) 18 (36.7) 33 (21.7) Gender Male 93 (57.8) 279 (58.4) 0.9 25 (51.0) 88 (57.9) 0.4 Female 68 (42.2) 199 (41.6) 24 (49.0) 64 (42.1) Monthly income (RMB) <1000 6 (3.7) 21 (4.4) 0.09 3 (6.1) 9 (5.9) 0.6 >=1000 46 (28.6) 124 (25.9) 9 (18.4) 36 (23.7) >=3000 46 (28.6) 186 (38.9) 15 (30.6) 54 (35.5) >=6000 63 (39.1) 147 (30.8) 22 (44.9) 53 (34.9) Delivery Spontaneous labor 78 (48.4) 190 (39.7) 0.048 29 (59.2) 84 (55.3) 0.6 Abdominal delivery 83 (51.6) 290 (60.7) 20 (40.8) 68 (44.7) Vaccination Yes 146 (90.7) 436 (91.2) 0.8 36 (73.5) 129 (84.9) 0.07 No 15 (9.3) 42 (8.8) 13 (26.5) 23 (15.1) Infant feeding Exclusively breastfed 75 (46.6) 225 (47.1) 0.9 16 (32.7) 79 (52.0) 0.03 Partially breastfed 38 (23.6) 117 (24.5) 11 (22.4) 32 (21.1) Not breastfed 48 (29.8) 136 (28.5) 22 (44.9) 41 (27.0) Asthma Yes 23 (14.3) 29 (6.1) 0.001 6 (12.2) 11 (7.2) 0.3 No 138 (85.7) 449 (93.9) 43 (87.8) 141 (92.8) Rickets Yes 44 (27.3) 133 (27.8) 0.9 7 (14.3) 43 (28.3) 0.04 No 117 (72.7) 345 (72.2) 42 (85.7) 109 (71.7) Food preference Yes 92 (57.1) 225 (47.1) 0.03 25 (51.0) 60 (39.5) 0.2 No 69 (42.99)% 253 (52.9) 24 (49.0) 92 (60.5) Snack Occasionally 63 (39.1) 225 (47.1) 0.01 13 (26.5) 70 (46.1) 0.001 Regularly 77 (47.8) 223 (46.7) 26 (53.1) 75 (49.3)

Yes 44 (27.3) 133 (27.8) 0.9 7 (14.3) 43 (28.3) 0.04 No 117 (72.7) 345 (72.2) 42 (85.7) 109 (71.7) Food preference Yes 92 (57.1) 225 (47.1) 0.03 25 (51.0) 60 (39.5) 0.2 No 69 (42.99)% 253 (52.9) 24 (49.0) 92 (60.5) Snack Occasionally 63 (39.1) 225 (47.1) 0.01 13 (26.5) 70 (46.1) 0.001 Regularly 77 (47.8) 223 (46.7) 26 (53.1) 75 (49.3) Frequently 21 (13.0) 30 (6.3) 10 (20.4) 7 (4.6) Drinking water Drink 46 (28.6) 122 (25.5) 0.001 11 (22.4) 36 (23.7) 0.5 Plain boiled water 97 (60.2) 338 (70.7) 33 (67.3) 108 (71.1) Fresh Fruit juice 18 (11.2) 18 (3.8) 5 (10.2) 8 (5.3) Sleeping habit With adults 120 (74.5) 313 (65.5) 0.003 27 (55.1) 84 (55.3) 0.8 With himself or herself 26 (16.1) 64 (13.4) 9 (18.4) 33 (21.7) Both 15 (9.3) 101 (21.1) 13 (26.5) 35 (23.0) Trip mode Car 97 (60.2) 344 (72.0) 0.005 27 (55.1) 105 (69.1) 0.1 Walking 44 (27.3) 106 (22.2) 18 (36.7) 33 (21.7) Bicycle 20 (12.4) 28 (5.9) 4 (8.2) 14 (9.2) Someone with CRD in the same room Yes 19 (11.8) 41 (8.6) 0.2 9 (18.4) 12 (7.9) 0.04 No 142 (88.2) 437 (91.4) 40 (81.6) 140 (92.1) Someone smoking in the same room Yes 90 (55.9) 207 (43.3) 0.006 27 (55.1) 62 (40.8) 0.1 No 71 (44.1) 271 (56.7) 22 (44.9) 90 (59.2) Sweeping the room frequently Yes 126 (78.3) 404 (84.5) 0.07 38 (77.6) 132 (86.8) 0.2 No 35 (21.7) 74 (15.5) 11 (22.4) 20 (13.2) CRD: chronic respiratory disease Table 4 Multivariate analysis on risk factors for recurrent respiratory infections in preschool children between resident and floating population in Yiwu, China, 2009-2010 Characteristics Resident population Mean SE Wald P value OR 95% CI Mean SE Wald P value OR 95% CI Age 0.004 0.10 0.002 0.9 1.00 0.82 - 1.22 -0.14 0.29 0.27 0.6 0.87 0.51 - 1.47

Table 4 Multivariate analysis on risk factors for recurrent respiratory infections in preschool children between resident and floating population in Yiwu, China, 2009-2010 Characteristics Resident population Mean SE Wald P value OR 95% CI Mean SE Wald P value OR 95% CI Age 0.004 0.10 0.002 0.9 1.00 0.82 - 1.22 -0.14 0.29 0.27 0.6 0.87 0.51 - 1.47 Gender (Male) 0.24 0.22 1.20 0.3 1.27 0.83 - 1.96 0.55 0.41 1.87 0.2 1.74 0.79 - 3.85 Mother’s age at child’s birth -0.07 0.03 6.01 0.01 0.93 0.88 - 0.99 Infant feeding (Exclusively breastfed) 0.80 0.27 8.77 0.003 2.24 1.31 - 3.81 Without Asthma 1.17 0.36 10.73 0.001 3.21 1.60 - 6.45 Without Rickets 1.67 0.50 11.17 0.001 5.30 1.99 - 14.09 1.29 0.77 2.84 0.09 3.65 0.81 - 16.46 No Food preference 0.45 0.22 4.15 0.04 1.57 1.02 - 2.43 Snack (Occasionally) 0.40 0.17 5.67 0.02 1.49 1.07 - 2.07 0.72 0.34 4.64 0.031 2.06 1.07 - 3.98 Trip mode with car 1.02 0.47 4.73 0.03 2.77 1.11 - 6.94 SE: Standard error; CI: Confidence interval Table5 Laboratory factors for recurrent respiratory infections in preschool children between resident and floating population in Yiwu, China, 2009-2010 IgE IgG IgA IgM C3 ALP Copper Zinc Calcium Magnesium Iron Lead Hgb Resident population Children with RRIs 96.8 (146.6) 8.2 (1.5) 0.9 (0.5) 1.2 (0.2) 1.2 (0.2) 247.2 (73.0) 23.5 (4.1) 85.5 (35.6) 1.6 (0.2) 1.6 (0.5) 9.1 (3.1) 41.8 (15.4) 129.0 (7.5) Control 86.5 (109.3) 8.2 (1.8) 1.0 (0.4) 1.2 (0.2) 1.2 (0.2) 242.9 (64.9) 23.7 (2.7) 84.8 (10.3) 1.7 (0.5) 1.5 (0.2) 8.8 (0.8) 40.6 (15.1) 129.2 (12.1) t 0.788 0.146 -0.499 -0.381 -0.381 0.665 -0.743 -0.234 -1,197 1.089 0.862 0.889 -0.244

Children with RRIs 96.8 (146.6) 8.2 (1.5) 0.9 (0.5) 1.2 (0.2) 1.2 (0.2) 247.2 (73.0) 23.5 (4.1) 85.5 (35.6) 1.6 (0.2) 1.6 (0.5) 9.1 (3.1) 41.8 (15.4) 129.0 (7.5) Control 86.5 (109.3) 8.2 (1.8) 1.0 (0.4) 1.2 (0.2) 1.2 (0.2) 242.9 (64.9) 23.7 (2.7) 84.8 (10.3) 1.7 (0.5) 1.5 (0.2) 8.8 (0.8) 40.6 (15.1) 129.2 (12.1) t 0.788 0.146 -0.499 -0.381 -0.381 0.665 -0.743 -0.234 -1,197 1.089 0.862 0.889 -0.244 P value 0.431 0.884 0.618 0.703 0.703 0.506 0.458 0.815 0.232 0.276 0.389 0.374 0.807 floating population Children with RRIs 98.7 (156.6) 8.2 (1.5) 0.9 (0.5) 1.2 (0.4) 1.1 (0.2) 243.92 (54.0) 24.2 (3.1) 85.0 (13.2) 1.6 (0.2) 1.6 (0.2) 8.7 (1.0) 40.7 (15.2) 127.9 (8,7) Control 70.9 (116.1) 8.2 (1.9) 0.9 (0.4) 1.2 (0.4) 1.1 (0.2) 224.8 (36.7) 23.2 (3.0) 101.3 (15.6) 1.8 (1.1) 1.6 (0.3) 9.2 (2.7) 42.7 (18.4) 126.7 (6.9) t 1. -0.039 -0.857 1.041 0.230 2.308 1.966 1.709 -1.822 0.492 -2.010 0.731 0.864 P value 0.3 0.9 0.4 0.3 0.82 0.02 -0.05 0.09 0.07 0.6 0.046 0.5 0.4 RRI: respiratory tract infection Ig: Immunoglobulin; Alk ph: Alkaline phosphatase; Cu: Hgb; Hemoglobin

Control 70.9 (116.1) 8.2 (1.9) 0.9 (0.4) 1.2 (0.4) 1.1 (0.2) 224.8 (36.7) 23.2 (3.0) 101.3 (15.6) 1.8 (1.1) 1.6 (0.3) 9.2 (2.7) 42.7 (18.4) 126.7 (6.9) t 1. -0.039 -0.857 1.041 0.230 2.308 1.966 1.709 -1.822 0.492 -2.010 0.731 0.864 P value 0.3 0.9 0.4 0.3 0.82 0.02 -0.05 0.09 0.07 0.6 0.046 0.5 0.4 RRI: respiratory tract infection Ig: Immunoglobulin; Alk ph: Alkaline phosphatase; Cu: Hgb; Hemoglobin Discussion In the present study, child characteristics such as age, gender, birth characteristics; socioeconomic factors, fostering factors and housing conditions associated with childhood recurrent respiratory infections were studied in resident and floating population respectively. There were significant differences on parents’ education, delivery, vaccination, sleeping habit, taking care between resident and floating population. These differences may be related to the influence of social and environmental factors, health behaviors, and level of awareness concerning certain health conditions. Child bearing age was associated with the recurrent respiratory infection. Consisted with the findings in Infante-Rivard C’s study[11], childbearing age was a slight protective factor for recurrent respiratory infections with OR of 0.93 in resident children in this study, but probably due to Socio-economic differences and health awareness, maternal age was not found to be an influence factor in floating population.

ings in Infante-Rivard C’s study[11], childbearing age was a slight protective factor for recurrent respiratory infections with OR of 0.93 in resident children in this study, but probably due to Socio-economic differences and health awareness, maternal age was not found to be an influence factor in floating population. Nutritional factors take up an important role for recurrent respiratory infections by influencing body immune status. The global child disease burden attributable to maternal and child undernutrition has recently been quantified [12]. Approximately 9 million children less than 5 years old die every year in the world. Acute lower respiratory tract infections (ALRI) account for approximately 20% of these deaths[13,14].

dy immune status. The global child disease burden attributable to maternal and child undernutrition has recently been quantified [12]. Approximately 9 million children less than 5 years old die every year in the world. Acute lower respiratory tract infections (ALRI) account for approximately 20% of these deaths[13,14]. Vitamin D plays an important role for human immune system [15]. Vitamin D deficiency could result to abnormal metabolism of calcium and phosphor, and could be harmful to body immune functions. Vitamin D deficiency is now presumptively linked to a range of infectious inflammatory diseases throughout the life course and around the world [16]. On the contrary, recurrent respiratory infections could result to the body stimulus status with nutrition storage exhausted; and with medicine for infection treatment, child had decreased appetite and limited outdoor exercises. These factors are also presumptively result to the deficient intake of vitamin D and calcium. In the present study, rickets was a risk factor for recurrent respiratory infections both in resident population with OR of 5.30. Also, we found that in floating population, preschool children with recurrent respiratory infection have high level of alkaline phosphatase; Alkaline phosphatase suggested Vitamin D deficiency. So interventions to improve child vitamin D status to improve the immune function could prevent recurrent respiratory infections and reduce the global burden of recurrent respiratory infections[17].

rrent respiratory infection have high level of alkaline phosphatase; Alkaline phosphatase suggested Vitamin D deficiency. So interventions to improve child vitamin D status to improve the immune function could prevent recurrent respiratory infections and reduce the global burden of recurrent respiratory infections[17]. Malnutrition is the underlying cause of approximately half of these fatal ALRIs. Four key nutritional risk factors for ALRI disease burden have been identified. These are macronutrient undernutrition, low birthweight, zinc deficiency and suboptimal breastfeeding[18]. In floating population, preschool children with recurrent respiratory infection have lower level of ferrum. This finding was probably due to socio-economic status such as family income, parent education and their awareness of nutrition et al in floating population. We also found non-breastfeeding was a risk factor for recurrent respiratory infections both in resident population with OR of 2.24. Probably due to maternal nutritional status, working and living pressure as well as education, floating population used to select non breastfeeding. A lack of exclusive breastfeeding in the first 6 months of life increases the frequency and severity of ALRI, and the risk of death from ALRIs[19-21]. Secondly, breastfeeding enhances the infant’s antibody responses to respiratory pathogens and influences maturation of the immune system[22,23].

elect non breastfeeding. A lack of exclusive breastfeeding in the first 6 months of life increases the frequency and severity of ALRI, and the risk of death from ALRIs[19-21]. Secondly, breastfeeding enhances the infant’s antibody responses to respiratory pathogens and influences maturation of the immune system[22,23]. In our study, snacks tend to be a risk factor for recurrent respiratory infections with OR of 1.50 and 2.06 in resident and floating pre-children respectively probably because of the component or addictives in the snack other than protein. Food preference was found to be a risk factor for recurrent respiratory infections with OR of 1.57 only in resident children. Food preference could result to macronutrient undernutrition or overweight which depends on the preferred food. Both macronutrient undernutrition and overweight could be the key risk factors[24]. So we should promote breastfeeding, and integrate nutrition into other aspects of well child care, and increase both public and health professional nutritional knowledge as essential components of policy development.

In our study, snacks tend to be a risk factor for recurrent respiratory infections with OR of 1.50 and 2.06 in resident and floating pre-children respectively probably because of the component or addictives in the snack other than protein. Food preference was found to be a risk factor for recurrent respiratory infections with OR of 1.57 only in resident children. Food preference could result to macronutrient undernutrition or overweight which depends on the preferred food. Both macronutrient undernutrition and overweight could be the key risk factors[24]. So we should promote breastfeeding, and integrate nutrition into other aspects of well child care, and increase both public and health professional nutritional knowledge as essential components of policy development. Although asthma and recurrent respiratory infections has different mechanism, asthma was reported to be a risk factor for respiratory infection in early life and the atopic or asthmatic background is a severe enough predisposing factor for the development of later recurrent respiratory infection[25]. Our study does find asthma associated with the risk of recurrent respiratory infections in resident preschool children, and probably because of airway hyper reactivity in asthma children. While in floating population, this association was not found, probably because of the awareness of disease and the medical counseling behavior of their parents.

ssociated with the risk of recurrent respiratory infections in resident preschool children, and probably because of airway hyper reactivity in asthma children. While in floating population, this association was not found, probably because of the awareness of disease and the medical counseling behavior of their parents. Nitrogen oxides, sulfur dioxide, carbon monoxide, particulate pollutants in the air could be important threats for respiratory system[26-28]. John D.Spengler found the relationship between housing characteristics and respiratory health[29, 30]. In the present study, trip mode was significant in multivariate analysis in floating preschool children with OR of 2.77. An explanation could be that going out by car could avoid the traffic-related pollution. But trip mode was not found to be significant in resident population probably because of the living behavior and living condition to avoid the traffic-related air pollution.

analysis in floating preschool children with OR of 2.77. An explanation could be that going out by car could avoid the traffic-related pollution. But trip mode was not found to be significant in resident population probably because of the living behavior and living condition to avoid the traffic-related air pollution. Conclusion childbearing age, asthma, rickets, food preference and snack were risk factors for recurrent respiratory infection in resident preschool children; infant feeding, snack, trip mode were risk factors for recurrent respiratory infection in floating preschool children. In floating population, preschool children with recurrent respiratory infection have lower level of ferrum and high level of alkaline phosphatase. These findings suggest a number of measures should be taken to reduce the effects of risk factors on children’s health. Nutrition status especially vitamin D early surveillance should be taken, and integrate nutrition should be integrate into other aspects of well child care in both resident and floating population. Breastfeeding should be promoted and nutritional knowledge should be considered as essential components of policy development, and outdoor air pollution should be avoided by improve their living conditions in floating population. Acknowledgment We wish to thank the staffs participating in this study for participation and providing support during investigation, laboratory work and the data collection phase. Conflict of Interest: None

Introduction Complementary and alternative Medicine (CAM) refers to a wide range of views and actions that have not been precisely defined in current standard medicine. It is neither taught in conventional medical schools, nor applied in hospitals. CAM includes various methods for disease treatment or prevention with a different efficacy approach from traditional medicine. Various types of CAM are quite diverse in different parts of the world and influenced by culture, history, level of education and individual interests. Countries such as China, South Korea and Vietnam have accepted complementary medicine into their health systems[1]. In Africa, about 85% of people tend to use alternative medicine to treat various diseases[2]. CAM is also applied at different extents in developed countries. American academy of pediatrics (AAP) reported that 20-40% of healthy and 50% of children suffering from chronic diseases receive CAM therapies[3]. Studies conducted in different countries suggest that variety and rate of CAM for children are quite different[4-9].

lso applied at different extents in developed countries. American academy of pediatrics (AAP) reported that 20-40% of healthy and 50% of children suffering from chronic diseases receive CAM therapies[3]. Studies conducted in different countries suggest that variety and rate of CAM for children are quite different[4-9]. Due to increasing rate of CAM application in children worldwide, it is highly recommended to recognize its worth appropriately among people. Therefore, through increasing people’s knowledge in this context, and appropriate application of it for patients, one would be able to promote the patient care for children worldwide. This research project was conducted under supervision of the Research Council of Birjand University of Medical Sciences on mothers to determine the prevalence, related factors, types, the information sources and knowledge for the possible side effects. Subjects and Methods The present study conducted on the mothers of 300 children who were admitted to the general pediatric clinic in Vali-Asr Hospital, Birjand during spring (April –June) 2012. This hospital, as the main hospital in town, accepts all referrals from different strata of the society.

Due to increasing rate of CAM application in children worldwide, it is highly recommended to recognize its worth appropriately among people. Therefore, through increasing people’s knowledge in this context, and appropriate application of it for patients, one would be able to promote the patient care for children worldwide. This research project was conducted under supervision of the Research Council of Birjand University of Medical Sciences on mothers to determine the prevalence, related factors, types, the information sources and knowledge for the possible side effects. Subjects and Methods The present study conducted on the mothers of 300 children who were admitted to the general pediatric clinic in Vali-Asr Hospital, Birjand during spring (April –June) 2012. This hospital, as the main hospital in town, accepts all referrals from different strata of the society. To avoid recall bias, initially the project objective has been explained for mothers. It was also mentioned that this is just a research project, the mother’s and child’s names will be not recorded. Moreover, the participation is done completely voluntarily. Then a trained nurse interviewed the mothers who were interested to participate, and a questionnaire was completed. First of all the nurse gave necessary information about CAM and its variants including medicinal herbs, prayer therapy, acupuncture, unction, cupping therapy, leech therapy, hydrotherapy, etc. The questionnaire included demographics information for children (age and sex) and for mothers (age, education level, employment status and number of children), the use of this kind of therapy at least once during the last year for their children, divers types of CAM used, the sources of maternal information and knowledge for the possible side effects of CAM.

nformation for children (age and sex) and for mothers (age, education level, employment status and number of children), the use of this kind of therapy at least once during the last year for their children, divers types of CAM used, the sources of maternal information and knowledge for the possible side effects of CAM. The face and content validity of the questionnaire were approved by consulting with several expert faculty members and its reliability was determined after completing 30 questionnaires and calculating the Cronbach's alpha coefficient. Data entered into the SPSS software (version 17) and were analyzed by descriptive statistics (mean and frequency) and analytical statistics (Chi-square test). P value less than 0.05 was considered as significant. Findings All 300 mothers agreed to participate in the study. Median age of mothers and children was 31.65±4.74 and 3.24±2.60 years, respectively. 66% of children were less than three years old and 52% were boys. The majority of mothers (87%) were housewives and most of them (61.2%) had one or two children. 35.6% of mothers had used CAM. There was a significant and direct correlation between using CAM with increased maternal age, decreased level of mother’s education, mother being as a housewife and having more than two children (Table 1). Regarding the type of CAM which was used, 105 mothers responded to this section. The most common treatment was medicinal herbs (93.3%), oil rub (26.6%) and prayer therapy (25.7%).

with increased maternal age, decreased level of mother’s education, mother being as a housewife and having more than two children (Table 1). Regarding the type of CAM which was used, 105 mothers responded to this section. The most common treatment was medicinal herbs (93.3%), oil rub (26.6%) and prayer therapy (25.7%). In terms of the information source for CAM, 196 mothers responded to this section. Family (72%) and neighbors (50%) were the most, while physicians consisted the least (2%) source of the information. Since many of the mothers had selected more than one response about types and information source, the overall percentage in these two sections was over 100%. Only 1.3% of total mothers (300 subjects) knew that complementary medicine may also exert side effects at some extent. Table 1 CAM usage for children based on children and mothers demographic characteristics Use of CAM in the treatment of children No Yes Total P -Value Child sex Girl 94 (65.3%) 50 (34.7%) 144 (100%) 0.74 Boy 99 (63.5%) 57 (36.5%) 156 (100%) Child age < 3 years 132 (66.7%) 66 (33.3%) 198 (100%) 0.24 ≥ 3 years 61 (59.8%) 41 (40.2%) 102 (100%) Number of children ≤ 2 child 137 (74.9%) 46 (25.1%) 183 (100%) <0.001* > 2 child 56 (47.9%) 61 (52.1%) 117 (100%) Maternal employment Housewife 157 (60.2%) 104 (39.8%) 261 (100%) <0.001* Employed 36 (92.3%) 3 (7.7%) 39 (100%) Maternal level of education Illiterate 3 (14.3%) 18 (85.7%) 21 (100%) <0.001* Primary 35 (37.6%) 58 (62.4%) 93 (100%) Middle school 61 (76.2%) 19 (23.8%) 80 (100%) Diploma 80 (95.1%) 6 (4.9%) 86 (100%) University 14 (70%) 6 (30%) 20 (100%)

Maternal employment Housewife 157 (60.2%) 104 (39.8%) 261 (100%) <0.001* Employed 36 (92.3%) 3 (7.7%) 39 (100%) Maternal level of education Illiterate 3 (14.3%) 18 (85.7%) 21 (100%) <0.001* Primary 35 (37.6%) 58 (62.4%) 93 (100%) Middle school 61 (76.2%) 19 (23.8%) 80 (100%) Diploma 80 (95.1%) 6 (4.9%) 86 (100%) University 14 (70%) 6 (30%) 20 (100%) Maternal age (years) 20 to 30 106 (71.1%) 43 (28.9%) 149 (100%) 0.03* 30 to 40 80 (58.8%) 56 (41.2%) 136 (100%) ≥ 40 7 (46.7%) 8 (53.3%) 15 (100%) Discussion Due to increasing rate of CAM application in children worldwide, determination of the rate of CAM application and the factors influencing the usage of it is very important. In the present study, about one-third of mothers have used CAM for their children. This rate was 58.6% and 57% in Turkey[4,5]; 33% at St. Louis[6]; 31% in Nigeria[7] and 12% in Pittsburgh[10]. Unlike other studies[4,5,9] that higher maternal education levels were usually associated with more CAM application, in the present study lower level of maternal education was correlated with higher CAM usage for their children. This could be due to the fact that educated mothers in Birjand have opposite views about using this type of medicine. In this study, there was a significant increase in the use of CAM in children with mother being as housewife, but in a study in Karachi[9], being employed or working as a housewife had no significant influence on the usage of CAM. Significant correlation with mothers being a house wife may be related to this fact that these mothers have low education levels.

ncrease in the use of CAM in children with mother being as housewife, but in a study in Karachi[9], being employed or working as a housewife had no significant influence on the usage of CAM. Significant correlation with mothers being a house wife may be related to this fact that these mothers have low education levels. In this study and In Cardiff[11] and Bass[12] studies, there was no significant association between children's age and gender with CAM usage. Honey followed by herbal tea in Karachi[9], biological products in Nigeria[7], herbal remedies (41%) and prayer therapy (37%) in Michigan[8] and herbal medicine in Turky[4,5], were the most used CAM. In the present study, the most common treatment was herbal medicine. It is due to its special place among the people of this region which is a rich region in terms of herbal medicine. Family and neighbors in the present, Turkey[4] and Karachi[9] studies and friends and neighbors in Nigerian study[7] were the most sources of maternal information for using CAM. In all of these studies physicians had no important role for giving information to people. Since information from family and neighbors may be more confusing rather than it is helpful, people reasonably must receive information and guidance on the efficacy and side effects of CAM from their doctors.

ation for using CAM. In all of these studies physicians had no important role for giving information to people. Since information from family and neighbors may be more confusing rather than it is helpful, people reasonably must receive information and guidance on the efficacy and side effects of CAM from their doctors. In the present study, only 1.3% of mothers were aware of the possible side effects of this treatment. Herbal medicine was the most common treatment in the present study. Since these herbs may have some side effects and interaction with modern medication, it is recommended that pediatricians ask parents about CAM using, warn them regarding the possible risk of using these drugs, and educate them to use the appropriate form. This study has several limitations. Information gathered just in one clinic and that is a hospital clinic which runs with modern medicine. Perhaps, if the study has been performed in a traditional medical center, it would have led to quite different results. Mothers, who refer to a modern medical clinic, may be afraid to be blamed by doctors if they give the actual information about the usage of CAM. This is a native and regional study and the results might be different from another city in a different geographical area.

enter, it would have led to quite different results. Mothers, who refer to a modern medical clinic, may be afraid to be blamed by doctors if they give the actual information about the usage of CAM. This is a native and regional study and the results might be different from another city in a different geographical area. Conclusion Considering the fact that about one third of mothers used CAM modalities, physicians were the least maternal source of CAM information, nearly all mothers were unaware of the side effects of CAM. It is recommended that physicians learn about the CAM to enable them providing information to parents regarding its benefits and disadvantages. It is also highly recommended to promote community knowledge about the proper use of different kinds of CAM. Acknowledgment This study was approved and funded by medicine research center of Birjand University of medical sciences. The author would like to thank all mothers who participated in this study. Conflict of Interest: None

Introduction Patent ductus arteriosus (PDA) is extremely common in very premature infants and untreated symptomatic PDA may be associated with chronic lung disease[1]. Clinical and epidemiological data strongly suggest that infections, either prenatal or nosocomial, and the presence of a patent ductus arteriosus (PDA) play a major role in the neonatal mortality and morbidity[2,3,4]. For this reason, efforts to prevent this complication in low birth weight infants should include an aggressive approach to the prevention and treatment of prenatal and neonatal infections and an early closure of the PDA. Pharmacological closure of PDA with indomethacin or with ibuprofen, that are both prostaglandin inhibitors, has remained the mainstay of treatment in premature infants over the last three decades[5,6]. During the search for an explanation of the interaction between neonatal infection and PDA, is observed that the presence of a systemic infection in the premature infant adversely affects permanent closure of the ductus, often inducing ductal opening after the first week of life and failure to respond to medical treatment with indomethacin[7]. A likely explanation for this interaction is the elevated serum levels of prostaglandins and tumor necrosis factor (TNF) observed in infants with infections. In addition, infants with serious infections frequently have complications that prevent or delay the medical or surgical treatment of the PDA.

th indomethacin[7]. A likely explanation for this interaction is the elevated serum levels of prostaglandins and tumor necrosis factor (TNF) observed in infants with infections. In addition, infants with serious infections frequently have complications that prevent or delay the medical or surgical treatment of the PDA. As a result, the ductus remains open for prolonged periods of time, maintaining an increased pulmonary blood flow, high capillary pressure, and increased lung fluid. Furthermore, when both complications (infection and PDA) occurred at the same time, they produced a synergistic interaction, further increasing the risk for developing chronic lung disease (CLD)[10]. As a consequence of the left-to-right shunting through the PDA, pulmonary blood flow and lung fluid increases, negatively affecting lung function and gas exchange, and thereby increasing the risk for CLD[9]. The presence of a PDA has also been associated with elevated concentrations of myeloperoxidase in the tracheobronchial fluid, suggesting that the increased pulmonary blood flow may result in damage of the pulmonary endothelium and adhesion and migration of polymorphonuclear cells (PMNs) into the lung tissue[7,8]. Considerable biological plausibility thus exists to explain the influence of significant PDA and sepsis on feed tolerance in preterm neonates. PDA and sepsis are possibly markers of prematurity, and a prolonged interval between starting feeding and full enteral nutrition simply reflects the reluctance to start or continue feeding in the presence of such perceived risk factors for food intolerance and necrotizing enterocolitis (NEC)[11,12].

n preterm neonates. PDA and sepsis are possibly markers of prematurity, and a prolonged interval between starting feeding and full enteral nutrition simply reflects the reluctance to start or continue feeding in the presence of such perceived risk factors for food intolerance and necrotizing enterocolitis (NEC)[11,12]. In this study, we compared the efficacy and safety of oral versus intravenous ibuprofen for the pharmacological closure of PDA in low birth weight (LBW) preterm infants. Subjects and Methods The study was designed as a prospective, randomized, single-blinded study. The study was conducted in the neonatal intensive care unit of the University Hospital for Obstetrics and Gynecology ”Koço Gliozheni”, Tirana, Albania, between January 2010 to December 2012. This study was approved by the Medicine University and Neonatology Department. The study enrolled preterm infants with a gestational age 28-32 weeks, birth weight ≤2000g, postnatal age 48-96 hours, respiratory distress syndrome (RDS) treated with mechanical ventilation (CPAP [continuous positive airway pressure] or IPPV [intermittent positive pressure ventilation]) with additional oxygen requirements above 30% and one of the following echocardiographic criteria: a ductal size >1.5 mm, a left atrium-to-aorta ratio >1.4, and a left-to-right shunting of blood in addition to signs of PDA. Several sources have used a similar cutoff in the value of PDA, with inclusion of patients whose ductus’ diameter was more than 1.5 mm, although no general consensus is achieved[4-6].

c criteria: a ductal size >1.5 mm, a left atrium-to-aorta ratio >1.4, and a left-to-right shunting of blood in addition to signs of PDA. Several sources have used a similar cutoff in the value of PDA, with inclusion of patients whose ductus’ diameter was more than 1.5 mm, although no general consensus is achieved[4-6]. 2D (two-dimensional) echocardiography was performed with an ALOKA ultrasound machine (Hitachi), with 5 and 7.5 MHz electronic sector transducers. Gestational age (GA) was assessed by obstetrical dating criteria or, when obstetrical data was inadequate, by Ballard examination. Exclusion criteria were major congenital abnormalities, right–to–left ductal shunting, life-threatening infection, grade 3 or 4 intraventricular hemorrhage, oliguria of less than 1 ml/kg/h during the preceding eight hours, serum creatinine concentration in excess of 1.6 mg/dl, blood urea nitrogen in excess of 60 mg/dl, thrombocyte count of less than 60 000/mm3, clinical bleeding tendency as revealed by hematuria, blood in the gastric aspirate or in the stools, blood in the endotracheal tube aspirate, oozing from venous or capillary puncture sites, hyperbilirubinemia for which exchange transfusion was required and pulmonary hypertension (Fig. 1). Fig. 1 Flow chart of the study

Exclusion criteria were major congenital abnormalities, right–to–left ductal shunting, life-threatening infection, grade 3 or 4 intraventricular hemorrhage, oliguria of less than 1 ml/kg/h during the preceding eight hours, serum creatinine concentration in excess of 1.6 mg/dl, blood urea nitrogen in excess of 60 mg/dl, thrombocyte count of less than 60 000/mm3, clinical bleeding tendency as revealed by hematuria, blood in the gastric aspirate or in the stools, blood in the endotracheal tube aspirate, oozing from venous or capillary puncture sites, hyperbilirubinemia for which exchange transfusion was required and pulmonary hypertension (Fig. 1). Fig. 1 Flow chart of the study All infants who met the inclusion criteria first underwent echocardiography and cranial ultrasonography, after which they were treated with oral ibuprofen (Brufen, Abbot S.r.l; Italy). Ibuprofen in a dose of 10 mg/kg was given via an orogastric tube, flushed with 1 mL of sterile water to ensure delivery of the drug, otherwise an intravenous route was used (Pedea, Orphan Europe; a vial of 2 mL containing 10 mg of ibuprofen), with ibuprofen infused over a 15-minute period with a syringe pump, and the line was subsequently flushed with saline.

via an orogastric tube, flushed with 1 mL of sterile water to ensure delivery of the drug, otherwise an intravenous route was used (Pedea, Orphan Europe; a vial of 2 mL containing 10 mg of ibuprofen), with ibuprofen infused over a 15-minute period with a syringe pump, and the line was subsequently flushed with saline. The two imaging procedures were performed again 24 hours after each ibuprofen dose. When the PDA was still hemodynamically significant, as demonstrated by echocardiography, and there was no evidence of deterioration in brain ultrasonography, a second dose of ibuprofen 5 mg/kg was administered. A third equivalent dose was given after another 24 hours if deemed necessary. Cranial ultrasound was repeated one week after the last ibuprofen dose and again before discharge from the ward. RDS was treated with respiratory support (CPAP, IPPV or with high-frequency ventilation), oxygen supplements, and surfactant (Curosurf, Chiesi, Italy; a vial of 1.5 mL containing 120 mg) was administered intratracheally at the dosage of 100 to 200 mg/kg. Prophylactic antibiotics were started on admission and stopped after five days if blood cultures were negative. Occurrence of any of the following conditions was enough to discontinue treatment: IVH intraventricular hemorrhage (IVH) grade 3–4, renal failure, NEC, and presence of GEB (gastrointestinal bleeding).

RDS was treated with respiratory support (CPAP, IPPV or with high-frequency ventilation), oxygen supplements, and surfactant (Curosurf, Chiesi, Italy; a vial of 1.5 mL containing 120 mg) was administered intratracheally at the dosage of 100 to 200 mg/kg. Prophylactic antibiotics were started on admission and stopped after five days if blood cultures were negative. Occurrence of any of the following conditions was enough to discontinue treatment: IVH intraventricular hemorrhage (IVH) grade 3–4, renal failure, NEC, and presence of GEB (gastrointestinal bleeding). Before and 24 hours after treatment, all patients were evaluated with a complete blood count, renal function tests (serum creatinine level, blood urea nitrogen and urine output), cranial ultrasonography, and echocardiography. All infants continued their current enteral feeding during the treatment. Findings A total of 168 premature infants at gestational age <32 weeks and birth weight <2000g and RDS were admitted to our NICU (neonatal intensive care unit), from January 2010 to December 2012 and underwent an echocardiographic Doppler ultrasound evaluation at the age of 48-96 hours. The entire study protocol was completed for 80 patients due to drop-out related to various reasons (Fig. 1). We had a minimum value of PDA of 1.5 mm (as inclusion criteria) and a maximum of 3.4 mm (average 2.2 mm; standard deviation ±0.6 mm).

Findings A total of 168 premature infants at gestational age <32 weeks and birth weight <2000g and RDS were admitted to our NICU (neonatal intensive care unit), from January 2010 to December 2012 and underwent an echocardiographic Doppler ultrasound evaluation at the age of 48-96 hours. The entire study protocol was completed for 80 patients due to drop-out related to various reasons (Fig. 1). We had a minimum value of PDA of 1.5 mm (as inclusion criteria) and a maximum of 3.4 mm (average 2.2 mm; standard deviation ±0.6 mm). All premature infants that resulted with a PDA during 48-96 hours of life were treated for three consecutive days with three doses of ibuprofen (dosages and routes described above). 24 hours after the third dose, an echocardiography was performed, and if PDA persisted, a second course of ibuprofen treatment with three other doses was given. Patients with persistent PDA even after the second course were surgically treated. As a result, the closure time was 4 days for the first responsive group (53 patients [30 treated orally and 23 intravenously], see graphics below); 7 days for the responsive group undergoing the second course of treatment (15 patients in total); and the time of surgery for completely non-responsive patients (3 patients undergoing ligation).

re time was 4 days for the first responsive group (53 patients [30 treated orally and 23 intravenously], see graphics below); 7 days for the responsive group undergoing the second course of treatment (15 patients in total); and the time of surgery for completely non-responsive patients (3 patients undergoing ligation). After the first course of the treatment, PDA was closed in 30 (83.3%) of the patients assigned to the oral ibuprofen group versus 23 (71.8%) of those enrolled in the intravenous ibuprofen group. Six (16.6%) patients in the oral ibuprofen group required a second course of drug therapy, compared with 9 (28.1%) in the intravenous ibuprofen group. 15 patients needed a second treatment course and they had all (100%) clinical signs of infection and positive blood culture. The cumulative closure rates were higher in both groups, and only three (9.3%) patients in the intravenous ibuprofen group had surgical ligation. There was no reopening of the ductus after closure was achieved. Baseline characteristics were similar between the two groups in the first 96 hours (Table 1). In the evaluation of renal tolerance, none of the patients had oliguria. The serum creatinine levels and plasma blood urea nitrogen after the treatment did not differ significantly between the groups (Table 2).

After the first course of the treatment, PDA was closed in 30 (83.3%) of the patients assigned to the oral ibuprofen group versus 23 (71.8%) of those enrolled in the intravenous ibuprofen group. Six (16.6%) patients in the oral ibuprofen group required a second course of drug therapy, compared with 9 (28.1%) in the intravenous ibuprofen group. 15 patients needed a second treatment course and they had all (100%) clinical signs of infection and positive blood culture. The cumulative closure rates were higher in both groups, and only three (9.3%) patients in the intravenous ibuprofen group had surgical ligation. There was no reopening of the ductus after closure was achieved. Baseline characteristics were similar between the two groups in the first 96 hours (Table 1). In the evaluation of renal tolerance, none of the patients had oliguria. The serum creatinine levels and plasma blood urea nitrogen after the treatment did not differ significantly between the groups (Table 2). Discussion Intravenous ibuprofen is not available in most countries (and in our country too), and is more expensive than the oral form. If oral ibuprofen was as efficient as intravenous ibuprofen with no greater adverse effects, the more simple administration and lower cost would be important advantages. Our study was designed with sufficient power for determining whether oral and intravenous ibuprofen treatments are equally efficacious and safe in PDA closure in premature infants with RDS. Our results showed oral ibuprofen to be effective and safe in PDA closure, with 30 of our 36 (83.3%) study infants achieving a successful outcome. The rate of closure in the group assigned to intravenous ibuprofen was similar to rates previously reported[4,11]. Some trials on the use of oral ibuprofen for closure of PDA have been recently published[14,15,21].

ffective and safe in PDA closure, with 30 of our 36 (83.3%) study infants achieving a successful outcome. The rate of closure in the group assigned to intravenous ibuprofen was similar to rates previously reported[4,11]. Some trials on the use of oral ibuprofen for closure of PDA have been recently published[14,15,21]. Table 1 Baseline characteristics of the low birth weight preterm infants in oral vs intravenous ibuprofen therapy Variable Oral (n = 36) Intravenous (n= 32) Gestational age, weeks 28.1 – 30 weeks 19 (52.7%) 18 (56.2%) 30.1 – 32 weeks 17 (47.2%) 14 (43.7%) Birth weight, grams <750g n (%) 2 (5.5%) 0 (0%) 751-1000g n (%) 7 (19.4%) 6 (18.7%) 1001-1500g n (%) 15 (41.6%) 19 (59.3%) 1501-2000g n (%) 12 (33.3%) 7 (21.8%) Gender Male, n (%) 22 (61.1%) 15 (46.8%) Female, n (%) 14 (38.8%) 17 (53.1%) Delivery by cesarean section, n (%) 20 (55.5%) 14 (43.7%) Antenatal glucocorticoid treatment, number and percentage (%) 28 (77.7%) 18 (56.2%) Perinatal asphyxia, n (%) 11 (30.5%) 9 (28.1%) Table 2 Biochemical values in two group low birth weight preterm infants in oral vs intravenous ibuprofen therapy Parameters Oral group (n=36) Intravenous group (n= 32) P. value Plasma blood urea nitrogen (mg/dl) (mean ± SD) Day 1 30.7 (14.8 ) 30.4 (13.7) .90 Day 2 30.3 (14.2) 30.6 (14.0) .89 Day 3 30.88 (7.76) 31.66 (9.90) .68 Mean plasma creatinine (mg/dL) (mean±SD) Day 1 1.07 (0.24) 1.09 (0.24) .06 Day 2 1.20 (0.95) 0.97 (0.45) .07 Day 3 0.76 (0.48 ) 0.79 (0.46) .07 Oligo/anuria (ml/kg/h) Day 1 0 (0%) 0 (0%) Day 2 0 (0%) 0 (0%) Day 3 0 (0%) 0 (0%) Infection and need for a second treatment course 6 (16.6%) 9 (28.1%)

Plasma blood urea nitrogen (mg/dl) (mean ± SD) Day 1 30.7 (14.8 ) 30.4 (13.7) .90 Day 2 30.3 (14.2) 30.6 (14.0) .89 Day 3 30.88 (7.76) 31.66 (9.90) .68 Mean plasma creatinine (mg/dL) (mean±SD) Day 1 1.07 (0.24) 1.09 (0.24) .06 Day 2 1.20 (0.95) 0.97 (0.45) .07 Day 3 0.76 (0.48 ) 0.79 (0.46) .07 Oligo/anuria (ml/kg/h) Day 1 0 (0%) 0 (0%) Day 2 0 (0%) 0 (0%) Day 3 0 (0%) 0 (0%) Infection and need for a second treatment course 6 (16.6%) 9 (28.1%) Need for surgical ligation 0 (0%) 3 (9.3%) All studies had small sample size. Aly[16] in a randomized pilot study, reported that PDA was closed in 7 of 9 premature (≤35 weeks) infants given oral ibuprofen and in 10 of 12 premature infants given intravenous indomethacin.

Infection and need for a second treatment course 6 (16.6%) 9 (28.1%) Need for surgical ligation 0 (0%) 3 (9.3%) All studies had small sample size. Aly[16] in a randomized pilot study, reported that PDA was closed in 7 of 9 premature (≤35 weeks) infants given oral ibuprofen and in 10 of 12 premature infants given intravenous indomethacin. Fakhraee[22] in a randomized study, reported that PDA was closed in all of 18 premature (≤34 weeks) infants given oral ibuprofen and in 15 of 18 premature infants given oral indomethacin (P>.05). Efficacy of oral ibuprofen compared with intravenous indomethacin, was reported by Supapannachart et al[23] and Chotigeat et al[24] as well. In nonrandomized open trials, Heyman et al[25] and Cherif et al[18] reported a ductal closure with oral ibuprofen respectively in 21 (95.4%) of 22 patients, 38 (95%) of 40 patients, and in 11 (84.6%) of 13 patients. The authors concluded that oral ibuprofen might constitute a feasible alternative in the treatment of PDA. Van Overmeire et al studied the efficacy of indomethacin and ibuprofen given to larger premature (≤32 weeks) infants at the age of 2-4 days. They reported that the closure rate was similar (66% and 70%, respectively) after the first course and that there was no significant difference in side effects, although ibuprofen was associated with significantly less impairment of renal function[11,25]. Two studies increase the number of infants randomized and expand the information about the safety and efficacy of oral ibuprofen in more mature VLBW (very low birth weight) infants[19,20]. We hope the same for our study.

cts, although ibuprofen was associated with significantly less impairment of renal function[11,25]. Two studies increase the number of infants randomized and expand the information about the safety and efficacy of oral ibuprofen in more mature VLBW (very low birth weight) infants[19,20]. We hope the same for our study. Since renal tolerability of ibuprofen on renal function in the neonate is a major argument in favor of its use in the treatment of PDA[19,20] our study expands our information about the safety and efficacy of oral ibuprofen in more mature VLBW infants. Serum creatinine and urea levels in our patients were within normal range at all times, so there was no contraindication for a second dose of ibuprofen when it was needed. This might be an explanation for the higher rate of pharmacologic ductal closure observed in our study. Gonzales at al showed that late PDA episodes were more frequent in infants with infection than in those without infection and were associated with an increased risk of PDA closure failure. Furthermore, when both factors were temporally related, they further increased the risk of CLD[5,7].

Serum creatinine and urea levels in our patients were within normal range at all times, so there was no contraindication for a second dose of ibuprofen when it was needed. This might be an explanation for the higher rate of pharmacologic ductal closure observed in our study. Gonzales at al showed that late PDA episodes were more frequent in infants with infection than in those without infection and were associated with an increased risk of PDA closure failure. Furthermore, when both factors were temporally related, they further increased the risk of CLD[5,7]. There are several limitations to our study. This was an open-label, one-arm study, and the physicians and nurses were aware of the nature of the study, although the cardiologist who supervised the echocardiographic studies was blinded as with regard to the status of the infants, and whether they were treated with oral ibuprofen or intravenous ibuprofen. On the other hand, being not a hemodynamic study, we could not obtain pressure values of the heart chambers (systolic and diastolic pressures, TR pressure gradients, pulmonary artery pressure); such a data collection would have surely enhanced the value of our conclusions. The exclusion of PDA patients whose ductal size was less than 1.5 mm might be another limitation, although several sources use such a cutoff[4-6]. This is not simply related to the fact that a smaller ductal size might be not important hemodynamically, and other authors do consider, although through the denomination ‘mild’, the hemodynamic importance even of a ductal size of less than 1.5 mm[26]. The problem of visualizing a patent ductus and thus differentiating it from a closed ductus when values are less than 1.5 mm remains however, a technical challenge[27]. This technicality might be overcome through using a 3D (three-dimensional) echocardiography, with higher spatial resolution and a better diagnostic accuracy[28].

m of visualizing a patent ductus and thus differentiating it from a closed ductus when values are less than 1.5 mm remains however, a technical challenge[27]. This technicality might be overcome through using a 3D (three-dimensional) echocardiography, with higher spatial resolution and a better diagnostic accuracy[28]. This is the first experience that we have had in Albania with ibuprofen (oral or intravenous) regarding the treatment of PDA in preterm infants. Conclusion Our data indicate that, for preterm infants especially for LBW infants, the rate of early ductal closure was comparable and the adverse effects were fewer with oral ibuprofen in comparison to the intravenous route. Association of PDA with perinatal infection has a negative impact on the pharmacological closure of the ductus, increasing need for a second course and for surgical ligation. The oral form was as safe as the intravenous form in terms of renal tolerance. Larger comparative studies are needed to validate these findings. Conflict of Interest: None Authors’ Contribution E. Pistulli concept and manuscript preparation, data analysis and interpretation. A. Hamiti, S. Buba, A. Hoxha and N. Kelmendi acquisition of data, data analysis and interpretation. G. Vyshka manuscript preparation and critical revision of the manuscript. All authors have approved the final version of the paper.

Introduction Lymphoblastic lymphoma (LBL) is a highly malignant tumor of variegated lymphoid reticuloendothelial cells, one common histologic subtype of non-Hodgkin’s lymphoma in childhood[1]. It is clear that along with the development of multimodality therapy, the prognosis of children with LBL have been markedly improved, where L-asparaginase is an important and universal component in chemotherapy[2,3]. It can significantly improve long-term event-free survival[4,5]. However, asparaginases are associated with a unique set of side effects[3]. Generally, asparaginase is cleared rapidly with an apparent half life of about 20 hours[6]. In previous clinical trials, the drug had to be administered intramuscularly 6-10 times every other day for maintaining its effective drug activity. Moreover, clinical hypersensitivity reactions and silent inactivation due to antibodies against E. coli-asparaginase, led to inactivation of E. coli-asparaginase in up to 60% of cases[3,7]. Polyethylene glycosylated–asparaginase (pegas-pargase), formed by covalently attaching polyethylene glycol to the native Escherichia coli enzyme, was developed for reducing the immunogenic potential. After one single dose of this drug, the potential therapeutic enzyme activity can be maintained for at least 2 weeks[8,9]. For these reasons, we explored to use pegaspargase instead of L-asparaginase to treat children with advanced-stage LBL in order to improve patient’s treatment compliance and reduce the risk of acute anaphylactic reaction. Subjects and Methods Patients

Introduction Lymphoblastic lymphoma (LBL) is a highly malignant tumor of variegated lymphoid reticuloendothelial cells, one common histologic subtype of non-Hodgkin’s lymphoma in childhood[1]. It is clear that along with the development of multimodality therapy, the prognosis of children with LBL have been markedly improved, where L-asparaginase is an important and universal component in chemotherapy[2,3]. It can significantly improve long-term event-free survival[4,5]. However, asparaginases are associated with a unique set of side effects[3]. Generally, asparaginase is cleared rapidly with an apparent half life of about 20 hours[6]. In previous clinical trials, the drug had to be administered intramuscularly 6-10 times every other day for maintaining its effective drug activity. Moreover, clinical hypersensitivity reactions and silent inactivation due to antibodies against E. coli-asparaginase, led to inactivation of E. coli-asparaginase in up to 60% of cases[3,7]. Polyethylene glycosylated–asparaginase (pegas-pargase), formed by covalently attaching polyethylene glycol to the native Escherichia coli enzyme, was developed for reducing the immunogenic potential. After one single dose of this drug, the potential therapeutic enzyme activity can be maintained for at least 2 weeks[8,9]. For these reasons, we explored to use pegaspargase instead of L-asparaginase to treat children with advanced-stage LBL in order to improve patient’s treatment compliance and reduce the risk of acute anaphylactic reaction. Subjects and Methods Patients Between 2000 and 2006, children younger than 16 years who had a previously untreated LBL on admission to our hospital with advanced features, defined as stage Ⅲ and stage Ⅳ diseases on the St Jude staging system, were eligible for the trial. The Institutional Review Boards approved the protocol before enrollment. Written informed consents were obtained from their parents or legal guardians before starting therapy based on the BFM-95 protocol.

anced features, defined as stage Ⅲ and stage Ⅳ diseases on the St Jude staging system, were eligible for the trial. The Institutional Review Boards approved the protocol before enrollment. Written informed consents were obtained from their parents or legal guardians before starting therapy based on the BFM-95 protocol. Pretreatment evaluation of stage and diagnosis The initial diagnostic workup for LBL included a detailed physical examination and bone marrow aspiration, computed tomography (CT) scan of the chest and abdomen, bone scintigraphy, immunophenotyping study and examination of the cerebrospinal fluid (CSF). Patients were classified according to the St Jude staging system[10]. Study Design All patients were randomly assigned to receive treatment with either BFM-95 protocol or modified BFM-95 protocol. In modified BFM-95 protocol L-asparaginase was replaced by pegaspargase, administered by intramuscular injection (2.500 IU/m2/d) on days 12, 28 in induction and on day 8 in reinduction phase, which replaced 8 doses of L-asparaginase (10000 U/m2/d) on days 12, 15, 18, 21, 24, 27, 30 and 33 in induction and 4 doses on days 8, 11, 15 and 18 in reinduction, respectively, as defined by the Non-Hodgkin Lymphoma– Berlin-Frankfurt-Munster-95 (NHL-BFM-95) protocol[2]. No single injection dose was more than 2.0 mL (1500 IU) at one injection site. The doses and administration modes of the other chemotherapeutic agents were the same as in BFM-95 protocol. Response definitions and toxicity assessment

All patients were randomly assigned to receive treatment with either BFM-95 protocol or modified BFM-95 protocol. In modified BFM-95 protocol L-asparaginase was replaced by pegaspargase, administered by intramuscular injection (2.500 IU/m2/d) on days 12, 28 in induction and on day 8 in reinduction phase, which replaced 8 doses of L-asparaginase (10000 U/m2/d) on days 12, 15, 18, 21, 24, 27, 30 and 33 in induction and 4 doses on days 8, 11, 15 and 18 in reinduction, respectively, as defined by the Non-Hodgkin Lymphoma– Berlin-Frankfurt-Munster-95 (NHL-BFM-95) protocol[2]. No single injection dose was more than 2.0 mL (1500 IU) at one injection site. The doses and administration modes of the other chemotherapeutic agents were the same as in BFM-95 protocol. Response definitions and toxicity assessment Complete response (CR) was defined as no evidence of tumor by physical examination and imaging studies (CT scans or magnetic resonance imaging). Bone marrow aspirate was needed. Patients were considered in partial response (PR) if there was a decrease of 50% or more in all measurable mass lesions. No response (NR) was defined as less than 50% reduction in the extent of disease[11]. Event-free survival (EFS) was defined as the interval between diagnosis and disease progression, relapse, or death; and overall survival (OS) was defined as the interval between diagnosis and death from any cause or last contact. During treatment, patients were closely monitored on peripheral blood cell counts, major organ functions (liver, renal and cardiac functions), serum electrolytes, and coagulation functions, etc. All toxicity were collected and graded based on the National Cancer Institute Common Terminology Criteria for Adverse Events, version 4.0[12].

patients were closely monitored on peripheral blood cell counts, major organ functions (liver, renal and cardiac functions), serum electrolytes, and coagulation functions, etc. All toxicity were collected and graded based on the National Cancer Institute Common Terminology Criteria for Adverse Events, version 4.0[12]. After completion of chemotherapy, patients entered the follow-up and received the periodic reevaluation on biochemical and imaging studies. Statistical Design and Analysis The primary outcome measures for this study were EFS and OS. Life-table estimates of survival time were calculated by the method of Kaplan and Meier for outcome comparison. The chi-square test was used to analyze significance between the two groups, comparing the treatment toxicity. Outcome was assigned to the randomized treatment, regardless of the therapy received. P<0.05 was considered significant. Finding Patients Between June 2000 and December 2006, 57 patients were assigned to this study. Table 1 Clinical characteristics of the 54 patients at presentation Variable Characteristic No Patients (%) Sex Female 19 (35.2) Male 35 (64.8) Immunophenotyping T-lineage 40 (74.1) B-lineage 14 (25.9) Stage category stage Ⅲ 30 (55.6) stage Ⅳ 24 (44.4) Chemotherapy NHL-BFM-95 protocol 23 (42.6) modified NHL-BFM-95 protocol 31 (57.4) NHL-BFM-95 protocol: Non-Hodgkin Lymphoma– Berlin-Frankfurt-Munster-95 protocol

Table 1 Clinical characteristics of the 54 patients at presentation Variable Characteristic No Patients (%) Sex Female 19 (35.2) Male 35 (64.8) Immunophenotyping T-lineage 40 (74.1) B-lineage 14 (25.9) Stage category stage Ⅲ 30 (55.6) stage Ⅳ 24 (44.4) Chemotherapy NHL-BFM-95 protocol 23 (42.6) modified NHL-BFM-95 protocol 31 (57.4) NHL-BFM-95 protocol: Non-Hodgkin Lymphoma– Berlin-Frankfurt-Munster-95 protocol Three patients were excluded because lack of complete documentation. Mean age of the remaining 54 patients was 5.1±2.58 (95%CI, 4.40 to 5.80) years, ranging from 2.5 years to 14.7 years. On immunophenotyping study, the tumor cells were classified as T-lineage in 40 patients and B-lineage in 14 patients. 17 stage Ⅲ patients and 6 stage Ⅳ patients were assigned to BFM-95 group (B-lineage 5, T-lineage 18), while 13 stage Ⅲ patients and 18 stage Ⅳ patients were assigned to modified BFM-95 group (B-lineage 9, T-lineage 22). There was no significant difference on histologic subtypes (P>0.05). The clinical detailed characteristics of the remaining 54 patients are listed in Table 1. Treatment outcomes

Three patients were excluded because lack of complete documentation. Mean age of the remaining 54 patients was 5.1±2.58 (95%CI, 4.40 to 5.80) years, ranging from 2.5 years to 14.7 years. On immunophenotyping study, the tumor cells were classified as T-lineage in 40 patients and B-lineage in 14 patients. 17 stage Ⅲ patients and 6 stage Ⅳ patients were assigned to BFM-95 group (B-lineage 5, T-lineage 18), while 13 stage Ⅲ patients and 18 stage Ⅳ patients were assigned to modified BFM-95 group (B-lineage 9, T-lineage 22). There was no significant difference on histologic subtypes (P>0.05). The clinical detailed characteristics of the remaining 54 patients are listed in Table 1. Treatment outcomes Patients’ treatment responses were evaluated on day 33 of induction and at the end of chemotherapy[13]. The CR rate was 97% in the modified BFM-95 group and 94% in the BFM-95 group on day 33 of induction. The 5-year overall survivals were 88%±8% (95% CI, 72.31% to 103.68%) with BFM-95 versus 91%±6% (95%CI, 79.24% to 102.76%) with modified BFM-95 (Fig 1), and the event-free survival rates were 80%±9% (95%CI, 62.36% to 97.64%) with BFM-95 versus 87±7%(95%CI, 73.28% to 100.72%) with modified BFM-95 (Fig 2)(median follow-up time, 5.75±1.33 years) (95%CI, 5.38 to 6.12) (Table 2). Patients with recurrence or progressive disease

Patients’ treatment responses were evaluated on day 33 of induction and at the end of chemotherapy[13]. The CR rate was 97% in the modified BFM-95 group and 94% in the BFM-95 group on day 33 of induction. The 5-year overall survivals were 88%±8% (95% CI, 72.31% to 103.68%) with BFM-95 versus 91%±6% (95%CI, 79.24% to 102.76%) with modified BFM-95 (Fig 1), and the event-free survival rates were 80%±9% (95%CI, 62.36% to 97.64%) with BFM-95 versus 87±7%(95%CI, 73.28% to 100.72%) with modified BFM-95 (Fig 2)(median follow-up time, 5.75±1.33 years) (95%CI, 5.38 to 6.12) (Table 2). Patients with recurrence or progressive disease A total of 8 randomly assigned patients had a documented relapse or disease progression: 4 (9%) in the BFM-95 group and 4 (10%) in the modified BFM-95 group. Two patients in the BFM-95 group and three in the modified BFM-95 group died. Five deaths in both groups were due to disease relapse or progression. Three remaining relapsed patients, two with pre-T LBL and one with pre-B LBL, were alive. Toxicity

A total of 8 randomly assigned patients had a documented relapse or disease progression: 4 (9%) in the BFM-95 group and 4 (10%) in the modified BFM-95 group. Two patients in the BFM-95 group and three in the modified BFM-95 group died. Five deaths in both groups were due to disease relapse or progression. Three remaining relapsed patients, two with pre-T LBL and one with pre-B LBL, were alive. Toxicity Among 31 patients receiving Pegaspargase during induction and reinduction phases, stage 4 myelosuppression was the most common complication. Besides, one experienced grade 3, the other four grade 2 coagulation defects. The patient with grade 3 coagulation defects had continuous capillary hemorrhage at the injection spot and prolonged APTT, the plasma FIX was lower than 30% of the normal values and the fibrinogen level was decreased to 1.0 g/L. The other four patients developing grade 2 coagulation defects had decreased fibrinogen levels which was lowered to 1.0 g/L, without any clinical symptoms for about two weeks. One developed grade 3 anaphylactic reaction (symptomatic broncho-spasm and urticaria). In addition, there was another grade 2 complication in one patient, who developed mild pancreatitis without any symptom. Table 2 Treatment outcomes according to stage and randomized treatment regimen Regimen 5-year Event-free survival 5-year overall survival Complete response NHL-BFM-95 83% 91% 94% Modified NHL-BFM-95 87% 90% 97% NHL-BFM-95 protocol: Non-Hodgkin Lymphoma– Berlin-Frankfurt-Munster-95 protocol; Table 3 Toxicity differences between the two protocols Modified-BFM-95 BFM-95 P value Total 31 23

Table 2 Treatment outcomes according to stage and randomized treatment regimen Regimen 5-year Event-free survival 5-year overall survival Complete response NHL-BFM-95 83% 91% 94% Modified NHL-BFM-95 87% 90% 97% NHL-BFM-95 protocol: Non-Hodgkin Lymphoma– Berlin-Frankfurt-Munster-95 protocol; Table 3 Toxicity differences between the two protocols Modified-BFM-95 BFM-95 P value Total 31 23 Stage 4 myelosupression 31 100% 23 100% Coagulation defects Grade 2 4 12.9% Grade 2 2 8.7% 0.627 Grade 3 1 3.2% Grade 3 - - 0.385 Anaphylactic reaction Grade 3 1 3.2% Grade 3 5 21.7% 0.032 Grade 4 - - Grade 4 1 4.3% 0.241 Pancreatitis Grade 2 2 6.5% Grade 2 2 8.7% 0.756 Increased pancreatic amylases were only found on routine screening. All of them achieved complete recovery after treatment. No hemorrhage or thrombosis was seen. There was no severe complication during the sequential phases. Among other 23 patients receiving BFM-95 protocol, beside the myelosuppression, complication of anaphylactic reaction was noted. Five of 23 patients experienced grade 3 anaphylactic reaction (urticarial lesions covering larger than 30% of body surface) during reinduction, including one experiencing a severe anaphylactic shock. Two experienced grade 2 pancreatitis without any symptoms. Two developed grade 2 coagulation defects, whose fibrinogen level was decreased to 1.2 g/L for 3-5 days without prolonged PT or APTT. There is no significant difference regarding coagulation defects of the two groups (P<0.05). The detailed data are shown in Table 3.

xperienced grade 2 pancreatitis without any symptoms. Two developed grade 2 coagulation defects, whose fibrinogen level was decreased to 1.2 g/L for 3-5 days without prolonged PT or APTT. There is no significant difference regarding coagulation defects of the two groups (P<0.05). The detailed data are shown in Table 3. Discussion Until now, most studies were associated with evaluating Pegaspargase on treating children with ALL, rather than lymphoma. As the common regimen for LBL is similar to leukemia’s, we attempted to use Pegaspargase to treat children with advanced-stage LBL, and the outcome showed that the response to modified BFM-95 protocol was similar to the previously reported[14,15]. To our knowledge, this is the first trial to treat LBL children with Pegaspargase. The result of this study indicated that Pegasparagse can be administered safely to children in combination with other chemotherapeutic agents. For both groups, stage 4 myelosuppression was the most common complication. However, it was considered to be mostly associated with the myelosuppressive activity of the other cytotoxic chemotherapy drugs.

dicated that Pegasparagse can be administered safely to children in combination with other chemotherapeutic agents. For both groups, stage 4 myelosuppression was the most common complication. However, it was considered to be mostly associated with the myelosuppressive activity of the other cytotoxic chemotherapy drugs. Of the 31 patients treated with Pegaspargase, only one experienced a grade 3 coagulation disorder, APTT was prolonged and the patient had continuous capillary hemorrhage at the injection spot. The study of coagulation factors revealed that the plasma FIX was lower than 30% of the normal values and the fibrinogen level was decreased to 1.0 g/L. Interestingly, after administering prothrombin complex concentrate (PCC) (20 IU/kg/d), vitamin K1 (5 mg/d), and fibrinogen (0.5g/d), APTT was still prolonged and getting worse and worse until plasma infusion was done. This indicated that coagulation disorder in patients receiving Pegaspargase was more sensitive to response to plasma infusion rather than administering PCC, which meant Pegaspargase might influence other coagulation factors such as FII and FVII.

till prolonged and getting worse and worse until plasma infusion was done. This indicated that coagulation disorder in patients receiving Pegaspargase was more sensitive to response to plasma infusion rather than administering PCC, which meant Pegaspargase might influence other coagulation factors such as FII and FVII. As we know, asparaginase preparations interfere with hepatic production of both coagulant and anti-coagulant proteins. Rytting reported that the incidence of thrombosis was generally 10% in pediatric patients[16]. Pancreatitis and thrombotic complications are less common in children than in adolescents and adults. In our study, no thrombosis was noted either in modified BFM-95 group or BFM-95 group. The incidences of other complications were similar to previously reported[11,16]. The major adverse reaction noted in the L-asparaginase group in our study was anaphylactic reaction. Because it is a bacterially derived protein, L-asparaginase can often induce anaphylaxis and immune responses which may lead to development of specific antibody, resulting in rapid elimination of enzyme and losing pharmacological activity. Development of the specific antibody of L-asparaginase is common, it was reported in over 50% of patients treated with multiple administrations of L-asparaginase[17,18].

and immune responses which may lead to development of specific antibody, resulting in rapid elimination of enzyme and losing pharmacological activity. Development of the specific antibody of L-asparaginase is common, it was reported in over 50% of patients treated with multiple administrations of L-asparaginase[17,18]. Anaphylactic reaction in the Pegasparagse group compared to L-asparaginase group is quite rare. Only 1 patient suffered from it. It was in accordance with some other studies in which Pegaspargase had a relatively lower immunogenicity due to the covalent conjugation to monomethoxy polythlene glycol and used to replace L-asparaginase in patients who had developed allergic reaction. In adults with newly diagnosed acute lymphoblastic leukemia, incidence of allergic reaction to Pegasparagse was strikingly lower compared to L-asparaginase, ranging from 0～15%[4,9,16,15,19], in pediatric patients it was also lower, around 9%[20]. In our study, an interesting thing was that all anaphylactic reactions were observed during induction with Pegasparagse, and no such a reaction was seen in reinduction. It was in contrast to L-asparaginase. The frequency of complications would increase with continued L-asparaginase treatment. Maybe, Pegasparagse could easily cause immune tolerance because of the less frequency of administration. On the number of observed patients, we could not rule out the possibility of coincidence. Further clinical observation is needed.

he frequency of complications would increase with continued L-asparaginase treatment. Maybe, Pegasparagse could easily cause immune tolerance because of the less frequency of administration. On the number of observed patients, we could not rule out the possibility of coincidence. Further clinical observation is needed. Recently, Children’s Oncology Group applied Erwinia asparaginase as an alternative drug in case of hypersensitivity to Pegaspargase, the outcome was excellent[21]. However, Erwinia asparaginase is not commercially available now.

he frequency of complications would increase with continued L-asparaginase treatment. Maybe, Pegasparagse could easily cause immune tolerance because of the less frequency of administration. On the number of observed patients, we could not rule out the possibility of coincidence. Further clinical observation is needed. Recently, Children’s Oncology Group applied Erwinia asparaginase as an alternative drug in case of hypersensitivity to Pegaspargase, the outcome was excellent[21]. However, Erwinia asparaginase is not commercially available now. In CALGB 9511 clinical trial, pegaspargase was used in lieu of the native enzyme, the aim was to compare the differences on overall survival and disease-free survival between patients who did and did not achieve asparagine depletion. They concluded that effective asparagine depletion with Pegaspargase was feasible as part of an intensive multiagent therapeutic regimen in adult ALL and appeared associated with improved outcomes[16]. Our study demonstrated that the antitumor activity of Pegaspargase is comparable to L-asparaginase. Neither the EFS nor OS was significantly different between the groups. Moreover, we observed that coagulation disorder was the most common complication in patients treated with modified BFM-95 protocol (there was no significant difference between the 2 groups, P>0.05), while anaphylactic reaction was the major complication in patients treated with BFM-95 protocol (there was significant difference between them, P<0.05). In all of 31 patients who received Pegaspargase, although a few patients experienced some complications, it was well tolerated and the patients recovered soon after associated symptomatic treatment.

the major complication in patients treated with BFM-95 protocol (there was significant difference between them, P<0.05). In all of 31 patients who received Pegaspargase, although a few patients experienced some complications, it was well tolerated and the patients recovered soon after associated symptomatic treatment. Conclusion Obviouslly, our study showed that L-asparaginase could be replaced by Pegaspargase safely to treat children with advanced-stage LBL with enhancing patient’s compliance to chemotherapy and without decreasing the OS and EFS. Even though our clinical trial was a small sample clinical trial, it can indicate the benefits of Pegaspargase. Further study is needed to evaluate the long-term outcome of Pegaspargase. Acknowledgment We thank Dr. William Henery Meyer for helpful discussion and editing. Conflict of Interest: None Authors Contribution Y. Zhang: conceptualized and designed the study, drafted the initial manuscript J. Chang: initial analyses, Critical revision of the manuscript L. Feng: designed the data collection instruments, and coordinated and supervised data collection, critically reviewed the manuscript X. Zhong: designed the data collection instruments and coordinated and supervised data collection, critically reviewed the manuscript L. Wang: designed the data collection instruments, and coordinated and supervised data collection, critically reviewed the manuscript All authors approved the final manuscript as submitted.

Introduction Marshall Syndrome was first defined by Marshal in 1987[1]. The acronym FAPA (fever, aphthous stomatitis, pharyngitis, cervical adenitis) was was later renamed to PFAPA (periodic fever, aphthous stomatitis, pharyngitis, and cervical adenitis)[2]. This clinical syndrome occurs among children less than 5 years old. It is a sporadic disease with periodic attacks of inflammation[3]. Periodic fevers usually last 3 to 6 days and occur with regular intervals about every 3 to 6 weeks. The diagnosis is established on the basis of clinical criteria that require the presence of a recurrent fever of early onset (<5 years) and ≥1 of the 3 associated symptoms (aphthosis, cervical adenitis, and pharyngitis) in the absence of upper respiratory tract infections and cyclic neutropenia[2]. Between episodes, the children usually are well. The exact cause of PFAPA is unknown and autoimmune or infectious processes in its pathogenesis are not proven. Moreover no geographical or ethnic tendencies have been shown. WBC and ESR are increased during acute episodes[4]. Because a single dose of corticosteroid can resolve attacks, inflammatory cytokines production in response to infectious factors is considered as a leading cause of the syndrome. IFN-γ, TNF and IL-6 increase during episodes[5,6]. It is more common in males and the prognosis is very good[7,8]. PFAPA has been reported in adult patients too[9,10]. Although PFAPA is a periodic fever, no specific gene mutation is known for it. Nevertheless it is possible that genes involved in other inflammatory diseases play a role in its pathogenesis[2].

The exact cause of PFAPA is unknown and autoimmune or infectious processes in its pathogenesis are not proven. Moreover no geographical or ethnic tendencies have been shown. WBC and ESR are increased during acute episodes[4]. Because a single dose of corticosteroid can resolve attacks, inflammatory cytokines production in response to infectious factors is considered as a leading cause of the syndrome. IFN-γ, TNF and IL-6 increase during episodes[5,6]. It is more common in males and the prognosis is very good[7,8]. PFAPA has been reported in adult patients too[9,10]. Although PFAPA is a periodic fever, no specific gene mutation is known for it. Nevertheless it is possible that genes involved in other inflammatory diseases play a role in its pathogenesis[2]. In Cazeneuve’s study among 12 allele analysis only one mutation of M694V was found[11]. Berkun study showed that in PFAPA patients with MEFV gene mutation the disease was less severe and aphthous stomatitis, duration of disease, number of attacks, and need to steroid were less common. Therefore they found MEFV gene effect as a modifier on PFAPA[12]. In a study on 393 children with PFAPA, the clinical features were compared in genetically negative and positive results, 82 had positive genetic results, 75 uncertain and 236 negative results. Jn patients with positive genetic results diarrhea, vomiting, arthralgia and rash was more frequent. In the genetically negative group exudative pharyngitis was encountered more frequently[13].

compared in genetically negative and positive results, 82 had positive genetic results, 75 uncertain and 236 negative results. Jn patients with positive genetic results diarrhea, vomiting, arthralgia and rash was more frequent. In the genetically negative group exudative pharyngitis was encountered more frequently[13]. Gattorno et al evaluated the relationship between PFAPA and other periodic fevers such as FMF, MVK, and TRAPS. They found a close relationship between PFAPA and other periodic fever syndromes[14]. FMF is an autosomal recessive disease characterized by acute attacks of fever and polyserositis. This disease mainly occurs in Mediterranean population and is caused by MEFV gene mutations[2]. This study was designed to analyse 12 most common MEFV gene mutations in PFAPA patients and also evaluating Gaslini score test based on detected mutations. Subjects and Methods Twenty one patients who had diagnostic criteria were enrolled in this study. Twelve common MEFV gene mutations were evaluated: P369S, F479L, M680I(G/C), M680I(G/A), I692del, M694V, M694I, K695R, V726A, A744S, R761H, E148Q. All the patients were screened for 12 common MEFV mutations by a reverse hybridization assay (FMF StripAssay, Vienna lab, Vienna, Austria) according to the instructions provided by the manufacturer. Statistical analysis was performed using SPSS 16.0. Comparison between the different genotypes was assessed using chi-square test. A P-value <0.05 was accepted as statistically significant.

erse hybridization assay (FMF StripAssay, Vienna lab, Vienna, Austria) according to the instructions provided by the manufacturer. Statistical analysis was performed using SPSS 16.0. Comparison between the different genotypes was assessed using chi-square test. A P-value <0.05 was accepted as statistically significant. Findings The age of patients was between 6 months and 14 years, and 15 were males. The mean age at onset of symptoms was 1.52±2.72 years (range 6 months - 5 years). The mean duration of fever was 3.57±1.64 days, and that of the disease 4.23±1.56 days. The interval of the attacks was 32.52±20.53 days. All of the patients had fever and 12 (57.14%) patients had abdominal pain, 19 (90.47%) children were breast fed infants. In family history one patient's brother, one patient's father and one patient's mother had FMF. Eight (38.09%) patients had MEFV gene mutations, seven of them were heterozygotes and one was combined heterozygous (K695R, V725A). (Table 1). Although abdominal pain, aphthous stomatitis and duration of attacks were more common in non-mutated patients, these findings in association with episodes of fever and response to treatment did not show meaningful difference between the two groups (P≤0.05). Table 1 Prevalence of MEFV gene mutations MEFV Gene Prevalence Percent M694V 4 45 V726A 3 33 K694R 1 11 E148Q 1 11 Table 2 Clinical findings in different studies Signs and symptoms Our study % Gattorno [ 14 ] % Stojanov [ 15 ] % Padeh [ 8 ] % Dagan[ 16 ] % Fever 100 -- 78 100 -- Weakness and lethargy 71.42 -- -- 100 -- Skin rash 14.28 24 18 -- -- Headache 28.6 46 6.5 17.8 -- Oral aphthosis 47.61 63 13 67.8 33.3

Prevalence Percent M694V 4 45 V726A 3 33 K694R 1 11 E148Q 1 11 Table 2 Clinical findings in different studies Signs and symptoms Our study % Gattorno [ 14 ] % Stojanov [ 15 ] % Padeh [ 8 ] % Dagan[ 16 ] % Fever 100 -- 78 100 -- Weakness and lethargy 71.42 -- -- 100 -- Skin rash 14.28 24 18 -- -- Headache 28.6 46 6.5 17.8 -- Oral aphthosis 47.61 63 13 67.8 33.3 Abdominal pain 57.14 68 42 17.8 35.1 Myalgia 61.9 3.5 -- -- -- Diarrhea -- 36 -- -- -- Vomiting -- 41 -- -- -- Arthralgia 13 49 -- 10.7 -- Discussion Padeh[8], Stojanov [15] and Dagan [16] studies in comparison with our results are shown in Table 2. Table 3 shows our results and those of Gattornos. Bonyadi revealed that the carrier rate of five common MEFV genes in the Azeri Turkish population was 25.5%, with E148Q being the most common (11.5%) mutation followed by V726A (1.75%). This study indicates that the FMF carrier rate and E148Q mutation frequency are high in the Iranian Azeri Turkish population[17] and one of our studies showed the frequency of E148Q mutation in FMF patients to be 9.95% (in press). It does not seem that these MEFV mutations in PFAPA patients are related to normal population varieties. The authors have known two siblings suffering from FMF with M694V – V726A gene mutation, and PFAPA syndrome, offspring of whom one had monozygotic mutation M694V and the other monozygotic mutation V726A. In other authors’ experience there were 2 patients who had FMF and PFAPA simultaneously, it was observed that the first patient had R761H-M694I combined mutations and the second one M680I monozygotic mutation.

The authors have known two siblings suffering from FMF with M694V – V726A gene mutation, and PFAPA syndrome, offspring of whom one had monozygotic mutation M694V and the other monozygotic mutation V726A. In other authors’ experience there were 2 patients who had FMF and PFAPA simultaneously, it was observed that the first patient had R761H-M694I combined mutations and the second one M680I monozygotic mutation. It seems that there is a genetic similarity, on the basis of pathogenesis, between FMF and PFAPA. This genetic similarity has led to suggest a scoring system in genetic analysis of periodic fever syndromes in PFAPA patients[13]. Table 3 MEFV genes mutations in two studies Heterozygote Our study Gattorno(14) No of Mutation= 8 No of Mutation= 40 Compound Heterozygote Number 1 7 Type of Mutation K695R/ V726A M694V/V726A M680I/V726A M694V/E148Q L110P/L110P V726A/E148Q V726A/S108R P369S/R408Q Heterozygote Number 7 33 Type of Mutation M694V (4) V726A (2) E148Q (1) E148Q (11) K695R (5) M694V (4) A744S (3) V726A (2) P369S (1) V487M (1) R408Q (1) M680I (1) R717L (1) Table 4 Gaslini score in our patients No Age Sex Age at Onset Abd pain Diarrhea Chest pain Aphtous stomatitis Genotype Family history Gaslini score

Heterozygote Number 1 7 Type of Mutation K695R/ V726A M694V/V726A M680I/V726A M694V/E148Q L110P/L110P V726A/E148Q V726A/S108R P369S/R408Q Heterozygote Number 7 33 Type of Mutation M694V (4) V726A (2) E148Q (1) E148Q (11) K695R (5) M694V (4) A744S (3) V726A (2) P369S (1) V487M (1) R408Q (1) M680I (1) R717L (1) Table 4 Gaslini score in our patients No Age Sex Age at Onset Abd pain Diarrhea Chest pain Aphtous stomatitis Genotype Family history Gaslini score 1 5 Boy 2y Yes No No No Wt- Wt No -0.027 2 7 Boy 3y Yes Yes No Yes Wt-M694V No +1.38 3 5 Boy 2y No No No No Wt- Wt No -1.608 4 13 Girl 6 mo Yes No No Yes Wt- Wt No +1.082 5 6 Boy 4y Yes No No No Wt- Wt No -0.228 6 7 Boy 6y Yes No No Yes Wt- Wt No -3.34 7 4 Girl 2y Yes No No Yes Wt- Wt No -0.124 8 5 Boy 3y No No No No Wt- Wt No -2.412 9 5 Girl 1y No No No Yes Wt-V726A Father: FMF -0.802 10 5 Boy 10 mo No No No No Wt- Wt No -0.67 11 12 Boy 2y No No No Yes Wt- Wt No -3.112 12 12 Boy 3y No No No No Wt-M694V No -2.412 13 12 Boy 2y Yes No No Yes Wt- Wt Brother: Periodic Fever +1.379 14 4 Boy 6 mo Yes No No Yes Wt- Wt No +1.082 15 14 Girl 6y Yes No No No Wt-E148Q No -1.836 16 9 Boy 3y No No No Yes Wt- Wt No -3.916 17 10 Boy 5y Yes No No Yes Wt-M694V No -2.536 18 13 Boy 5y No No No Yes K695R-V726A No -5.524 19 8 Boy 2y No No No Yes Wt-M694V No -3.112 20 7 Girl 3y Yes No No Yes Wt-V726A Mother: FMF +0.575 21 8 Girl 3y Yes No No Yes Wt- Wt No -0.928 This scoring is based on clinical criteria and is called Gaslini diagnostic score. If a PFAPA patient has scored above 1.32, he/she will be in a high risk of having others periodic fever syndromes.

Yes Wt-M694V No -3.112 20 7 Girl 3y Yes No No Yes Wt-V726A Mother: FMF +0.575 21 8 Girl 3y Yes No No Yes Wt- Wt No -0.928 This scoring is based on clinical criteria and is called Gaslini diagnostic score. If a PFAPA patient has scored above 1.32, he/she will be in a high risk of having others periodic fever syndromes. In Gattorno study it was observed that Gaslini diagnostic score is a useful diagnostic tool in evaluating differential diagnosis of PFAPA syndrome. Because 38 percent of PFAPA patients in this study had MEFV gene mutations, it seems that genetic basis of FMF has a role in pathogenesis of PFAPA; however there is no significant difference between mutation positive and negative patients in duration of disease, episodes of disease and response to treatment. Table 4 shows patients’ Gaslini diagnostic scoring test. Conclusion This scoring was not helpful to predict the probability of MEFV gene mutation (12 common mutations) in our study and in these patients it has also been shown that MEFV gene mutation doesn't have any specific effect in patient’s clinical manifestations. We should find another way to predict the presence of MEFV gene mutations in this syndrome. Conflict of Interest: None Authors’ Contribution Concept / Design: F. Salehzadeh Acquisition of Data: M. Vahedi, M. Hosseini-Khotbesara Genetic Analysis: S. Hosseini-Asl Data Analysis / Interpretation: F. Salehzadeh Drafting of the Manuscript: S. Habibzadeh; S. Jahangiri Critical Revision of the Manuscript: S. Hosseini-Asl; F. Salehzadeh All authors approved final version of the Article

Although percutaneous placement of intravascular stents in congenital heart disease is a common practice, there are few reports regarding native right ventricular outflow tract (RVOT) stenting in children[1]. Stenting of conduit stenoses are more commonly reported[2]. In the preoperative setting, palliation of significant cyanosis by balloon valvuloplasty or RVOT stenting has been advocated by some as a means for reducing symptomatic cyanosis in patients with severe annular hypoplasia. Improvement in antegrade flow is thought to simultaneously enhance pulmonary arterial growth by augmenting pulmonary blood flow. Most of transcatheter interventions for relieving RVOT were done for conduit stenosis. There are few reports about native RVOT stenting, and to the best of our knowledge there are very few reports on native RVOT stenting in tetralogy of fallot (TOF) with absent left pulmonary artery[3]. A 9-year-old child was admitted with cyanosis noted from birth with failure to thrive, cyanotic spells and worsening cyanosis. The patient had undergone central modified Blalock- Tausig (MBT) shunt and right MBT shunt at the ages of three and six respectively. At this admission the child weighed 17 kg, had severe systemic desaturation (<55%) and severe cyanosis, digital clubbing and a New York Heart Association (NYHA) classification of class IV. Clinical examination revealed unremarkable pulmonary examination and 3/6 systolic heart murmur at pulmonary focus.

ix respectively. At this admission the child weighed 17 kg, had severe systemic desaturation (<55%) and severe cyanosis, digital clubbing and a New York Heart Association (NYHA) classification of class IV. Clinical examination revealed unremarkable pulmonary examination and 3/6 systolic heart murmur at pulmonary focus. EKG revealed normal sinus rhythm, right axis deviation and severe right ventricular hypertrophy (RVH). Echocardiography showed TOF anatomy, severe RVOT stenosis with 75 mmHg pressure gradient, RVH, abscent left pulmonary artery branch (LPA), non-functioning previous MBT shunts and major collateral arteries originating from descending aorta. On catheterization, both previous MBT shunts were occluded, the right pulmonary artery (RPA) was small with fairly acceptable arborization. Left lung was supplied by major collateral arteries originating from descending aorta and severe RVOT stenosis was present (Fig. 1). RVOT stenting was performed by two consecutive stents (17×7 mm Express LD, Boston Scientific. USA and 18×5 mm Racer renal stents, Medtronic USA). Post stenting angiography showed significant increase in pulmonary blood flow to the right lung (Fig. 2).

from descending aorta and severe RVOT stenosis was present (Fig. 1). RVOT stenting was performed by two consecutive stents (17×7 mm Express LD, Boston Scientific. USA and 18×5 mm Racer renal stents, Medtronic USA). Post stenting angiography showed significant increase in pulmonary blood flow to the right lung (Fig. 2). The arterial oxygen saturation rose to 83%. At six-month follow-up, his arterial oxygen saturation was maintained at above 80% and NYHA functional class was improved to class II. Palliative procedures for relieving RVOT stenosis include either surgical or transcatheter interventions. The indications for RVOT stenting are RV-to-PA conduit stenosis, residual infundibular stenosis after intracardiac repair, TOF with hypoplastic branch pulmonary arteries after palliative shunt surgery, pulmonary atresia after perforation of atretic segment, and RV hypertrophic cardiomyopathy and also as a bridge to surgery. RVOT stenting is also usually indicated in cases in those patients who are not amenable to total surgical repair. Our patient had absent LPA, small RPA and poor clinical condition. Thus, we carried out percutaneous RVOT stenting. Fig. 1 Right ventricular injection in anteroposterior view shows severe right ventricular outflow tract stenosis and absent left pulmonary artery RVOT stenting in such patients theoretically can lead to two major problems: overflow and edema of the right lung with resultant vasculopathy in the long term, and free pulmonary valve regurgitation with its consequences that include right ventricular dilatation and dysfunction.

Fig. 1 Right ventricular injection in anteroposterior view shows severe right ventricular outflow tract stenosis and absent left pulmonary artery RVOT stenting in such patients theoretically can lead to two major problems: overflow and edema of the right lung with resultant vasculopathy in the long term, and free pulmonary valve regurgitation with its consequences that include right ventricular dilatation and dysfunction. In our case, none of these complications occurred, because we chose the stent size with scrutiny and we did not sacrifice the pulmonary valve. Also we did not have complications such as stent migration, ventricular arrhythmias, collapse or fracture of the stent and recurrent stenosis during follow up. To our knowledge the patient has not undergone any curative operation by now. RVOT stenting provides an effective palliative modality for children with TOF and unfavorable pulmonary artery anatomy. Although re-stenosis can occur but it responds to re-dilatation.

In our case, none of these complications occurred, because we chose the stent size with scrutiny and we did not sacrifice the pulmonary valve. Also we did not have complications such as stent migration, ventricular arrhythmias, collapse or fracture of the stent and recurrent stenosis during follow up. To our knowledge the patient has not undergone any curative operation by now. RVOT stenting provides an effective palliative modality for children with TOF and unfavorable pulmonary artery anatomy. Although re-stenosis can occur but it responds to re-dilatation. In high risk patients such as our patient with severe cyanosis and high hemoglobin level and blood viscosity, the RVOT stenting decreases perioperative morbidity and mortality[4]. In conclusion native RVOT stenting is an effective and safe procedure in appropriately selected patients, especially in whom total correction is not possible. This procedure causes better growth of pulmonary artery branches, decreases right ventricular hypertrophy and increases left ventricular volume. Although many of these stents cannot be dilated to adult size, their efficacy in small infants and children in whom further surgery will ultimately be required is remarkable. Fig. 2 Right ventricular injection in anteroposterior view after right ventricular outflow tract stenting shows significant resolved stenosis and increased right lung blood flow without pulmonary valve involvement.

Sir, We would like to present an idiopathic sigmoid colon perforation revealed with a rare manifestation of pneumoscrotum. A forty two days old boy was referred to us because of agitation, progressive abdominal distention and swollen scrotum (Fig. 1). There were not any symptoms of other gastrointestinal tract such as nausea, vomiting, diarrhea, fever and the change of the stool color. On physical examination, the infant had tachycardia and tachypnea. His abdomen had distention but was soft and lax, and his scrotum had swelling with red skin that led us into a consideration of strangulated inguinal hernia. Portable CXR was normal but plain abdominal x-ray three hours later revealed bowel distention, specially in the distal colon, with free intra-peritoneal gas (Rigler's sign)(Fig 2). A small bubble of gas was seen in the left scrotal region. According to the Rigler's sign, we made a tentative diagnosis of bowel perforation resulting from left sided inguinal hernia. Fig. 1 A 42 day old boy with agitation, progressive abdominal distention and an erythematous swollen scrotum Fig. 2 Abdominal x-ray shows free intraperitoneal gas and Rigler sign (gas in the inner and outer side of the bowel loop- arrow).The open arrow shows gas in the left scrotal sac.

Portable CXR was normal but plain abdominal x-ray three hours later revealed bowel distention, specially in the distal colon, with free intra-peritoneal gas (Rigler's sign)(Fig 2). A small bubble of gas was seen in the left scrotal region. According to the Rigler's sign, we made a tentative diagnosis of bowel perforation resulting from left sided inguinal hernia. Fig. 1 A 42 day old boy with agitation, progressive abdominal distention and an erythematous swollen scrotum Fig. 2 Abdominal x-ray shows free intraperitoneal gas and Rigler sign (gas in the inner and outer side of the bowel loop- arrow).The open arrow shows gas in the left scrotal sac. An urgent ultrasound of the scrotum revealed only reverberation artefact, consistent with gas. Both testes were normal. An exploratory laparotomy was done through midline incision. The peritoneum was grossly contaminated and there was perforation of the descending and sigmoid colon on the antimesenteric side . Rest of the large and small bowel was normal and there were not any vesiculation or ulceration and the mucosal surfaces were intact. The mucosal biopsy specimen from the edge of perforation had a normal appearance under light microscopy, and there were not any characteristic CMV inclusions or HSV cytopathic changes. However, we had not immunohistochemistry, FISH, and PCR in our center to detect latent infection. Also multiple distal biopsies revealed normal ganglion cells and Hirschprung's disease was excluded.

normal appearance under light microscopy, and there were not any characteristic CMV inclusions or HSV cytopathic changes. However, we had not immunohistochemistry, FISH, and PCR in our center to detect latent infection. Also multiple distal biopsies revealed normal ganglion cells and Hirschprung's disease was excluded. The incidence of spontaneous colonic perforation is extremely rare. As in one study, it was reported only ten cases, seven of ten in very-low-birth-weight (VLBW) infants and three of ten in term or near-term neonates (one with Hirschsprung disease and the two cases without clear etiology during 10 years (2000-2009)[1]. Prompt diagnosis and early vigorous management are mandatory for neonatal colonic perforation, because of the highest mortality rate of it among neonatal gastrointestinal perforations which was reported in some studies[2]. Since 1912 (the first reported case)[3], there are few reported pneumoscrotum due to procedural or pathologic processes[4]. We think that our patient was the first case which reported spontaneous colonic perforation with unknown etiology, which presented with pneumoscrotum during the last decades. Endoscopic (upper and lower) and laparoscopic procedures were the second and third most common causes, respectively [5]. Therefore, as mentioned above, the finding of gas in the scrotal sac may be an early sign of a life-threatening condition like visceral perforation or may represent an incidental finding associated with more benign conditions like laparoscopic procedures.

Endoscopic (upper and lower) and laparoscopic procedures were the second and third most common causes, respectively [5]. Therefore, as mentioned above, the finding of gas in the scrotal sac may be an early sign of a life-threatening condition like visceral perforation or may represent an incidental finding associated with more benign conditions like laparoscopic procedures. In conclusion, although a combination of free abdominal gas and distended abdomen usually make the diagnosis evident and surgical intervention should be the rule[9], this case presentation reminds us that eventhough pneumoscrotum is a benign, rare condition, its mere presence should signal the possibility of a severe, life-threatening disease process within the peritoneum like bowel perforation or retroperitoneum like pancreatic or renal abscess. Acknowledgment The authors would like to thank the staff of Non-Communicable Pediatric Diseases Research Center of Amirkola Children Hospital.

Introduction Allergic diseases, especially food allergy, have been on a rise in the recent years[1]. An estimated 10%-15% of the population report symptoms of food allergy[2] whereas the actual prevalence of cow's milk allergy (CMA) in children varies and seems to range between 0.1 and 4.2% in different countries worldwide[3-4]. Cow's milk protein allergy is one of the most common allergies which is of great importance in many aspects. In addition to its clinical symptoms and limited diagnostic approach, when omitted from the child's daily diet causes major nutritional concerns in the child and occasionally in the mother. Therefore, primary prevention of cow's milk allergy, reducing clinical symptoms, disease duration and nutritional deficits are of special concern[5]. Recently, food allergy preventive approaches in pregnancy have shifted from avoidance towards exposure, because in some countries, mainly Australia, the occurrence of food allergy in infants whose mothers avoided common food allergens, was higher than in those whose mothers had a free daily diet[6]. As a good example, in comparison to introduction at 4–6 months, introducing egg into the infant’s diet later has been associated with higher rates of egg allergy[7]. On the other hand, the importance of the infantile immune system maturation especially in the first few months of life and its effect on developing allergic diseases makes these early years the ideal time for prevention of such diseases[3,4,8,9].

Therefore, primary prevention of cow's milk allergy, reducing clinical symptoms, disease duration and nutritional deficits are of special concern[5]. Recently, food allergy preventive approaches in pregnancy have shifted from avoidance towards exposure, because in some countries, mainly Australia, the occurrence of food allergy in infants whose mothers avoided common food allergens, was higher than in those whose mothers had a free daily diet[6]. As a good example, in comparison to introduction at 4–6 months, introducing egg into the infant’s diet later has been associated with higher rates of egg allergy[7]. On the other hand, the importance of the infantile immune system maturation especially in the first few months of life and its effect on developing allergic diseases makes these early years the ideal time for prevention of such diseases[3,4,8,9]. Previous studies have proven the efficacy of probiotics on clinical symptoms of cow's milk protein allergy (CMPA)[10-11] as well as several other diseases[12-16], but some recent studies have shown no benefits[17]. As both prebiotics and probiotics have immunomodulation effects, their mixture as synbiotic may be more effective in tolerance induction. In this study we evaluated the effect of a newly introduced symbiotic, a mixture of seven probiotic bacteria and fructo oligo sacharide (FOS), on the infants’ growth rate, clinical symptoms and disease course of cow's milk protein allergy in this age group.

ure as synbiotic may be more effective in tolerance induction. In this study we evaluated the effect of a newly introduced symbiotic, a mixture of seven probiotic bacteria and fructo oligo sacharide (FOS), on the infants’ growth rate, clinical symptoms and disease course of cow's milk protein allergy in this age group. Subjects and Methods This was a randomized double blind clinical trial performed from February 2009 to December 2010 in the pediatric allergy and gastrointestinal clinics of Ghaem education and research center, Mashhad, Iran. Totally 32 infants were randomly divided into the study and placebo groups each consisting of 16 cases (Fig. 1). The inclusion criteria consisted of 1-12 month-old infants with the clinical symptoms of cow's milk protein allergy including rectal bleeding, diarrhea, vomiting and evidences of colitis with complete resolution of symptoms following exclusion of dairy products from diet of mother and infant and confirmed by reappearance of symptoms after reintroducing dairy products, a good general condition, breast milk feeding and cow's milk consuming mother. The infants were excluded from the study if they had a known immune deficiency, gastrointestinal disease, positive stool culture, coagulopathy or were receiving any type of antibiotic therapy in the past two weeks (Fig. 1). Fig.1 Flow diagram of the study Table 1 Baseline characteristics of the studied cases Baseline characteristics Study group (SD) Placebo group (SD) P- value Age (month) 6 (2.4) 5.5 (2.8) 0.3 Sex 5/11 8/8 0.2 Head Circumference (cm) 41.68 (2.25) 42.46 (1.91) 0.3

The infants were excluded from the study if they had a known immune deficiency, gastrointestinal disease, positive stool culture, coagulopathy or were receiving any type of antibiotic therapy in the past two weeks (Fig. 1). Fig.1 Flow diagram of the study Table 1 Baseline characteristics of the studied cases Baseline characteristics Study group (SD) Placebo group (SD) P- value Age (month) 6 (2.4) 5.5 (2.8) 0.3 Sex 5/11 8/8 0.2 Head Circumference (cm) 41.68 (2.25) 42.46 (1.91) 0.3 Height (cm) 66.93 (4.5) 65.9 (4.25) 0.5 Weight (gr) 7028 (1038.5) 7728 (1271.7) 0.1 SD: Standard Deviation The patients' data including name, clinical symptoms, growth indices (height, weight and head circumference) were recorded in a questionnaire on entering the study. At the end of the first 72 hours, first week, second week and third week the clinical symptoms were obtained by phone and recorded. At the end of the first and final month the patients visited at the child's GI clinic where their clinical symptoms and growth indices were recorded. Clinical diagnosis and follow up visits were performed by the same person. After fully explaining the study protocol and obtaining an informed consent from the child's guardian, a synbiotic containing 1 billion Colony Forming Units (CFU) of ProtexinR Restore: a mixture of Lactobacillus casei, Lactobacillus rhamnosus, Streptococcus thermophilus, Bifidobacterium breve, Lactobacillus acidophilus, Bifidobacterium infantis, Lactobacillus bulgaricus and FOS (Protexin healthcare, Somerset, UK), in the form of freeze dried powder was fed daily for 4 weeks.

FU) of ProtexinR Restore: a mixture of Lactobacillus casei, Lactobacillus rhamnosus, Streptococcus thermophilus, Bifidobacterium breve, Lactobacillus acidophilus, Bifidobacterium infantis, Lactobacillus bulgaricus and FOS (Protexin healthcare, Somerset, UK), in the form of freeze dried powder was fed daily for 4 weeks. The placebo group received placebo with the same shape, color and packaging for the same duration. During this time the consumption of all kinds of food containing cow's milk protein was prohibited in all infants and their mothers. Ethics committee of Mashhad University of Medical Sciences has approved the proposal. For data analysis the R software language was used. Shapiro-Wilk normality test, t-test, Wilcoxon rank sum test and Chi-square tests were applied and a P-value <0.05 was considered as statistically significant. Findings Placebo and study groups, each included 16 cases, with mean age of 6 and 5.5 months, respectively. Baseline characteristics of the studied cases are shown in Table 1. At the beginning of study, all patients in both groups had diarrhea. Rectal bleeding was seen in 81.25% and 75% in placebo and study groups, respectively. 18.5% in placebo group and 25.5% in study group had colics as a symptom. 12.5% of patients in both groups had vomiting. There was no significant difference in reduction of the daily vomiting or diarrhea between case and placebo groups (Table 2 and 3).

as seen in 81.25% and 75% in placebo and study groups, respectively. 18.5% in placebo group and 25.5% in study group had colics as a symptom. 12.5% of patients in both groups had vomiting. There was no significant difference in reduction of the daily vomiting or diarrhea between case and placebo groups (Table 2 and 3). During the study, there were no significant differences in rectal bleeding between two groups. Intestinal colic got better in both groups after 72 hours and no patient had colic after 2 weeks but the difference was not significant. After 1, 2, 3 and 6 moths follow up there was no significant difference between symptomatic patients (who had at least one symptom) in the two groups. Table 4 shows that at the end of the first and third month, the difference of head circumference weight between the two groups was statistically significant, but not significant for height. Table 2 Comparing the mean reduction in daily vomiting between the study and placebo group Time Study group Placebo group P. value Mean (SD) Mean (SD) After 72 hrs -0.25 (0.77) -0.31 (0.84) 0.5 End of week 1 -0.25 (0.77) -0.37 (1.09) 0.5 End of week 2 to end of the 3 rd month -0.25 (0.77) -0.75 (2.05) 0.6 SD: Standard Deviation Table 3 Comparing the mean reduction in daily defecation times between the study and placebo group Time Study group Placebo group P. value Mean (SD) Mean (SD) After 72 hrs -1.62 (1.15) -1.87 (2.30) 0.4 End of week 1 -2.06 (1.29) -2.56 (2.68) 0.4 End of week 2 -2.69 (1.25) -2.97 (2.71) 0.3 End of week 3 -2.97 (1.60) -3.56 (2.68) 0.8 End of the 1 st month -3.38 (2.06) -3.80 (2.76) 0.7 End of the 2 nd month -3.83 (2.23) -4.41 (2.92) 0.8

Table 3 Comparing the mean reduction in daily defecation times between the study and placebo group Time Study group Placebo group P. value Mean (SD) Mean (SD) After 72 hrs -1.62 (1.15) -1.87 (2.30) 0.4 End of week 1 -2.06 (1.29) -2.56 (2.68) 0.4 End of week 2 -2.69 (1.25) -2.97 (2.71) 0.3 End of week 3 -2.97 (1.60) -3.56 (2.68) 0.8 End of the 1 st month -3.38 (2.06) -3.80 (2.76) 0.7 End of the 2 nd month -3.83 (2.23) -4.41 (2.92) 0.8 End of the 3 rd month -4.15 (2.11) -4.43 (2.77) 0.7 SD: Standard Deviation Discussion In this clinical trial we showed that using synbiotics for four weeks in infants with symptoms of CMA may increase growth indices but had no significant effect on gastrointestinal symptoms and disease course. There are several studies about effect of probiotics on rapid resolution of allergy symptoms.

End of the 3 rd month -4.15 (2.11) -4.43 (2.77) 0.7 SD: Standard Deviation Discussion In this clinical trial we showed that using synbiotics for four weeks in infants with symptoms of CMA may increase growth indices but had no significant effect on gastrointestinal symptoms and disease course. There are several studies about effect of probiotics on rapid resolution of allergy symptoms. Vandenplas et al revealed that both casein and whey hydrosylates formula enriched with probiotics can improve symptom scores in cow’s milk allergy[18]. Baldassarre et al showed that adding Lactobacillus GG to extensive hydrolyzed formula compared to taking this type of formula alone in cow’s milk induced colitis can improve colonic symptoms as well as reducing stool calprotectin and occult blood[19]. Ivakhnenko et al in an open randomized prospective clinical trial using Bifidobacterium lactis BB-12 (1х109 CFU) and Streptococcus thermophilus TH-4 (1х108 CFU) for four weeks showed that probiotics in addition to elimination diet in children with atopic dermatitis and cow's milk allergy may decrease gastrointestinal symptoms including diarrhea, constipation and infantile colic[20]. According to our study, we could not find any significant reduction in diarrhea and colic symptoms between the two groups. Used strains in our study were different to those of Vandenplas and Baldassarrre’s. Doses and duration of probiotics were similar in our study and Ivakhnenko et al. Their sample size was larger than ours and it may explain the difference between these findings[20].

in diarrhea and colic symptoms between the two groups. Used strains in our study were different to those of Vandenplas and Baldassarrre’s. Doses and duration of probiotics were similar in our study and Ivakhnenko et al. Their sample size was larger than ours and it may explain the difference between these findings[20]. The effect of probiotics on tolerance acquisition in patients with CMA is an unexplored area of research. Some studies have shown that supplementation of probiotics suppressed the allergic reaction mainly through increased intestinal secretary IgA and regulatory T cell induction[21]. West et al showed that a tolerogenic environment could be formed by dietary factors, including polyunsaturated fatty acids, probiotics, oligosaccharides, antioxidants, folic acid, and other vitamins such as vitamin D[9]. Once allergies are developed, the consumption of synbiotics may to some extent reduce symptoms and complications. In a recent interesting study, Canani et al showed supplementation of an extensively hydrolyzed casein formula with Lactobacillus GG accelerated tolerance acquisition to cow’s milk protein (CMP)[11]. Table 4 Changes of weight, height and HC in the studied cases after 1 and 3 months of follow up Variable Study group Mean (SD) Placebo group Mean (SD) P. value Head circumference (after 1 st month) 0.02 (0.01) 0.02 (0.01) 0.048 Head circumference (after 3 rd month) 0.06 ( 0.03) 0.05 (0.02) 0.03 Height (after 1 st month) 0.02 (0.01) 0.03 (0.02) 0.7 Height (after 3 rd month) 0.06 (0.03) 0.08 (0.05) 0.9 Weight (after 1 st month) 0.12 (0.07) 0.06 (0.06) 0.008

Mean (SD) Placebo group Mean (SD) P. value Head circumference (after 1 st month) 0.02 (0.01) 0.02 (0.01) 0.048 Head circumference (after 3 rd month) 0.06 ( 0.03) 0.05 (0.02) 0.03 Height (after 1 st month) 0.02 (0.01) 0.03 (0.02) 0.7 Height (after 3 rd month) 0.06 (0.03) 0.08 (0.05) 0.9 Weight (after 1 st month) 0.12 (0.07) 0.06 (0.06) 0.008 Weight (after 3 rd month) 0.25 (0.15) 0.15 (0.10) 0.02 SD: Standard Deviation As our study design did not include long term follow up, we could not have a reliable judgment about tolerance acquisition in our patients. But increasing growth indices in our study may indicate improving disease state.

Weight (after 1 st month) 0.12 (0.07) 0.06 (0.06) 0.008 Weight (after 3 rd month) 0.25 (0.15) 0.15 (0.10) 0.02 SD: Standard Deviation As our study design did not include long term follow up, we could not have a reliable judgment about tolerance acquisition in our patients. But increasing growth indices in our study may indicate improving disease state. Gut microbiota may play a role in weight gain by different mechanisms such as improvement of mineral bioavailability, synthesis of vitamins, regulation of gastrointestinal secretion and motility, digestion of macronutrients and regulation of energy extraction from the diet[3,22,23]. Some studies showed that changing microflora can increase weight gain[24,25] but it was inconsistent in other studies[26] taking into consideration several other factors including the probiotic type, dosage and the patient‘s age. In a recent meta-analysis, Lactobacillus gasseri was associated with weight loss both in obese humans and in animals, Lactobacillus fermentum and Lactobacillus ingluviei were associated with weight gain and Lactobacillus plantarum was associated with weight loss in animals. The authors concluded that different Lactobacillus species are associated with different effects on weight change that are host-specific[27]. This emphasizes that functional capabilities of probiotics are strain-dependent. In our study we could show that combinations of seven probiotics with fructooligosaccharides increase weight and other growth indices.

Lactobacillus species are associated with different effects on weight change that are host-specific[27]. This emphasizes that functional capabilities of probiotics are strain-dependent. In our study we could show that combinations of seven probiotics with fructooligosaccharides increase weight and other growth indices. As the therapeutic effects of synbiotics depend on type of probiotic strains, single-strain or multi-strain, dosage, duration and study population, these variables are the main causes of different results in clinical trials[12,28]. Ideal commercial product should contain probiotics or synbiotic strains that along with tolerance induction can increase growth status as well. Moreover, this study had its own limitations; the number of studied cases was not large enough. Future studies should be carried out on a greater number focusing on the early prescription of synbiotics (even during pregnancy), and at the critical age, first few months of life, when the tolerogenic environment can affect the incidence of allergic diseases. Finally, as different factors contribute to the development of food allergy and other types of allergies, a successful model of prevention should consider additional measures in the mother and child diet which affect tolerance induction including vitamin D status, antioxidants and folic acid[9]. Conclusion This study showed that synbiotics may increase growth indices (weight and head circumference) in children with cow’s milk protein allergy.

Future studies should be carried out on a greater number focusing on the early prescription of synbiotics (even during pregnancy), and at the critical age, first few months of life, when the tolerogenic environment can affect the incidence of allergic diseases. Finally, as different factors contribute to the development of food allergy and other types of allergies, a successful model of prevention should consider additional measures in the mother and child diet which affect tolerance induction including vitamin D status, antioxidants and folic acid[9]. Conclusion This study showed that synbiotics may increase growth indices (weight and head circumference) in children with cow’s milk protein allergy. Acknowledgment We would like to thank Fatemeh Chaji, for her invaluable assistance. This study was supported by a grant from the Vice Chancellor for Research of the Mashhad University of Medical Sciences for the research project as a medical student thesis with approval number of 87357.This study is registered in Iranian Registry of Clinical Trials (IRCT) ID: IRCT201301314976N2. Conflict of Interest: None

Acknowledgment We would like to thank Fatemeh Chaji, for her invaluable assistance. This study was supported by a grant from the Vice Chancellor for Research of the Mashhad University of Medical Sciences for the research project as a medical student thesis with approval number of 87357.This study is registered in Iranian Registry of Clinical Trials (IRCT) ID: IRCT201301314976N2. Conflict of Interest: None Authors Contribution H. Ahanchian and H.R. Kianifar conceptualized and designed the study, drafted the initial manuscript. SA Jafari, MH Amirian, H.R Kianifar and H. Ahanchian contributed to patient selection and follow up, reviewed and revised the manuscript. Z. Noori, T. Moghiman, H.R. Kianifar and A. Ezzati designed the data collection instruments and supervised data collection. S.A. Jafari and H. Ahanchian critically reviewed the manuscript. All Authors approved the final manuscript as submitted.

In developing countries, there is an increasing trend of chronic diseases such as tooth decay, obesity and diabetes especially in children[1]. Tooth decay in its severe form which is called nursing caries or early childhood caries (ECC) has impact on child growth and development. Children diagnosed with ECC weighed less than the matched control group due to the impact of pain and dysfunction on child’s eating and sleeping habit[2]. Oral health related quality of life (OHRQoL) is measured by different tools[3,4]. In pre-school age children, Pahel developed ECOHIS questionnaire[5], which was validated in local languages[6]. Severe form of ECC (S-ECC) is defined as the presence of any sign of smooth-surface caries in children younger than 3 years of age or detecting 1 or more cavitated lesions in smooth surfaces of maxillary anterior teeth or more than 4 decayed, missing, or filled surfaces in 3-5-year old children[7]. Parents of children under 5 years old attending three clinics providing complete dental treatment under general anesthesia (GA) consisting of Imam Khomeini Hospital, Mofid Children’s Hospital, and Shayamehr Clinics in Tehran, entered the study after ethical clearance and asked to fill in the validated Farsi version of ECOHIS questionnaire (n=81). Cases referred had severe caries, however were re-examined by two independent pediatric

thesia (GA) consisting of Imam Khomeini Hospital, Mofid Children’s Hospital, and Shayamehr Clinics in Tehran, entered the study after ethical clearance and asked to fill in the validated Farsi version of ECOHIS questionnaire (n=81). Cases referred had severe caries, however were re-examined by two independent pediatric dentists to confirm the need for treatment under GA. Those children having severe medical conditions and mental retardation were excluded. On the 1st and 2nd follow-up sessions 4 weeks and 3-months after dental rehabilitation the same family member who spent most of the time with the child was asked to fill in the same questionnaire. The questionnaire contains 13 questions in two sections. Child section consists questions in four domains: (i) Child Symptom Domain (CSD) includes pain, (ii) Child Function Domain (CFD) includes child trouble in eating and drinking, pronouncing words and missing preschool or day care, (iii) Child Psychology Domain (CPD) includes trouble sleeping and being irritable, (iv) Child Self-image and Social Interaction Domain (CSID) includes avoiding smile or talk. Parent section was comprised (i) Family Distress Domain (FDD) and (ii) Family Function Domain (FFD)[8]. Responses were from “never” to “very often” scored 0 to 4 and summed up in each domain for mean value calculation. Higher value in each domain demonstrates perceived negative impact of the oral health condition and compared on the baseline and follow up sessions.

The questionnaire contains 13 questions in two sections. Child section consists questions in four domains: (i) Child Symptom Domain (CSD) includes pain, (ii) Child Function Domain (CFD) includes child trouble in eating and drinking, pronouncing words and missing preschool or day care, (iii) Child Psychology Domain (CPD) includes trouble sleeping and being irritable, (iv) Child Self-image and Social Interaction Domain (CSID) includes avoiding smile or talk. Parent section was comprised (i) Family Distress Domain (FDD) and (ii) Family Function Domain (FFD)[8]. Responses were from “never” to “very often” scored 0 to 4 and summed up in each domain for mean value calculation. Higher value in each domain demonstrates perceived negative impact of the oral health condition and compared on the baseline and follow up sessions. In this study, significant improvement was observed in both child and parent sections. In the CFD domain the mean score 5.08±3.5 was significantly improved in the follow up sessions (Table 1). Abanto et al, have also reported high mean score of 4.15±3.92 in this domain among Brazilian children with S-ECC[9]. Table 1 Comparison of the Mean (Standard Deviation) ECOHIS score in each domain in the whole sample, pre and post treatment [minimum and maximum score of each domain] Domain Pre- anesthetic 1-Month follow up 3-month follow up CSD [0-4] 1.8 (1.2) (n=79) 0.29* (0.71) (n=75) 0.061‡ (0.31) (n=49) CFD [0-16] 5.08 (3.5) (n=71) 0.48* (0.95) (n=70) 0.02 (0.14) (n=48) CPD [0-8] 3.2 (2.4) (n=79) 0.26* (0.95) (n=70) Not reported CSID [0-8] 0.97 (1.8) (n=76) 0.13*(0.68) (n=75) Not reported

Table 1 Comparison of the Mean (Standard Deviation) ECOHIS score in each domain in the whole sample, pre and post treatment [minimum and maximum score of each domain] Domain Pre- anesthetic 1-Month follow up 3-month follow up CSD [0-4] 1.8 (1.2) (n=79) 0.29* (0.71) (n=75) 0.061‡ (0.31) (n=49) CFD [0-16] 5.08 (3.5) (n=71) 0.48* (0.95) (n=70) 0.02 (0.14) (n=48) CPD [0-8] 3.2 (2.4) (n=79) 0.26* (0.95) (n=70) Not reported CSID [0-8] 0.97 (1.8) (n=76) 0.13*(0.68) (n=75) Not reported FDD [0-8] 5.5 (2.1) (n=77) 0.34* (1.08) (n=76) Not reported Child [0-36] 10.88 (7.6) (n=67) 0.98* (2.3) (n=65) 0.4 (0.2) (n=46) Parent [0-16] 9.5 (3.5) (n=74) 2.4* (2.2) (n=73) 0.8 (1.32) (n=50) * P<0.05; wilcoxon Signed Ranks Test ‡ P<0.05; Friedman Test This implies caries in its severe form has impact on children’s usual daily function. There was also improvement in CSD domain after treatment. It was reported in the study of Acs et al, that improvement in painwas predominant followed by improved abilities to eat and sleep, reported by 86, 69, and 41% of parents, respectively[10]. Higher mean score of family distress domain, compared with child related domains was observed in this study in line with the study of Abanto et al, who reported the mean score of 3.43±2.59[9]. Filstrup et al, using the Michigan scale parents of children with ECC, evaluated their children’s oral health-related quality of life worse than that of children without ECC and they were able to realize that dental treatment of ECC improved children’s oral health-related quality of life[11].

ean score of 3.43±2.59[9]. Filstrup et al, using the Michigan scale parents of children with ECC, evaluated their children’s oral health-related quality of life worse than that of children without ECC and they were able to realize that dental treatment of ECC improved children’s oral health-related quality of life[11]. This study showed significant improvement in oral-health related quality of life of children and especially their parents after dental rehabilitation. However, this should not be considered as a good reason to postpone prevention and promote treatment under general anesthesia in children.

Introduction Breast-feeding with the proper technique, frequency, duration, exclusive breast-feeding up to 6 months of age, and continued breast-feeding along with appropriate complementary foods up to 2 years of age are the principal conditions of ideal nutrition for infants[1]. According to the data of the Turkish Population and Health Surveys, 68.9% of the newborns from 0 to 1 month of age, 42% of the infants between 4 and 5 months of age, and 21.9% of the infants between 4 and 5 months of age are exclusively fed with breast milk[2]. In Turkey, every year, approximately 1.4 million infants are born. According to the research, 10% of those infants are born with low-birth-weight[3].

rom 0 to 1 month of age, 42% of the infants between 4 and 5 months of age, and 21.9% of the infants between 4 and 5 months of age are exclusively fed with breast milk[2]. In Turkey, every year, approximately 1.4 million infants are born. According to the research, 10% of those infants are born with low-birth-weight[3]. The ideal nourishment for newborns with low birth-weights is breast milk. The breast milk of mothers who gave birth to low-birth-weight infants is specially adjusted to the infants’ needs. Although low-birth-weight infants have sucking and swallowing difficulties, those who are born in the 32nd week of pregnancy or later can easily suckle breast milk[4]. However, nursing difficulties are still commonly seen in feeding such infants for reasons related to mothers, such as their doubting that their milk will suffice, lacking the support of healthcare personnel, and having incorrect traditional beliefs[5]. Although the fact that low-birth-weight infants have a greater need for breast milk in comparison to infants with normal birth-weight, they are more frequently and more commonly bottle-fed with artificial formulas[1]. The studies carried out in this field exhibited that the breast-feeding rates of mothers of low-birth-rate infants are significantly lower than that of those who had gave birth at term[6]. In addition it was found that the breast-feeding duration of the low-birth-weight and preterm infants is considerably shorter than that for full-term normal-weight infants[7,8].

d that the breast-feeding rates of mothers of low-birth-rate infants are significantly lower than that of those who had gave birth at term[6]. In addition it was found that the breast-feeding duration of the low-birth-weight and preterm infants is considerably shorter than that for full-term normal-weight infants[7,8]. Perception of self-efficacy plays an important role in determining the actions an individual will take or prevent. Breast-feeding self-efficacy is the confidence the mother feels pertaining to breast-feeding[9]. It was determined that while mothers with low breast-feeding self-efficacy tend to wean far earlier than the recommended period, mothers having high breast-feeding self-efficacy experience fewer problems in starting and sustaining breast-feeding[10]. It was determined that the actions of healthcare professionals working in newborns’ units of hospitals, such as enlightening mothers of low-birth-weight infants on breast-feeding, encouraging them to participate in their babies’ care, giving them emotional support, and conducting regular follow-up home visits after their release from hospital, increase the mothers’ postpartum self-efficacy and breast-feeding success[11,12]. No study that examines the effects of natural-feeding states of the mothers of low-birth-weight infants, their self-efficacy levels, and the interferences to promote natural feeding on breast-feeding success could be found in Turkey.

tal, increase the mothers’ postpartum self-efficacy and breast-feeding success[11,12]. No study that examines the effects of natural-feeding states of the mothers of low-birth-weight infants, their self-efficacy levels, and the interferences to promote natural feeding on breast-feeding success could be found in Turkey. The present study was conducted with the purpose of determining the effect of natural-feeding education given to the mothers of low-birth-weight infants treated in newborns’ clinics, on the mothers’ breast-feeding self-efficacy levels, and the breast-feeding success of the infants. Subjects and Methods The following is a description of the subjects and methods of the study. Universe and Sample of the Study

The present study was conducted with the purpose of determining the effect of natural-feeding education given to the mothers of low-birth-weight infants treated in newborns’ clinics, on the mothers’ breast-feeding self-efficacy levels, and the breast-feeding success of the infants. Subjects and Methods The following is a description of the subjects and methods of the study. Universe and Sample of the Study The universe of the study, which was carried out between January 2010 and January 2011, was comprised of the low-birth-weight infants and their mothers, who received treatment in the neonatal clinics of two hospitals located in the eastern Turkey. Families included in this study had lower levels of income and education, according to the average levels of income and education in Turkey. The region of the study has a cold climate and hard geographical conditions compared to other regions. However, the region where the study was carried out is an important health center that accepts patients from other provinces. The sample size of the study was determined through power analysis. The sample of the study consisted of 86 low-birth-weight infants and their mothers, with 42 for test group and 44 for control group, who were selected from the mentioned universe by means of the nonprobability accidental sampling method. The sample size for the study, 95% test power with the effect size of 0.8 represents the power of the universe, and 85% were identified as a result of the power analysis. In order to prevent the mothers from influencing each other with the education they received, first, the data of the control group, and later, the data of the test group were collected.

a court order[23]. Additionally, youths under the age of 18 generally have limited rights to make certain medical decisions independently, for instance regarding treatment for sexually transmitted diseases, substance and alcohol abuse treatment, blood donation, mental health treatment and family planning services[23]. It appears that in the above-mentioned countries legislators have clearly specified the legal age for giving valid consent and at the same time have taken every measure to safeguard the right to medical decision-making for people just under the age of maturity who possess proper decision-making capacity. The responsibility to ascertain this capacity in underage patients would naturally fall to the physician. In the Iranian legislation and according to Shi’a Fiqh, stages of capacity – or competency in legal texts[9] – are as follow[24,25]: 1) Stage of Gheare Momayyez - Unawareness -: In this stage the child has limited understanding and powers of discernment and cannot distinguish between benefit and loss, and therefore is not legally considered to have a will. The stage lasts from 2 to 7 years and corresponds to early childhood in the ethical classification.

est power with the effect size of 0.8 represents the power of the universe, and 85% were identified as a result of the power analysis. In order to prevent the mothers from influencing each other with the education they received, first, the data of the control group, and later, the data of the test group were collected. Inclusion Criteria Two sets of criteria were considered in the study: infant-related criteria and mother-related criteria. Infant-related criteria: Infants who had following criteria were included in the study: had no conditions preventing nutrition, had birth weight of less than 2500 g, were born at more than 32 weeks of gestational age, and were hospitalized for at least 5 days Mother-related criteria: Mothers who had following criteria were included in the sample: were between the ages of 18 and 35, had no seeing or hearing problems, were living in the city center, and were open to communication and cooperation Examining the demographic attributes of the mothers and infants included within the scope of the study (mother's age, level of education, employment status, income level, whether or not the pregnancy was planned, number of children, delivery method, breast-feeding experience, breast-feeding education, duration she intends to breastfeed her baby, gender of the baby, gestational week of birth, and when breast-feeding started) showed that there is no significant difference between the groups (P>0.05) and that both groups were similar in terms of demographic attributes (Table 1). Data Collection Tools

Examining the demographic attributes of the mothers and infants included within the scope of the study (mother's age, level of education, employment status, income level, whether or not the pregnancy was planned, number of children, delivery method, breast-feeding experience, breast-feeding education, duration she intends to breastfeed her baby, gender of the baby, gestational week of birth, and when breast-feeding started) showed that there is no significant difference between the groups (P>0.05) and that both groups were similar in terms of demographic attributes (Table 1). Data Collection Tools For collecting the data the “Personal Information Form,” “Breast-feeding Self-Efficacy Form,” “LATCH Breast-feeding Assessment Tool,” “Breast-feeding Follow-Up Card,” “Infant Anthropometric Measurement Form,” infant weighing scale, infant height measurement ruler, and head circumference tape were used. Personal Information Form: The Personal Information Form used in data collection consists of 13 questions focused on the information pertaining to the mother (mother’s age and employment status and the income level of the family), information about the pregnancy (planning of pregnancy, number of children, and delivery method) information on the newborn (gender and gestational week) and breast-feeding-related information (breast-feeding experience, if breast-feeding education was received, when breast-feeding started, and intention regarding feeding exclusively with breast milk).

lanning of pregnancy, number of children, and delivery method) information on the newborn (gender and gestational week) and breast-feeding-related information (breast-feeding experience, if breast-feeding education was received, when breast-feeding started, and intention regarding feeding exclusively with breast milk). Breast-feeding Self-Efficacy Scale: The initial form of the Breast-feeding Self-Efficacy Scale, which was developed by Dennis[13] (2003) for the purpose of assessing mothers’ breast-feeding self-efficacy, had 33 items. Table 1 Comparison of experimental and control group characteristics of mothers regarding their breastfeeding and demographic features Variable Features Experimental Group [ n (%)] Control Group Total P. value n (%) n (%) Age group (year) 19-29 30 (71.4) 28 (65.1) 58 (68.2) 0.5 30 and above 12 (28.6) 15 (34.9) 27 (31.8) Education Primary education 21 (50.0) 23 (53.5) 44 (51.8) 0.7 High school 14 (33.3) 11 (25.6) 25 (29.4) University 7 (16.7) 9 (20.9) 16 (18.8) Work status Housewife 36 (85.7) 36 (83.7) 72 (84.7) 0.8 Working 6 (14.3) 7 (16.3) 13 (15.3) Income status Less expense to income 1 (2.4) 5 (11.6) 6 (7.1) 0.2 Expense to be equivalent income 23 (54.8) 21 (48.8) 44 (51.8) More income to expense 18 (42.9) 17 (39.5) 35 (41.2) Pregnancy planning status Planned 36 (85.7) 35 (81.4) 71 (83.5) 0.6 Not planned 6 (14.3) 8 (18.6) 14 (16.5) Number of children First 18 (42.9) 17 (39.5) 35 (41.2) 0.7 Two or more 24 (57.1) 26 (60.5) 50 (58.8) Mode of birth Normal 17 (40.5) 16 (37.2) 33 (38.8) 0.8 Cesarean 25 (59.5) 27 (62.8) 52 (61.2) Mother Breastfeeding

Income status Less expense to income 1 (2.4) 5 (11.6) 6 (7.1) 0.2 Expense to be equivalent income 23 (54.8) 21 (48.8) 44 (51.8) More income to expense 18 (42.9) 17 (39.5) 35 (41.2) Pregnancy planning status Planned 36 (85.7) 35 (81.4) 71 (83.5) 0.6 Not planned 6 (14.3) 8 (18.6) 14 (16.5) Number of children First 18 (42.9) 17 (39.5) 35 (41.2) 0.7 Two or more 24 (57.1) 26 (60.5) 50 (58.8) Mode of birth Normal 17 (40.5) 16 (37.2) 33 (38.8) 0.8 Cesarean 25 (59.5) 27 (62.8) 52 (61.2) Mother Breastfeeding Experience Yes 24 (57.1) 26 (48.8) 50 (58.8) 0.5 No 18 (42.9) 17 (51.2) 35 (41.2) The condition of breastfeeding Education Yes 8 (19.0) 7 (16.3) 15 (17.6) 0.7 No 34 (81.0) 36 (83.7) 70 (82.4) Time for giving breast milk 4 months 17 (40.5) 19 (44.2) 36 (42.4) 0.7 6 months 15 (35.7) 12 (27.9) 27 (31.8) Mixed (breast milk+ mama) 10 (23.8) 12 (27.9) 22 (25.9) Sex of the baby Female 28 (66.7) 26 (60.5) 54 (63.5) 0.5 Male 14 (33.3) 17 (39.5) 31 (36.5) Gestational week 32-35 13 (31.0) 15 (34.9) 28 (32.9) 0.8 36-38 24 (57.1) 22 (51.2) 46 (54.1) 39 and above 5 (11.9) 6 (14.0) 11 (12.9) Baby’ first breastfeeding time Shortly after birth 8 (19.0) 4 (9.3) 12 (14.1) 0.5 Within 60 minutes 13 (31.0) 11 (25.6) 24 (28.2) 61 minutes or more 6 (14.3) 7 (16.3) 13 (15.3) Breastfeeding did not happen 15 (35.7) 21 (48.8) 36 (42.4) Later, in 2003, a 14-item short form of the scale was developed. Dennis suggests this short form for use. While the lowest point that can be scored in the scale is 14, the maximum is 70. Higher points scored indicate high breast-feeding self-efficacy.

13 (15.3) Breastfeeding did not happen 15 (35.7) 21 (48.8) 36 (42.4) Later, in 2003, a 14-item short form of the scale was developed. Dennis suggests this short form for use. While the lowest point that can be scored in the scale is 14, the maximum is 70. Higher points scored indicate high breast-feeding self-efficacy. The validity and reliability study of the scale for the Turkish was carried out by Tokat[10] (2009), and its Cronbach’s alpha value was found to be 0.86. In the present study, the Cronbach’s alpha value of the scale found for immediate postpartum was 0.74, and it was 0.98 by the end of the 6th month following the birth. LATCH Breast-feeding Assessment Tool: The LATCH Breast-feeding Assessment Tool was developed by Jensen et al[14]. Turkish validity of the assessment tool was examined two times, and it was found to be a reliable tool[17,18]. The Cronbach’s alpha value of the LATCH Breast-feeding Assessment Tool was found to be 0.95, by Yenal and Okumus[15], and 0.96 by Koyun[16]. In the present study, the Cronbach’s alpha value of the LATCH Breast-feeding Assessment Tool was found to be 0.93. Whereas the maximum point that can be scored in the LATCH Breast-feeding Assessment Tool is 10, the minimum is 0[15,16]. Higher points scored in the tool indicate higher success in breast-feeding. Breast-feeding Follow-Up Form: This is a form developed by the researcher to assess breast-feeding activities observed through the LATCH breast-feeding scoring system and to record the results. Data Collection

Whereas the maximum point that can be scored in the LATCH Breast-feeding Assessment Tool is 10, the minimum is 0[15,16]. Higher points scored in the tool indicate higher success in breast-feeding. Breast-feeding Follow-Up Form: This is a form developed by the researcher to assess breast-feeding activities observed through the LATCH breast-feeding scoring system and to record the results. Data Collection The researcher collected the data by conducting one-on-one interviews with the mothers in hospitals and in their homes. Collection of Pretest Data: In order to collect pretest data, on the date of their hospitalization, the mothers of low-birth-weight infants of both test and control groups were given the Personal Information Form, Breast-feeding Self-Efficacy Scale and LATCH Breast-feeding Assessment Tool.

The researcher collected the data by conducting one-on-one interviews with the mothers in hospitals and in their homes. Collection of Pretest Data: In order to collect pretest data, on the date of their hospitalization, the mothers of low-birth-weight infants of both test and control groups were given the Personal Information Form, Breast-feeding Self-Efficacy Scale and LATCH Breast-feeding Assessment Tool. Collection of Posttest Data: For collecting posttest data, the mothers of both test and control groups were visited once in the hospital on the 5th day of their hospitalization and then once a month in their homes for a period of 6 months. During the visits, the mothers were asked to breastfeed their babies. By observing their breast-feeding statuses, the Breast-feeding Follow-Up Forms were filled. At the end of the 6th month, the Breast-feeding Self-Efficacy Scale and the LATCH Breast-feeding Assessment Tool were once again applied at the mothers’ home. During the home visits, necessary reminders were made to the mothers of the test group, and their questions were answered. The visits made to the homes of the participants of the test group were limited to 30 minutes, whereas they were limited to 10 minutes for the control group. Immediately after the application of the final tests at the end of the 6th month, the mothers in the control group were provided with an informative booklet. Interference Tools The following is a description of the interference tools used during the study. Educatory booklet – Every Mother Can Breastfeed; Wanting to Is Enough

Collection of Posttest Data: For collecting posttest data, the mothers of both test and control groups were visited once in the hospital on the 5th day of their hospitalization and then once a month in their homes for a period of 6 months. During the visits, the mothers were asked to breastfeed their babies. By observing their breast-feeding statuses, the Breast-feeding Follow-Up Forms were filled. At the end of the 6th month, the Breast-feeding Self-Efficacy Scale and the LATCH Breast-feeding Assessment Tool were once again applied at the mothers’ home. During the home visits, necessary reminders were made to the mothers of the test group, and their questions were answered. The visits made to the homes of the participants of the test group were limited to 30 minutes, whereas they were limited to 10 minutes for the control group. Immediately after the application of the final tests at the end of the 6th month, the mothers in the control group were provided with an informative booklet. Interference Tools The following is a description of the interference tools used during the study. Educatory booklet – Every Mother Can Breastfeed; Wanting to Is Enough The educatory booklet covers information regarding the importance of self-efficacy in breast-feeding and of breast milk for infants with low birth-weight, the things breast-feeding and breast milk are good for, preparation of the mother for breast-feeding, risks of artificial feeding, breast-feeding positions, breast-feeding duration and intervals, nutrition of the nursing mothers, extraction and preservation of breast milk, feeding methods of low-birth-weight infants, and the most common problems the mothers experience in the process of starting and sustaining breast-feeding. In addition, the participant mothers received no formula-feeding training or information, and no instruction was given to them at the hospitals because these hospitals were “baby-friendly” hospitals.

t infants, and the most common problems the mothers experience in the process of starting and sustaining breast-feeding. In addition, the participant mothers received no formula-feeding training or information, and no instruction was given to them at the hospitals because these hospitals were “baby-friendly” hospitals. Low-Birth-Weight Infant Dummy A training dummy of approximately 750 g weight, with movable head, neck and limbs for representing a low-birth-weight infant was used in the study for training purposes. Breast-feeding Pillow The pillow used in the study was designed for comfortable use during breast-feeding. The pillow was 150 cm long, 25 cm wide, and 8 l in volume. Produced with materials that are free of carcinogenic and allergenic substances, the pillow was made of 100% cotton, with a soft, light, washable cover. In order to prevent infections, the cover of the pillow was changed with a new one before each breast-feeding session. Nursing Interference

The pillow used in the study was designed for comfortable use during breast-feeding. The pillow was 150 cm long, 25 cm wide, and 8 l in volume. Produced with materials that are free of carcinogenic and allergenic substances, the pillow was made of 100% cotton, with a soft, light, washable cover. In order to prevent infections, the cover of the pillow was changed with a new one before each breast-feeding session. Nursing Interference For 5 days starting from their first day of hospitalization, the mothers included in the test group were given education in line with the content of the book titled Every Mother Can Breastfeed; Wanting to Is Enough, at the hours convenient for the mothers and their babies. At the end of each training session, breast-feeding was first demonstrated by the researcher with the use of the breast-feeding pillow and the training dummy. Afterwards, the mothers were first asked to practice breast-feeding with the dummy, and then asked to breastfeed their own babies. At the end of 5 days of training, which took about 30 minutes every day, the mothers in the test group were provided with the booklet Every Mother Can Breastfeed; Wanting to Is Enough. One week after their release from the hospital, the mothers of the test group were contacted via phone and asked whether they were having any problems regarding breast-feeding. They were informed again regarding their inadequate or wrong practices.

ded with the booklet Every Mother Can Breastfeed; Wanting to Is Enough. One week after their release from the hospital, the mothers of the test group were contacted via phone and asked whether they were having any problems regarding breast-feeding. They were informed again regarding their inadequate or wrong practices. During the study period, the nurse who worked day-shift provided breast-feeding counseling in line with the hospital policy, but the counseling could not be given daily and regularly, and no equal amount of counseling-time was given for each patient. The counseling did not include home visits after hospital discharge at all. Yet, our counseling was regularly provided 5 days a week, with demonstrations on baby-dolls, and mothers were made to practice. In addition, monthly regular home visits were paid to the babies. As for the control group, they were given only breast-feeding training by the breast-feeding trainer. Evaluation of Data Evaluation of the data obtained from the study was carried out in a computerized environment with the use of the SPSS 15.0 (Statistical Package for Social Science) package software. In the evaluation of the data, percentage distribution, chi-square test, t-test, and Cronbach’s alpha coefficient calculations were used. The significance value has been established as P<0.05 in this study. Ethical Principles of the Study

Evaluation of the data obtained from the study was carried out in a computerized environment with the use of the SPSS 15.0 (Statistical Package for Social Science) package software. In the evaluation of the data, percentage distribution, chi-square test, t-test, and Cronbach’s alpha coefficient calculations were used. The significance value has been established as P<0.05 in this study. Ethical Principles of the Study Before commencing the study, both written and verbal permissions of the hospitals, in which the study was to be carried out, were obtained. The study was also submitted to the ethical committee of the university, where the study was to be conducted, and the approval was granted. Before collecting any data, the purpose and duration of the study, and the things expected to be realized within the scope of the study were clearly explained to the mothers; their questions were answered, and then their written and verbal consents were obtained. The hospitals that were not “baby friendly” were excluded from the study and were asked to provide routine breast-feeding counseling for the control group, too. Findings As for the intergroup–intragroup comparison of the breast-feeding self-efficacy point averages of the test and control groups, it was found that the posttest point average of the mothers included in the test group was considerably higher than the posttest point average of the mothers from the control group, and that difference was statistically significant (P<0.001, Table 2). Table 2 Intra-group and intergroup comparison of breast feeding self-efficacy scale

Findings As for the intergroup–intragroup comparison of the breast-feeding self-efficacy point averages of the test and control groups, it was found that the posttest point average of the mothers included in the test group was considerably higher than the posttest point average of the mothers from the control group, and that difference was statistically significant (P<0.001, Table 2). Table 2 Intra-group and intergroup comparison of breast feeding self-efficacy scale Breast Feeding Self-Efficacy Scale Pretest Posttest t P. Value Mean (SD) Mean (SD) Test group 42.00 (9.99) 65.81 (7.37) 12.617 <0.001 Control Group 41.77 (11.40) 39.70 (12.21) 0.898 0.4 t 0.100 11.898 P. Value 0.9 <0.001 SD: Standard Deviation Table 3 Comparison of the LATCH breastfeeding assessment tool point averages scored of mothers Tagged with Day and Month Test Grup Mean (SD) Control Grup Mean (SD) t P. value First day of hospitalization 5.90 (1.62) 5.81 (1.72) 0.060 0.8 5 th day 8.76 (1.90) 6.35 (2.29) 5.287 <0.001 1 st month 9.60 (1.23) 7.65 (2.05) 5.293 <0.001 2 nd month 9.79 (0.95) 8.19 (1.94) 4.803 <0.001 3 rd month 9.83 (0.93) 8.12 (2.24) 4.595 <0.001 4 th month 9.86 (0.93) 7.67 (.41) 5.492 <0.001 5 th month 9.86 (0.93) 7.53 (2.40) 5.851 <0.001 6 th month 9.86 (0.93) 7.51 (2.42) 5.567 <0.001 SD: Standard Deviation

5 th day 8.76 (1.90) 6.35 (2.29) 5.287 <0.001 1 st month 9.60 (1.23) 7.65 (2.05) 5.293 <0.001 2 nd month 9.79 (0.95) 8.19 (1.94) 4.803 <0.001 3 rd month 9.83 (0.93) 8.12 (2.24) 4.595 <0.001 4 th month 9.86 (0.93) 7.67 (.41) 5.492 <0.001 5 th month 9.86 (0.93) 7.53 (2.40) 5.851 <0.001 6 th month 9.86 (0.93) 7.51 (2.42) 5.567 <0.001 SD: Standard Deviation In the intergroup comparison of the point averages of mothers from both the test and control groups obtained from the LATCH Breast-feeding Assessment Tool, it was determined that the LATCH point averages of the test group mothers increased from the control group scored (P<0.005, Table 3). While there were no babies that refused breast milk or “weaned prematurely” from the test group, it was determined that 85.7% of the babies were being fed exclusively with breast milk and that there was a statistically significant difference between the test and control groups in these terms (P<0.001, Table 4). Discussion Despite the facts that the importance of breast milk in infant nutrition is emphasized at both national and international levels, and that it is accepted by many countries and announced in several declarations that healthy nutrition is a right for all children, it is believed that throughout the world today only 39% of the newborns are exclusively fed with breast milk in the first 6 months of life[17]. According to the data of the TNSA [2], in Turkey, 68.9% of the newborns from 0 to 1 month of age, 42% of the infants between 4 and 5 months of age, and 21.9% of the infants between 4 and 5 months of age are exclusively fed with breast milk.

ewborns are exclusively fed with breast milk in the first 6 months of life[17]. According to the data of the TNSA [2], in Turkey, 68.9% of the newborns from 0 to 1 month of age, 42% of the infants between 4 and 5 months of age, and 21.9% of the infants between 4 and 5 months of age are exclusively fed with breast milk. At the end of the study, it was understood that the mothers from the test group had considerably higher perceptions of self-efficacy, and they experienced fewer breast-feeding problems in comparison to those in the control group (Table 2). There are many studies emphasizing that the education and training offered regarding breast-feeding have positive effects on mothers’ willingness to breastfeed[2,10,18,19], and, that as a modifiable perception, breast-feeding self-efficacy can be increased[11,13,20]. In the literature it is stated that the high breast-feeding self-efficacy level of the mother has considerable effects on her behavior of sustaining breast-feeding[12,16,20]. However, Barnes and Adamson[21] (2004) determined in their study that mothers of preterm infants have considerably low perceptions of self-efficacy, and these mothers need much more support. Table 4 Distribution of the ways with which the babies from test and control groups were being fed at the end of the sixth month Nutrition Type Test Group Control Group t P. value n (%) n (%) Only breast milk 36 (85.7) 5 (11.6) <0.001 Mixed feeding 6 (14.3) 22 (51.2) <0.001 Milk was interrupted before the sixth month 0 6 (14.3) <0.001

At the end of the study, it was understood that the mothers from the test group had considerably higher perceptions of self-efficacy, and they experienced fewer breast-feeding problems in comparison to those in the control group (Table 2). There are many studies emphasizing that the education and training offered regarding breast-feeding have positive effects on mothers’ willingness to breastfeed[2,10,18,19], and, that as a modifiable perception, breast-feeding self-efficacy can be increased[11,13,20]. In the literature it is stated that the high breast-feeding self-efficacy level of the mother has considerable effects on her behavior of sustaining breast-feeding[12,16,20]. However, Barnes and Adamson[21] (2004) determined in their study that mothers of preterm infants have considerably low perceptions of self-efficacy, and these mothers need much more support. Table 4 Distribution of the ways with which the babies from test and control groups were being fed at the end of the sixth month Nutrition Type Test Group Control Group t P. value n (%) n (%) Only breast milk 36 (85.7) 5 (11.6) <0.001 Mixed feeding 6 (14.3) 22 (51.2) <0.001 Milk was interrupted before the sixth month 0 6 (14.3) <0.001 Did not take breast milk 0 7 (16.3) <0.001 The breast-feeding successes of the mothers from both test and control groups in the 6 months of the study exhibited that the LATCH points of the test group are higher than those of the control group and that the difference between the groups is statistically significant (Table 3). The literature has many studies indicating that education has an important effect on breast-feeding[10,12,22,23].

s in the 6 months of the study exhibited that the LATCH points of the test group are higher than those of the control group and that the difference between the groups is statistically significant (Table 3). The literature has many studies indicating that education has an important effect on breast-feeding[10,12,22,23]. In the study it was found that 85.7% of the mothers of the test group sustained exclusive breast-feeding during the 6 months following their release from the hospital, whereas only 38.9% of the mothers from the routine control group sustained exclusive breast-feeding in the same period (Table 4). The reason why the mothers from the test group sustained exclusive breast-feeding for a longer period than the control group did, is believed to be because within the first 5 days following the birth, mothers from the test group were given education by the researcher on breast-feeding techniques and the benefits of breast milk, which were controlled and supported again after 1 week, and the education was maintained through the 6 months postpartum.

s believed to be because within the first 5 days following the birth, mothers from the test group were given education by the researcher on breast-feeding techniques and the benefits of breast milk, which were controlled and supported again after 1 week, and the education was maintained through the 6 months postpartum. In previous research, it was determined that the breast-feeding periods of low-birth-weight and premature infants are considerably lower than those of normally born infants[7,8]. Moreover, in another study carried out in this field, it was exhibited that the breast-feeding durations of mothers of low-birth-weight infants are significantly lower than those who had given birth at term[6]. In their study carried out with the participation of mothers of low-birth-weight infants, Santoro-Junior and Martinez[24] (2007) determined that the education and support given to the mothers has positive effects on the duration of feeding their babies exclusively with breast milk. In addition, the study of Varol and Yıldız[12] (2006) emphasized that breast-feeding education and follow-up maintained for 6 months starting from the date of birth increases the duration of exclusive breast-feeding. Koskinen et al[25] confirmed that early breast-feeding experiences and maternity hospital practices have an association with maternal breast-feeding self-efficacy. Masters et al[26] determined that the program (a theory-based booklet and pre- and postnatal home visits by trained assistants) significantly improved exclusive breast-feeding rates at 6 months postpartum.

eding experiences and maternity hospital practices have an association with maternal breast-feeding self-efficacy. Masters et al[26] determined that the program (a theory-based booklet and pre- and postnatal home visits by trained assistants) significantly improved exclusive breast-feeding rates at 6 months postpartum. Because the hospital where the study was conducted was a baby-friendly hospital, no training and guidance was given to the mothers about formula feeding. That babies who received formula feeding or mixed feeding had lower weights in our study, may have resulted from the fact that mothers did not receive any training about formula feeding before hospital discharge and they tried to feed their babies according to their feeding-standards. Conclusion Breast-feeding education and home visits are affective for the promotion and implementation of breast-feeding. Our results show that an interactive education can increase the mother’s knowledge, management of breast-feeding practice, breast-feeding self-efficacy, and baby growth. The effect of introducing an interactive supportive breast-feeding education program may well be stronger in countries with less developed health care systems. It is necessary to increase the awareness of the nurses by providing them with on-the-job trainings so that they can train mothers and fathers who have low-birth-weight infants, about the importance of breast-feeding self-sufficiency and breast milk. Moreover, having nurses who are exclusively breast-feeding trainers will increase the quality of the training for mothers.

es by providing them with on-the-job trainings so that they can train mothers and fathers who have low-birth-weight infants, about the importance of breast-feeding self-sufficiency and breast milk. Moreover, having nurses who are exclusively breast-feeding trainers will increase the quality of the training for mothers. Acknowledgment The authors thank the personnel of the neonatal unit of Ataturk University Health Research and Application Hospital for their efforts in collection and documentation of the data. Conflict of Interest: None Authors’ Contribution All authors have carried out (design, acquisition of data, data analysis, manuscript preparation, critical revision of the manuscript) the study. All authors read and approved the final manuscript.

Introduction In Part I of this paper, we reviewed familial Mediterranean fever (FMF) and periodic fever, aphthous stomatitis, pharyngitis, and cervical adenitis (PFAPA) and repoted on our experience in Iran[1]. In Part II we try to review some other periodic fever syndromes known as autoinflammatory disorders. There are few reports of experience on these disorders in Iran. To establish diagnosis in autoinflammatory disorders is very difficult because there are no definite diagnostic criteria or a gold standard, so it depends on both clinical findings and genetic study to get the diagnosis confirmed (Table 1). All patients usually have a history of repeated hospitalizations and antibiotic therapy. It is due to similar symptoms and signs in autoinflammatory disorders and infectious diseases, immuno-deficiency and/or hematologic disorders. So in each patient, these disorders, especially infectious diseases, should be ruled out[1]. Hyperimmunoglobulin D syndrome (HIDS) HIDS or mevalonate kinase deficiency is a hereditary periodic fever syndrome that occurs in unpredictable intervals[2-4]. Dutch-type periodic fever is another name for this autosomal recessive disorder[5]. Epidemiology: First cases of this syndrome were reported in Dutch population in 1984 but later several patients were reported from other countries and in other races[2,6,7]. The syndrome usually starts in childhood and 70% of patients are younger than 1 year old at the beginning of the disease[3,5-7]. Sometimes fever attacks are stimu-lated with immunization, trauma, and stress[2].

h population in 1984 but later several patients were reported from other countries and in other races[2,6,7]. The syndrome usually starts in childhood and 70% of patients are younger than 1 year old at the beginning of the disease[3,5-7]. Sometimes fever attacks are stimu-lated with immunization, trauma, and stress[2]. Pathophysiology: This syndrome results from a mutation in a gene which is located on chromosome 12q24 and encodes mevalonate kinase (MVK) enzyme[2-4]. This enzyme contributes to cholesterol synthesis and it has an anti-inflammatory effect[2,3]. Mutated enzyme has a decreased function with a pro-inflammatory response[2,8]. Clinical manifestations: The typical symptoms of the disease are fever, malaise, chills, cervical lymphadenopathy, and diarrhea. Abdominal pain, arthritis or arthralgia, skin rash, and hepatosplenomegaly are other signs[2,5,6]. Fever and other symptoms resolve after 3 to 7 days with an afebrile interval lasting 6-8 weeks, but skin rash and arthritis may last longer[5,8,9]. Arthritis in HIDS is transient and non-destructive[3,10,11]. Diagnosis and differential diagnosis: Diagnosis of HIDS is based on clinical manifestations, elevated serum IgD level (>100), and low activity of MVK enzyme and is confirmed with detection of gene mutation[3]. Immunoglobulin D level should be measured twice with one month interval[7]. Level of IgD might not be very high in children younger than 3 years of age. Immunoglobulin A level in 80% of patients is high[6].

vated serum IgD level (>100), and low activity of MVK enzyme and is confirmed with detection of gene mutation[3]. Immunoglobulin D level should be measured twice with one month interval[7]. Level of IgD might not be very high in children younger than 3 years of age. Immunoglobulin A level in 80% of patients is high[6]. Treatment: There are no absolutely effective drugs for treatment of HIDs, thalidomide, etanercept (anti-tumor necrosis factor), and corticosteroids are used successfully in some patients, but efficacy of these drugs is doubt-ful[12-14]. Anti interleukine-1 (IL-1) (anakinra or canakinumab) has been effective in severe cases unresponsive to other treatments[10]. Prognosis: Prognosis of HIDS is excellent even without treatment[6], and amyloidosis is very rare in these patients[7,15]. Episodes of fever decreases by aging[16]. Hyper IgD in Iran: In our registry, we reported a HIDS case with concurrent intermittent neutropenia in a 15-month-old boy with good control under intermittent steroid therapy[17]. Tumor Necrosis Factor Receptor–Asso-ciated Periodic Syndrome (TRAPS) TRAPS or familial Hibernian fever (FHF)[7,10,18], is an autosomal dominant disease that occurs in irregular intervals and is characterized by episodes of fever, pain in muscle groups and painful skin lesions on trunk or extremi-ties[6,9,18,19].

Hyper IgD in Iran: In our registry, we reported a HIDS case with concurrent intermittent neutropenia in a 15-month-old boy with good control under intermittent steroid therapy[17]. Tumor Necrosis Factor Receptor–Asso-ciated Periodic Syndrome (TRAPS) TRAPS or familial Hibernian fever (FHF)[7,10,18], is an autosomal dominant disease that occurs in irregular intervals and is characterized by episodes of fever, pain in muscle groups and painful skin lesions on trunk or extremi-ties[6,9,18,19]. Epidemiology: For the first time, TRAPS was described in a large Irish family, but then it was reported in all countries[7,18]. The male to female ratio is 3:2[7] or 1:1[18] and FHF is more common in Caucasians and Arabs but less common in Indians[20]. This syndrome has been reported in patients 2 weeks to 53 years of age[8,9]. Etiology and Pathophysiology: The cause of this syndrome is mutation in the gene that encodes the type 1 TNF receptor[6,8,18,19]. The gene is TNFRSF1A and is located on chromosome 12p13.2[3,18,19,21]. TNFR1 is a trans-membrane receptor and the main mediator of signaling by TNF-α[22]. As binding of TNF-α to mutate TNFR1 lessens, as a result, TNF induced apoptosis decreases[3]. Consequently, following increase the activity of TNF, inflammatory response enhances[8]. Clinical manifestations: Similar to other periodic syndromes, fever is the main symptom in children, in adults attack may be without fever[3,9,18]. Fever duration in TRAPS usually is longer than in other periodic fever syndromes such as FMF or in hyper IgD syndrome and it may last up to 3 weeks[2,7,18,21,23,24].

Etiology and Pathophysiology: The cause of this syndrome is mutation in the gene that encodes the type 1 TNF receptor[6,8,18,19]. The gene is TNFRSF1A and is located on chromosome 12p13.2[3,18,19,21]. TNFR1 is a trans-membrane receptor and the main mediator of signaling by TNF-α[22]. As binding of TNF-α to mutate TNFR1 lessens, as a result, TNF induced apoptosis decreases[3]. Consequently, following increase the activity of TNF, inflammatory response enhances[8]. Clinical manifestations: Similar to other periodic syndromes, fever is the main symptom in children, in adults attack may be without fever[3,9,18]. Fever duration in TRAPS usually is longer than in other periodic fever syndromes such as FMF or in hyper IgD syndrome and it may last up to 3 weeks[2,7,18,21,23,24]. Table 1 Characteristics of classic periodic fever and cryopyrin associated periodic syndromes Autoinflmatory disorder Characteristics Hyper IgD syndrome Hereditary periodic fever syndromes with good prognosis. Clinical presentation is not specific and diagnosis is based on detection of gene mutation. TRAPS Recurrent fever with migratory muscle spasms with or without transient blanching erythematous skin lesions. Genetic assessment is necessary for definite diagnosis. CINCA or NOMID Recurrent fever, relapsing arthritis, urticaria, chronic meningitis and brain atrophy. Anti interlukine-1 drugs control the symptoms effectively Muckle-Wells Syndrome Fever, limb pain, recurrent urticaria-like lesions, conjunctivitis and deafness. Canakinumab was very effective in patients (complete remission). Familial Cold Autoinflammatory Syndrome

Recurrent fever, relapsing arthritis, urticaria, chronic meningitis and brain atrophy. Anti interlukine-1 drugs control the symptoms effectively Muckle-Wells Syndrome Fever, limb pain, recurrent urticaria-like lesions, conjunctivitis and deafness. Canakinumab was very effective in patients (complete remission). Familial Cold Autoinflammatory Syndrome Mildest disease of CAPS, clinical symptoms occur after exposure to cold. Fever, arthralgia, conjunctivitis, non-itching urticarial like rash, and headache. Treatment with anakinra before exposure to cold may be effective in prevention of inflammatory symptoms IgD: Immunoglobulin D; TRAPS: Tumor Necrosis Factor Receptor–Asso-ciated Periodic Syndrome; CINCA: Chronic Infantile Neurologic Cutaneous and Articular Syndrome; NOMID: Neonatal Onset Multisystem Inflammatory Disease; CAPS: Cryopyrin Associated Periodic Syndromes

Treatment with anakinra before exposure to cold may be effective in prevention of inflammatory symptoms IgD: Immunoglobulin D; TRAPS: Tumor Necrosis Factor Receptor–Asso-ciated Periodic Syndrome; CINCA: Chronic Infantile Neurologic Cutaneous and Articular Syndrome; NOMID: Neonatal Onset Multisystem Inflammatory Disease; CAPS: Cryopyrin Associated Periodic Syndromes Interval between fever episodes has variable frequency but in average it occurs every 5 to 6 weeks[8,18]. The onset of the disease occurs along with fever and muscular spasm[6,8,18]. No known triggers for the onset of the illness are identified[8]. The muscle pain is migratory and affected muscles are tender in palpation and warm[3,7,21,25]. This finding is usually associated with erythematous patches that are warm and tender and are resolved with pressure[2,3,9,18,25]. Other symptoms of TRAPS are abdominal pain (due to inflammation in peritoneal cavity or muscular cramp in abdominal wall) that is seen in 92% of patients, conjunctivitis or periorbital edema, which is a characteristic for TRAPS and is seen in over 80% of patients, chest pain (in 57% of patients), and arthralgia[3,6-8,18,19,24]. Other less common symptoms are arthritis, scrotal pain and lymphadenopathy[3,7].

scular cramp in abdominal wall) that is seen in 92% of patients, conjunctivitis or periorbital edema, which is a characteristic for TRAPS and is seen in over 80% of patients, chest pain (in 57% of patients), and arthralgia[3,6-8,18,19,24]. Other less common symptoms are arthritis, scrotal pain and lymphadenopathy[3,7]. Laboratory findings: Increased acute phase reactants such as erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) and normal muscle enzyme are laboratory findings during episodes but ESR may remain high in intervals[2,3,6,8,18,19]. Level of soluble TNFRSF1A may reduce during or between attacks, although normal level does not rule out TRAPS[7,19]. Diagnosis and differential diagnosis: Although Alvarez-Lobos et al introduced a diagnostic criteria for TRAPS, it has not been validated in adults or pediatric patients[26]. When the above mentioned symptoms are associated with the family history of similar symptoms and the response to glucocorticoids (but not to colchicine) TRAPS should be considered, but definite diagnosis is made with genetic testing[8,9,18]. Nowadays, more than 80 mutations of TNFRSF1A gene have been identified, but there are several patients with clinical symptoms of TRAPS without specific mutation[8,9,18]. FHF should be differentiated from other periodic fevers especially from FMF[3,9,18]. Other differential diagnoses are chronic infections and malignancies (lymphoma)[8,6,18].

mutations of TNFRSF1A gene have been identified, but there are several patients with clinical symptoms of TRAPS without specific mutation[8,9,18]. FHF should be differentiated from other periodic fevers especially from FMF[3,9,18]. Other differential diagnoses are chronic infections and malignancies (lymphoma)[8,6,18]. Treatment: Non steroidal anti inflammatory drugs (NSAIDs) are used for treatment of fever but musculoskeletal and abdominal pain do not respond to NSAIDs, so they should be treated with corticosteroids; neither of these drugs can prevent episodes[3]. Etanercept is an anti-TNF agent that decreases severity, duration, and frequency of the attacks[27]. Treatment with anakinra, which is an IL-1 blocker, may be also effective[22]. Treatment with infliximab, an anti-TNF antibody, may lead to paradoxical inflammatory reactions, so it should not be used[27,22]. Prognosis: The most severe complication of FHF is amyloidosis that occurs in less than 25% of untreated patients and manifests with dysfunction of involved organs[8,9]. So, prognosis of FHF is related to subsequent amyloidosis[3,7]. Risk of amyloidosis increases in mutation that affects cysteine amino acids[18].

Treatment: Non steroidal anti inflammatory drugs (NSAIDs) are used for treatment of fever but musculoskeletal and abdominal pain do not respond to NSAIDs, so they should be treated with corticosteroids; neither of these drugs can prevent episodes[3]. Etanercept is an anti-TNF agent that decreases severity, duration, and frequency of the attacks[27]. Treatment with anakinra, which is an IL-1 blocker, may be also effective[22]. Treatment with infliximab, an anti-TNF antibody, may lead to paradoxical inflammatory reactions, so it should not be used[27,22]. Prognosis: The most severe complication of FHF is amyloidosis that occurs in less than 25% of untreated patients and manifests with dysfunction of involved organs[8,9]. So, prognosis of FHF is related to subsequent amyloidosis[3,7]. Risk of amyloidosis increases in mutation that affects cysteine amino acids[18]. TRAPS in Iran: There is no report on TRAPS from Iran in the literature. We have a not reported case under observation in our periodic fever clinic. A 13 year old patient with recurrent fever and abdominal pain and a few patchy erythematous rashes on the trunk and face. Rashes appear with fever and disappear after fever attack. Hyper-IgD and FMF was ruled out with genetic study and trial treatment with colchicine. After starting of low dose oral prednisolone, duration and interval of the febrile attacks decreased significantly. Based on clinical presentation and response to treatment, we suggested TRAPS but genetic study has not been yet performed in this case. Cryopyrin Associated Periodic Syndromes (CAPS)

TRAPS in Iran: There is no report on TRAPS from Iran in the literature. We have a not reported case under observation in our periodic fever clinic. A 13 year old patient with recurrent fever and abdominal pain and a few patchy erythematous rashes on the trunk and face. Rashes appear with fever and disappear after fever attack. Hyper-IgD and FMF was ruled out with genetic study and trial treatment with colchicine. After starting of low dose oral prednisolone, duration and interval of the febrile attacks decreased significantly. Based on clinical presentation and response to treatment, we suggested TRAPS but genetic study has not been yet performed in this case. Cryopyrin Associated Periodic Syndromes (CAPS) CAPS refer to a group of diseases which have similar phenotypes all of which result from mutation in the CIAS1 (cold-induced autoinflammatory syndrome) gene, which encodes cryopyrin[28]. This protein regulates generation of pro-inflammatory cytokines[2,29]. Chronic infantile neurologic cutaneous and articular syndrome (CINCA), Muckle-Wells syndrome (MWS), and familial cold autoinflammatory syndrome (FCAS), which will be explained respectively, belong to CAPS. These syndromes usually have overlapping symptoms the severity of which are affected by other genetic mutations and environmental factors[28]. After ruling out infection and malignancies, auto-inflammatory disorders should be considered. However, physician needs to reevaluate each patient with periodic disorder for non-inflammatory syndromes. CINCA Syndrome

CAPS refer to a group of diseases which have similar phenotypes all of which result from mutation in the CIAS1 (cold-induced autoinflammatory syndrome) gene, which encodes cryopyrin[28]. This protein regulates generation of pro-inflammatory cytokines[2,29]. Chronic infantile neurologic cutaneous and articular syndrome (CINCA), Muckle-Wells syndrome (MWS), and familial cold autoinflammatory syndrome (FCAS), which will be explained respectively, belong to CAPS. These syndromes usually have overlapping symptoms the severity of which are affected by other genetic mutations and environmental factors[28]. After ruling out infection and malignancies, auto-inflammatory disorders should be considered. However, physician needs to reevaluate each patient with periodic disorder for non-inflammatory syndromes. CINCA Syndrome CINCA syndrome is the most severe one among the cryopyrin associated periodic syndromes[30]. These syndromes occur in all ethnic groups without any superiority[15]. CINCA syndrome that is also named neonatal onset multisystem inflammatory disease (NOMID) is a congenital inflammatory disorder that starts in neonatal period and it has various symptoms such as recurrent fever, central nervous system (CNS) involvement, cutaneous manifestations, chronic arthropathy, and morphologic feature[2,30,31]. Skin manifestations which are the first presentation at birth in 75% of patients, are similar to urticaria and vary during the day[31,32]. Urticaria is stimulated with cold. Central nervous system involvement exists in almost all patients and hydrocephalus and ventriculomegaly may be present prenatally[2]. Neurologic manifestations result from chronic meningitis and brain atrophy[30,32,33], and include seizure, sensory organ involvement such as ocular manifestations, progressive sensory neural hearing loss, and headache[3,31,32,33]. Mental retardation exists in some patients[30,33] but it is not an initial manifestation[32]. Relapsing joint involvement is also seen in CINCA patients[32] and severity of arthropathy is related to the age of onset, i.e. patients with a later onset have milder manifestations without destruction[2]. Knees are the most common involved joints and then ankles, feet, and elbows[2]. Patients with CINCA have a typical morphology. They have a short stature, macrocephalus, hoarseness, saddle nose, short extremities, and clubbing[2,3,31,33].

.e. patients with a later onset have milder manifestations without destruction[2]. Knees are the most common involved joints and then ankles, feet, and elbows[2]. Patients with CINCA have a typical morphology. They have a short stature, macrocephalus, hoarseness, saddle nose, short extremities, and clubbing[2,3,31,33]. Diagnosis of CINCA is based on the above mentioned specific features[24,30,31]. Triad of continuous rash, CNS involvement, and relapsing joint involvement is helpful for diagnosis but this syndrome should be differentiated from other hereditary periodic fever syndromes such as FMF, TRAPS, and HIDS. The majority of these syndromes occur sporadically[32] but there is a mutation in the CIAS1 (now named NLRP3) gene on chromosome 1q44 [2] that encodes the cryopyrin protein in 60% of patients[29,32,34,35]. However, this mutation can be found in two other syndromes including MWS and FCAS[3,7,21,30]. Treatment with anti IL-1 drugs especially with anakinra is effective in CINCA and results in significant improvement in symptoms of the disease[3,30,34,36]. Nonsteroidal anti-inflammatory drugs and glucocorticosteroids are effective on fever and pain but not on skin lesions [3].

The majority of these syndromes occur sporadically[32] but there is a mutation in the CIAS1 (now named NLRP3) gene on chromosome 1q44 [2] that encodes the cryopyrin protein in 60% of patients[29,32,34,35]. However, this mutation can be found in two other syndromes including MWS and FCAS[3,7,21,30]. Treatment with anti IL-1 drugs especially with anakinra is effective in CINCA and results in significant improvement in symptoms of the disease[3,30,34,36]. Nonsteroidal anti-inflammatory drugs and glucocorticosteroids are effective on fever and pain but not on skin lesions [3]. CINCA in Iran: Similar to some other rare autoinflammatory disorders, there is no report on CINCA from Iran in the literature. We have 3 suggestive cases to CINCA in our periodic fever clinic. One of them was a 3 month girl with recurrent fever form neonatal period, skin rashes, seizure and hepatosplenomegaly. There was no positive culture for infection. Evaluation for primary immunodeficiency and hemophagocytic lymphohistiocytosis syndrome was negative. After 2 months, she was referred to our clinic. In physical examination, we found a prominent joint. Anakinra was not available in Iranian pharmacopeia. This case was treated with methylprednisolone pulse for 3 days followed by oral prednisolone and ibuprofen. During 1 year follow-up she had 2 fever attacks and she was hospitalized for more evaluation. We did not find any new clinical and paraclinical findings. After 1 year follow-up we tapered prednisolone and now she is on low dose prednisolone (2.5 mg/day) and ibuprofen. Now, she is 18 months old with a small delay in motor development. She had a few episodes of fever and skin rash during last 6 months.

valuation. We did not find any new clinical and paraclinical findings. After 1 year follow-up we tapered prednisolone and now she is on low dose prednisolone (2.5 mg/day) and ibuprofen. Now, she is 18 months old with a small delay in motor development. She had a few episodes of fever and skin rash during last 6 months. Second case was a 6 month old girl referred to our pediatric rheumatology division. She had a history of fever and allergic skin rash from infancy and 2 times admission with osteomyelitis or arthritis suggestion. There was no positive findings for infection but she was treated with antimicrobial agents in the hospital. Urticaria-like skin rashes appeared with fever and disappeared after fever control. We consider autoinflamatory disorders for this patient. Hearing and visual evaluations did not show any significant impairment. She was treated with prednisolone and ibuprofen and she was candidate for treatment with anakinra. After discharge, she had recurrent periodic fever, urticaria-like skin rashes, failure to thrive and motor developmental delay. After 4 months she was treated with anakinra and after 5-10 doses of injection of ankinra, symptoms and signs disappeared. Fever attack was continuing with reduced severity after treatment with ankinra. After 14 months follow-up, she is 20 months old and has attacks of recurrent fever and skin rashes. Acute phase reactants (ESR, CRP) are increased in each attack and reduce after fever control but ESR has not been normal between attacks. Genetic study was performed in our cases, we are waiting for the result.

ankinra. After 14 months follow-up, she is 20 months old and has attacks of recurrent fever and skin rashes. Acute phase reactants (ESR, CRP) are increased in each attack and reduce after fever control but ESR has not been normal between attacks. Genetic study was performed in our cases, we are waiting for the result. Third case was a 3 month old boy with a history of recurrent arthritis and multifocal osteomyelitis. There were no positive serologic findings, positive culture for infection or findings for primary immunodeficiency disorders. Continuous fever, skin rash and arthritis without any response to antibiotic therapy leaded us to autoinflammatory disorders. Based on history and examination and a delay in developmental millstones CINCA was suggested, so he was treated with prednisolone plus ibuprofen and he was candidate for treatment with anakinra. After control of fever, he was discharged. He did not appear for follow-up. Genetic study was not performed in these cases. Muckle-Wells Syndrome (MWS)

Third case was a 3 month old boy with a history of recurrent arthritis and multifocal osteomyelitis. There were no positive serologic findings, positive culture for infection or findings for primary immunodeficiency disorders. Continuous fever, skin rash and arthritis without any response to antibiotic therapy leaded us to autoinflammatory disorders. Based on history and examination and a delay in developmental millstones CINCA was suggested, so he was treated with prednisolone plus ibuprofen and he was candidate for treatment with anakinra. After control of fever, he was discharged. He did not appear for follow-up. Genetic study was not performed in these cases. Muckle-Wells Syndrome (MWS) MWS also called urticarial deafness amyloidosis syndrome, is a rare autosomal dominant syndrome[9,21]. It is of middle severity between CAPS. Episodes of this syndrome are presented with fever, limb pain, recurrent or sub-chronic urticaria-like lesions and conjunctivitis. Episcleritis and optic disc edema are other eye related findings[24]. Fever is usually mild and disappears in 12 to 36 hours[22]. Two-thirds of patients have sensory neural hearing loss that usually starts in the second decade of life[2]. Amyloidosis occurs in one-fourth of patients which can lead to renal failure[36] and peripheral neuropathy that occurs later than renal amyloidosis[2]. Unlike other CAPS, urticarial rash is not always stimulated with cold[7,9]. The time of onset is during infancy and sometimes in adolescence. Lifelong arthralgia and non-erosive arthritis may exist[34]. There is a mutation in NLRP3 gene in MWS as in other CAPS, but some mutations are typically found in MWS[7,35,37]. Incidence of CAPS syndromes is not subject to sex [7,34].

ated with cold[7,9]. The time of onset is during infancy and sometimes in adolescence. Lifelong arthralgia and non-erosive arthritis may exist[34]. There is a mutation in NLRP3 gene in MWS as in other CAPS, but some mutations are typically found in MWS[7,35,37]. Incidence of CAPS syndromes is not subject to sex [7,34]. Three anti-IL-1 drugs including anakinra (an IL-1 receptor antagonist), rilonacept (an IL-1 soluble receptor), and canakinumab (a human monoclonal antibody against IL-1β) are effective on CAPS. Canakinumab and rilonacept have FDA approval for treatment of FCAS and MWS[2,7,37]. Treatment with canakinumab was very effective and in 96% of patients complete remission was achieved[38]. Studies show that treatment cannot eradicate existing damages but hearing loss may return[2]. Muckle-Wells Syndrome in Iran: There is no report on Muckle-Wells Syndrome from Iran. Familial Cold Autoinflammatory Syndrome (FCAS)

Three anti-IL-1 drugs including anakinra (an IL-1 receptor antagonist), rilonacept (an IL-1 soluble receptor), and canakinumab (a human monoclonal antibody against IL-1β) are effective on CAPS. Canakinumab and rilonacept have FDA approval for treatment of FCAS and MWS[2,7,37]. Treatment with canakinumab was very effective and in 96% of patients complete remission was achieved[38]. Studies show that treatment cannot eradicate existing damages but hearing loss may return[2]. Muckle-Wells Syndrome in Iran: There is no report on Muckle-Wells Syndrome from Iran. Familial Cold Autoinflammatory Syndrome (FCAS) This syndrome, formerly called familial cold urticaria, is the mildest CAPS disease[2,8,29,39]. As other CAPS, FCAS is inherited in an autosomal dominant fashion[9]. FCAS usually begins before 6 months of age[23,33] and in 60% of patients symptoms are seen during the first days of life[2]. Clinical symptoms occur within 8 hours[9,40] after generalized exposure to cold[33] and last for 12 to 48 hours[8,9]. Fever, arthralgia, conjunctivitis, non-itching urticarial like rash, and headache are indicators of FCAS[8,9,29]. There are no signs of CNS involvement[28]. Audiology symptoms and deafness is absent[34]. Localized exposure to cold is not a trigger in this disease and it distinguishes FCAS from acquired cold urticaria[33].

er, arthralgia, conjunctivitis, non-itching urticarial like rash, and headache are indicators of FCAS[8,9,29]. There are no signs of CNS involvement[28]. Audiology symptoms and deafness is absent[34]. Localized exposure to cold is not a trigger in this disease and it distinguishes FCAS from acquired cold urticaria[33]. Treatment with anakinra before exposure to cold may be effective in prevention of inflammatory symptoms[23]. Moving to warm places is recommended to these patients[41]. In all three diseases of CAPS, treatment with anakinra is effective[25]. Rilonacept and canakinumab as long acting IL1 blockers have been approved by FDA for treatment of FCAS and MWS[23,28,37]. Long time prognosis of FCAS is good and amyloidosis is uncommon [9]. Autoinflammatory Bone Disorders Autoinflammatory Bone Disorders are a group of autoinflammatory disorders with sterile bone inflammation as a hallmark finding. Some of the disorders of this group are monogenic such as pyogenic arthritis, pyoderma gangraenosum and acne (PAPA) syndrome, the deficiency of IL-1 receptor antagonist (DIRA) and Majeed syndrome (or familial chronic multifocal osteomyelitis). Some other sterile osteitises have polygenic background or are without specific genetic basis, so these disorders are classified as sporadic group,. In which chronic recurrent multifocal osteomyelitis (CRMO) and synovitis, acne, pustulosis, hyperostosis and osteitis (SAPHO) syndrome are also classified[42]. Pyogenic arthritis, Pyoderma gangaenosum and Acne (PAPA) syndrome

Autoinflammatory Bone Disorders are a group of autoinflammatory disorders with sterile bone inflammation as a hallmark finding. Some of the disorders of this group are monogenic such as pyogenic arthritis, pyoderma gangraenosum and acne (PAPA) syndrome, the deficiency of IL-1 receptor antagonist (DIRA) and Majeed syndrome (or familial chronic multifocal osteomyelitis). Some other sterile osteitises have polygenic background or are without specific genetic basis, so these disorders are classified as sporadic group,. In which chronic recurrent multifocal osteomyelitis (CRMO) and synovitis, acne, pustulosis, hyperostosis and osteitis (SAPHO) syndrome are also classified[42]. Pyogenic arthritis, Pyoderma gangaenosum and Acne (PAPA) syndrome PAPA syndrome is a rare autosomal-dominant disease that characterizes with sterile inflammation in joints and cutaneous manifestation. The cause of the disease is mutation in PSTPIP1 (proline/serine/threonine phosphatase–interacting protein 1) located on chromosome 15q24-25[34]. This gene produces a protein that mostly exists in hematopoietic cells and regulates T cell activation, cytoskeletal organization and releases of interleukin-1B (IL-1B). Mutation in this gene leads to dysregulation of IL-1B production 77 and macrophages. PAPA syndrome patients have impaired invasive motility 77 and might have dysregulated apoptosis[34].

ostly exists in hematopoietic cells and regulates T cell activation, cytoskeletal organization and releases of interleukin-1B (IL-1B). Mutation in this gene leads to dysregulation of IL-1B production 77 and macrophages. PAPA syndrome patients have impaired invasive motility 77 and might have dysregulated apoptosis[34]. Two major manifestations of PAPA syndrome are erosive arthritis and cutaneous syndromes both of which start in childhood. Joints involvement is recurrent, sterile and destructive and may develop spontaneously or after minor trauma. Skin symptoms of the disease are severe acne, recurrent non healing sterile ulcer called pyoderma gangraenosum and pathergy. For treatment, the disease responds well to TNFα blockers such as Infliximab, adalimumab and anakinra. Some new immunosuppressive drugs may also be used to treat PAPA syndrome. Majeed Syndrome Majeed Syndrome is an autosomal recessive disease caused from a mutation in LPIN243. The disease was firstly reported in 1989[9]. Majeed syndrome is a chronic disease which affects the whole life and is characterized by early onset of multifocal osteomyelitis. Other features of Majeed syndrome are congenital anemia and transient inflammatory dermatosis which result from infiltration of neutrophils into dermis and is called Sweet syndrome. Diagnosis is suspected based on clinical manifestations and is confirmed with genetical studies[34]. Treatment of the disease includes non-steroidal anti-inflammatory drugs eventually in combination with corticosteroids or interferon-α. Deficiency of the Interlukin-1 Receptor Antagonist (DIRA)

Majeed Syndrome is an autosomal recessive disease caused from a mutation in LPIN243. The disease was firstly reported in 1989[9]. Majeed syndrome is a chronic disease which affects the whole life and is characterized by early onset of multifocal osteomyelitis. Other features of Majeed syndrome are congenital anemia and transient inflammatory dermatosis which result from infiltration of neutrophils into dermis and is called Sweet syndrome. Diagnosis is suspected based on clinical manifestations and is confirmed with genetical studies[34]. Treatment of the disease includes non-steroidal anti-inflammatory drugs eventually in combination with corticosteroids or interferon-α. Deficiency of the Interlukin-1 Receptor Antagonist (DIRA) Deficiency of the IL-1 receptor antagonist is a new TNF-receptor-associated periodic syndrome which was primarily described in 2009[33]. The disease results from mutation in the IL1RN (2q14)[27]. It is an autosomal recessive disease. Signs of the disease usually start in prenatal period and include systemic inflammation, multifocal osteolytic lesions, periostitis, and a pustular rash. Other manifestations are heterotopic bone formation around the proximal femur, thrombosis, and rarely vasculitis. Some manifestations which are seen in NOMID but not in DIRA are meningitis, cochlear inflammation, hearing loss, and conjunctivitis or uveitis. DIRA patients have a good response to treatment with the recombinant IL-1 receptor antagonist anakinra[29].

mation around the proximal femur, thrombosis, and rarely vasculitis. Some manifestations which are seen in NOMID but not in DIRA are meningitis, cochlear inflammation, hearing loss, and conjunctivitis or uveitis. DIRA patients have a good response to treatment with the recombinant IL-1 receptor antagonist anakinra[29]. Autoinflammatory Bone Disorders in Iran: There is no report on autoinflammatory bone disorders with monogenic pattern from Iran. In our experience, a 5 months old boy was referred to our periodic fever clinic. He had a history of soft tissue abscess, arthritis and osteomyelitis after 2 series of vaccination. All evaluation for infection and primary immunodeficiency was negative. Periodic fever, osteomyelitis and prominent articular joints were other symptoms. Visual and hearing evaluation was normal but he had an episode of otitis media with otorrhea. No hepatosplenomegaly or skin rashes were present. The symptoms were compatible with those of auto-inflammatory bone disorders, although we considered CINCA as a diagnosis. Genetic study for DIRA was negative. Further study to rule out/in autoinflammatory disorders is not available now. The symptoms were reduced with prednisolone and Iboprofen, but the disease is not fully controlled. Synovitis, Acne, Pustulosis, Hyperostosis and Osteitis (SAPHO) syndrome or Chronic Recurrent Multifocal Osteomyelitis (CRMO)

Autoinflammatory Bone Disorders in Iran: There is no report on autoinflammatory bone disorders with monogenic pattern from Iran. In our experience, a 5 months old boy was referred to our periodic fever clinic. He had a history of soft tissue abscess, arthritis and osteomyelitis after 2 series of vaccination. All evaluation for infection and primary immunodeficiency was negative. Periodic fever, osteomyelitis and prominent articular joints were other symptoms. Visual and hearing evaluation was normal but he had an episode of otitis media with otorrhea. No hepatosplenomegaly or skin rashes were present. The symptoms were compatible with those of auto-inflammatory bone disorders, although we considered CINCA as a diagnosis. Genetic study for DIRA was negative. Further study to rule out/in autoinflammatory disorders is not available now. The symptoms were reduced with prednisolone and Iboprofen, but the disease is not fully controlled. Synovitis, Acne, Pustulosis, Hyperostosis and Osteitis (SAPHO) syndrome or Chronic Recurrent Multifocal Osteomyelitis (CRMO) SAPHO and CRMO are chronic nonbacterial or sterile osteomyelitises due to an auto-inflammatory process. Some authorities believe this disorders are different diseases with similar pathway and they have common clinical, radiological and maybe histological findings[43,44]. Innate immune system is involved and any auto-antibodies or autoreactive T-cells have not been fouind in these disorders. SAPHO is usually described in adult patients with similar symptoms of spondylarthropathies[45]. Vertebral bone is the most common site of involvement and it may lead to vertebral collapse. In SAPHO syndrome, skin involvement such as acne and pustulosis is prominent. CRMO is suggested to be a variant of SAPHO syndrome in children[46], but in CRMO skin manifestations are rare findings[47]. Exacerbations and remissions of the inflammation is a hallmark for differentiation of these disorders from similar disorders. Increasing number of lesions over the time and self-limiting course without major sequelae are other characteristics. The patients present symptoms of CRMO at school age that include bone lesions. One fourth of the patients had spinal involvement and half of them developed vertebral deformities[29].

similar disorders. Increasing number of lesions over the time and self-limiting course without major sequelae are other characteristics. The patients present symptoms of CRMO at school age that include bone lesions. One fourth of the patients had spinal involvement and half of them developed vertebral deformities[29]. Treatment is based on NSAIDs and/or corticosteroids. In cases with resistance to these drugs methotrexate, sulfasalazine, and TNF inhibitors are used. CRMO is usually self-limited but a large number of patients experience a prolonged disease[29]. In patients with bone lesions, treatment with bisphosphonates and pamidronate may be helpful and pain resolves 3 months after treatment[48]. MRI is the most sensitive way of diagnosing bone lesions and finding those lesions that are not observed in a simple radiography. CRMO in Iranian patients: There are 2 reports on CRMO from Iran. Both were girls 4.5 and 12 years old. In the latter patient the disease had association with ulcerative colitis[49,50]. We have a 3.5 years old patient with recurrent osteomyelitis and arthritis of the elbow, wrist and humerus. This case will be published in near future in detail[51]. SWEET’s Syndrome

CRMO in Iranian patients: There are 2 reports on CRMO from Iran. Both were girls 4.5 and 12 years old. In the latter patient the disease had association with ulcerative colitis[49,50]. We have a 3.5 years old patient with recurrent osteomyelitis and arthritis of the elbow, wrist and humerus. This case will be published in near future in detail[51]. SWEET’s Syndrome In 1964, Sweet described a syndrome with fever and neutrophilic dermatitis[52]. Three types of disease were described by Cohen and Kurzrock[53-55]. Malignancy-associated form can be associated with hematologic malignancy (such as acute leukemia) and solid tumors. It can be first manifestation of a malignant disorder or first presentation of recurrent malignancy[109,110]. Second form is classical form which usually is seen following a respiratory or gastrointestinal infection, pregnancy or rheumatologic disorders (such as irritable bowel disease, systemic lupus erythematosus, Behcet’s disease, FMF and Sarcoidosis)[53,54,56]. Third form of Sweet’s syndrome is drug induced following antibiotics (trimethoprim-sulfamethoxazole, nitrofurantoin), NSAIDs (celecoxib and diclofenac), antiepileptics (diazepam and carbamazepine), antihypertensives (hydralazine), oral contraceptives, propylthiouracil and retinoids[53,56]. Pathogenesis of this syndrome is unknown. Some Cytokines (such as Interleukin 1, TNF-alpha, IL-8, IL-17), human leukocyte antigen serotypes have a possible role in the pathogenesis of this syndrome[57-59]. The innate immune is involved in this syndrome, so recently it is classified as a autoinflmmatory disorder[57].

In 1964, Sweet described a syndrome with fever and neutrophilic dermatitis[52]. Three types of disease were described by Cohen and Kurzrock[53-55]. Malignancy-associated form can be associated with hematologic malignancy (such as acute leukemia) and solid tumors. It can be first manifestation of a malignant disorder or first presentation of recurrent malignancy[109,110]. Second form is classical form which usually is seen following a respiratory or gastrointestinal infection, pregnancy or rheumatologic disorders (such as irritable bowel disease, systemic lupus erythematosus, Behcet’s disease, FMF and Sarcoidosis)[53,54,56]. Third form of Sweet’s syndrome is drug induced following antibiotics (trimethoprim-sulfamethoxazole, nitrofurantoin), NSAIDs (celecoxib and diclofenac), antiepileptics (diazepam and carbamazepine), antihypertensives (hydralazine), oral contraceptives, propylthiouracil and retinoids[53,56]. Pathogenesis of this syndrome is unknown. Some Cytokines (such as Interleukin 1, TNF-alpha, IL-8, IL-17), human leukocyte antigen serotypes have a possible role in the pathogenesis of this syndrome[57-59]. The innate immune is involved in this syndrome, so recently it is classified as a autoinflmmatory disorder[57]. Clinical features of this syndrome are a recurrent and acute illness with high fever, peripheral neutrophillia, and skin involvement such as erythematous plaques, erythema nodosum and ulcers. Other manifestations are general malaise, headache, and myalgia, arthralgia, and conjunctivitis[53,54]. Central nervous system (aseptic meningitis), kidneys, intestines, liver, heart, and lungs may be involved[54]. Diagnostic criteria for classic and drug-induced Sweet syndrome is shown in Table 2. Criteria for malignancy-associated Sweet’s syndrome are similar to classic Sweet’s syndrome.

gia, and conjunctivitis[53,54]. Central nervous system (aseptic meningitis), kidneys, intestines, liver, heart, and lungs may be involved[54]. Diagnostic criteria for classic and drug-induced Sweet syndrome is shown in Table 2. Criteria for malignancy-associated Sweet’s syndrome are similar to classic Sweet’s syndrome. In histopathology, diffuse and neutrophil-rich infiltrate is seen in the dermis[54,60]. There is usually no evidence of vasculitis (fibrin deposition or neutrophils) in the vessel wall[61]. There are some reports on successful treatment of Sweet’s syndrome with antibiotics, oral potassium iodide, NSAIDs and colchicines; oral corticosteroids or metylprednolone pulse is very useful in patients with refractory disease[54,62]. Other treatments are infusion of intravenous immunoglobulin and biologic agents such as anakinra and anti TNF-α agants[62]. Sweet’s syndrome in Iranian patients: There is a report on 15 adult cases from Iran with classic form of Sweet’s syndrome[63]. All of these patients were female. In our experience, we have a 7 year old boy with recurrent fever, hyper leukocytosis (>40000 in febrile episode) and skin abscess formation or panniculitis. Evaluations for immunodeficiency and leukocyte adhesion deficiency, all were negative. Diagnosis was confirmed by histopathology study. The disease attack was controlled with oral steroid in combination with colchicine. This case is under report[64]. Table 2 Diagnostic criteria for classic and drug-induced Sweet’s syndrome[65] Classical Sweet’s syndrome

Sweet’s syndrome in Iranian patients: There is a report on 15 adult cases from Iran with classic form of Sweet’s syndrome[63]. All of these patients were female. In our experience, we have a 7 year old boy with recurrent fever, hyper leukocytosis (>40000 in febrile episode) and skin abscess formation or panniculitis. Evaluations for immunodeficiency and leukocyte adhesion deficiency, all were negative. Diagnosis was confirmed by histopathology study. The disease attack was controlled with oral steroid in combination with colchicine. This case is under report[64]. Table 2 Diagnostic criteria for classic and drug-induced Sweet’s syndrome[65] Classical Sweet’s syndrome Major Criteria (both are mandatory) Abrupt onset of painful erythematous plaques or nodules. Histopathologic evidence of a dense neutrophilic infiltrate without evidence of leukocytoclastic vasculitis. Minor Criteria Pyrexia: T>38°C. Association with an underlying hematologic or visceral malignancy, inflammatory disease, pregnancy, or preceded by an upper respiratory or gastrointestinal infection or vaccination. Excellent response to treatment with systemic corticosteroids or potassium iodide. Abnormal laboratory findings (3 of 4): ESR>20mm/hr / positive CRP / WBC>8000 /Neutrophilia > 70%. Drug-induced Abrupt onset of painful erythematous plaques or nodules. Histopathologic evidence of a dense neutrophilic infiltrate. Pyrexia: T>38°C. Temporal relationship between drug ingestion and clinical presentation, or temporally related recurrence after oral challenge.

ESR>20mm/hr / positive CRP / WBC>8000 /Neutrophilia > 70%. Drug-induced Abrupt onset of painful erythematous plaques or nodules. Histopathologic evidence of a dense neutrophilic infiltrate. Pyrexia: T>38°C. Temporal relationship between drug ingestion and clinical presentation, or temporally related recurrence after oral challenge. Temporally related resolution of lesions after drug withdrawal or treatment with systemic corticosteroids. Diagnosis of classic Sweet’s syndrome is confirmed by 2 major criteria and two of the four minor criteria but for diagnosis of drug-induced Sweet’s syndrome all five criteria are mandatory. Blau Syndrome Blau syndrome or “juvenile systemic granulomatosis”[34] or early onset sarcoidosis[9,33] was reported for the first time in 1985[9]. It is a rare disease related to mutation in the CARD15/NOD2 gene localized on chromosome 16q12[34]. The disease is autosomal dominant and begins in early childhood usually before 4 years of age[9,35].

or “juvenile systemic granulomatosis”[34] or early onset sarcoidosis[9,33] was reported for the first time in 1985[9]. It is a rare disease related to mutation in the CARD15/NOD2 gene localized on chromosome 16q12[34]. The disease is autosomal dominant and begins in early childhood usually before 4 years of age[9,35]. The first episode of Blau syndrome is in the first 2 years of age with continuous attack[33]. The most common sites of this syndrome are skin, joints and eyes but not lungs or hilar lymph nodes. Skin lesions which are rarely seen in sarcoidosis in adults include brown colored scaly maculopapules with tapioca-like appearance which are lichenoid-like. Another skin manifestation is erythema nodosum[9,33]. Destructive arthritis and tendinitis are other manifestations of musculoskeletal involvement[33]. Pneumonitis, eye involvement (uveitis and iritis) and involvement of other organs such as liver and kidney is seen in this syndrome[33]. Characteristic feature of the disease is non-caseating granulomatous lesions involving joints, skin and eyes that lead to symmetric polyarthritis and cataract in 50% of patients. Differentiation of Blau syndrome from sarcoidosis based on histological findings is difficult but clinical symptoms are clearly different[9]. Treatment options for Blau syndrome are corticosteroids, immunosuppressant drugs (such as methotrexate and cyclosporine), infliximab or anakinra[34]. Blau in Iran: There is no report on Blau syndrome from Iran in the literature and we have no confirmed or suggestive cases with this syndrome in our system registry.

The first episode of Blau syndrome is in the first 2 years of age with continuous attack[33]. The most common sites of this syndrome are skin, joints and eyes but not lungs or hilar lymph nodes. Skin lesions which are rarely seen in sarcoidosis in adults include brown colored scaly maculopapules with tapioca-like appearance which are lichenoid-like. Another skin manifestation is erythema nodosum[9,33]. Destructive arthritis and tendinitis are other manifestations of musculoskeletal involvement[33]. Pneumonitis, eye involvement (uveitis and iritis) and involvement of other organs such as liver and kidney is seen in this syndrome[33]. Characteristic feature of the disease is non-caseating granulomatous lesions involving joints, skin and eyes that lead to symmetric polyarthritis and cataract in 50% of patients. Differentiation of Blau syndrome from sarcoidosis based on histological findings is difficult but clinical symptoms are clearly different[9]. Treatment options for Blau syndrome are corticosteroids, immunosuppressant drugs (such as methotrexate and cyclosporine), infliximab or anakinra[34]. Blau in Iran: There is no report on Blau syndrome from Iran in the literature and we have no confirmed or suggestive cases with this syndrome in our system registry. What is new in autoinflammatory disorders?

Treatment options for Blau syndrome are corticosteroids, immunosuppressant drugs (such as methotrexate and cyclosporine), infliximab or anakinra[34]. Blau in Iran: There is no report on Blau syndrome from Iran in the literature and we have no confirmed or suggestive cases with this syndrome in our system registry. What is new in autoinflammatory disorders? A new group of autoinflammatory disorders are recently reported with no response to anti IL-1 agents[66,67]. Three anti IL-1 agents have been approved by FDA for autoinflammatory disorders. Rilonacept and canakinumab are long acting and anakinra is a short acting IL-1 inhibitor[67]. New classification of monogenic autoinflammatory syndromes has been suggested based on completely responsive, variably responsive and unresponsive to anti IL-1 agents (Table 3)[66]. Another classification is based on clinical features of autoinflammatory disorders[67]. In this classification 6 groups of autoinflammatory disorders have been described. The first group is classic periodic fever syndromes that present with recurrent fever, skin rashes and abdominal pain (FMF, TRAPS, HIDS). The second group is cryopyrinopathies that present with neutrophilic urticaria rashes (CINCA, FCAS, Muckle-Wells syndrome). Blau syndrome is authinflammatory syndrome in the third group that presents with granulomatous skin lesions and minimal or low-grade fever[67].

, skin rashes and abdominal pain (FMF, TRAPS, HIDS). The second group is cryopyrinopathies that present with neutrophilic urticaria rashes (CINCA, FCAS, Muckle-Wells syndrome). Blau syndrome is authinflammatory syndrome in the third group that presents with granulomatous skin lesions and minimal or low-grade fever[67]. The fourth group is presented with pustular skin rashes and episodic or continuous fever. Autoinflammatory bone disorders (including DIRA, Majeed syndrome), early onset inflammatory bowel disease, deficiency of interleukin 36 receptor antagonist and CARD14-mediated psoriasis are categorized in this group. The fifth group includes poteasome associated autoinflammatory diseases that present with atypical neutrophilic dermatosis and histiocytic-like infiltrate. Autoinflammation disorders with immunodeficiency are the sixth group of autoinflammatory diseases. Autoinflammation and PLCγ 2-associated antibody deficiency (1) and immune dysregulation, PLCγ 2-associated antibody deficiency (2), and immune dysregulation, and HOIL-1 deficiency (3) are 3 known disorders in this group[67]. Table 3 New classification of autoinflammatory syndromes Category Disorder Genetic mutation Responder to IL-1 agents (IL-1 mediated) Classic autoinflammatory diseases Familial Mediterranean fever TRAPS HIDS/mevalonate kinase deficiency Monogenic autoinflammatory bone diseases DIRA Majeed syndrome Cryopyrinopathies Familial cold autoinflammatory syndrome Muckle-Wells syndrome NOMID or CINCA MEFV gene TNFRSF1A gene MVK gene IL1RN gene LPIN2 gene NLRP3 gene NLRP3 gene NLRP3 gene Partially or variable responses to Anti-IL-1 agents PAPA

Familial Mediterranean fever TRAPS HIDS/mevalonate kinase deficiency Monogenic autoinflammatory bone diseases DIRA Majeed syndrome Cryopyrinopathies Familial cold autoinflammatory syndrome Muckle-Wells syndrome NOMID or CINCA MEFV gene TNFRSF1A gene MVK gene IL1RN gene LPIN2 gene NLRP3 gene NLRP3 gene NLRP3 gene Partially or variable responses to Anti-IL-1 agents PAPA Blau syndrome HOIL-1 deficiency APLAID PSTPIP1 gene CARD15/NOD2 gene HOIL1 PLCG2 Unresponsive to IL-1 (Non- IL-1 Mediated) Deficiency of IL-36 receptor antagonist PRAAS/CANDLE CARD14-mediated psoriasis Early-Onset IBD IL36RN gene PSMB8 CARD14 gene IL10, IL10RA and/or IL10RB TRAPS: Tumor necrosis factor receptor–associated periodic syndrome; HIDS: Hyperimmunoglobulin D syndrome; DIRA: deficiency of interleukin 1 receptor antagonist; NOMID: Neonatal-onset multisystem inflammatory disease; CINCA: Chronic infantile neurologic cutaneous and articular; PAPA: Pyogenic sterile arthritis, pyoderma gangraenosum, and acne syndrome; APLAID: Autoinflammation and PLCγ 2-associated antibody deficiency and immune dysregulation; PRAAS/CANDLE: Proteasome-associated autoinflammatory syndromes or chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature. Acknowledgment After the first part of this paper was published, authors received a letter from Dr. F. Salehzadeh indicating that the first pediatric case series of FMF in Iranian children was presented in the 7th International Congress of Pediatrics (in Tehran) with 13 cases[68] and PFAPA was reported in the 13th International Congress of Pediatrics (in Tehran) with 4 cases[69].

d, authors received a letter from Dr. F. Salehzadeh indicating that the first pediatric case series of FMF in Iranian children was presented in the 7th International Congress of Pediatrics (in Tehran) with 13 cases[68] and PFAPA was reported in the 13th International Congress of Pediatrics (in Tehran) with 4 cases[69]. Increasing number of mutations of MEFV gene in the registry of hereditary auto-inflammatory disorders was another comment of him. Total current number of mutations for MEFV is 296 mutations up to now (Apr 2014)[70]. The authors wish to thank Dr. N. Parvaneh and F. Salehzadeh for his valuable comments. Conflict of Interest: None

Introduction In modern medical ethics, patient autonomy is considered a major principle in making decisions about an individual’s health, and those who receive healthcare should have the right to practice their autonomy consciously and freely; healthcare providers, on the other hand, are obligated to respect this right and allow patients to practice their autonomy in the course of their treatment[1]. In cases where a patient cannot exercise this right due to his or her limited ability to make medical decisions – a condition referred to as lack of capacity – a qualified person will proceed to make such decisions as the patient’s surrogate based on his or her best interests[2]. It should be noted that an individual’s autonomy in legitimate matters is a logical notion acknowledged by the Islamic Fiqh, as long as it does not jeopardize the sanctity of human life[3]. For this reason, according to the Paragraph 2 of Article 59 of the Islamic Penal Code of Iran, all medical and surgical procedures must be performed with the approval of the patients, their parents, guardians or legal representatives, and with due consideration for technical, scientific, and government regulations.

3]. For this reason, according to the Paragraph 2 of Article 59 of the Islamic Penal Code of Iran, all medical and surgical procedures must be performed with the approval of the patients, their parents, guardians or legal representatives, and with due consideration for technical, scientific, and government regulations. In pediatrics, physicians face a wide range of intellectual and cognitive difficulties on the side of the patient concerning participation in medical decision-making; this, combined with the presence of parents, who have the right and responsibility to maintain their children according to the Article 1168 of the Civil Code of the Islamic Republic of Iran, exposes the doctor to ethical challenges while making medical decisions. These challenges are not limited to treatment alone and extend to pediatric research in particular, which we cannot afford to discuss in this paper[4,5]. The present paper aimed to examine the scope of the autonomy of children and adolescents and the extent of their parents’ authority in medical decision-making based on ethical principles and jurisprudential and legal basis. In order to make it more tangible, we have used two common cases to provide practical advices. Case 1: Commencing Treatment without Parents’ Consent

The present paper aimed to examine the scope of the autonomy of children and adolescents and the extent of their parents’ authority in medical decision-making based on ethical principles and jurisprudential and legal basis. In order to make it more tangible, we have used two common cases to provide practical advices. Case 1: Commencing Treatment without Parents’ Consent A 12-year-old girl is taken to an internal medicine specialist by her teacher. Her complaints include polyuria and polydipsia, and severe weight loss. Strong suspicious to uncontrolled diabetes was made based on the history. Although the school contacted her parents many times to emphasize that she needs medical care, they ignored that as they seemed not to believe in modern medicine, and believed that herbal teas can cure her condition. The teacher asked the doctor to start proper pharmaceutical treatment and was willing to meet all the expenses. Case 2: Father Does Not Consent to Child’s Surgery A surgeon in the emergency ward had to persuade the father of a 5-year-old, in whom an acute appendicitis was diagnosed, to sign a consent form for operation. Clinical examinations and the patient’s CBC pointed to the necessity of an emergency surgery, but the father insisted on non-surgical treatment and all the surgeon’s efforts to convince him otherwise have failed. The Decision-Making Capacity Necessary to Exercise Patient Autonomy

A surgeon in the emergency ward had to persuade the father of a 5-year-old, in whom an acute appendicitis was diagnosed, to sign a consent form for operation. Clinical examinations and the patient’s CBC pointed to the necessity of an emergency surgery, but the father insisted on non-surgical treatment and all the surgeon’s efforts to convince him otherwise have failed. The Decision-Making Capacity Necessary to Exercise Patient Autonomy In order to exercise patients’ autonomy and preserve their integrity throughout a particular course of treatment, they need to possess the appropriate capability and decisional capacity[6]. In the ethical approach, decision-making capacity is a relative matter and by no means a black and white situation. A patient’s decision-making capacity can only be assessed in light of his or her specific condition, including the nature and degree of potential risks[7]. Assessment of decision-making capacity is influenced by the challenge between the right to autonomy on the part of the patient and principles of beneficence and non-malficence on the part of the physician[8]. In cases of disagreement or where the patient is not cooperating properly in the assessment process of his or her decision-making capacity, it is recommended to seek help from experts such as psychiatrists, or consultation from hospital ethics committees[9,10].

icence and non-malficence on the part of the physician[8]. In cases of disagreement or where the patient is not cooperating properly in the assessment process of his or her decision-making capacity, it is recommended to seek help from experts such as psychiatrists, or consultation from hospital ethics committees[9,10]. In the interaction between physician and patient, if the patient is a child and therefore not completely autonomous in making medical decisions, it is notwithstanding the physician’s duty to give the patient the opportunity to participate in the process in a manner appropriate to his or her capacity[11]. There have been numerous recommendations regarding the issue of pediatric patients’ participation in medical decision-making in different countries [12-14]. Naturally, in circumstances where a pediatric patient lacks the capacity required to make a particular medical decision, it appears only logical to assign the parents the right to make medical decisions, as they are responsible for raising and maintaining their children and such responsibility entails the right to make decisions for them. On the other hand, parents’ love for their children, the responsibility they feel for their children’s life and future, and their sensitivity to their best interests, makes them the best surrogates for recognizing the pediatric patients’ best interests[15]. Children’s Age and Their Right of Decision-Making from the Medical Ethics Point of View

Naturally, in circumstances where a pediatric patient lacks the capacity required to make a particular medical decision, it appears only logical to assign the parents the right to make medical decisions, as they are responsible for raising and maintaining their children and such responsibility entails the right to make decisions for them. On the other hand, parents’ love for their children, the responsibility they feel for their children’s life and future, and their sensitivity to their best interests, makes them the best surrogates for recognizing the pediatric patients’ best interests[15]. Children’s Age and Their Right of Decision-Making from the Medical Ethics Point of View Assessment of pediatric patients’ decision-making capacity should be based on their ability to evaluate their condition and the consequences of the medical decisions made, and their power to make accurate and logical deductions[16]. Nevertheless, it would facilitate pediatricians’ ethical decisions in assessing their patients’ capacity if the latter could be classified according to their age in such a way that patients in each group would possess similar capacity. Such classification would naturally be based on the customary assessment of the decision-making capacity of each age group; therefore in each group, a specific level of capacity can be assumed, unless proved otherwise. Ethical guidelines often recognize three stages of childhood: early childhood, middle childhood and adolescence[14]. In the first group, parents are basically the only decision-makers and the child is not allowed to participate in the process. In the second group, however, parents are the final decision-makers, although it is considered ethical to gain the child’s approval by offering treats and to take his or her persistent and severe resistance seriously when possible. The child’s assent is obviously sufficient in this group, and there is no need for his or her informed consent [14,16].

arents are the final decision-makers, although it is considered ethical to gain the child’s approval by offering treats and to take his or her persistent and severe resistance seriously when possible. The child’s assent is obviously sufficient in this group, and there is no need for his or her informed consent [14,16]. The most complicated situation pertains to autonomy of adolescents before they reach full capacity. It seems adolescents’ range of capacity in medical decision-making is quite broad and may vary from complete lack thereof to perfect capacity. In this age group, it is typical to assess the patient’s capacity and base all judgments on the assessment, and physicians are ethically obliged to involve adolescent patients in medical decision-making to the extent appropriate to their capacity[17]. Children’s Age and Their Right of Decision-Making from the Point of View of Law and Fiqh

The most complicated situation pertains to autonomy of adolescents before they reach full capacity. It seems adolescents’ range of capacity in medical decision-making is quite broad and may vary from complete lack thereof to perfect capacity. In this age group, it is typical to assess the patient’s capacity and base all judgments on the assessment, and physicians are ethically obliged to involve adolescent patients in medical decision-making to the extent appropriate to their capacity[17]. Children’s Age and Their Right of Decision-Making from the Point of View of Law and Fiqh In the legal realm, certain levels of capacity are presumed for each specific age group with the turning point being the age of maturity. In an overview of the current legislations of the Western countries one can see that in the years immediately before age of maturity there are special considerations regarding a youth’s valid consent in personal matters. In Australia, the age of maturity is 18, but in case the physician determines that a patient younger than 16 is fully capable of decision-making, his or her consent is considered valid, provided that another doctor who has examined the patient prior to treatment also confirms his or her capacity in writing[18]. In Canada, the age of maturity is 16, although a younger patient’s consent may be considered valid under specific circumstances, where his or her physician and another independent and legally qualified medical doctor verifies the patient’s capacity and the necessity of the procedure based on the patient’s best interests[19]. In Ireland, the situation is more or less the same, although in the period between 16 and 18 years of age, a patient can consent to treatment, but his or her right to refuse treatment remains in doubt[20]. The legal age for giving consent to treatment is 16 in England, before which a patient’s consent is considered valid if his or her capacity is confirmed[21]. In the American legal system, the age of maturity is 18, while youths under 18 cannot make healthcare decisions without their parents’ consent[22]. One exception, however, is the case of emancipated minors; these are youths under 18 that have obligations similar to adults, that is, they are financially independent, are married or have children, are enlisted in the military, or have been granted the status of adulthood by a court order[23]. Additionally, youths under the age of 18 generally have limited rights to make certain medical decisions independently, for instance regarding treatment for sexually transmitted diseases, substance and alcohol abuse treatment, blood donation, mental health treatment and family planning services[23].

age of Gheare Momayyez - Unawareness -: In this stage the child has limited understanding and powers of discernment and cannot distinguish between benefit and loss, and therefore is not legally considered to have a will. The stage lasts from 2 to 7 years and corresponds to early childhood in the ethical classification. 2) Stage of Momayyez – Awareness -: This stage lies between the stage of unawareness and legal competency, and pertains to the period in which a minor is believed to have partial powers of discernment and can distinguish between benefit and loss to some extent. The law in Iran does not specify a certain age for this stage and there are no strict criteria for recognition of awareness in people as this is a faculty that can only be determined through relevant customary assessments. Based on anecdotes about Imam Ali and Imam Jafar Sadiq (PBUT), this stage may begin between ages 7 and 9[26]. 3) Age of Taklif: according to Shiite jurisprudents boys and girls after age of taklif are accountable for any actions they do and such they should act in accordance with God's order and avoid his prohibitions. In Iranian legislation, based on the Article 1210 of the Civil Code of the Islamic Republic of Iran and according to Shiite jurisprudents, age of Taklif is 15 full lunar years for boys and 9 full lunar years for girls[27], and upon reaching this age, no one can be treated as incompetent based on mental immaturity unless his or her insanity or mental immaturity is proved.

the Civil Code of the Islamic Republic of Iran and according to Shiite jurisprudents, age of Taklif is 15 full lunar years for boys and 9 full lunar years for girls[27], and upon reaching this age, no one can be treated as incompetent based on mental immaturity unless his or her insanity or mental immaturity is proved. Based on the Consistency Clause No. 30 of the Iran Supreme Court issued on December 31st, 1985, upon reaching the age of Taklif, a minor will automatically be considered competent in non-financial matters such as divorce and giving evidence in non-monetary proceedings and can begin to act independently; as regards financial matters, however, reaching age of Taklif is not sufficient grounds for competence and the minor’s mental maturity needs to be established in court as well[28]. Consequently, if medical decisions do not entail disposition of the minor’s property, the terms above apply and age of Taklif will be considered the criterion for the right to make medical decisions[29]. It is needless to say that in these cases, as in the case of adult patients, the right to make decisions is dependent upon demonstrating the necessary intellectual capacity. In other words, after reaching the age of Taklif, patients are presumed to possess the capacity for making decisions and physicians can validate their decisions and proceed with proper treatments, unless evidence is found to the contrary. Thus the child in the first above-mentioned case is legally free to make an independent decision regarding her condition, provided she possesses the required capacity to do so.

for making decisions and physicians can validate their decisions and proceed with proper treatments, unless evidence is found to the contrary. Thus the child in the first above-mentioned case is legally free to make an independent decision regarding her condition, provided she possesses the required capacity to do so. The Shi’a Fiqh appears to have a similar stance toward such cases. Following an inquiry by the authors from a number of religious leaders (personal communications with Ayatollahs Safi Golpaygani, Makarem Shirazi and Moosavi Ardebili), all stated that the age criterion for possessing the capacity to give informed consent is reaching the age of Taklif as clarified by the Sharia, and neither believed there was need for any other person’s consent, parents included. In their written response, Ayatollahs Safi Golpaygani and Makarem Shirazi had also emphasized the necessity to ascertain the patient’s mental maturity and the ability to distinguish between what is in his or her interests and what is not. In other words, they were of the opinion that it is necessary for the physician to establish the patient’s capacity for medical decision-making. Some contemporary Faqihs believe that if a patient is of age, but is not mentally mature and therefore lacks the decision-making capacity, his or her parents’ consent is required in addition to the patient’s permission or assent[30].

ary for the physician to establish the patient’s capacity for medical decision-making. Some contemporary Faqihs believe that if a patient is of age, but is not mentally mature and therefore lacks the decision-making capacity, his or her parents’ consent is required in addition to the patient’s permission or assent[30]. Stage of Maturity – Roshd-: Adulthood is the stage when a patient’s capacity and competence in all legal financial or non-financial matters is presumed, and supervision or consent of parents in those matters is seemingly unnecessary. According to the Article 1209 of the Civil Code of the Islamic Republic of Iran prior to the 1982 reforms, a person was considered legally mature upon reaching 18 full solar years and his or her parents’ guardianship would expire with no court proceedings. After the Civil Code reforms the age specification has been repealed, although anyone 18 years and older is considered independent and accountable for his/her actions and decisions as common legal practice[31,32]. Based on the facts stated above, it can be inferred that according to the Iranian legal system, in cases where a form of payment is not required for medical treatment, reaching the age of Taklif indicates a patient’s independence in making medical decisions, unless he or she is proven to lack capacity. However, if the patient needs to have access to his or her property in order to pay for the treatment, he or she needs to have reached the age of 18. The Challenges a Physician Faces Regarding Determination of a Patient’s Capacity

Based on the facts stated above, it can be inferred that according to the Iranian legal system, in cases where a form of payment is not required for medical treatment, reaching the age of Taklif indicates a patient’s independence in making medical decisions, unless he or she is proven to lack capacity. However, if the patient needs to have access to his or her property in order to pay for the treatment, he or she needs to have reached the age of 18. The Challenges a Physician Faces Regarding Determination of a Patient’s Capacity In Iran, according to the law and Fiqh, patients have the right to make medical decisions, if their decision-making capacity is established, or in terms of Fiqh, they should have the ability to distinguish between what is in their interests and what is not. The fact that for girls the age of Taklif, after which a person is considered competent, is lower raises concern about their mental capacity to make decisions in the years immediately following the age of Taklif as clarified by the Sharia.

e the ability to distinguish between what is in their interests and what is not. The fact that for girls the age of Taklif, after which a person is considered competent, is lower raises concern about their mental capacity to make decisions in the years immediately following the age of Taklif as clarified by the Sharia. Unwary acceptance of the Taklif age as the criterion for young patients’ right to make medical decisions regardless of the sensitivity of such decisions can sometimes cause problems for doctors and the healthcare system. Considering that making medical decisions bears directly or indirectly upon the patients’ control over their body, which is no less significant than their control over their finances, it seems essential to raise the age of decision making capacity, particularly in case of young girls. As the author of the acclaimed book on Fiqh, entitled Orvatolvosqa states, in marriage, which is a non-financial matter like medical treatment, a young girl’s consent is necessary in addition to that of her parent only when apart from being of age, she has reached the stage of mental maturity necessary to make an independent decision in this respect. Otherwise she can only get married with the guidance and supervision of her parent and by his permission[33]. This point along with some other considerations caused the legal age for girls to be married to be raised from 9 to 13[34]. It seems that in medical decision-making likewise there should be no discrimination between girls and boys with regard to the age of decision making capacity; it is therefore recommended that the age requirement for the right to make medical decisions be raised to 15 for girls as well, and the laws regarding decision-making capacity be duly reformed.

n-making likewise there should be no discrimination between girls and boys with regard to the age of decision making capacity; it is therefore recommended that the age requirement for the right to make medical decisions be raised to 15 for girls as well, and the laws regarding decision-making capacity be duly reformed. On the other hand, it should be noted that emotional and responsible nature of the Iranian family does not allow parents to be indifferent to their young children’s fate, especially their daughters, even though they may have reached the age of maturity and possess the decision-making capacity. Even if the reform suggested above regarding the age requirement for girls to make medical decisions is enacted, the significant role of the young patient’s family cannot be overlooked. Proper use of communication skills in interactions with patients and their families seems to be essential in order to reach medical decisions that not only ensure individuals’ right to autonomy, but also produce the best results with the least amount of anxiety and stress for young patients and their families. Ethical Solutions for Parents’ Wrong Decisions regarding their Children

On the other hand, it should be noted that emotional and responsible nature of the Iranian family does not allow parents to be indifferent to their young children’s fate, especially their daughters, even though they may have reached the age of maturity and possess the decision-making capacity. Even if the reform suggested above regarding the age requirement for girls to make medical decisions is enacted, the significant role of the young patient’s family cannot be overlooked. Proper use of communication skills in interactions with patients and their families seems to be essential in order to reach medical decisions that not only ensure individuals’ right to autonomy, but also produce the best results with the least amount of anxiety and stress for young patients and their families. Ethical Solutions for Parents’ Wrong Decisions regarding their Children In the course of treatment, physicians may come across instances of wrong decisions made by parents that are clearly not in the child’s best interests. Deliberations on the reasons why parents are selected as surrogates in medical decision-making highlight two basic presumptions in this respect that parents are responsible and caring, when it comes to their children, and that they have their children’s best interests in heart. If the two above-mentioned presumptions no longer apply, it does not seem morally appropriate that parents continue to enjoy their right to surrogacy[14,35]. There are inevitably disagree-ments as to what the exact criteria for determination of a child’s best interests may be, but one valuable measure for pediatricians could be the principle of protecting children against serious harm, pain and death[36,37]. In guidelines offered in medical ethics literature there are restrictions on the scope of autonomy of adult patients in medical decision-making, for instance a patient’s wish to commit suicide is invalid; likewise, multiple restrictions apply to the autonomy of parents regarding their children[38]. For example, in most countries people have the right to refuse life-saving treatments, while no one is granted this same right regarding their children[15]. Similarly, parents cannot refuse their children life-sustaining treatments on account of their own religious beliefs[36]. Parents are morally obliged to make medical decisions based on their children’s best interests, not their own wishes and well-being, and if the physician decides that their decision is not in the best interests of the child, a reliable authority, that is, the court or ethical committee, can preserve the child’s rights in an unbiased manner.

liged to make medical decisions based on their children’s best interests, not their own wishes and well-being, and if the physician decides that their decision is not in the best interests of the child, a reliable authority, that is, the court or ethical committee, can preserve the child’s rights in an unbiased manner. In situations like the second case above, where the parents’ refusal or consent to a certain treatment is clearly in conflict with the child’s best interests, the doctor should offer adequate explanations and make the parents aware of the consequences of their decision so they can eventually reach an agreement[11]. In case the physician does not succeed in changing the parents’ mind, the situation can be resolved by referring the matter to the ethical committee. In rare occasions, where child abuse or neglect is suspected, it might be necessary to take the matter to court[15]. These approaches apply only if the child is not in an emergency or a life-threatening situation. In emergent conditions, the physician can disregard parents’ refusal and proceed with urgent medical intervention until the situation is no longer life-threatening[21]. From the point of view of Fiqh, parents or legal guardians are obligated to have only the child’s best interests in mind and not reject measures upon which the child’s life depends within reasonable limits, as this is the responsibility they have been charged with[29,39].

These approaches apply only if the child is not in an emergency or a life-threatening situation. In emergent conditions, the physician can disregard parents’ refusal and proceed with urgent medical intervention until the situation is no longer life-threatening[21]. From the point of view of Fiqh, parents or legal guardians are obligated to have only the child’s best interests in mind and not reject measures upon which the child’s life depends within reasonable limits, as this is the responsibility they have been charged with[29,39]. Such ethical views on the scope of parents’ rights to make decisions regarding their children appear to be acceptable in the legal system of Iran. According to the Article 1184 of the Civil Code of the Islamic Republic of Iran “if the natural guardian of a child is unmindful of his ward’s well-being and manages his or her property in a manner that brings about loss, the court will, on application by the relatives of the child or by request of the Public Prosecutor, and after the establishment of the incapacity or dishonesty of the guardian, discharge the guardian of his duties and conclude his management of the child’s properties, and will appoint a proper financial trustee in his place.” This article may appear to be related to children’s properties and financial affairs, but can extend to more important matters by the same token.

onesty of the guardian, discharge the guardian of his duties and conclude his management of the child’s properties, and will appoint a proper financial trustee in his place.” This article may appear to be related to children’s properties and financial affairs, but can extend to more important matters by the same token. Moreover, based on the Article 1173 of the Civil Code of the Islamic Republic of Iran, “If the physical health or moral education of a child is endangered as a result of carelessness or moral degradation of the father or mother who have custody of the child, the court can take any decision it deems appropriate regarding custody of the child upon request of his or her relatives, guardian or the Public Prosecutor.” Such laws indicate that custody is contingent upon preservation of the child’s well-being or, in other words, best interests. It is noteworthy that in the Shi’a Fiqh the judge can dismiss a parent’s guardianship of his child if his incapacity or dishonesty is established. The author of the book Javaherulkalaam who is also a renowned expert on the Shi’a Fiqh maintains that even if evidence and circumstances indicate that the father or paternal grandfather of a minor or an insane person has caused them to incur material loss, the judge needs to take away their control over their ward’s property[27]. It should be noted that in cases like this, called non-litigious cases, the court should start proceedings directly even if there is no dispute or complaint[40].

grandfather of a minor or an insane person has caused them to incur material loss, the judge needs to take away their control over their ward’s property[27]. It should be noted that in cases like this, called non-litigious cases, the court should start proceedings directly even if there is no dispute or complaint[40]. In response to our inquiry regarding whether it is permissible to perform a necessary but not urgent procedure on a minor, examples varying from administration of antibiotics to surgeries, most responses (personal communications with Ayatollahs Safi Golpaygani, Makarem Shirazi and Moosavi Ardebili) pointed to the necessity of seeking parents’ consent, although they considered it acceptable to disregard the parents’ refusal to consent to procedures in case of life-threatening situations.

ics to surgeries, most responses (personal communications with Ayatollahs Safi Golpaygani, Makarem Shirazi and Moosavi Ardebili) pointed to the necessity of seeking parents’ consent, although they considered it acceptable to disregard the parents’ refusal to consent to procedures in case of life-threatening situations. Conclusion One of the most important missions of medical ethics is to protect the rights of all individuals and ensure that they exercise autonomy within their intellectual ability and capacity. According to the Shi’a Fiqh and Iran’s written laws, the age requirement for the right to make medical decisions is reaching the age of Taklif, which is 15 full lunar years for boys and 9 full lunar years for girls. This sex-based difference in age of decision making capacity seems to be in discordance with the current medical ethics guidelines and is therefore unjustifiable. The concern is mostly regarding young girls’ vulnerability due to the fact that they can practice their medical decision-making right at an earlier age. In order to exercise this right, an adolescent’s capacity needs to be established, as is the case with adults; as regards young girls who have just come of age, more care needs to be taken in establishing their capacity.

ility due to the fact that they can practice their medical decision-making right at an earlier age. In order to exercise this right, an adolescent’s capacity needs to be established, as is the case with adults; as regards young girls who have just come of age, more care needs to be taken in establishing their capacity. Physicians, health policy makers and legislators should reach an agreement regarding the age of capacity for medical decision-making based on their estimations of the average age for attaining capacity in the general public and the society’s overall welfare, as has been done in case of age of maturity for getting married and taking control over one’s property. The authors do not believe there should be any discrimination between the sexes in this regard. In many cases, it is crucial that medical decisions be made in the shortest time possible so that treatment can begin immediately. A physician’s determination of an adult patient’s capacity puts him or her in a complicated situation, particularly if the patient disagrees with the doctor or his or her parents. Under these circumstances, physicians are supposedly authorized to proceed with life-sustaining measures, but the moral and legal liabilities of such decisions are substantial and therefore stressful for doctors, especially if they occur frequently over time. Moreover, complexities of these decisions oftentimes necessitate exchange of views among various people.

pposedly authorized to proceed with life-sustaining measures, but the moral and legal liabilities of such decisions are substantial and therefore stressful for doctors, especially if they occur frequently over time. Moreover, complexities of these decisions oftentimes necessitate exchange of views among various people. It seems that it would be helpful for health centers to have access to an easily reachable committee charged with the responsibility to make such decisions. Such a committee should be endorsed by the nation’s legal system and be answerable to any possible grievances. On account of the authority bestowed by Fiqh and the law on physicians regarding distinguishing lifethreatening situations and medical emergencies, and determination of an adult patient’s mental maturity, such a committee can ensure patients’ best interests with the lowest mental, emotional and legal consequences for doctors. Authors’ Contribution A. Parsapoor: Concept, Acquisition of data, Interpretation, Drafting of the manuscript M. Bagher Parsapoor: concept, critical revision of the manuscript and approval of the article. N. Rezaei: concept and approval of the article. F. Asghari: critical revision of the manuscript and approval of the article. All authors approved the final version of the article. Conflict of Interest: None

Introduction The genus Enterobacter in a family of Enterobacteriaceae was first described by Hormaeche and Edwards in 1960 and has undergone considerable taxonomic modification over the last 50 years[1,2]. The name Enterobacter cowanii is proposed for a group of microorganisms referred to as NIH group 42. The G + C content of its DNA ranges from 52.5% to 53.6%[3,4]. Because of distinct differentiation of E. cowanii by DNA hybridization methods from other members of Enterobacteriaceae family as well as its unique phenotypic and genotypic properties and since the DNA relatedness (5-38%) is closer to species of the genus Enterobacter than to other species of the Enterobacteriaceae, the members of NIH group 42 were placed in the genus Enterobacter[3-5]. The majority of E. cowanii strains were isolated from clinical and plant specimens[3]. E. cowanii is a genus of common gram-negative, facultatively anaerobic, rod-shaped, oxidase-negative, catalase positive, non-spore-forming bacterium of the Enterobacteriaceae family. It is motile by peritrichous flagella[2,4]. This organism grows well on selective media for gram negative bacteria such as MacConkey agar and is facultatively anaerobic. E. cowanii is commonly found in ecological niches. This opportunistic pathogen is isolated from clinical specimens such as urine, sputum, blood, and pus. It was isolated by blood culture from a patient with gastric cancer. It may also be found in foods[4,5]. E. cowanii has been isolated from eucalyptus trees[6,7].

bic. E. cowanii is commonly found in ecological niches. This opportunistic pathogen is isolated from clinical specimens such as urine, sputum, blood, and pus. It was isolated by blood culture from a patient with gastric cancer. It may also be found in foods[4,5]. E. cowanii has been isolated from eucalyptus trees[6,7]. In recent years a remarkable increase in nosocomial infections has been reported especially in neonatal intensive care units, intensive care units and oncology departments. Underlying diseases, low-birth-weight, immunocompromised immune system, cancer chemotherapy, and intravenous catheterization can be predisposing factors in cases of infections due to unusual microorganisms, including Enterobacter spp. in neonates[8-11]. Infants admitted to NICUs, especially ones who have undergone surgery or have congenital abnormalities, are often at high risk for developing nosocomial infections. Clinical manifestations are often misleading and, in some circumstances, it may be difficult or even impossible to distinguish the source of the infection[9,10]. Enterobacteriaceae family members are potential PIF-borne pathogens.

ve congenital abnormalities, are often at high risk for developing nosocomial infections. Clinical manifestations are often misleading and, in some circumstances, it may be difficult or even impossible to distinguish the source of the infection[9,10]. Enterobacteriaceae family members are potential PIF-borne pathogens. Neonates and young children are exclusively vulnerable to infections caused by foodborne pathogens[9,12]. Contamination of PIF with E. cowanii will be associated with many diseases in neonates. Therefore, the microbiological safety of PIF is very important. Because PIF is not a sterile product, it is an excellent medium to support the bacterial growth. Bovine milk and plant materials are essential ingredients of PIF and a potential source of various bacteria that are pathogenic to neonates and adults[12-14]. The aim of our study was to isolate and identify antimicrobial susceptibility pattern of E. cowanii isolated from PIF in NICUs in Tehran hospitals. Subjects and Methods Place and Duration of Study: Department of Pathobiology, School of Public Health, Tehran University of Medical Sciences, Tehran, between Jun 2011 and March 2012. Sampling: A cross-sectional study was carried out on 125 samples of powdered infant formula milk (PIF) purchased from hospital drug stores in Tehran between Jun 2011 and March 2012.

Place and Duration of Study: Department of Pathobiology, School of Public Health, Tehran University of Medical Sciences, Tehran, between Jun 2011 and March 2012. Sampling: A cross-sectional study was carried out on 125 samples of powdered infant formula milk (PIF) purchased from hospital drug stores in Tehran between Jun 2011 and March 2012. Isolation and Identification: PIF cans were surface sanitized with 70% ethanol and were opened in a laminar flow cabinet. Samples were taken from each product under aseptical conditions. E. cowanii was isolated according to FDA method[15,16]. We prepared 3 Erlenmeyer flasks of sterile distilled water (pre-warmed to 45ºC) at 9, 90 and 900 ml containing 1, 10 and 100 g of PIF, respectively. After the PIF was completely mixed and dissolved in distilled water, it was incubated at 37ºC for 18-24 h. Following incubation, 10 ml of each sample was added to 90 ml of Enterobacteriaceae enrichment (EE) broth medium and incubated at 37ºC for 18-24 h. After incubation, a lapful of the enrichment culture was streaked onto duplicates violet red bile glucose agar (VRBGA) plates and cultured at 37ºC for 18-24 h. A total of 4 suspicious colonies were picked from each VRBGA plate and pure culture was achieved. For detection of non-lactose fermenting isolates, presumptive colonies were streaked onto MacConkey agar and incubated at 37ºC for 72 h. For final confirmation biochemical tests were embedded in the API-20E biochemical kit system (Bio-Mérieux) and manual biochemical tests were used according to directions of the manufacturer. For long term storage, the purified isolates were stored in tryptic soy broth (TSB) with 20% glycerol (Merck Co.) at -20ºC.

72 h. For final confirmation biochemical tests were embedded in the API-20E biochemical kit system (Bio-Mérieux) and manual biochemical tests were used according to directions of the manufacturer. For long term storage, the purified isolates were stored in tryptic soy broth (TSB) with 20% glycerol (Merck Co.) at -20ºC. Antibiotic sensitivity testing: Antibiotic sensi-tivity testing was performed using Kirby-Bauer disk diffusion method on Mueller Hinton agar according to CLSI guidelines[17]. Antimicrobial agents used in this study are listed in Table 1. Statistical analysis: The calculation of sample size was performed by using McNemar's test. Data were analyzed using SPSS software, version 19. Findings Out of the 125 PIF investigated samples, 4 (3.2%) samples were positive for Enterobacter cowanii. The gram staining of the colony of organism showed gram negative rods. On VRBGA agar selective medium purple/pink colored colonies, and on MacConkey agar lactose fermenting, smooth, convex, punctuate, umbilicated, glistening colonies were grown during 16 to 20 hours. Isolated strains were oxidase negative, catalase positive, motile and produced other biochemical reactions which are characteristic of E. cowanii (Table 2). Fifty percent of isolates were resistant to ampicillin, amoxicillin, and cotrimoxazole. Susceptibility patterns of isolates are listed in Table 1. Discussion

Out of the 125 PIF investigated samples, 4 (3.2%) samples were positive for Enterobacter cowanii. The gram staining of the colony of organism showed gram negative rods. On VRBGA agar selective medium purple/pink colored colonies, and on MacConkey agar lactose fermenting, smooth, convex, punctuate, umbilicated, glistening colonies were grown during 16 to 20 hours. Isolated strains were oxidase negative, catalase positive, motile and produced other biochemical reactions which are characteristic of E. cowanii (Table 2). Fifty percent of isolates were resistant to ampicillin, amoxicillin, and cotrimoxazole. Susceptibility patterns of isolates are listed in Table 1. Discussion E. cowanii shows the common characteristics of the genus Enterobacter, mostly isolated from different sources such as clinical specimens, foods, plants, in developed and developing countries worldwide[3-6]. One large difficulty in preventing NICU and hospital-acquired infections is to find the source of the infectious agent and its route of transmission. The consumption of powdered infant milk formula is wide-spreading; however, few studies have been undertaken to evaluate the role of E. cowanii in food safety.

wide[3-6]. One large difficulty in preventing NICU and hospital-acquired infections is to find the source of the infectious agent and its route of transmission. The consumption of powdered infant milk formula is wide-spreading; however, few studies have been undertaken to evaluate the role of E. cowanii in food safety. To the best of our knowledge, there is no previous report on the isolation and identification of E. cowanii from PIF and determination of antimicrobial susceptibility pattern of this bacterium in Iran. In this study, we demonstrated that E. cowanii strains are widely spread in PIF. The results showed that all E. cowanii strains isolated from PIF samples were sensitive to meropenem, ceftazidime, ciprofloxacin, imipenem, chloram-phenicol, cefepime, levofloxacin, piperacillin, gentamicin, moxifloxacin, and colistin. In the present study 50% of isolates were resistant to ampicillin, amoxicillin, and cotrimoxazole. Table 1 Antimicrobial susceptibility pattern of Enterobacter cowanii strains isolated from PIF (n=4) Antibiotic Sensitive (%) Intermediate (%) Resistant (%) Ampicillin (AP) 1 (25) 1 (25) 2 (50) Amoxicillin (A) 1 (25) 1 (25) 2 (50) Aztreonam (ATM) 3 (75) 1 (25) - Cefotaxime (CTX) 1 (25) 2 (50) 1 (25) Amikacin (AK) 3 (75) 1 (25) - Streptomycin (S) 3 (75) 1 (25) - Meropenem (MEM) 4 (100) - - Mezlocillin (MEZ) 2 (50) 2 (50) - Nalidixic acid (NA) 3 (75) 1 (25) - Tigecycline (TGC) 3 (100) 1 (25) - Tetracycline (T) 3 (75) 1 (25) - Ticarcillin (TC) 3 (75) 1 (25) - Chloramphenicol (C) 4 (100) - - Ceftazidime (CAZ) 4 (100) - - Ciprofloxacin (CIP) 4 (100) - - Cefepime (CPM) 4 (100) - - Imipenem (IMI) 4 (100) - -

Streptomycin (S) 3 (75) 1 (25) - Meropenem (MEM) 4 (100) - - Mezlocillin (MEZ) 2 (50) 2 (50) - Nalidixic acid (NA) 3 (75) 1 (25) - Tigecycline (TGC) 3 (100) 1 (25) - Tetracycline (T) 3 (75) 1 (25) - Ticarcillin (TC) 3 (75) 1 (25) - Chloramphenicol (C) 4 (100) - - Ceftazidime (CAZ) 4 (100) - - Ciprofloxacin (CIP) 4 (100) - - Cefepime (CPM) 4 (100) - - Imipenem (IMI) 4 (100) - - Levofloxacin (LEV) 4 (100) - - Minocycline (MN) 3 (75) 1 (25) - Piperacillin (PRL) 4 (100) - - Piperacillin-tazobactam (PTZ) 2 (50) 2 (50) - Carbenicillin (PY) 1 (25) - 3 (75) Tobramycin (TN) 3 (75) 1 (25) - Cotrimoxazole (TS) 2 (50) - 2 (50) Moxifloxacin (MFX) 4 (100) - - Gentamicin (GM) 4 (100) - - Colistin (CO) 4 (100) - - Table 2 Biochemical reactions of Enterobacter cowanii Test Reaction/Result Gram stain Gram-negative, rod Triple sugar iron agar Acid (Yellow) slant/Acid (Yellow) butt. No H2S Motility + Oxidase - Catalase + Nitrate reduction + Simmons citrate’s at 37°C + Gas from glucose + Acid from glucose + Lactose + Maltose + Sucrose + Sorbitol + Mannitol + Xylose + Raffinose + Arabinose - D-Cellobiose + Malonate - Adonitol - Rhamnose + a-Methyl-D-glucoside - Salicin + Inositol - Indole - Methyl red (MR) - Voges proskauer (VP) + Urease - Lysine decarboxylase - Ornithine decarboxylase - Arginine dehydrolase - Phenylalanine deaminase - Esculin hydrolysis + Gelatine hydrolysis - ONPG + DNAase -

Maltose + Sucrose + Sorbitol + Mannitol + Xylose + Raffinose + Arabinose - D-Cellobiose + Malonate - Adonitol - Rhamnose + a-Methyl-D-glucoside - Salicin + Inositol - Indole - Methyl red (MR) - Voges proskauer (VP) + Urease - Lysine decarboxylase - Ornithine decarboxylase - Arginine dehydrolase - Phenylalanine deaminase - Esculin hydrolysis + Gelatine hydrolysis - ONPG + DNAase - To the best of our knowledge, there is no previous report on the isolation and identification of E. cowanii from PIF and determination of antimicrobial susceptibility pattern of this bacterium in Iran. In this study, we demonstrated that E. cowanii strains are widely spread in PIF. The results showed that all E. cowanii strains isolated from PIF samples were sensitive to meropenem, ceftazidime, ciprofloxacin, imipenem, chloram-phenicol, cefepime, levofloxacin, piperacillin, gentamicin, moxifloxacin, and colistin. In the present study 50% of isolates were resistant to ampicillin, amoxicillin, and cotrimoxazole.

owed that all E. cowanii strains isolated from PIF samples were sensitive to meropenem, ceftazidime, ciprofloxacin, imipenem, chloram-phenicol, cefepime, levofloxacin, piperacillin, gentamicin, moxifloxacin, and colistin. In the present study 50% of isolates were resistant to ampicillin, amoxicillin, and cotrimoxazole. E. cowanii is an opportunistic organism and, when introduced into the human organs or other fauna, may cause infection. Disease caused by this organism can occur in individuals with underlying diseases especially patients hospitalized in NICU[10]. Confirmed virulence of E. cowanii is difficult to reveal, because clinical reports involving E. cowanii are typically of polymicrobial nature, often involve patients that are already affected by diseases of other origin, lack pathogenicity confirmation, and diagnostic isolates are rarely conserved for confirmatory analysis. The results of the present study helps to better understanding of the role of E. cowanii as an opportunistic pathogen-causing disease in NICU.

involve patients that are already affected by diseases of other origin, lack pathogenicity confirmation, and diagnostic isolates are rarely conserved for confirmatory analysis. The results of the present study helps to better understanding of the role of E. cowanii as an opportunistic pathogen-causing disease in NICU. Neonates and high risk groups (e.g. immunosuppressed and HIV positive individuals, senile persons) are particularly assailable to foodborne opportunistic pathogens[18,19]. Low birth weight infants in neonatal intensive care units are typically immunocompromised patients and their immunity is not fully mature. Therefore they are susceptible to different hospital-acquired infections[10,20-22]. In newborn infants, local natural barriers against bacterial infections are compromised and the production of secretory immunoglobin A is absent during the first days of life[9]. The decreased production and function of local and systemic defense depends on antigen exposure and contributes to greater susceptibility to bacterial infection during the neonatal period[9,23]. From our results we conclude that E. cowanii may be able to start a hospital outbreak, through powdered infant milk formula. The inherent capability of this organism to remain viable and grow well at room temperature may contribute to such contamination. Caregivers in hospital neonatal units should be constantly alerted to the fact that powdered infant formula products are not sterile and may be colonized with different microorganisms.

mula. The inherent capability of this organism to remain viable and grow well at room temperature may contribute to such contamination. Caregivers in hospital neonatal units should be constantly alerted to the fact that powdered infant formula products are not sterile and may be colonized with different microorganisms. In addition, infant formula producers must accomplish guidelines aimed to decrease the risks of products contamination with foodborne pathogens. Controlling the primary populations of E. cowanii during the PIF production process and preventing post processing contamination by using suitable microbiological guidelines, is accessible. Sanitary practices for the preparation of infant formula in both the home and hospitals should be carefully controlled. Conclusion Powdered infant milk formula containing members of the Enterobacteriaceae might impose additional risk of infection to the neonates and especially to the low birth weight premature babies. There is very little information about virulence factors and pathogenicity of E. cowanii in human, so complementary studies are necessary to clarify the possible role of E. cowanii as a food contaminant, in common NICU infections and high risk groups including persons with underlying disease and immunocompromised individuals. Authors’ Contribution: All authors listed have contributed sufficiently to the project to be included as authors, and all those who are qualified to be authors are listed in the author byline. Acknowledgment This work was supported by Tehran University of Medical Sciences, Tehran, Iran (Project No. 13382).

Conclusion Powdered infant milk formula containing members of the Enterobacteriaceae might impose additional risk of infection to the neonates and especially to the low birth weight premature babies. There is very little information about virulence factors and pathogenicity of E. cowanii in human, so complementary studies are necessary to clarify the possible role of E. cowanii as a food contaminant, in common NICU infections and high risk groups including persons with underlying disease and immunocompromised individuals. Authors’ Contribution: All authors listed have contributed sufficiently to the project to be included as authors, and all those who are qualified to be authors are listed in the author byline. Acknowledgment This work was supported by Tehran University of Medical Sciences, Tehran, Iran (Project No. 13382). Conflict of Interest: None

Introduction Delivery is an important phenomenon and may be one of the most painful and stressful events that mothers experience during their lifetime[1]. The hormones, such as catecholamines, cortisol, epinephrine, and beta-endorphins, which are secreted in response to tension and anxiety, interfere in the progress of cervical dilatation. On the other hand, they influence the uterine smooth muscles and reduce the contractile power of the uterus as well as its efficiency in the delivery process eventually lengthening the delivery, increasing the pain, and leading to anxiety[2]. During long-term anxiety, the autonomous nervous system is stimulated leading to an increase in the contraction of the smooth muscles of the arterial system. This subsequently reduces the uteroplacental blood flow as well as oxygenation to the uterus, leads to fetal hypoxia, and increases abnormal fetal heart rates[3-5]. Intrauterine hypoxia is an important risk factor of infant mortality and abnormal outcomes during infancy which causes problems in 5-10% of pregnancies[6,7]. Recently, counting nucleated red blood cells (NRBCs) in every 100 white blood cells (WBCs) in infants' umbilical venous blood is considered as a sign of prenatal hypoxia. Each hypoxia event leads to a compensatory fetal reaction increasing hematopoiesis and entrance of immature red blood cells into the fetal blood flow level of which is associated with prenatal hypoxia[8]. Thus, the relationship between chronic stress and delivery outcomes, shows the necessity for intervention in order to reduce it[9]. In general, non-pharmacological methods are among the most effective methods of coping with stress and delivery pain. One of these methods involves continuous support of the mother by the doula[10]. Hung conducted a semi-experimental study on the effect of a supportive companion during labor on the delivery outcomes. The study showed that supporting the women during labor was effective in improving the uterine dysfunction and reduced the length of labor as well as the cesarean rate. Besides, the supported women felt less exhausted and were more satisfied after the delivery[11].

rtive companion during labor on the delivery outcomes. The study showed that supporting the women during labor was effective in improving the uterine dysfunction and reduced the length of labor as well as the cesarean rate. Besides, the supported women felt less exhausted and were more satisfied after the delivery[11]. Teixeira et al (1991) revealed a relationship between the mother's anxiety score and increase in the vascular resistance of uterine arteries[12]. Labor anxiety and release of cortisol and catecholamines may lengthen the labor, decrease the placental blood flow, and lead to fetal hypoxia[13].

rtive companion during labor on the delivery outcomes. The study showed that supporting the women during labor was effective in improving the uterine dysfunction and reduced the length of labor as well as the cesarean rate. Besides, the supported women felt less exhausted and were more satisfied after the delivery[11]. Teixeira et al (1991) revealed a relationship between the mother's anxiety score and increase in the vascular resistance of uterine arteries[12]. Labor anxiety and release of cortisol and catecholamines may lengthen the labor, decrease the placental blood flow, and lead to fetal hypoxia[13]. Acupressure is another non-pharmacological intervention which is used in order to decrease anxiety. Acupressure is an acupuncture branch in which finger pressure is used instead of needles[14]. Studies have shown that acupuncture is effective in releasing oxytocin from the pituitary gland and stimulates the secretion of oxytocin which directly stimulates the uterine contractions leading to beginning and progress of the delivery and reduction of the labor length[15,16]. In general, various acupuncture points are used to induce and control the delivery and BL32 acupoint [Shang Liao (urinary bladder or bladder meridian)][17] was used in the present investigation. In this study, acupressure which is a simple, non-invasive method and doula supportive care was used in order to reduce the mothers' pain and anxiety during labor and prevent undesirable fetal outcomes, such as hypoxia. Thus, the present study aims to assess the effects of decreasing maternal anxiety on fetal oxygenation and f NRBCs count in the cord blood.

imple, non-invasive method and doula supportive care was used in order to reduce the mothers' pain and anxiety during labor and prevent undesirable fetal outcomes, such as hypoxia. Thus, the present study aims to assess the effects of decreasing maternal anxiety on fetal oxygenation and f NRBCs count in the cord blood. Subjects and Methods This randomized clinical trial was conducted in the delivery ward of the selected educational center of Shiraz University of Medical Sciences (Shoushtari Hospital) in 2012. Considering d=5, α=0.05, 1-β=0.90, SD=7, and the following formula, a 150-subject sample size (50 subjects in each group) was determined for the study: n=2(Z1-α2+Z1-β)2SD2d2 Then, the subjects were selected through simple random sampling and were divided into supportive care, acupressure, and control groups using stratified block randomization. In doing so, a number was randomly selected from the table of random numbers and the researcher moved toward the right or left column or row and wrote the 5 digit numbers down. Since the participants were divided into 3 groups in this study, 3-therapy method was used and classification was performed as follows: A: supportive care group, B: acupressure group, and C: control group. Accordingly, ABC: 1, ACB: 2, BAC: 3, BCA: 4, CAB: 5, and CBA: 6. It should be noted that numbers 0, 7, and 9 were ignored.

e participants were divided into 3 groups in this study, 3-therapy method was used and classification was performed as follows: A: supportive care group, B: acupressure group, and C: control group. Accordingly, ABC: 1, ACB: 2, BAC: 3, BCA: 4, CAB: 5, and CBA: 6. It should be noted that numbers 0, 7, and 9 were ignored. The inclusion criteria of the study were first or second pregnancy, single and term pregnancy, vertex presentation, spontaneous beginning of the delivery process, 3-4 cm cervical dilatation, being 18-35 years old, and having at least middle school degree. The exclusion criteria of the study were: suffering from any physical or mental problems, preeclampsia, having a history of smoking, maternal diabetes, Rh incompatibility, oligohyd-ramnios, and thick meconium,.

elivery process, 3-4 cm cervical dilatation, being 18-35 years old, and having at least middle school degree. The exclusion criteria of the study were: suffering from any physical or mental problems, preeclampsia, having a history of smoking, maternal diabetes, Rh incompatibility, oligohyd-ramnios, and thick meconium,. The study data were collected using a questionnaire including demographic, clinical, and pregnancy information and an observation form including the information about the labor stages and the laboratory results. Also, Spielberger's questionnaire was employed in order to assess the anxiety level. This questionnaire includes 40 questions in two 20-item scales. The first 20 questions measure the state anxiety which is defined as a feeling of worry and tension resulting from situational stress. Yet, the second 20 questions assess the trait anxiety which refers to individual differences in the tendency toward evaluating the situations as threatening or dangerous[18]. In Iran, Aghamohammadi et al (2007) used this questionnaire on 150 patients undergoing surgery and reported its reliability as 97%[19] which is the basis for the present study. The supportive care and acupressure groups completed this questionnaire before and after the intervention. In the control group, on the other hand, the questionnaire was completed at the beginning of the active phase of the labor and the end of the first stage.

ility as 97%[19] which is the basis for the present study. The supportive care and acupressure groups completed this questionnaire before and after the intervention. In the control group, on the other hand, the questionnaire was completed at the beginning of the active phase of the labor and the end of the first stage. In the first group, the researcher as the doula was beside the mother from her entering the department. She reassured the mother, encouraged her, and gave her correct and appropriate information which led to the mother's tranquility, understanding the origin of pain, developing a positive attitude toward the pain experience, and increase in her cooperation in the delivery progress. In the second group, in 3-4 and 7-8cm dilatation, the mother was placed in a proper position and BL32 point was pressed. This point lays approximately one index finger length above the top of the buttock crease, approximately one thumb width either side of the spine (Fig. 1)[20]. The pressure was continuously and gently applied by both thumbs for 20 minutes. It was applied by the beginning and stopped at the end of the contractions. On the other hand, the control group only received the department's routine care and underwent no interventions.

thumb width either side of the spine (Fig. 1)[20]. The pressure was continuously and gently applied by both thumbs for 20 minutes. It was applied by the beginning and stopped at the end of the contractions. On the other hand, the control group only received the department's routine care and underwent no interventions. After birth, the infants' cord was double-clamped before or simultaneous with the first breath. In order to facilitate the sampling, the blood was directed toward the cord by the fingers so that the umbilical cord vessels were full of blood. Then, in order to count the total number of the blood cells (CBC diff), 2cc cord blood was poured into a bottle containing Ethylen Diamine Tetracetic Acid (EDTA) and transferred to the hospital laboratory for hematological analysis. After preparing and staining the peripheral blood smear, the number of NRBCs was determined per 100 WBCs and the results were recorded in the related form. Finally, the data were entered into the SPSS statistical software (v. 16) and analyzed using Chi-square, one-way ANOVA, LSD, and logistic regression analysis. P<0.05 was considered as statistically significant. Findings The study results revealed no significant difference between the study groups regarding the demographic variables including mother's age, level of education, occupation, gestational age, and number of pregnancies. In addition, the results of one-way ANOVA showed no significant difference between the three groups regarding the anxiety score before the intervention (P=0.4). Fig. 1 Location of the BL 32 acupoint

The study results revealed no significant difference between the study groups regarding the demographic variables including mother's age, level of education, occupation, gestational age, and number of pregnancies. In addition, the results of one-way ANOVA showed no significant difference between the three groups regarding the anxiety score before the intervention (P=0.4). Fig. 1 Location of the BL 32 acupoint Table 1 The mothers’ mean score of anxiety (state and trait) before and after the intervention in the three groups Anxiety assessment time Supportive care (n=50) Mean (SD) Acupressure (n=50) Mean (SD) Control (n=50) Mean (SD) P . value Before intervention 56.8 (2.6) 57.6 (3.6) 57.6 (2.2) 0.4 After intervention 35.5 (5.8) 37.5 (9.1) 69.8 (6.6) <0.001 Changes in anxiety score * -20.5 (8.0) -20.0 (7.2) 11.1 (9.5) <0.001 SD: standard deviation * Anxiety scores after intervention-Anxiety scores before intervention After the intervention, however, the anxiety mean score was higher in the control group compared to the supportive care and acupressure groups and the difference was statistically significant (P<0.001) (Table 1).

Changes in anxiety score * -20.5 (8.0) -20.0 (7.2) 11.1 (9.5) <0.001 SD: standard deviation * Anxiety scores after intervention-Anxiety scores before intervention After the intervention, however, the anxiety mean score was higher in the control group compared to the supportive care and acupressure groups and the difference was statistically significant (P<0.001) (Table 1). According to the results of Chi-square test regarding the distribution of NRBCs in the peripheral blood smear, the control group showed 68% and 66% difference with the supportive care and acupressure groups, respectively which were statistically significant (P<0.001). In all the three study groups, 41.3% (0-10) of NRBCs of the blood cord were observed in the peripheral blood smear. Among the 9 cases with NRBCs in the supportive care group, 14% had 1 and 4% had 2 NRBCs. Also, among the 10 cases with NRBCs in the acupressure group, 10% had 3 and 2% had 1 NRBC. In the control group, on the other hand, among the 43 cases with NRBCs, 18% had 2 and 2% had 10 NRBCs. Overall, a larger number of NRBCs were detected in the peripheral blood smear in the control group compared to the two intervention groups (Table 2).

s with NRBCs in the acupressure group, 10% had 3 and 2% had 1 NRBC. In the control group, on the other hand, among the 43 cases with NRBCs, 18% had 2 and 2% had 10 NRBCs. Overall, a larger number of NRBCs were detected in the peripheral blood smear in the control group compared to the two intervention groups (Table 2). Using logistic regression analysis and eliminating the effect of group, a significant relationship was observed between the length of the first and second labor stages and the frequency of NRBCs in the cord blood (P=0.01) (OR=1.01). In all the three groups, the mean length of the first and second stages of labor was higher in the cases with NRBCs detected in their peripheral blood. In the control group, the mean length of labor in the cases with NRBCs detected in their peripheral smears was 190.0 and 175.2 minutes higher in comparison to the supportive care and acupressure groups, respectively. Among these cases, the highest and lowest mean length of labor was related to the control and supportive care groups, respectively (Table 3).

of labor in the cases with NRBCs detected in their peripheral smears was 190.0 and 175.2 minutes higher in comparison to the supportive care and acupressure groups, respectively. Among these cases, the highest and lowest mean length of labor was related to the control and supportive care groups, respectively (Table 3). Based on the results of Chi-square test, a larger number of NRBCs were detected in the peripheral smears in cesarean deliveries. This showed a significant relationship between the type of delivery and the number of NRBCs in the cord blood in both the intervention groups (P<0.001) and the control group (P=0.03) (Table 4). Moreover, the highest number of NRBCs was observed in cesarean deliveries and this measure in the intervention groups was 14.3% lower in comparison to the control group. In natural vaginal delivery also, the number of cases without NRBCs was 62.7% higher in the intervention groups compared to the control group. Discussion The findings of the present study confirmed the effectiveness of applying pressure at BL32 point and psychologically supporting the mother during labor on reduction of maternal anxiety and improvement of fetal oxygenation. Table 2 Comparison of distribution of cord blood nucleated red blood cells in the intervention and control groups NRBC in cord blood count, peripheral smear Supportive care (n=50) n (%) Acupressure (n=50) n (%) Control (n=50) n (%) Total (n=150) n (%) Unobserved 41 (82) 40 (80) 7 (14) 88 (58.7) Observed 9 (18) 10 (20) 43 (86) 62 (41.30 Rang 0-2 0-3 0-10 0-10 P. value<0.001; NRBC: Nucleated red blood cells

Table 2 Comparison of distribution of cord blood nucleated red blood cells in the intervention and control groups NRBC in cord blood count, peripheral smear Supportive care (n=50) n (%) Acupressure (n=50) n (%) Control (n=50) n (%) Total (n=150) n (%) Unobserved 41 (82) 40 (80) 7 (14) 88 (58.7) Observed 9 (18) 10 (20) 43 (86) 62 (41.30 Rang 0-2 0-3 0-10 0-10 P. value<0.001; NRBC: Nucleated red blood cells Table 3 The relationship between duration of the first and second stages of labor and frequency of cord blood nucleated red blood cells in the intervention and control groups NRBC in cord blood count, peripheral smear Supportive care (n=50) Mean (SD) Acupressure (n=50) Mean (SD) Control (n=50) Mean (SD) Total (n=150) Mean (SD) Unobserved 127.0 (52.0) 212.2 (49.1) 337.8 (68.9) 224.4 (61.5) Observed 227.7 (33.7) 242.5 (56.9) 417.7 (107.1) 361.6 (125.3) P. value<0.01; According to logistic regression analysis and eliminating the effect of group [OR=1.009; 95% CI (1.002-1.017)] NRBC: Nucleated red blood cells; SD: Standard deviation

Mean (SD) Total (n=150) Mean (SD) Unobserved 127.0 (52.0) 212.2 (49.1) 337.8 (68.9) 224.4 (61.5) Observed 227.7 (33.7) 242.5 (56.9) 417.7 (107.1) 361.6 (125.3) P. value<0.01; According to logistic regression analysis and eliminating the effect of group [OR=1.009; 95% CI (1.002-1.017)] NRBC: Nucleated red blood cells; SD: Standard deviation After the intervention, the anxiety score decreased by 20.5 points in the acupressure group and by 20 points in the supportive care group, but increased by 11.1 points in the control group. Chao et al (2007) also performed acupressure at various points and revealed this non-pharmacological method to be effective in reduction of women's pain and anxiety during labor[21]. In the same line, the results of the study by Fassoulaki et al (2003) showed the effectiveness of acupressure in reduction of pain and stress[22]. These results were in line with those of the present study in which, the anxiety score decreased by 34.8% in the acupressure group after the intervention. Considering the supportive care group, the present study results were in agreement with those of the study by Hofmeyr et al (1991) reporting the anxiety mean score as 28.2 in the supported group and 37.8 in the routine care group[23]. In the current study also, the mean score of anxiety reduced by 37.5% in the supportive care group after the intervention which might be due to accompanying and supporting the mother leading to reduction of her pain and anxiety. Pilkington et al (2007) also expressed the effect of acupressure on the patients’ anxiety level based on Spielberger’s scale[24]. Various studies have shown that acupressure, without any complications, controls and reduces anxiety by stimulating brain responses and hormonal activities through increasing the blood flow and mediating the metabolism[25]. Therefore, it can be concluded that mother’s anxiety which is accompanied by increased uterine vessels' resistance and decreased oxygenation plays a major role in fetal as well as maternal outcomes. This emphasizes the necessity for performing interventions through labor in order to reduce the mothers’ anxiety. On the contrary, Langer et al (1998) showed that supporting the women through labor was not effective in the need for medical interventions and mothers' anxiety, pain, self-confidence, and satisfaction. In that study, the mean score of anxiety was 49.1 in the supported group and 49.2 in the control group and the difference was not statistically significant[26].

that supporting the women through labor was not effective in the need for medical interventions and mothers' anxiety, pain, self-confidence, and satisfaction. In that study, the mean score of anxiety was 49.1 in the supported group and 49.2 in the control group and the difference was not statistically significant[26]. The difference between that study and the present one might be due to intervention methods, hospital policies, women's cultural background, short period of support, and types of doulas' activities. Yet the support provided by the nurses and the staff might have also been effective in reducing the difference between the two groups. In this study, distribution of the NRBCs in the peripheral blood smears was 18%, 20%, and 86% in the supportive care, acupressure, and control groups, respectively. Stress and anxiety lead to secretion of epinephrine. Table 4 The relationship between the frequency of NRBC in the cord blood and delivery mode in the intervention (supportive care and acupressure) and control groups Delivery mode NRBC in cord blood count, peripheral smear Intervention (n=100) n (%) Control (n=50) n (%) Vaginal delivery Unobserved 80 (86.0) 7 (23.3) Observed 13 (14.0) 23 (76.7) Caesarean section Unobserved 1 (14.3) 0 Observed 6 (85.7) 20 (100) P. value<0.001 in intervention group and 0.3 in control group; NRBC: Nucleated red blood cells

Table 4 The relationship between the frequency of NRBC in the cord blood and delivery mode in the intervention (supportive care and acupressure) and control groups Delivery mode NRBC in cord blood count, peripheral smear Intervention (n=100) n (%) Control (n=50) n (%) Vaginal delivery Unobserved 80 (86.0) 7 (23.3) Observed 13 (14.0) 23 (76.7) Caesarean section Unobserved 1 (14.3) 0 Observed 6 (85.7) 20 (100) P. value<0.001 in intervention group and 0.3 in control group; NRBC: Nucleated red blood cells Epinephrine through beta-adrenergic receptors in the uterus leads to uterine muscle hypoxia, disruption in uteroplacental blood perfusion, and fetal hypoxia[27]. Basically, intrauterine hypoxia is one of the main factors in increasing erythropoietin which stimulates the fetal hematopoietic system and increases the production of NRBCs. Since NRBCs can change their shape, size, and expansibility, they are released from the bone marrow into the peripheral blood. In the current study, the number of the cases with NRBCs detected in the peripheral blood smears was higher in the control group compared to the two intervention groups. Thus, fetal hypoxia was probably higher in the control group which received no interventions for reduction of pain and anxiety during labor. Several studies have indicated a significant relationship between the number of NRBCs and fetal hypoxia and have considered the number of NRBCs as an important index for identifying hypoxia[28-30]. Predicting the importance of fetal hypoxia needs awareness of its duration and grade[31]. NRBC count increases in acute and chronic hypoxia but the more the fetal hypoxia, the more increases NRBC count[29]. The average rate of nucleated red blood cells is 500 in mm3 in the first hours of healthy term infant’s life (more than 1000 is abnormal)[32]. 1 to 2 percent of healthy infants have increased nucleated red blood cells without any reason[33]. For instance, in the study by Tomar et al (2011), the mean number of NRBCs was 10.34+3.87 in the fetal distress group and 5.7+2.33 in the control group[34]. In addition, Hanion-Lundberg et al (1999) measured the mean number of NRBCs as 9.2+18.1 in 1561 hypoxic infants[35]. In the study by Kovalak et al (2011) the mean number of NRBCs was 13 (range: 0-37) in the case group and 8 (range: 0-21) in the control group[36]. In the present study, providing the mothers with doula supportive care and applying pressure at BL32 acupoint were performed to decrease the effective factors in fetal hypoxia including anxiety, and prevent the undesirable fetal and maternal outcomes. However, no such interventions were carried out in the studies by Hanion-Lundberg and Kovalak.

iding the mothers with doula supportive care and applying pressure at BL32 acupoint were performed to decrease the effective factors in fetal hypoxia including anxiety, and prevent the undesirable fetal and maternal outcomes. However, no such interventions were carried out in the studies by Hanion-Lundberg and Kovalak. In Hanion-Lundberg’s study, NRBC count in infants with first minute’s Apgar 0-3 was significantly and statistically higher than that in infants with Apgar >7; this study included also infants with diabetic mothers and meconium stained cases. In our study, these cases and also infants with basis problem were excluded and pregnancy was terminated immediately after detection of abnormal heart models. Also, ill neonates with low Apgar that needed NICU were not in our study. Our neonates despite the abnormal heart rate model in control group had Apgar scores higher than 7 and did not need resuscitative measures and NICU care, and distress and intrauterine hypoxia intensity and duration were not high enough to cause over increase in NRBC count.

Apgar that needed NICU were not in our study. Our neonates despite the abnormal heart rate model in control group had Apgar scores higher than 7 and did not need resuscitative measures and NICU care, and distress and intrauterine hypoxia intensity and duration were not high enough to cause over increase in NRBC count. In this study, the labor was longer in the cases with fetal NRBCs detected in the peripheral blood smears. Besides, the highest and lowest labor length was related to the control (417.7+107.1) and supportive care group (227.7+33.7), respectively. As the labor length increases, mother's anxiety increases, as well. Thus, due to secretion of stress hormones, including cortisol, epinephrine, and norepinephrine, the vessels are contracted and less oxygen is delivered to the fetus. Then, the production of erythropoietin is increased in response to hypoxia and NRBCs enter the peripheral blood. A large number of studies have also shown that the infants exposed to intrauterine hypoxia have more NRBCs in their cord blood [28-30,37,38]. In the studies by Ferns[39] and Lim[40] also, a significant correlation was found between the length of labor and the number of NRBCs, which is consistent with the findings of the present study. However, Kovalak[36] reported no significant relationship between the number of NRBCs and the length of labor. This difference might result from the fact that most of the control group women in Kovalak's study underwent cesarean section and, consequently, the length of labor was shorter in the control group compared to the case group (distressed infants). In contrast to the current study, Ghosh[8] did not consider hypertension, preeclampsia, tobacco use, meconium, growth restriction, and chorioamnionitis which can increase the number of NRBCs as the exclusion criteria.

the length of labor was shorter in the control group compared to the case group (distressed infants). In contrast to the current study, Ghosh[8] did not consider hypertension, preeclampsia, tobacco use, meconium, growth restriction, and chorioamnionitis which can increase the number of NRBCs as the exclusion criteria. In this study, a larger number of NRBCs were detected in peripheral blood smears in cesarean delivery in all the three groups. This implies a significant relationship between the type of delivery and the number of NRBCs in the blood cord in both the intervention groups and the control group. In line with these results, Ferns[39] also believed that the number of NRBCs was affected by the delivery mode. On the contrary, in the study conducted by Saracoglu et al (2000), the mean number of NRBCs was 8.25+3.39 and 7.58+4.04 in the cesarean and natural delivery, respectively and the difference was not statistically significant[41]. Kovalak[36], Axt[33], Ghosh[8], Hanion-Lundberg[35], and Korst[28] also found no significant relationship between the number of NRBCs and type of delivery. This might be due to the fact that the above-mentioned studies had not exclude hypertension, preeclampsia, tobacco use, meconium, growth restriction, and chorioamnionitis. These factors increase the number of NRBCs regardless of the delivery mode and labor length because by uterine vessels contraction in these patients, perfusion and oxygenation to the fetus is decreased and more NRBCs are produced and entered into the peripheral blood in response to hypoxia.

ction, and chorioamnionitis. These factors increase the number of NRBCs regardless of the delivery mode and labor length because by uterine vessels contraction in these patients, perfusion and oxygenation to the fetus is decreased and more NRBCs are produced and entered into the peripheral blood in response to hypoxia. Conclusion The findings of the present study showed that supportive care and acupressure were effective, safe, simple, and inexpensive techniques which could be used in order to reduce the delivery pain, mother’s anxiety, and length of labor, resulting in improvement of oxygenation to the fetus and prevention of asphyxia. Asphyxia, regardless of duration, does not always increase the NRBC count and NRBC count is far more increased in some cases without asphyxia. Therefore, NRBC count is not the only decisive factor of intrauterine asphyxia intensity and duration. In case a large number of NRBCs are detected in infants, they should be provided with more care in order to prevent them from probable nervous damages. Although the number of NRBCs alone cannot be a prognostic factor, further studies on this issue can confirm or reject the results obtained in the current study. Overall, considering the national and international approach toward physiological delivery, this non-pharmacological method is recommended to be used to reduce the mothers' anxiety during labor and improve the maternal and fetal outcomes.

es on this issue can confirm or reject the results obtained in the current study. Overall, considering the national and international approach toward physiological delivery, this non-pharmacological method is recommended to be used to reduce the mothers' anxiety during labor and improve the maternal and fetal outcomes. One of the limitations of this study was interference of gynecologists and midwives in the normal process of labor. In case their interventions affected the results, the patient was excluded from the study. Moreover, the participants’ mental conditions while completing the questionnaire, family problems, and undiagnosed diseases could affect the study results, which was out of the researcher’s control. Authors’ Contribution Z. Masoudi, M Akbarzadeh, F. Vaziri and M. Ramzi conceived of the study and participated in its design and coordination and helped to draft the manuscript. Z. Masoudi collected the data. Z. Masoudi and N. Zare performed the statistical analysis. Z. Masoudi and M. Akbarzadeh made critical revisions to the paper and translated it into English language. All authors read and approved the final version of the paper.

s design and coordination and helped to draft the manuscript. Z. Masoudi collected the data. Z. Masoudi and N. Zare performed the statistical analysis. Z. Masoudi and M. Akbarzadeh made critical revisions to the paper and translated it into English language. All authors read and approved the final version of the paper. Acknowledgment The present article was extracted from Ms. Zahra Masoudi’s MSc thesis (proposal No. 6356, IRCT 2013081411706N3). The study was financially supported by the Research Vice-chancellor of Shiraz University of Medical Sciences, Shiraz, Iran. Hereby, the authors would like to thank the authorities of the college of nursing and midwifery and physicians and midwives of Shoushtari Hospital, Shiraz, Iran. They are also grateful for Ms. A. Keivanshekouh at Research Improvement Center of Shiraz University of Medical Sciences for improving the use of English in the manuscript. Conflict of Interest: None

Introduction Sydenham’s chorea (SC) is a late manifestation of acute rheumatic fever (ARF) and can occur several months after group A β-hemolytic Streptococcus infections[1]. Although the incidence of ARF and SC has declined significantly in developed countries, they are still serious health concerns in developing countries. Sydenham’s chorea is characterized by involuntary choreiform movements with or without other motor symptoms including facial grimacing, hypotonia, muscle weakness, gait disturbance, difficulty in writing and speaking. Generalized weakness and hypotonia can be so severe that the patient becomes bedridden. This special form of SC is called “chorea paralytica”. Neuropsychiatric symptoms are also commonly observed[2]. There is no specific laboratory test for SC. The diagnosis relies on a careful clinical history and laboratory assessment to rule out other causes of the symptoms, such as systemic lupus erythematosus, drug intoxication, Wilson’s disease, familial chorea and hyperthyroidism[3-5]. The neuroimaging findings of SC have been rarely reported. In this study, the clinical and cranial MRI findings of 17 patients with SC were evaluated.

assessment to rule out other causes of the symptoms, such as systemic lupus erythematosus, drug intoxication, Wilson’s disease, familial chorea and hyperthyroidism[3-5]. The neuroimaging findings of SC have been rarely reported. In this study, the clinical and cranial MRI findings of 17 patients with SC were evaluated. Subjects and Methods Seventeen patients with acute SC who were admitted to the Pediatric Neurology Unit of the Eskisehir Osmangazi University Faculty of Medicine were retrospectively evaluated between 2010 and 2012. Sydenham’s chorea was diagnosed according to the 1992 revision of the Jones criteria[1]. Patient medical histories were collected. Physical and neurological examinations were performed. Anti-streptolysin O (ASO), erythrocyte sedimentation rate and the total blood count were routinely obtained from patients at the time of their admission to the hospital. An ASO titer greater than 250 Todd U/mL was considered to be elevated. Tandem MS, urine organic acid analysis and screening for lysosomal enzymes were performed in one patient. Cranial MRI was performed in all patients during the acute phase of SC. MRI examinations were performed on 1.5 T MR scanner (Siemens, VisionPlus, Germany) equipped with the head coil. The MRI protocol included T1 and T2 spin echo sequences in axial planes, fluid attenuation inversion recovery in coronal plane. After 0.1 mmol/kg body weight intravenous gadolinium injection, axial and sagittal T1-weighted image were obtained. The DWI was performed with spin–echo EPI sequence with three b values (0, 500 and 1000).

ocol included T1 and T2 spin echo sequences in axial planes, fluid attenuation inversion recovery in coronal plane. After 0.1 mmol/kg body weight intravenous gadolinium injection, axial and sagittal T1-weighted image were obtained. The DWI was performed with spin–echo EPI sequence with three b values (0, 500 and 1000). The other causes of chorea were excluded. If SC was the only major feature of ARF, an ophthalmic examination, a thyroid function test (TFT), antinuclear antibody (ANA) and anti-dsDNA antibody, serum ceruloplasmin and tests of liver enzymes were performed. Chorea was classified as follows: a)    mild (minimal movements), b)    moderate (inconvenient movements for the patient that do not interfere with his or her personal care), c)    severe (incapacitating movements requiring the patient to have help for daily activities). Sydenham chorea was also classified according to localization as follows: generalized (movements affecting the whole body) or hemichorea (movements affecting one side of the body). If the patient became bedridden because of generalized hypotonia, they were diagnosed as chorea paralytica[2]. Patients were followed up at one month intervals. Walking, speech and swallowing disorders, muscle weakness, behavioral disorders, treatment, symptom recovery time and recurrence were evaluated. All patients received ARF prophylaxis with benzathine G penicillin. Chorea treatment was also investigated in all patients. Table 1 The clinical findings and treatment of the patients Patient No Age years/gender Chorea Chorea scale Muscle weakness Speech disorder Dysphagia Behavioral changes Carditis treatment

Sydenham chorea was also classified according to localization as follows: generalized (movements affecting the whole body) or hemichorea (movements affecting one side of the body). If the patient became bedridden because of generalized hypotonia, they were diagnosed as chorea paralytica[2]. Patients were followed up at one month intervals. Walking, speech and swallowing disorders, muscle weakness, behavioral disorders, treatment, symptom recovery time and recurrence were evaluated. All patients received ARF prophylaxis with benzathine G penicillin. Chorea treatment was also investigated in all patients. Table 1 The clinical findings and treatment of the patients Patient No Age years/gender Chorea Chorea scale Muscle weakness Speech disorder Dysphagia Behavioral changes Carditis treatment 1 14/F G 3 + + - + + VA 2 9.5/F G 3 CP + + + + - VA, HIVMP 3 8/F G 3 + + + + - VA 4 9.5/M H 2 - + - - + PHB 5 11.5/M H 3 - + - + - VA 6 11/F H 2 - + - - - VA 7 8.5/M G 3 + + + + - VA 8 8/F G 3 + + - + + VA 9 15/F G 3 + + - + - VA 10 6/F G 3 + + - + - VA 11 13/M H 2 - - - - + VA 12 8/F H 3 + + - + - VA 13 12/F G 3 + + - + - VA 14 15/F G 1 - - - - - HL 15 13.5/F G 3 CP + + + + - VA, HIVMP 16 16.5/F H 2 - - - - - PHB 17 12/F G 3 + + - + - VA G: generalized; H: hemichorea; CP: chorea paralytica; VA: valproic acid; PHB: phenobarbital; HL: haloperidol; MP: methylprednisolone; HIVMP: high-dose intravenous methylprednisolone Table 2 The neuroimaging findings of the patients Patient No MRI finding Control MRI Other findings Finding Time (months)

16 16.5/F H 2 - - - - - PHB 17 12/F G 3 + + - + - VA G: generalized; H: hemichorea; CP: chorea paralytica; VA: valproic acid; PHB: phenobarbital; HL: haloperidol; MP: methylprednisolone; HIVMP: high-dose intravenous methylprednisolone Table 2 The neuroimaging findings of the patients Patient No MRI finding Control MRI Other findings Finding Time (months) 1 Hyperintensity within the caudate nuclei Regression 4 - 2 Confluent hyperintense lesions in the bilateral periventricular-subcortical white matter No change 6 and 12 - 3 Hyperintense foci in white matter on T2-WI Normal 6 - 4 Hyperintense foci in the white matter on T2-WIs Normal 6 - 5 Hyperintense foci in the white matter on T2-WIs No change 6 MR angiography: Normal 6 Hyperintense foci in the white matter on T2-WIs Normal 6 - 10 Hyperintense foci in the brain stem on T2-WIs - - - 15 A hyperintense cortical lesion in the left temporal region and no contrast enhancement No change 6 and 12 MR spectroscopy: Lactate peak in the left lateral temporal gyrus MRI: Magnetic Resonance Imaging Findings

1 Hyperintensity within the caudate nuclei Regression 4 - 2 Confluent hyperintense lesions in the bilateral periventricular-subcortical white matter No change 6 and 12 - 3 Hyperintense foci in white matter on T2-WI Normal 6 - 4 Hyperintense foci in the white matter on T2-WIs Normal 6 - 5 Hyperintense foci in the white matter on T2-WIs No change 6 MR angiography: Normal 6 Hyperintense foci in the white matter on T2-WIs Normal 6 - 10 Hyperintense foci in the brain stem on T2-WIs - - - 15 A hyperintense cortical lesion in the left temporal region and no contrast enhancement No change 6 and 12 MR spectroscopy: Lactate peak in the left lateral temporal gyrus MRI: Magnetic Resonance Imaging Findings The patients were predominantly female (13/17). The average age at onset of Sydenham’s chorea was 11.23 years (SD=3.01 years, range 6-16.5 years). Three patients had been previously diagnosed as ARF and the other 14 patients (82%) had a history of throat infection within two months. The serum ASO level was elevated in all patients (ASO >250 Todd U/mL). Sydenham’s chorea was the only major finding of ARF in 76% of the patients and 24% of the patients presented with accompanying carditis. Chorea was mild in 1 (5.9%) patient, moderate in 4 (23.5%) patients and severe in 12 (70.6%) patients. Two (11%) patients were diagnosed with chorea paralytica. Chorea was generalized in 11 (64%) patients and 6 patients had hemichorea (36%). Fourteen (82%) patients had a speech disorder. Behavioral changes, muscle weakness and dysphagia occurred in 70%, 64% and 23% of the patients, respectively. The clinical findings and treatments are summarized in Table 1. Cranial MRI was performed in all patients and abnormalities were observed in 8 (47%) patients. The neuroimaging findings are shown in Table 2. Nonspecific signal hyperintensities were observed on T2-weighted images in the white matter, brain stem and caudate nucleus (Fig. 1). In patient 15, MR spectroscopy demonstrated a hyperintense cortical lesion in the left temporal region, which was suggestive of a dysembryoplastic neuroepithelial tumor (DNET). Control MRI was performed after 4-12 months of follow up in 7 of the patients. Three of 8 patients with neuroimaging findings demonstrated complete resolution after 6 months, while 1 patient’s MRI findings regressed after 4 months. In two patients with chorea paralytica, the hyperintense lesions observed using MRI were unchanged after 12 months of follow up. Fifteen patients were administered valproic acid (VA), phenobarbital (PHB), or haloperidol (HL) treatment for 1-8 months. After the disappearance of the patient's choreiform movements, the medication was tapered over a period of two months. Two patients who had chorea paralytica were administered high-dose intravenous methylprednisolone (HIVMP) (30 mg/kg/day, 5 days).

VA), phenobarbital (PHB), or haloperidol (HL) treatment for 1-8 months. After the disappearance of the patient's choreiform movements, the medication was tapered over a period of two months. Two patients who had chorea paralytica were administered high-dose intravenous methylprednisolone (HIVMP) (30 mg/kg/day, 5 days). Bedridden and dysphagia improved within the first week and chorea and the other symptoms recovered within 1 month. Choreiform movements of the 15 patients who were not given HIVMP, recovered completely within 1-7 months. Muscle weakness, speech disorders, dysphagia and behavioral changes disappeared in the first and second controls. The mean follow-up period was 9.6 (5-21) months and there was no recurrence during the follow-up period. Fig. 1 ( A,B) A hyperintense lesion within the right caudate nuclei is observed on both T2-weighted axial image and the FLAIR coronal image. (C,D) In the follow-up MRI studies, lesion regression was evident after 4 months (Patient 1). Fig. 2 (A,B) Both T2-weighted axial image and the FLAIR coronal image show confluent hyperintense lesions in the bilateral periventricular-subcortical white matter. These lesions persist in the follow-up images (Patient 2). Fig. 3 T2-weighted axial image shows a single hyperintense focus in the left frontal white matter (Patient 5). Fig. 4 (A,B) The hyperintense cortical lesion in the left temporal region on a T2-weighted axial image and a FLAIR coronal image (Patient 15).

Fig. 2 (A,B) Both T2-weighted axial image and the FLAIR coronal image show confluent hyperintense lesions in the bilateral periventricular-subcortical white matter. These lesions persist in the follow-up images (Patient 2). Fig. 3 T2-weighted axial image shows a single hyperintense focus in the left frontal white matter (Patient 5). Fig. 4 (A,B) The hyperintense cortical lesion in the left temporal region on a T2-weighted axial image and a FLAIR coronal image (Patient 15). Discussion Sydenham’s chorea is a poststreptococcal autoimmune disease and it is the most common cause of chorea during childhood[2]. Sydenham’s chorea manifests in children aged 5-15 years, with a female predominance[6]. The major neurological features of SC are involuntary movements, which are exacerbated by stress and disappear during sleep. Chorea is defined as distal, rapid, purposeless movements. They can be generalized or observed on one side of the body (hemichorea). Incoherent speech, dysarthria, muscle weakness and hypotonia are commonly associated features. Hypotonia and weakness have a range of severity from mild to severe. Flaccid quadriparesis, which is also known as “chorea paralytica”, can develop. Neuropsychiatric symptoms, such as emotional lability, crying spells, irritability, tics and obsessive-compulsive signs, are often observed in SC[2,7]. In this study, 76% of the patients were female and the mean age was 11.2 years. These findings are consistent with the literature[7,8]. Chorea was generalized in 64% of our patients. More than half of the patients had a speech disorder, behavioral changes and muscle weakness and one fourth of the patients had dysphagia. Our two patients with chorea paralytica had severe weakness and dysphagia, hospitalization was therefore required.

he literature[7,8]. Chorea was generalized in 64% of our patients. More than half of the patients had a speech disorder, behavioral changes and muscle weakness and one fourth of the patients had dysphagia. Our two patients with chorea paralytica had severe weakness and dysphagia, hospitalization was therefore required. Approximately 35% of patients with ARF develop chorea. Cardiac involvement occurs in 42-70.5% of cases with chorea[9,10]. In this study, SC was the only finding of ARF in 76% of the patients and 24% of the patients presented with accompanying carditis. The serum ASO level was elevated in all patients and 82% of the patients had a history of throat infection[11]. In our study the serum ASO level was elevated in all patients. The ASO titer peaks 3 to 5 weeks after the onset of GABHS pharyngitis and then gradually declines over the following weeks[2]. Although SC is a late complication of GABHS, ASO titers were higher in most patients in some studies. Fusco et al12 reported that serum ASO titers were elevated in nine of ten patients with SC. And also elevated ASO was demonstrated in 80% of cases[13]. In the study of Ridel et al14, ASO and anti-DNase B titers were elevated in all patients with SC as in our study. It may be speculated that depending on the initial very high titers, ASO titers may still be high at diagnosis of SC.

ine of ten patients with SC. And also elevated ASO was demonstrated in 80% of cases[13]. In the study of Ridel et al14, ASO and anti-DNase B titers were elevated in all patients with SC as in our study. It may be speculated that depending on the initial very high titers, ASO titers may still be high at diagnosis of SC. Sydenham’s chorea is thought to be an autoimmune disorder, but the exact pathophysiology is still unclear. It is believed that antibodies against group A β-hemolytic Streptococcus (GABHS) cross-react with neurons of the basal ganglia. These anti-basal ganglia antibodies react with the surface of neuronal cells and signal the induction of calcium calmodulin-dependent protein kinase II. Thus, the tyrosine hydroxylase level is elevated and dopamine is released, leading to the movement disorder[15,16]. Pathological studies have demonstrated neuronal loss, cytoplasmic and nuclear cell changes, gliosis, endothelial swelling, perivascular round cell infiltration and petechial hemorrhages within the cerebral cortex, basal ganglia and thalamus[17].

levated and dopamine is released, leading to the movement disorder[15,16]. Pathological studies have demonstrated neuronal loss, cytoplasmic and nuclear cell changes, gliosis, endothelial swelling, perivascular round cell infiltration and petechial hemorrhages within the cerebral cortex, basal ganglia and thalamus[17]. The diagnosis of SC is difficult without carditis or other manifestations of ARF. Other causes of chorea should be excluded[3-5]. Although increased ASO titers exist in two thirds of cases, it is not helpful in the diagnosis of SC[18]. Magnetic resonance imaging is generally studied to exclude other causes of chorea. There are no typically defined MRI features in SC. MRI may show varying degrees of signal hyperintensity on T2-weighted images in focal regions, such as the corpus striatum, caudate nucleus, putamen and multiple other areas. These abnormalities may be localized to the basal ganglia, but they are often not consistent with the patient’s clinical signs. The MRI findings of SC typically disappear over time. Therefore, it is thought to develop as a result of vasculitis or inflammation. However, the persistence of these hyperintensities has been reported rarely[8,19-21]. In this study, increased signal intensities were detected in the white matter, brain stem and caudate nuclei. Some lesions completely regressed during follow up. We believe that the lesion regression may support an inflammatory origin. In patient 2, the confluent hyperintense lesions on the bilateral periven-tricular-subcortical white matter were still present at the follow-up MRI. These lesions might be secondary to ischemia or demyelination due to vasculitis or inflammation. In patient 15, the hyperintense cortical lesion was most likely consistent with DNET and we considered it to be an incidental finding. In our cases, to interpret all hyperintensities to be related to acute phase of SC may be speculative. However, based on data from other published reports, one or multiple increased signals in the basal ganglia or white matter have been known. In our cases, increased signal intensities were considered suggestive of vasculitis because none of the patients had other previously known causes of vasculitis and our findings were similar to previous reports.

r published reports, one or multiple increased signals in the basal ganglia or white matter have been known. In our cases, increased signal intensities were considered suggestive of vasculitis because none of the patients had other previously known causes of vasculitis and our findings were similar to previous reports. Sydenham’s chorea is a self-limiting condition with a mean duration of 2-4 months[22]. However, treatment is necessary for patients whose chorea is not mild. Antiepileptics, neuroleptics and phenothiazines have been reported to reduce the abnormal movements by affecting the dopaminergic or alpha-aminobutyric acid (GABA) pathways[2,23,24]. Intravenous immunoglobulin, plasma exchange and corticosteroids effectively reduce involuntary movements due to the pathogenesis of autoimmune SC[12,25]. However, because of the side effects of corticosteroids, their use is recommended only in chorea paralytica[26]. In our study, two patients with chorea paralytica who had MRI findings, persistent choreiform movements of the 15 patients not given HIVMP, received steroids and recovered completely within 1-7 months. Our two patients recovered more quickly with corticosteroids, which is in agreement with the literature[27]. We think that HIVMP appears to be a good choice for bedridden patients with serious dysphagia. Otherwise, classical drug therapy would seem to be more appropriate. In a previous study, recurrence was observed in approximately 30% of patients[28] and was associated with discontinuation of the antibiotic prophylactic therapy or poor compliance and perhaps with subclinical damage to the basal ganglia following the initial SC episode[29,30]. There was no recurrence in our study.

ate. In a previous study, recurrence was observed in approximately 30% of patients[28] and was associated with discontinuation of the antibiotic prophylactic therapy or poor compliance and perhaps with subclinical damage to the basal ganglia following the initial SC episode[29,30]. There was no recurrence in our study. Conclusion SC might be associated with nonspecific hyperintense white matter abnormalities in MRI. These abnormalities may be due to the inflammatory process associated with a longer duration of clinical signs. Because of the autoimmune role in the pathogenesis of SC, immunomodulatory therapies may be effective. To explain mechanisms behind the MRI findings and the pathogenesis of SC, comprehensive studies are needed. Authors’ Contribution Concept / Design: A. Ekici, A. Yakut, K.B. Carman Acquisition of Data: A. Ekici, S. Yimenicioğlu, S. Saylısoy Data Analysis / Interpretation: A. Ekici, A. Yakut, S. Yimenicioğlu, K.B. Carman, S. Saylısoy Manuscript Preparation: A. Ekici, A. Yakut Critical Revision of the Manuscript: A. Ekici, S. Saylısoy All authors approve final version of the paper. Acknowledgment This study was presented as a poster in 24th Annual Meeting of European Academy of Childhood Disability and published as an abstract in Development Medicine and Child Neurology, May 2012, Volume 54, Supplement 3, pp 43. Conflict of Interest: None

Introduction Metabolic syndrome (MetS) is a prevalent complex disorder consisting of concurrent metabolic abnormalities[1]. The prevalence of the syndrome is 1-2% in Iranian children and adolescents, much higher than that reported for other ethnicities[2-4]. It markedly increases the risk of developing cardiovascular diseases and type 2 diabetes[5]. According to the ATPIII criteria, MetS can be diagnosed based on hypertension, central obesity, insulin resistance and dyslipidemia. However, the most frequent components of the MetS in Iranian children and adolescents are low high-density lipoprotein cholesterol (HDL-C) and high triglycerides (TG)[2]. Among the genetic variants associated with the development of MetS, there is a naturally occurring variant (-1131T>C) in the promoter region of apolipoprotein A5 gene (APOA5). The mature APOA5 protein expresses exclusively in the liver and secretes into the plasma to modulate TG metabolism[6]. The -1131T>C (rs662799) variant of APOA5 gene is associated with increased triglyceride levels and confers risk for metabolic syndrome in adult populations[7-13]. However, little is known about the APOA5 variants in pediatric MetS. In the present study, we evaluated the association of rs662799 variant of the APOA5 gene with MetS in Isfahanian children and adolescents.

ssociated with increased triglyceride levels and confers risk for metabolic syndrome in adult populations[7-13]. However, little is known about the APOA5 variants in pediatric MetS. In the present study, we evaluated the association of rs662799 variant of the APOA5 gene with MetS in Isfahanian children and adolescents. Subjects and Methods A total of 50 cases of MetS and 50 controls were recruited to this study. Controls had normal weight and were healthy looking, without any clinical, laboratory or history records for MetS, diabetes or cardiovascular disorders. To diagnose the individuals with MetS, we used the modified ATPIII definition[2]. In brief, a MetS subject fulfilled at least three of the following components: waist circumference >75th percentile for age and gender in the studied population; fasting TG ≥100mg/dl; HDL-C <50 mg/dl (except in 15-19 year boys in whom the cut off was <45 mg/dl); systolic blood pressure/diastolic blood pressure >90th percentile recommended cut off points by the National Heart, Lung and Blood institute for gender, age and height[14]; fasting blood sugar (FBS) ≥100 mg/dl[2]. This study was approved by the Ethic Committee of Isfahan University. Informed consent was obtained from parents. The experimental design was approved by the Ethics Committee of Shahrekord University.

y the National Heart, Lung and Blood institute for gender, age and height[14]; fasting blood sugar (FBS) ≥100 mg/dl[2]. This study was approved by the Ethic Committee of Isfahan University. Informed consent was obtained from parents. The experimental design was approved by the Ethics Committee of Shahrekord University. Blood specimens were collected in EDTA-treated tubes and were stored at –70°C for further analysis. Extraction of DNA from total blood was performed using the Diatom DNA Prep 100 kit (Isogen Laboratory, Russia). mPCR-RFLP was used to genotype T>C polymorphism with primers described elsewhere[7]. The amplified fragment possessed an obligatory cleavage site for TruI restriction endonuclease to check if the digestion has occurred. Thermal conditions of the PCR cycles were as follow: initial denaturation at 95°C followed by 35 amplification cycles consisting of denaturation at 95°C for 40 sec; annealing at 61°C for 40 sec; extension at 72°C for 40 sec and a final extension at 72°C for 10 min. 10 μl of the PCR product was digested with 5 U of TruI enzyme. According to the digestion patterns, three genotypes were determined: TT genotype resulted in 21, 108 and 267 bp fragments, TC genotype created 21, 108, 267 and 288 bp products and CC genotype produced 108 and 288 bp fragments.

extension at 72°C for 10 min. 10 μl of the PCR product was digested with 5 U of TruI enzyme. According to the digestion patterns, three genotypes were determined: TT genotype resulted in 21, 108 and 267 bp fragments, TC genotype created 21, 108, 267 and 288 bp products and CC genotype produced 108 and 288 bp fragments. Statistical analyses were done with SPSS software (version 20.0). P values less than 0.05 were considered significant. A Chi-square statistic was calculated to compare the frequencies of genotypes and alleles. One way ANOVA was conducted to evaluate any differences between different groups, regarding the levels of biochemical factors and the distribution of different genotypes. Logistic regression analysis was performed to derive the odds ratios. Findings The major relevant clinical and biochemical characteristics of the study participants are presented in Table 1. The main risk factors for MetS were significantly augmented in the MetS cases except for HDL-C (P<0.01). Table 1 Clinical and biochemical data of the MetS and control subjects Parameter Cases (n=50) Controls (n=50) P value Boys/Girls 22(44%)/28(56%) 25(50%)/25(50%) 0.5 Body mass index (kg/m 2 ) 26.4 (0.45) 17.85 (0.94) <0.001 Age (years) 12.18 (0.24) 13.48 (0.44) 0.01 Triglyceride (mg/dl) 109.64 (8.24) 78.44 (5.12) 0.002 Total Cholesterol (mg/dl) 165.12 (3.80) 151.48 (14.14) 0.02 High Density Lipoprotein-C (mg/dl) 43.08 (0.74) 50.44 (1.74) <0.001 Low Density Lipoprotein -C (mg/dl) 95.66 (3.32) 79.32 (1.88) <0.001

Body mass index (kg/m 2 ) 26.4 (0.45) 17.85 (0.94) <0.001 Age (years) 12.18 (0.24) 13.48 (0.44) 0.01 Triglyceride (mg/dl) 109.64 (8.24) 78.44 (5.12) 0.002 Total Cholesterol (mg/dl) 165.12 (3.80) 151.48 (14.14) 0.02 High Density Lipoprotein-C (mg/dl) 43.08 (0.74) 50.44 (1.74) <0.001 Low Density Lipoprotein -C (mg/dl) 95.66 (3.32) 79.32 (1.88) <0.001 Fasting Blood Suger (mg/dl) 102.56 (1.35) 91.24 (1.77) <0.001 Table 2 Biochemical factor levels in MetS and control subjects according to the APOA5 -1131T>C genotypes Biochemical factors Cases P. value Controls P. value TT (n=43) TC+CC (n=7) TT (n=35) TC+CC (n=15) TG (mg/dl) 111.93 (59.55) 95.57 (51.33) 0.2 77.37 (36.14) 80.93 (37.59) 0.4 TC (mg/dl) 164.05 (24.99) 171.71 (38.74) 0.2 152.51 (30.52) 149.07 (27.07) 0.3 HDL-C (mg/dl) 43.37 (4.99) 41.29 (5.67) 0.2 49.97 (13.39) 51.53 (9.97) 0.3 FBS (mg/dl) 102.30 (8.64) 104.14 (15.08) 0.3 91.60 (14.51) 90.40 (6.35) 0.4 LDL-C (mg/dl) 93.93 (21.41) 106.29 (33.93) 0.1 80.71 (12.56) 76.07 (14.80) 0.1 TG: Triglyceride; TC: Total cholesterol; HDL-C: High density lipoprotein-cholesterol; LDL-C: Low density lipoprotein-cholesterol; FBS: Fasting blood sugar

HDL-C (mg/dl) 43.37 (4.99) 41.29 (5.67) 0.2 49.97 (13.39) 51.53 (9.97) 0.3 FBS (mg/dl) 102.30 (8.64) 104.14 (15.08) 0.3 91.60 (14.51) 90.40 (6.35) 0.4 LDL-C (mg/dl) 93.93 (21.41) 106.29 (33.93) 0.1 80.71 (12.56) 76.07 (14.80) 0.1 TG: Triglyceride; TC: Total cholesterol; HDL-C: High density lipoprotein-cholesterol; LDL-C: Low density lipoprotein-cholesterol; FBS: Fasting blood sugar Table 2 shows the comparison of the laboratory parameters of the control and MetS subjects stratified based on rs662799 genotypes. The data demonstrated that there was no evidence of an association of APOA5 genotypes with any of laboratory parameters. Genotype and allele frequencies are shown in Table 3. The data revealed that the frequency of C allele was greater in controls compared with that of the MetS. The odds ratio of the C allele in MetS versus control subjects in the unadjusted model was OR=0.38, 95%CI: 0.14-1.03, P=0.05. The Odds ratio adjusted for sex, age, HDL-C and TC was also calculated (OR=0.29, 95%CI: 0.08-1.07, P=0.06) and no significant differences between adjusted and unadjusted models were found (Table 4). Discussion We did not find an association between the -1131C polymorphism and elevated triglyceride levels. A number of studies have reported a significant association between triglyceride levels and the -1131T>C variant in MetS adults and obese children[7-10,15-18]. Our finding was in agreement with Mattei 's[19] results which showed no association between TG levels and the -1131T>C. Table 3 Genotype and allele frequencies Group Cases (n=50) Controls (n=50) Allele frequencies C 6 (12%) 9 (18%) T 44 (88%) 41 (82%)

Discussion We did not find an association between the -1131C polymorphism and elevated triglyceride levels. A number of studies have reported a significant association between triglyceride levels and the -1131T>C variant in MetS adults and obese children[7-10,15-18]. Our finding was in agreement with Mattei 's[19] results which showed no association between TG levels and the -1131T>C. Table 3 Genotype and allele frequencies Group Cases (n=50) Controls (n=50) Allele frequencies C 6 (12%) 9 (18%) T 44 (88%) 41 (82%) Genotype frequencies TT 43 (86%) 35 (70%) TC 2 (4%) 12 (24%) CC 5 (10%) 3 (6%) This inconsistency may be due to ethnicity influences[20]. Moreover, no association was found between the -1131T>C and MetS in our study. Significant correlation of -1131T>C APOA5 variant with MetS in adults has been indicated in several[8-13] but not all previous studies[7,17-19]. The findings of the current study are consistent with a recent meta-analysis[21] which showed that C allele carriers of -1131T>C had overall a significantly higher risk of MetS. After performing subgroup analysis according to ethnicity, the association was only significant in Asians, but not in white populations. Conclusion We showed that -1131T>C variant was neither associated with triglyceride levels nor MetS. Our results call for further studies to explore the effect of other APOA5 SNPs and haplotypes in Iranian children and adolescents. Table 4 Binary logistic regression analysis of the association between carrying APOA5 -1131C allele and the risk for MetS Group TT TC+CC Cases (n=50) 43 (86%) 7 (14%) Controls (n=50) 35 (70%) 15 (30%) Unadjusted model Odds ratio 0.38

Conclusion We showed that -1131T>C variant was neither associated with triglyceride levels nor MetS. Our results call for further studies to explore the effect of other APOA5 SNPs and haplotypes in Iranian children and adolescents. Table 4 Binary logistic regression analysis of the association between carrying APOA5 -1131C allele and the risk for MetS Group TT TC+CC Cases (n=50) 43 (86%) 7 (14%) Controls (n=50) 35 (70%) 15 (30%) Unadjusted model Odds ratio 0.38 95% CI 0.14-1.03 P. value 0.05 Adjusted model Odds ratio 0.29 95%CI 0.08-1.07 P. value 0.06 CI: Confidence Interval Authors’ Contribution S.G. Fatemi: Doing experiments, analysis of data, drafting the article. M. Emadi-Baygi: Conception, designing the study, analysis and interpretation of data the article. P. Nikpour: Conception, designing the study, analysis and interpretation of data and drafting and revising the article. R. Kelishadi: Sample collection, designing the study, analysis and interpretation of data the article. M. Hashemipour: Sample collection, analysis and interpretation of data the article. All authors approved final version of the manuscript. Acknowledgment This study was supported in part by a research grant to MEB from the Research Institute of Biotechnology, Shahrekord University, Shahrekord, Iran. Conflict of Interest: None

Introduction Neurogenic bladder results from neurological lesions that end in a lower urinary tract dysfunction[1], Abnormal bladder function can cause the bladder to be underactive (not emptying completely) or overactive (emptying too frequently/quickly). Children with NGB have a higher risk of urinary tract infection (UTI) and kidney damage. Neurogenic lower urinary tract dysfunction (NLUTD) may be caused by various diseases and events affecting the nervous system controlling the lower urinary tract. NLUTD depends grossly on the location and the extent of the neurologic lesion. Both in congenital and acquired NLUTD, early diagnosis and treatment is essential as irreversible changes may occur in paticular in children with myelomeningocle (MMC), but also in patients with traumatic spinal cord injury, even if the related neuropathologic signs were normal[2,3]. Diagnosis is based on history, physical examination, urodynamics, and typical manifestation of NLUTD. Treatment is based on non-invasive conservative measures such as catheterization, intravesical drug application (botulinum toxin injection, laser sphinctrotomy and urethral bulking agents). Surgical treatment can consist of urethral and bladder neck procedures, detrusor myectomy (auto-augmentation), denervation, covering bladder by striated muscle, bladder augmentation or substitution, and urinary diversion. Subjects and Methods In a retrospective and descriptive study, 33 patients who have been treated for NGBD were evaluated in Mofid Children’s Hospital from January 2007 to December 2012.

Introduction Neurogenic bladder results from neurological lesions that end in a lower urinary tract dysfunction[1], Abnormal bladder function can cause the bladder to be underactive (not emptying completely) or overactive (emptying too frequently/quickly). Children with NGB have a higher risk of urinary tract infection (UTI) and kidney damage. Neurogenic lower urinary tract dysfunction (NLUTD) may be caused by various diseases and events affecting the nervous system controlling the lower urinary tract. NLUTD depends grossly on the location and the extent of the neurologic lesion. Both in congenital and acquired NLUTD, early diagnosis and treatment is essential as irreversible changes may occur in paticular in children with myelomeningocle (MMC), but also in patients with traumatic spinal cord injury, even if the related neuropathologic signs were normal[2,3]. Diagnosis is based on history, physical examination, urodynamics, and typical manifestation of NLUTD. Treatment is based on non-invasive conservative measures such as catheterization, intravesical drug application (botulinum toxin injection, laser sphinctrotomy and urethral bulking agents). Surgical treatment can consist of urethral and bladder neck procedures, detrusor myectomy (auto-augmentation), denervation, covering bladder by striated muscle, bladder augmentation or substitution, and urinary diversion. Subjects and Methods In a retrospective and descriptive study, 33 patients who have been treated for NGBD were evaluated in Mofid Children’s Hospital from January 2007 to December 2012. Detailed history was taken and paraclinical examinations were performed and diagnosis was confirmed by ultrasonography (US), voiding-cysto-urethrography (VCUG), urodynamic study and lumbo-sacral MRI in myelodysplastic cases. We performed urodynamic evaluation without uroflowmetry in neonates after 3 months.

Subjects and Methods In a retrospective and descriptive study, 33 patients who have been treated for NGBD were evaluated in Mofid Children’s Hospital from January 2007 to December 2012. Detailed history was taken and paraclinical examinations were performed and diagnosis was confirmed by ultrasonography (US), voiding-cysto-urethrography (VCUG), urodynamic study and lumbo-sacral MRI in myelodysplastic cases. We performed urodynamic evaluation without uroflowmetry in neonates after 3 months. The patients were treated medically with anticholinergic and antibiotics and all had clean intermittent catheterization (CIC). Urine culture and sonography were checked every three months and diethylene triamine pentaacetic acid and dimercaptosuccinic acid renal scan every six months to one year, if necessary, during follow-up. Reduction in grade of hydronephrosis or vesicouretral reflux (VUR) was considered as improvement, and absence of disorders on evaluation was considered as cure. All records were evaluated and patients followed by personal visits in clinic or per phone call. Data regarding age, sex, clinical and paraclinical findings, sonography, imaging, renal scan, associated anomalies, treatment and outcomes were collected, entered in SPSS software version18 and analyzed by descriptive statistica. Table 1 Age distribution of our patients Age group Frequency (%) <1 m 8 (24.2%) 1-12 m 18 (54.6%) 1-2 years 5 (15.2%) 2-4 years 2 (6%) Total 33 (100%) Findings

The patients were treated medically with anticholinergic and antibiotics and all had clean intermittent catheterization (CIC). Urine culture and sonography were checked every three months and diethylene triamine pentaacetic acid and dimercaptosuccinic acid renal scan every six months to one year, if necessary, during follow-up. Reduction in grade of hydronephrosis or vesicouretral reflux (VUR) was considered as improvement, and absence of disorders on evaluation was considered as cure. All records were evaluated and patients followed by personal visits in clinic or per phone call. Data regarding age, sex, clinical and paraclinical findings, sonography, imaging, renal scan, associated anomalies, treatment and outcomes were collected, entered in SPSS software version18 and analyzed by descriptive statistica. Table 1 Age distribution of our patients Age group Frequency (%) <1 m 8 (24.2%) 1-12 m 18 (54.6%) 1-2 years 5 (15.2%) 2-4 years 2 (6%) Total 33 (100%) Findings Totally 33 patients aged three days to four years (mean 6.8 months) were included in this study (Table 1). There were 20 (61%) males and 13 (39%) females. Twenty five cases (76%) had UTI. Frequency and severity of hydronephrosis and VUR have been shown in Table 2. Blood urea nitrogen (BUN) in 4 patients (12.1%) and creatinine in 5 cases (15.2%) were normal. Four cases (12.1%) had creatinine more than 2 and 7 cases (21.2%) had BUN more than 30.

males and 13 (39%) females. Twenty five cases (76%) had UTI. Frequency and severity of hydronephrosis and VUR have been shown in Table 2. Blood urea nitrogen (BUN) in 4 patients (12.1%) and creatinine in 5 cases (15.2%) were normal. Four cases (12.1%) had creatinine more than 2 and 7 cases (21.2%) had BUN more than 30. The most common associated anomaly was meningomyelocle in 8 patients (24.2%). Kidney anomaly, bladder anomaly, club foot and ureteropelvic junction obstruction were seen each one in 1 patient and 21 cases (63.6%) didn’t have any anomaly. Urodynamic study was performed in 20 cases (61%), and after three months in neonates without uroflowmetry, which confirmed NGB. Table 2 Types and sites of hydronephrosis and vesicouretral reflux in our study group Complication Mild Moderate Severe Hydronephrosis Bilateral 5 18 4 Unilateral (Rt:4, Lt:2) 3 2 1 Vesicouretral reflux Bilateral 6 9 7 Unilateral(Rt:6, Lt:5) 0 6 5 Rt: Right; Lt: Left All patients received antibiotherapy and CIC, the treatment was succesful in 11 (33.3%) cases and 9 (27.3%) patients improved, but CIC was not successful in 13 cases (39.4%).

Moderate Severe Hydronephrosis Bilateral 5 18 4 Unilateral (Rt:4, Lt:2) 3 2 1 Vesicouretral reflux Bilateral 6 9 7 Unilateral(Rt:6, Lt:5) 0 6 5 Rt: Right; Lt: Left All patients received antibiotherapy and CIC, the treatment was succesful in 11 (33.3%) cases and 9 (27.3%) patients improved, but CIC was not successful in 13 cases (39.4%). Diversion was performed in 22 (67%) cases who did not respond to medical treatment and were in high risk position for diversion. High levels of creatinine and BUN decreased to normal in 90% of patients after medical treatment, CIC and vesicostomy during follow-up. Kidney scan showed scar in 10 patients (30.3%) at follow-up study, 7 patients in left, 2 patients in right kinney and 1 patient bilatral. Complete continence on follow-up was achieved in 24 (71%) patients, and improved in 6 (18%) of cases. 9% (3 cases) died. Mean follow-up period was 3.4±1.2 years (1.5 months to 5 years). Cure rate was 85% in UTI (based on negative u/c), 82.7% in hydronephrosis (based on AP pelvic diameter), 80% in VUR (from grade V and IV to II and I), and 86.5% in kidney function (based on GFR/creatinine).

improved in 6 (18%) of cases. 9% (3 cases) died. Mean follow-up period was 3.4±1.2 years (1.5 months to 5 years). Cure rate was 85% in UTI (based on negative u/c), 82.7% in hydronephrosis (based on AP pelvic diameter), 80% in VUR (from grade V and IV to II and I), and 86.5% in kidney function (based on GFR/creatinine). Discussion The diagnosis and treatment of NLUTD, which is a complex field, needs experience and requires up-to-date knowledge. The aims of treatment in NGB/NLUTD are the preservation of the upper urinary tract, bladder and bowel continence, independence, autonomy, and facilitation of self-estream[4]. After brief physical examination, urodynamic tests, uroflowmetry and ultrasound assessment are needed to clarify fine pathologic causes[5]. In cases with high detrusor pressure, the principal aim of treatment is conversion of high pressure bladder into a low-pressure reservoir. Alpha-blockers have been successful in decreasing bladder-outlet resistance, residual urine and automatic dysfunction[6,7]. Neurogenic bladder pathologies that commonly occur in patients with MMC include an elevated detrusor leak point pressure, VUR, and detrusor-external sphincter dyssynergia[8-10]. McGuire and colleagues[11] first showed increased risk for upper tract dilatation in children with MMC. Shapiro and colleagues[12] published outcomes of a 10-year therapy on 90 children with MMC treated with ileal loop diversion, and showed stable renal units in 69% of the patients. Kasabian and colleagues[13] demonstrated normal renal function in 92% of children with MMC with voiding dysfunction treated with Oxybutynin and CIC. We peformed vicicostomy as a diversion In those patients who did not respond to medical treatment, were in risk position and had severe VUR and hydronephrosis., So we achieved a cure rate of 85% in UTI, 82.7% in hydronephrosis, 80% in VUR, and 86.5% in kidney function[14,15]. After failure of conservative treatment in patients with NGB urinary diversion represents a safe long-term compromise. In Stein et al[16] study, upper urinary tract improved or remained stable in 97% of the renal units in patients with diversion, We performed too vesicostomy protection for urinary tract in some of our patients. Rawashdeh and colleagues[17] in a retrospective study in children younger than 16 years old with NGB who had undergone detrusor myotomy showed a safe and effective alternative for the management of pharmacologically intratable NGBD in children.

ormed too vesicostomy protection for urinary tract in some of our patients. Rawashdeh and colleagues[17] in a retrospective study in children younger than 16 years old with NGB who had undergone detrusor myotomy showed a safe and effective alternative for the management of pharmacologically intratable NGBD in children. Murphy and colleagues[18] reported obtaining total continency or major improvement with conservative care in 91% of 214 cerebral palsy patients with neurogenic bladder. Nue and colleagues[19] evaluated the management of acquired NGB in children using intradetrusor botulinum toxin type A injection and achieved five (62.5%) patients completely dry. Complete continence on follow-up was achieved in 24 (71%) patients, and it was improved in 6 (18%) of cases in our study, but in Rawashdeh et al[17] study it was reported in 8 (73%) patients, and improved only in one case. Stein et al[16] reported 98% complete continence in those with a continent stoma. Murphy et al[18] reported 91% total continence or major improvement with conservative care. Conclusion Although anticholinergic medications and CIC have proved an effective treatment method for many children with NGB dysfunction, it was not effective in all our patients. We believe that permanent vesicostomy is an effective and acceptable surgical intervention for protection of upper urinary tract decompression, especially in those who do not respond to medical treatment and have high risk position. Authors’ Contribution F. Roshanzamir: Acquisition of data, manuscript prepation and surgery.

Conclusion Although anticholinergic medications and CIC have proved an effective treatment method for many children with NGB dysfunction, it was not effective in all our patients. We believe that permanent vesicostomy is an effective and acceptable surgical intervention for protection of upper urinary tract decompression, especially in those who do not respond to medical treatment and have high risk position. Authors’ Contribution F. Roshanzamir: Acquisition of data, manuscript prepation and surgery. M. Rouzrokh: Concept, design, data analysis and surgery. A. Mirshemirani: Design, data analysis and surgery. A. Khaleghnejad: Critical revision of manuscript, surgery and funds collection. L. Mohajerzadeh: Data collection and analysis R. Dalirani: Data collection, analysis and medical management. All authors approved the final version of the manuscript. Acknowledgment This study was financially supported by the office of the Vice Chancellor for Clinical Research Development Center (CRDC) of Mofid Children's Hospital Conflict of Interest: None

To the Editor Fetal gallbladder stone is a rare phenomenon, which is observed by chance during third trimester ultrasonography and does not cause significant clinical symptoms. Even though the etiology of stones found in the fetal period are yet unknown; apnea of prematurity, sepsis, parenteral nutrition, motile diseases, blood group incompatibilities, metabolic diseases and dehydration of newborns are among the causes of formation of gallstones in pediatric age groups. We present a case where the mother had used high dose flaxseed oil capsules, which are widely used as sources of omega-3 and omega-6 during pregnancy. Antenatal ultrasonography determined an echogenic focus beneath the liver position of the fetus. This was found to be gallbladder mud and stone with abdominal ultrasonography on the 2nd day of life. This term (38 weeks and 5 days, 3300 g), appropriate for gestational age, male infant was born to a 33-year-old mother who used linseed (flaxseed) oil capsules 2-3 times (2000-3000 mg) a day during pregnancy for a period of 7 months. During antenatal period, when the fetus had gestanional age of 36 weeks; a hyperechogenic focus was showed beneath the liver position of the fetus. After birth, patient was evaluated and abdominal ultrasonography revealed “gallbladder mud and stone”(Fig .1). Biochemical parameters including liver, function test, total cholesterol and triglycerides were in normal range but total bilirubin and direct bilirubin were 6 mg/dl and 0.6 mg/dl, respectively. Fig. 1 Hyperechogenic foci beneath the liver

This term (38 weeks and 5 days, 3300 g), appropriate for gestational age, male infant was born to a 33-year-old mother who used linseed (flaxseed) oil capsules 2-3 times (2000-3000 mg) a day during pregnancy for a period of 7 months. During antenatal period, when the fetus had gestanional age of 36 weeks; a hyperechogenic focus was showed beneath the liver position of the fetus. After birth, patient was evaluated and abdominal ultrasonography revealed “gallbladder mud and stone”(Fig .1). Biochemical parameters including liver, function test, total cholesterol and triglycerides were in normal range but total bilirubin and direct bilirubin were 6 mg/dl and 0.6 mg/dl, respectively. Fig. 1 Hyperechogenic foci beneath the liver In full blood count, Hemoglobin was 16.7 g/dl, reticulocytes 5.81%, platelet count 245000/mm3 and white blodd cells 14400/mm3. C-reactive protein value and direct Coombs test were negative. A peripheral blood smear examination did not include hemolytic findings. There had been no pyruvate kinase and pyrimidine 5’ nucleotides deficiency of patient whose glucose 6 phosphate dehydrogenase level was 12 U/g HB. Tandem mass spectrometry, urine organic acid and urine blood amino acid tests were normal. Phototherapy was administered for three days due to indirect hyperbilirubinemia. After discharge on the 7th postnatal day, patient was found well-fed and had gained weight. During follow-up, abdominal ultrasonography revealed a normal gallbladder without gallstones at 1 month of age.

In full blood count, Hemoglobin was 16.7 g/dl, reticulocytes 5.81%, platelet count 245000/mm3 and white blodd cells 14400/mm3. C-reactive protein value and direct Coombs test were negative. A peripheral blood smear examination did not include hemolytic findings. There had been no pyruvate kinase and pyrimidine 5’ nucleotides deficiency of patient whose glucose 6 phosphate dehydrogenase level was 12 U/g HB. Tandem mass spectrometry, urine organic acid and urine blood amino acid tests were normal. Phototherapy was administered for three days due to indirect hyperbilirubinemia. After discharge on the 7th postnatal day, patient was found well-fed and had gained weight. During follow-up, abdominal ultrasonography revealed a normal gallbladder without gallstones at 1 month of age. Fetal gallbladder stone was defined for the first time by Beretski and Lankin in 1983[1]. Its prevalence is not known precisely. Its occurrence rate in the literature is low. Agnifili et al reported fetal gallbladder stone incidence as 0.39%[2]. The widest series was reported by Brown and colleagues, who detected echogenic foci in 25 fetus’ gallbladder[3]. None of the many hypotheses suggested can give a possible explanation for fetal gallbladder stone formation. Fanaroff and colleagues represent the idea that bilirubin, a breakdown product of hemoglobin, causes indirect bilirubin level increase by accessing the fetus through the placenta and that this causes fetal gallbladder stone formation[4].

s suggested can give a possible explanation for fetal gallbladder stone formation. Fanaroff and colleagues represent the idea that bilirubin, a breakdown product of hemoglobin, causes indirect bilirubin level increase by accessing the fetus through the placenta and that this causes fetal gallbladder stone formation[4]. It has been asserted that smoking during pregnancy, hematologic diseases, blood incompatibilities between mother and fetus, and structural anomalies like choledochal cysts may cause formation of fetal gallbladder stones[5]. On the other hand, Brown and colleagues proposed that high estrogen levels increase cholesterol secretion and decrease bile acid production and that this mechanism might cause pigment stones in gallbladder [3]. Flaxseed (Linum usitatissimum L., Linaceae) is a vegetable product, which contains high quantity of alpha linolenic acid that can be converted into omega-3. Phytoestrogens are polyphenolic nonsteroidal vegetable origin compounds which structurally and functionally resemble β-estradiol found in mammals[6]. Phytoestrogens have both agonistic and antagonistic effects on estrogen receptors. These behave like endogen estrogens as agonists and trigger estrogenic effects. Flaxseed contains high level phytoestrogen which is in lignan structure (8 mg/g secoisolariciresinol dry weight) which mimics the structure of 17 β-estradiol and synthetic estrogen “diethyl-stilbestrol”. Breakdown products of lignans also have estrogenic activities[6].

rogens as agonists and trigger estrogenic effects. Flaxseed contains high level phytoestrogen which is in lignan structure (8 mg/g secoisolariciresinol dry weight) which mimics the structure of 17 β-estradiol and synthetic estrogen “diethyl-stilbestrol”. Breakdown products of lignans also have estrogenic activities[6]. This is more likely due to high omega-3 content, flaxseed is generally preferred during pregnancy. But there is controversy in consuming flaxseed during pregnancy and lactation. It has been proved that flaxseed would pass from mother to baby during pregnancy and from breast milk following birth. However it is believed that its phytoestrogen content may increase rates of breast, prostate and endometrium cancer, as well as cause truncal obesity, hypertension, anabolic activity increase and stone formation in gallbladder due to its estrogenic activity[7]. For the patient we mentioned, high consumption of flaxseed may have caused stone formation in the gallbladder because of its phytogenic lignan structure which increases estrogenic activity.

Juvenile idiopathic arthritis (JIA) belongs to a group of arthritis with unknown etiology that occurs in children under 16 years old. Pathogenesis of this disease proposes the role of autoimmune process which is induced by antigens and results in inflammation of synovials and cartilage[1]. DiGeorge syndrome (DGS) or velo-cardio-facial-syndrome (VCFS) is a genetic disorder due to a defect in 22q11.2 chromosome[2]. Patients with 22q11.2 DS usually have characteristic facies including retrognathia or micrognathia, long face, downturned mouth, short philtrum low-set, malformed ears and hypertelorism. Congenital heart defects, either a cleft palate or incompetence of the soft palate, and immune deficiencies are common. Patients may have short stature and occasional instances of growth hormone deficiency[3]. Anomalies related to 22q11 monosomy have a wide range[4]. Renal, pulmonary, gastrointestinal, skeletal, and ophthalmologic abnormalities can also occur. Children and adults with 22q11.2DS have high rates of behavioral, psychiatric, and communication disorders. In children, these include attention-deficit/hyperactivity disorder, anxiety, and affective disorders. Adults have a high rate of psychotic disorders, particularly schizophrenia[3]. Parathyroid dysfunction may cause hypocalcemia and seizures in the neonatal period. Most patients with DGS have a partial form of the syndrome and thymic hypoplasia[5].

ention-deficit/hyperactivity disorder, anxiety, and affective disorders. Adults have a high rate of psychotic disorders, particularly schizophrenia[3]. Parathyroid dysfunction may cause hypocalcemia and seizures in the neonatal period. Most patients with DGS have a partial form of the syndrome and thymic hypoplasia[5]. This defect results in cellular immuno-deficiency, although humoral defects have also been described. Autoimmune diseases have been associated with DGS, probably in consequence of T cell regulatory defects and impaired central tolerance[5,6]. These diverse immunoregulatory defects might predispose to the development of autoimmune disease such as hemolytic anemia, thyroiditis and inflammatory arthropathies[5,6]. This study reports association of JIA and DGS in a 12 year old boy. He suffered from common cold like symptoms, overnight fever, weakness, and diffuse pain in limbs since 5 months ago. His right ankle had arthralgia and was swollen. Two weeks later his right wrist and left ankle also showed signs of arthritis. During the past two months he had intermittent fever with more joint involvement and disability which finally led to hospitalization of the patient.

nd diffuse pain in limbs since 5 months ago. His right ankle had arthralgia and was swollen. Two weeks later his right wrist and left ankle also showed signs of arthritis. During the past two months he had intermittent fever with more joint involvement and disability which finally led to hospitalization of the patient. The patient is the first child of family. Because of cleft palate he had a surgery during infancy, and due to cardiac murmur, PDA was detected in echocardiography. He had normal developmental and intellectual process. His 10 year old sister had a palatoplasty when she was 2 years old because of cleft palate. In physical examination, he had T: 38°C, no skin rash and/or petechiae and purpura and ophthalmologic examination was normal. He had micrognathia clearly. A holosystolic murmur was heard over heart. A bilateral axiliary adenopathy in size of 2×1.5×2 cm was removed for pathologic study. Positive Laboratory findings were: WBC: 15900 (Neut: 88%), Hgb: 10.6 g/dL, Plt: 549000, ESR: 70, CRP: 2+, Ca: 8 mg/dl, RF and ANA: negative and normal immunoglobulines. CT scan of thorax as well as sonography of abdomen and pelvis were normal. Radiography of the joints showed osteoporosis and soft tissue swelling. Lymph node biopsy revealed sinus histiocytosis pattern as a reactive adenopathy. Bone marrow aspiration was normal. Final diagnosis was oligoarthritic JIA. With regard to cleft palate, micrognathia, and PDA, DGS was considered and genetic evaluation, Fish test, was done. Results showed elimination in 22q11.2, according to DGS.

Positive Laboratory findings were: WBC: 15900 (Neut: 88%), Hgb: 10.6 g/dL, Plt: 549000, ESR: 70, CRP: 2+, Ca: 8 mg/dl, RF and ANA: negative and normal immunoglobulines. CT scan of thorax as well as sonography of abdomen and pelvis were normal. Radiography of the joints showed osteoporosis and soft tissue swelling. Lymph node biopsy revealed sinus histiocytosis pattern as a reactive adenopathy. Bone marrow aspiration was normal. Final diagnosis was oligoarthritic JIA. With regard to cleft palate, micrognathia, and PDA, DGS was considered and genetic evaluation, Fish test, was done. Results showed elimination in 22q11.2, according to DGS. Onset of JIA may be related to various types of chromosomal and genetic abnormalities with particular autoimmune process[6]. Association of VCFS syndrome with increased predisposition to JIA has been discussed by Rasmussen in three patients with polyarthritis and an evidence of impaired T cell function[7]. Two of the patients with polyarthritis also had IgA deficiency[8]. In a cohort study on patients with 22q11.2 deletion, etiologic relationship between DGS and JIA has been shown. JIA in this syndrome was 50 times more common than in normal population[9].

ith polyarthritis and an evidence of impaired T cell function[7]. Two of the patients with polyarthritis also had IgA deficiency[8]. In a cohort study on patients with 22q11.2 deletion, etiologic relationship between DGS and JIA has been shown. JIA in this syndrome was 50 times more common than in normal population[9]. Davies et al reported 5 new patients and analyzed 8 previously reported patients’ findings with the 22q11 deletion syndrome, who developed chronic inflammatory polyarticular arthritis. The arthritis in all these cases was moderate to severe, but at least partially responsive to methotrexate and/or corticosteroids, and was clinically indistinguishable from JIA. Of particular interest in his study was the high prevalence of IgA deficiency in this association[10]. Chromosome 22q11.2 deletion syndrome is associated with immunodeficiency, especially a mild to moderate deficiency in peripheral blood T cells. However, these patients could have normal laboratory evaluations[11,12]. Thymic hypoplasia or aplasia leading to defective T-cell function is the hallmark of DiGeorge anomaly. Depending on T-cell proliferative response to mitogens, DiGeorge anomaly can be classified as partial or complete. IL-7 may play a critical role in T-cell homeostasis in patients with partial DiGeorge anomaly[13].

2]. Thymic hypoplasia or aplasia leading to defective T-cell function is the hallmark of DiGeorge anomaly. Depending on T-cell proliferative response to mitogens, DiGeorge anomaly can be classified as partial or complete. IL-7 may play a critical role in T-cell homeostasis in patients with partial DiGeorge anomaly[13]. Although DiGeorge anomaly is commonly associated with T-lymphocyte immunodeficiency, B-lymphocyte defects also occur. A recently published review of 1023 patients with DiGeorge anomaly revealed 6% of patients older than 3 years had hypogammaglobulinemia and 3% of patients with DiGeorge anomaly were receiving immunoglobulin replacement therapy[14]. On the degree of T-cell deficiency patients have been divided into partial and complete forms of DGA. Whereas immunodeficiency was seen in the complete form, dysfunction of T-cell regulation was observed in milder forms of DGA, possibly related to abnormal T-suppressor cell function[7]. Peripheral blood may show the normal counts of blood T cell[15, 16]. The JIA disease is an antigen-driven autoimmune process, and despite HLA gene, inflammatory mediators are involved in this disease. It has been shown that the antigen-specific T-cells, especially T helper1 (Th1) are B cells, macrophages and monocytes stimulators, following that, pathogenic immune mediators induce disease process[17]. Immunologic disorders in DGS especially T cell type abnormality with the main role of T cells on JIA immunopathogenesis, could explain association of these two diseases.

Dear Editor, Accidents and incidents are mainly the third cause of death in all ages and are so first cause of death in ages under 40 years in the entire world, while, these are second cause of death after coronary and heart diseases in Iran[1]. Accidents are responsible for the annual death of more than 10,000 people on an international scale and the cause of approximately 10 percent of all children’s admissions to hospitals[2]. In this cross sectional study we studied 33204 patients referred to the health and treatment centers, emergency rooms and hospitals of Isfahan Medical University of Sciences, between 2006 and 2010; and analyzed collected data by χ2 test in SPSS-16 software. From 33204 children under 5 years, 61.8% were males and 38.2% females, 12.67% were under 1 year and 87.33% 1-5 years old. Impact was the most prevalent of the accident types (Table 1). The most frequent places in which accidents occurred were: in the house (66.37%), and on the streets (21.25%) and they occurred least frequently in working places (0.59%). Drowning had the highest death rate (35.48 %) followed by automobile accident (1.12%). This study indicates that different variables are responsible in accident occurrence and the relationship between variables such as gender, age, location and the result of the accident with the accident type is quite obvious. As mentioned before, the relationship between some of the variables are shown from a statistical viewpoint. Boys had approximately twice as many accidents as girls.

ent occurrence and the relationship between variables such as gender, age, location and the result of the accident with the accident type is quite obvious. As mentioned before, the relationship between some of the variables are shown from a statistical viewpoint. Boys had approximately twice as many accidents as girls. Also accident occurrence was much higher in urban than in rural areas, may be because of higher population in the city. The largest amount of accidents resulted in recovering of the patients after treatment, a small number (almost 0.03%) were disabled and 0.27% died. Each year averagely 6640 accidents involve children less than 5 years of age which mostly could be prevented. To prevent accidents it is necessary to constantly educate individuals, families, schools and society to avoid encountering dangers related to injuries. Also it is essential to engage in aiding and emergencies, to teach first aids and basics in all levels in accidents which have more risk of death. Table 1 Accident types in children less than 5 years old Accident < 1 year 1-5 years n (%) n (%) Motorcycle rder 136 (0.41) 1432 (4.31) Automobile driver 280 (0.84) 1592 (4.79) Impact 1383 (4.17) 11630 (35.03) Falling from heights 1102 (3.32) 7176 (21.61) Violence 9 (0.03) 121 (0.36) Burning 627 (1.89) 1445 (4.35) Pedestrian 101 (0.30) 1863 (5.61) Snake and scorpion bite 10 (0.03) 53 (0.16) Animal bite 6 (0.02) 214 (0.64) Poisoning 232 (0.70) 978 (2.95) Suicide 4 (0.01) 24 (0.07) Electrocution 9 (0.03) 46 (0.14) Drowning 2 (0.01) 29 (0.09) Other 306 (0.92) 2394 (7.21) Total 4207 (12.67) 28997(87.33)

Trichorhinophalangeal syndrome (TRPS), as the name suggests is a rare genetic disorder which affects the tricho (hair), rhino (nose), phalanges (digits)[1] and has been classified into three types. Trichorhinophalangeal syndrome type II (TRPS2) is also known as Langer-Giedion syndrome (LGS). It was first described by Andreas Giedion, a Swiss pediatric radiologist and Leonard O Langer Jr, an American radiologist. TRPS2 combines features of trichorhinophalangeal syndrome type I (TRPS1) and multiple exostoses[2]. It is characterized by sparse hair, multiple cone shaped epiphyses, multiple cartilaginous exostoses, bulbous nasal tip, thickened alar cartilage, upturned nares, prominent philtrum, large protruding ears and mild mental retardation[3]. Multiple cartilaginous exostoses distinguishes TRPS2 from TRPS1. Exostoses are multiple projections of bone capped by cartilage, mostly seen in the metaphyses as well as diaphyses of long bones. Flat bones, vertebrae and the ribs may also be affected. As the bone continues to grow, the exostoses appear to migrate towards the diaphysis. At puberty as the growth plate fuses, the linear growth ceases and no new exostoses develop[4]. There are no trigger factors for development of exostoses.

aphyses of long bones. Flat bones, vertebrae and the ribs may also be affected. As the bone continues to grow, the exostoses appear to migrate towards the diaphysis. At puberty as the growth plate fuses, the linear growth ceases and no new exostoses develop[4]. There are no trigger factors for development of exostoses. Microdeletions involve loss of small chromosome regions, the largest of which are detectable only with prophase chromosome studies or molecular methods. When such a deletion involves more than a single gene, the condition is referred to as a contiguous gene deletion syndrome[5]. TRPS2 is a true contiguous gene deletion syndrome with deletions in both TRPS1 and EXT1 genes on chromosome 8q24.1-q24.13. A 7 yr old female child born of a non-consanguineous marriage presented to a tertiary care hospital with failure to thrive and multiple cartilaginous exostoses. Mother had two first trimester abortions and two live healthy children. Child was immunized till date and was developmentally normal. On examination, her weight and height were below the 3rd percentile for age. Facial dysmorphism was noted which included microcephaly, sparse hair, bushy eyebrows, hypertelorism, long philtrum, micrognathia, high arched palate, poor dentition, low set ears, deformed ear cartilage and auricular sinus (Fig. 1). Limb deformities in the form of clinodactyly and overlapping of toes were observed. Multiple cartilaginous exostoses were noted over the ribs, elbows, wrists, knees and back (Fig. 2). Systemic examination revealed no abnormality.

palate, poor dentition, low set ears, deformed ear cartilage and auricular sinus (Fig. 1). Limb deformities in the form of clinodactyly and overlapping of toes were observed. Multiple cartilaginous exostoses were noted over the ribs, elbows, wrists, knees and back (Fig. 2). Systemic examination revealed no abnormality. Her intelligence quotient evaluation was within normal range. Radiograph of the hands revealed delayed bone age, cone shaped epiphysis and exostoses bilaterally (Fig. 3). Chromosomal analysis in the form of karyotyping was done which was normal. Fig. 1 Classical dysmorphic features showing hypertelorism, bushy eyebrows, low set ears, large philtrum, prognathism, upturned nares Fig. 2 Exotoses over the scapulae Fig. 3 Radiograph of the wrist joint showing exostoses crests These patients need a long-term follow-up throughout their life. Currently, our patient is 9 years old with normal intelligence and without any complications due to exostoses. The child is being followed-up monthly as these patients are known to develop complications such as Perthes disease, osteomas causing cervical nerve compression, growth hormone deficiency, infertility and malignant transformation[6].

s 9 years old with normal intelligence and without any complications due to exostoses. The child is being followed-up monthly as these patients are known to develop complications such as Perthes disease, osteomas causing cervical nerve compression, growth hormone deficiency, infertility and malignant transformation[6]. Differential diagnosis includes Trichorhino-phalangeal syndrome type 1 which has similar dysmorphic features to Langer-Giedion syndrome but without exostoses. Others include metachondromatosis in which the exostoses are primarily seen in the hands and feet along with enchondromata in the ends of long bones and iliac crests. Most of the metachondromatoses regress spontaneously[7]. The other differential diagnosis is 11p11 deletion syndrome (OMIM 601224) which is also characterised by the presence of multiple exostoses but is differentiated from Langer-Giedion syndrome by the presence of cutaneous syndactyly, skull abnormalities like brachycephaly, turricephaly, enlarged parietal foramina and craniofacial dysostosis. Treatment in Langer-Giedion syndrome is usually supportive but in case of complications due to exostoses such as pain, limited range of joint movement, pressure on nerves, blood vessels, the spinal cord, and tissues, surgical intervention is necessary. To conclude, our case is an unusual one, as the child presented as a classical LGS phenotype with multiple exostoses and typical dysmorphic features but without a gene deletion in the TRPS1 and EXT1 genes. (A similar case was presented by Pereza et al, 2012)[8]

Introduction Considering the high mortality in intensive care units (ICUs) in hospitals compared with other units as well as high costs of inpatient treatment in these units, mortality prediction has long been a concern[1]. Several tools are designed for mortality prediction in ICUs[2]. One of the most widely used tools for examining patients’ consciousness level and disease outcome prediction is Glasgow Coma Scale (GCS)[3,4]. This scale was first developed in 1974 to evaluate the consciousness level of head injury patients[5], and then was widely used for evaluating the consciousness level of other patients admitted to ICU[6]. Several studies have indicated that GCS provides the guideline for primary care and disease outcome prediction (mortality and morbidity)[7-9]. Because of the failure of GCS in examining the verbal responses of intubated patients and evaluating brainstem reflexes, several other scales have become popular for assessment of intubated patients’ consciousness level and disease outcome prediction during the past decade. However, none of the other scales have been used widely[3,10-12]. During recent years, many efforts to improve CGS have been made so that it can be used more easily. One of these tools is the Full Outline of Unresponsiveness (FOUR) score that was designed by Wijdicks et al in 2005[12]. This scale includes four considerable components: eye responses, motor responses, brainstem reflexes, and breathing pattern. Each component receives a score between 0 and 4 (the lowest and highest scores are 0 and 4, respectively)[13]. Several studies have investigated the validity of the FOUR score and suggest that it is a good alternative for GCS in disease outcome prediction[14-17].

or responses, brainstem reflexes, and breathing pattern. Each component receives a score between 0 and 4 (the lowest and highest scores are 0 and 4, respectively)[13]. Several studies have investigated the validity of the FOUR score and suggest that it is a good alternative for GCS in disease outcome prediction[14-17]. The results of the research by Cohen on 60 children admitted to an ICU in California (2009) indicated that the FOUR score is a powerful tool in disease outcome prediction for pediatric patients admitted to ICU and the inter-rater reliability for the FOUR score was excellent[14]. Because a large percentage of patients admitted to ICU are comatose[18], their examination is an important part of work in the ICU, and the most widely used tool for assessing patients’ level of consciousness and predicting disease outcome is GCS. Because of the weaknesses of GCS and its failure in assessing verbal responses in intubated patients, the brainstem reflexes and also the strengths of the FOUR score in brainstem reflex assessment, we decided to compare the ability of GCS and FOUR score in predicting the mortality and discharge of patients admitted to pediatric ICU (PICU).

sses of GCS and its failure in assessing verbal responses in intubated patients, the brainstem reflexes and also the strengths of the FOUR score in brainstem reflex assessment, we decided to compare the ability of GCS and FOUR score in predicting the mortality and discharge of patients admitted to pediatric ICU (PICU). Subjects and Methods This prospective study was conducted in the PICU of Ali Ibn Abitalib Hospital, Zahedan. The Children and Adolescents Health Research Center of Zahedan University of Medical Sciences approved the study. Convenience purposive sampling was used. Written consent was obtained from family members of patients, and they were assured that patient’s personal information would be safe and would be used only for research and they could withdraw from the study any time. Sample size was calculated at 200 according to the sample size formula. Inclusion criteria were all children with neurological or neurosurgery disorders admitted to PICU of the Hospital. Exclusion criteria included patients receiving sedating drugs and neuromuscular blockers including midazolam, fentanyl, sufentanil, morphine, pancuronium bromide, atracurium, nesdonal, and propofol, or had recognized vision, hearing, speech, or limb paralysis problems. In addition, patients under the age of two years and above 12 years (because of an inability to communicate verbally ill patients less than 2 years and because of lack of PICU admission in patients over 12 years) were excluded. Data collection lasted from February to November 2012. Data was collected using the FOUR score and GCS using questionnaires. The patients’ level of consciousness was routinely controlled by nurses using GCS after entering the PICU and recorded in a special flowchart for consciousness level measurement. To measure the patients’ level of consciousness using the FOUR score, it was primarily translated to Persian and then back-translated to English, and the accordance of English versions were examined by an individual fluent in both languages. Content validity index (CVI) was used for measuring the validity of the data collection tool. Ten faculty members of the department of neurology and neurosurgery were provided with the tool, and their comments and corrections were applied. The new coma scale (FOUR score) was taught to nurses participating in this study by the specialty pediatric neurology during three 30–45 min sessions on each item (Table 1)[16].

faculty members of the department of neurology and neurosurgery were provided with the tool, and their comments and corrections were applied. The new coma scale (FOUR score) was taught to nurses participating in this study by the specialty pediatric neurology during three 30–45 min sessions on each item (Table 1)[16]. And each participant was given an instruction booklet regarding the FOUR score. And again after a week of personal training, clinical training in the PICU was given by a pediatric neurologist. At the end of the course, each of the nurses participating in the study were allowed to practice on 2–3 patients and all the problems were resolved in relation to working with this scale. Sixteen nurses participated in the study. All of these 16 nurses had bachelor’s degree in nursing. Table 1 Full Outline of Unresponsiveness (FOUR) score[16]

And each participant was given an instruction booklet regarding the FOUR score. And again after a week of personal training, clinical training in the PICU was given by a pediatric neurologist. At the end of the course, each of the nurses participating in the study were allowed to practice on 2–3 patients and all the problems were resolved in relation to working with this scale. Sixteen nurses participated in the study. All of these 16 nurses had bachelor’s degree in nursing. Table 1 Full Outline of Unresponsiveness (FOUR) score[16] Nurses had different working shifts (morning, evening, and night) and different working experience including recruiting, contract, and formal nurses, and the average work experience was 8.31±7.14 years. To assess inter-rater reliability of the FOUR score, each patient was rated on the FOUR score by two differently trained nurses. The raters performed their examination on arrival of the patient to the PICU without knowledge of the other rater’s scores. To study the predictive ability of mortality and discharge rate of both scales, scores of the FOUR score were compared to those of GCS, which were routinely controlled by nurses and recorded in the special flowchart for measuring GCS scores. For patients who had undergone intubation, the lowest GCS verbal score was used both for scoring and for data analysis. Ultimately, both tools were compared regarding their predictability of patient mortality or discharge. Afterwards, obtained data was analyzed using SPSS 16.

special flowchart for measuring GCS scores. For patients who had undergone intubation, the lowest GCS verbal score was used both for scoring and for data analysis. Ultimately, both tools were compared regarding their predictability of patient mortality or discharge. Afterwards, obtained data was analyzed using SPSS 16. Findings Of the 200 patients that participated in this research, 55% (n=110) were males and 45% (n=90) females. The mean age of patients was 4.4 years. Of the 200 patients, 76% (n=152) had spontaneous respiration and 24% (n=48) were ventilated with a mechanical ventilator. The cause of patients’ admission to ICU was mostly intracranial hemorrhage. The admission diagnoses of patients are listed in Table 2. Of the 200 patients who participated in this study, 143 (71.5%) patients were discharged after recovery and 57 (28.5%) patients died in ICU. According to the results of the independent t test, patients’ age did not affect the outcome (discharge or death) (P=0.5). Also, results of the chi square test did not show any differences with regard to the outcome and patients’ sex (P=0.5). The inter-rater reliability of the FOUR score was evaluated using the weighted kappa (κw) coefficient. Table 2 Admission diagnosis of patients Diagnosis Number (Percent) Intracranial hemorrhage 36 (18) Intracranial infection 31 (15.5) Hydrocephaly 29 (14.5) Aneurism 28 (14) Seizure 27 (13.5) Brain tumor 22 (11)

Findings Of the 200 patients that participated in this research, 55% (n=110) were males and 45% (n=90) females. The mean age of patients was 4.4 years. Of the 200 patients, 76% (n=152) had spontaneous respiration and 24% (n=48) were ventilated with a mechanical ventilator. The cause of patients’ admission to ICU was mostly intracranial hemorrhage. The admission diagnoses of patients are listed in Table 2. Of the 200 patients who participated in this study, 143 (71.5%) patients were discharged after recovery and 57 (28.5%) patients died in ICU. According to the results of the independent t test, patients’ age did not affect the outcome (discharge or death) (P=0.5). Also, results of the chi square test did not show any differences with regard to the outcome and patients’ sex (P=0.5). The inter-rater reliability of the FOUR score was evaluated using the weighted kappa (κw) coefficient. Table 2 Admission diagnosis of patients Diagnosis Number (Percent) Intracranial hemorrhage 36 (18) Intracranial infection 31 (15.5) Hydrocephaly 29 (14.5) Aneurism 28 (14) Seizure 27 (13.5) Brain tumor 22 (11) Other causes 27 (13.5) A κw statistic of ≤0.4 is considered poor, values between 0.4 and 0.6 are considered fair to moderate, those between 0.6 and 0.8 suggest good inter-observer agreement, and values greater than 0.8 suggest excellent agreement. The rater agreement is shown in Table 3. The inter-rater reliability for the FOUR score was good to excellent (weighted κ: eye, 0.72; respiration, 0.82; brainstem, 0.74; motor, 0.78).

r to moderate, those between 0.6 and 0.8 suggest good inter-observer agreement, and values greater than 0.8 suggest excellent agreement. The rater agreement is shown in Table 3. The inter-rater reliability for the FOUR score was good to excellent (weighted κ: eye, 0.72; respiration, 0.82; brainstem, 0.74; motor, 0.78). The mean score of the FOUR and GCS at the time of ICU admission for all patients was 10.5±4.1 (range: 0–16) and 10.4±3.9 (range: 3–15), respectively. Mean of the FOUR score at the time of admission was 12.5±2.1 and 5.1±2.8 for discharged and dead patients, respectively (cut-off point 8) (Table 4). The differences between the two groups were statistically significant (P=0.001). The mean GCS at the time of admission was 11.4±3.5 and 7.9±3.8 for discharged and dead patients, respectively (cut-off point 9; P=0.001). Logistic regression analysis was performed to determine the ability of the two scales (GCS and FOUR score) to predict the outcome. Results of this test showed that odds ratios for the FOUR score are somewhat lower than those for the GCS (FOUR score=OR: 0.13; 95%CI: 0.06–0.29; P<0.001; GCS=OR: 2.49; 95%CI: 1.44–4.32; P<0.001). In previous studies, lower odds ratios have been related to a positive predictive value for a higher chance of a positive outcome with increased total score values[13,16].

the FOUR score are somewhat lower than those for the GCS (FOUR score=OR: 0.13; 95%CI: 0.06–0.29; P<0.001; GCS=OR: 2.49; 95%CI: 1.44–4.32; P<0.001). In previous studies, lower odds ratios have been related to a positive predictive value for a higher chance of a positive outcome with increased total score values[13,16]. Discussion The purpose of establishing a PICU is to obtain the best results and better outcomes for severely ill children. One of the ways to achieve that goal is to predict the mortality risk of the patients admitted to the PICU to provide them with the best care available[20]. It is necessary to develop models that predict the mortality risk in PICU to monitor the effectiveness of the care carried out[21]. For this purpose, the neurological examination tools or coma examination scales of patients are accepted as effective scales for disease outcome examination[2]. To be an effective tool, a coma scale must be practical for use in a wide variety of settings and by healthcare providers with diverse experience[14]. In this regard, the FOUR score is designed to remedy the deficiencies of GCS to show more neurological details in unconscious patients and predict the final result more accurately and easily[4,16]. Research results indicated that the inter-rater agreement with the FOUR score was good to excellent (weighted κ: eye, 0.72; respiration, 0.82; brainstem, 0.74; motor, 0.78). These results are consistent with those by Wolf et al[11] and Wijdicks et al[16]. The high level of agreement between nurse raters using the FOUR score suggests that the application of the FOUR score and assessment of the level of consciousness is easier and requires minimal facilities, and nurses with differing levels of experience and expertise are more likely to correctly assess the patient and assign the same score using the FOUR score.

raters using the FOUR score suggests that the application of the FOUR score and assessment of the level of consciousness is easier and requires minimal facilities, and nurses with differing levels of experience and expertise are more likely to correctly assess the patient and assign the same score using the FOUR score. Table 3 Kappa values, Standard Error and 95% Confidence Intervals for Inter-rater agreement on the Full Outline of Unresponsiveness score Eye Motor Brainstem Respiration Kappa 0.72 0.78 0.74 0.82 Standard Error 0.037 0.035 0.039 0.032 Confidence Intervals (CI) 0.67-0.77 0.73-0.84 0.69-0.80 0.77-0.87 Table 4 Mean score of FOUR coma sub score in discharged and deceased patients FOUR coma sub scale Number Mean P. value Eye opening Discharged 143 2.7 (0.97) 0.001 Deceased 57 0.73 (0.76) Motor Discharged 143 3.2 (0.82) 0.001 Deceased 57 1.6 (1.01) Brainstem Discharged 143 3.4 (0.7) 0.001 Deceased 57 1.6 (0.88) Respiration Discharged 143 3.1 (0.74) 0.001 Deceased 57 1.1 (0.88) Total score Discharged 143 12.5 (2.1) 0.001 Deceased 57 5.1 (2.8) FOUR: Full Outline of Unresponsiveness Although the GCS has been widely used in hospital settings, because of the failure in examining the verbal responses of intubated patients and evaluating brainstem reflexes, the FOUR score was developed. By these advantages, the FOUR score can show patients’ real state of consciousness. Therefore, it is better at predicting patients’ future state[3,14].

ed in hospital settings, because of the failure in examining the verbal responses of intubated patients and evaluating brainstem reflexes, the FOUR score was developed. By these advantages, the FOUR score can show patients’ real state of consciousness. Therefore, it is better at predicting patients’ future state[3,14]. Our results demonstrate that mortality in PICU patients with the lowest FOUR score is higher than in patients with the lowest GCS. The mortality rate for patients with the lowest FOUR score of 0 (100%) was higher than that for patients with the lowest GCS score of 3 (85.7%). With this finding, the FOUR score would have great value for outcomes prediction than the GCS. These results are consistent with those by Cohen[14], Wijdicks et al[16], and Iyer et al[15]. In the research by Büyükcam et al in Turkey, no significant difference was observed between these tools for predicting the mortality of children admitted to the ICU[19]. This difference is probably because of the fact that the participants of the research by Büyükcam et al were only children with a medical diagnosis of stroke, but in the present research a group of children with different medical neurology and neurosurgery diagnoses were investigated. This new coma scale, unlike the GCS, does not include a verbal response, and thus is more valuable in PICU that typically has a large number of intubated patients. In our study, 24% of patients were intubated, and GCS was less useful for verbal response.

Our results demonstrate that mortality in PICU patients with the lowest FOUR score is higher than in patients with the lowest GCS. The mortality rate for patients with the lowest FOUR score of 0 (100%) was higher than that for patients with the lowest GCS score of 3 (85.7%). With this finding, the FOUR score would have great value for outcomes prediction than the GCS. These results are consistent with those by Cohen[14], Wijdicks et al[16], and Iyer et al[15]. In the research by Büyükcam et al in Turkey, no significant difference was observed between these tools for predicting the mortality of children admitted to the ICU[19]. This difference is probably because of the fact that the participants of the research by Büyükcam et al were only children with a medical diagnosis of stroke, but in the present research a group of children with different medical neurology and neurosurgery diagnoses were investigated. This new coma scale, unlike the GCS, does not include a verbal response, and thus is more valuable in PICU that typically has a large number of intubated patients. In our study, 24% of patients were intubated, and GCS was less useful for verbal response. Research results indicated that cut off point 8 correlated with worse outcome, while the research by Wijdicks et al[16] indicated that a cut-off point of 9 and that by Akavipat et al[2] a cut-off point of 10 correlated with worse outcome,. This difference may be due to deterioration of the patients’ health participating in the study.

s indicated that cut off point 8 correlated with worse outcome, while the research by Wijdicks et al[16] indicated that a cut-off point of 9 and that by Akavipat et al[2] a cut-off point of 10 correlated with worse outcome,. This difference may be due to deterioration of the patients’ health participating in the study. A limitation of this study was that the population in this study included only patients with neurological problems and the results of this study cannot be extended to all patients admitted to PICU. Conclusion It is important to assess the consciousness level of patients admitted to PICU using an accurate, easy-to-use tool that is better at showing disease outcome. With respect to the results, the FOUR score is more capable than GCS in assessing patients’ level of consciousness and disease outcome predictability. Acknowledgment The authors wish to express their profound gratitude to the research council of the Research and Technology Department of Zahedan’s University of Medical Sciences for approving the research topic as well as the nursing staff working in the PICU for their collaboration in the research. Conflict of Interest: None Authors’ Contribution H. Askari, A. Khajeh: Conception and design, acquisition of data, analysis and interpretation of data A. Fayyazi, G. Miri-Aliabad, N.M. Noori: Critical Revision of the Manuscript, Funds Collection B Khajeh: Drafting of the manuscript All Authors approved final version of the manuscript.

Introduction Dermatoglyphics is the science of fingerprints and palm prints analysis which are constant throughout life[1-3]. It is believed that they may represent different genetically determined congenital abnormalities. Several genes are involved in the inheritance of the dermal traits[1]. Dermal ridges form in 6th week of gestation and reach their maximum size between 12th and 13th week. Historically, dermatoglyphics was a classical model to consider the polygenic inheritance. Therefore, some specific dermatoglyphic patterns may be accompanied with different genetic abnormalities[1,3]. Recent studies on dermatoglyphics have shown especial patterns in various congenital abnormalities such as skeletal maturation, diabetes type 1, schizophrenia, albinism, cleft lip and palate, rheumatoid arthritis, congenital spinal cord anomalies and Klinefelter syndrome[4-10]. Cystic Fibrosis (CF) is a common fatal, autosomal recessive disease in Caucasians[11,12]. CF is a progressive disease that involves exocrine glands, lungs, gastrointestinal system, pancreas, liver, kidneys and reproductive system[3,12,13]. CF prevalence varies from 1 to 2500 among Caucasians[13]; 10 million carriers of the recessive gene exist in the world[14]. CF can be diagnosed antenatally using genetic tests and in early childhood with sweat test[15]. The main idea of this paper was to determine the differences in dermatoglyphic patterns in CF and normal children.

Cystic Fibrosis (CF) is a common fatal, autosomal recessive disease in Caucasians[11,12]. CF is a progressive disease that involves exocrine glands, lungs, gastrointestinal system, pancreas, liver, kidneys and reproductive system[3,12,13]. CF prevalence varies from 1 to 2500 among Caucasians[13]; 10 million carriers of the recessive gene exist in the world[14]. CF can be diagnosed antenatally using genetic tests and in early childhood with sweat test[15]. The main idea of this paper was to determine the differences in dermatoglyphic patterns in CF and normal children. Subjects and Methods This is a case-control study performed between October 2012 and March 2013. The study group included 46 children with CF, confirmed by two positive sweat tests, referred to CF clinic in Dr. Sheikh Pediatric Hospital, Mashhad University of Medical Sciences, Iran. The control group consisted of 341 healthy participants. Exclusion criteria included children less than one year of age, individuals with current atopic dermatitis and any finger or palm abnormalities which interfered with the procedure. All subjects were informed about the process and consent forms were obtained.

ran. The control group consisted of 341 healthy participants. Exclusion criteria included children less than one year of age, individuals with current atopic dermatitis and any finger or palm abnormalities which interfered with the procedure. All subjects were informed about the process and consent forms were obtained. We used red powder blush to roll finger and palm separately. The powder was rubbed on palm and fingers of both hands by means of a brush. A piece of adhesive tape was placed on a rolling plate and the participant pressed his/her palm on the tape in distal to proximal direction. It was then transformed onto a labeled paper indicating right and left side. The same process was carried out for fingerprints as each finger was pressed separately to the adhesive tape. Fingers were named with roman numbers from I (thumb) to V (little finger). Fingerprint pattern types were categorized into three groups, arch (A), loop (L) and whorl (W). Whorl was divided into two subgroups named simple whorl (W) and double whorl (2W) (Fig. 1). Patterns of homolog fingers in right and left hand were given 0 or 1 score due to their similarity or dissimilarity. Finally, these scores were added and final scores ranged 0 to 5[2]. After the center point of each fingerprint was joined to triradius (i.e. the conjunction point of three opposing ridge systems), TRC was calculated by summing the total numbers of ridges between the two centers[16].

Fingerprint pattern types were categorized into three groups, arch (A), loop (L) and whorl (W). Whorl was divided into two subgroups named simple whorl (W) and double whorl (2W) (Fig. 1). Patterns of homolog fingers in right and left hand were given 0 or 1 score due to their similarity or dissimilarity. Finally, these scores were added and final scores ranged 0 to 5[2]. After the center point of each fingerprint was joined to triradius (i.e. the conjunction point of three opposing ridge systems), TRC was calculated by summing the total numbers of ridges between the two centers[16]. Total ridge count of arch pattern is always zero because it has no triradius. Loop pattern has one triradius, but whorl pattern has two triradii. The higher calculated number of lines was reported for whorl pattern TRC (Fig. 1). On the following step, we found the triradii (a, b, c, d) at the base of each finger (Fig. 2). a-b line was formed by joining a to b triradius. Fig. 1 Palm patterns are labeled a, b, c, d and t a: triradius "a" at the base of the 2nd finger b: triradius "b" at the base of the 3rd finger c: triradius "c" at the base of the 4th finger d: triradius "d" at the base of the 5th finger t: triradius "t" at the proximal part of the palm Fig. 2 Fingerprint pattern types are categorized into three groups: arch (A,B), loop (F,G) and whorl. Whorls are divided into two subgroups named simple whorl (C) and double whorl (D). c: center point of fingerprint, t: triradius The ridges on the a-b line in both hands were calculated with magnifying glass.

t: triradius "t" at the proximal part of the palm Fig. 2 Fingerprint pattern types are categorized into three groups: arch (A,B), loop (F,G) and whorl. Whorls are divided into two subgroups named simple whorl (C) and double whorl (D). c: center point of fingerprint, t: triradius The ridges on the a-b line in both hands were calculated with magnifying glass. After detection of triradius a (at the base of 2nd finger on palm print), t (on the proximal of palm) and d (at the base of 5th finger on the palm), atd angle was drawn and measured by protractor in both hands. All the data was collected twice by one person and the mean of these two numbers was finally reported. We used chi-square test for comparing frequency of pattern types, Fisher's exact test for comparing the fingerprint pattern types and the disease phenotype and Mann-Whitney test to compare a-b ridge count, TRC, atd angle and asymmetry of right and left hand between control and study groups. P-values less than 0.05 were considered statistically significant. Findings A total of 387 participants, 46 CF and 341 healthy children were enrolled in this study. Table 1 shows the baseline characteristics of studied group. Number and percentage of disease phenotype(failure to thrive, steatorrhea, clubbing, liver disease, other gastrointestinal symptoms and respiratory symptoms) and dermatoglyphic pattern types of the studied group are shown in Table 2. There was no significant difference in dermatoglyphic pattern types and phenotype. Table 1 Baseline characteristics of the studied groups Variable Mean (SD) Variable n (%)

Number and percentage of disease phenotype(failure to thrive, steatorrhea, clubbing, liver disease, other gastrointestinal symptoms and respiratory symptoms) and dermatoglyphic pattern types of the studied group are shown in Table 2. There was no significant difference in dermatoglyphic pattern types and phenotype. Table 1 Baseline characteristics of the studied groups Variable Mean (SD) Variable n (%) Age (month) 105.9 (43.0) Relative parents 30 (85.7) Weight 22.49 (8.8) FTT 16 (57.1) Height (cm) 116.4 (20.0) Steatorrhea 13 (46.4) HC a (cm) 50.98 (1.9) Clubbing 1 (3.6) MAC b (cm) 16.4 (2.3) Respiratory symptoms c 15 (53.6) Sweat test Cl 101.7 (38.8) Other gastrointestinal symptoms d 17 (60.7) Na 104.7 (47.6) Age of onset (month) 3.42 (6.29) Liver disease 2 (7.1) Age at diagnosis (month) 11.8 (17.2) a :Head Circumference b : Mid arm circumference c : Other gastrointestinal symptoms include vomiting, GER, rectal prolapse, chronic diarrhea d : Respiratory symptoms include pneumonia, cough, wheezing, dyspnea Table 2 Comparing the number and percentage of disease phenotype and dermatoglyphic pattern types in CF patients Digit Pattern type FTT Steatorrhea Clubbing Liver disease Other gastrointestinal symptoms a Respiratory symptoms b Right hand n (%) Left hand n (%) Right hand n (%) Left hand n (%) Right hand n (%) Left hand n (%) Right hand n (%) Left hand n (%) Right hand n (%) Left hand n( %) Right hand n (%) Left hand n (%) I Loop 9 (60) 9 (75) 8(53.3) 6 (50) 0(0) 1(100) 1(6.7) 0(0) 10(66.7) 5(41.7) 5(33.3) 5 (41.7) Arch ___ 0 (0) ___ 0 (0) __ 0(0) __ 0(0) __ 0(0) __ 1 (100) Whorl 7 (53.8) 7 (46.7) 5 (38.5) 7 (46.7) 1(100) 0(0) 1(7.7) 2(13.3) 7(53.8) 12(80) 10(76.9) 9 (60)

n (%) Right hand n (%) Left hand n (%) Right hand n (%) Left hand n( %) Right hand n (%) Left hand n (%) I Loop 9 (60) 9 (75) 8(53.3) 6 (50) 0(0) 1(100) 1(6.7) 0(0) 10(66.7) 5(41.7) 5(33.3) 5 (41.7) Arch ___ 0 (0) ___ 0 (0) __ 0(0) __ 0(0) __ 0(0) __ 1 (100) Whorl 7 (53.8) 7 (46.7) 5 (38.5) 7 (46.7) 1(100) 0(0) 1(7.7) 2(13.3) 7(53.8) 12(80) 10(76.9) 9 (60) P. value 0.7 0.17 0.43 1 0.46 0.46 1 0.52 0.7 0.05 0.05 0.4 II Loop 5 (50) 8 (57.1) 4 (40) 7 (50) 0(0) 1(7.1) 1(10) 1(7.1) 6(60) 9(64.3) 6(60) 6 (42.9) Arch 2 (100) 2 (66.7) 1 (50) 1(33.3) 0(0) 0(0) 0(0) 0(0) 0(0) 0(0) 1(50) 2 (66.7) Whorl 9 (56.3) 6 (54.5) 8 (50) 5(45.5) 1(6.3) 0(0) 1(6.3) 1(9.1) 11(68.8) 8(72.7) 8(50) 7 (63.6) P. value 0.5 1 0.84 1 1 1 1 1 0.05 0.8 0.5 III Loop 12 (63.2) 9 (52.9) 8 (42.1) 6(35.3) 1(100) 1(100) 2(10.5) 2(11.8) 12(63.2) 10(58.8) 9(47.4) 7 (41.2) Arch 2 (100) 3 (75.0) 1 (50) 2(50) 0(0) 0(0) 0(0) 0(0) 0(0) 1(25) 1(50) 3 (75) Whorl 2 (28.6) 4 (57.1) 4 (57.1) 5(71.4) 0(0) 0(0) 0(0) 0(0) 5(71.4) 6(85.7) 5(71.4) 5 (71.4) P. value 0.13 0.8 0.82 0.31 1 1 0.64 0.68 0.2 0.16 0.6 0.2 IV Loop 8 (61.5) 7 (58.3) 6 (46.2) 4(33.3) 1(7.7) 1(8.3) 1(7.7) 1(8.3) 7(53.8) 7(58.3) 8(61.5) 7 (58.3) Arch 1 (100) 1 (100) 0 (0) 0 (0) 0(0) 0(0) 0(0) 0(0) 0(0) 0(0) 0(0) 0 (0) Whorl 7 (50) 8 (53.3) 7 (50) 9 (60) 0(0) 0(0) 1(7.1) 1(6.7) 10(71.4) 10(66.7) 7(50) 8 (53.3) P. value 0.8 1 1 0.25 0.5 0.4 1 1 0.2 0.42 0.5 0.8 V Loop 9 (50) 8 (50 6 (33.3) 6(37.5) 1(100) 1(6.3) 2(11.1) 1(6.3) 11(61.1) 9(56.3) 10(55.6) 11 (68.8) Arch ___ 1 (100) ___ 0(0) __ 0(0) __ 0(0) __ 0(0) __ 0 (0) Whorl 7 (70) 7 (70) 7 (70) 7(70) 0(0) 0(0) 0(0) 1(10) 6(60) 7(70) 5(10) 4 (40)

Whorl 7 (50) 8 (53.3) 7 (50) 9 (60) 0(0) 0(0) 1(7.1) 1(6.7) 10(71.4) 10(66.7) 7(50) 8 (53.3) P. value 0.8 1 1 0.25 0.5 0.4 1 1 0.2 0.42 0.5 0.8 V Loop 9 (50) 8 (50 6 (33.3) 6(37.5) 1(100) 1(6.3) 2(11.1) 1(6.3) 11(61.1) 9(56.3) 10(55.6) 11 (68.8) Arch ___ 1 (100) ___ 0(0) __ 0(0) __ 0(0) __ 0(0) __ 0 (0) Whorl 7 (70) 7 (70) 7 (70) 7(70) 0(0) 0(0) 0(0) 1(10) 6(60) 7(70) 5(10) 4 (40) P. value 0.4 0.53 0.11 0.16 1 1 0.52 1 1 0.4 1 0.1 a : other gastrointestinal symptoms include vomiting, GER, rectal, Chronic diarrhea b : respiratory symptoms include Pneumonia, Cough, wheezing, dyspnea Table 3 Number and percentage of dermatoglyphic patterns in CF patients and control group Digit pattern type Right hand Left hand CF group n (%) Control n (%) Total (%) CF group n (%) Control n (%) Total (%) I Loop 25 (54.3) 162 (48.1) 187 (48.8) 24 (52.2) 173 (52.0) 197 (52.0) Arch 2 (4.3) 8 (2.4) 10 (2.6) 2 (4.3) 17 (5.1) 19 (5.0) Whorl 19 (41.3) 167 (49.6) 186 (48.6) 20 (43.5) 143 (42.9) 163 (43.0) P . value 0.5 1 II Loop 21 (45.7) 162 (48.1) 183 (47.8) 24 (52.2) 159 (47.7) 183 (48.3) Arch 5 (10.9) 29 (8.6) 34 (8.9) 6 (13.0) 28 (8.4) 34 (9.0) Whorl 20 (43.5) 146 (43.3) 166 (43.3) 16 (34.8) 146 (43.8) 162 (42.7) P. value 0.9 0.4 III Loop 32 (69.6) 222 (65.9) 254 (66.3) 29 (63.0) 212 (63.5) 241 (63.4) Arch 4 (8.7) 24 (7.1) 28 (7.3) 6 (13.0) 27 (8.1) 33 (8.7) Whorl 10 (21.7) 91 (27.0) 101 (26.4) 11 (23.9) 95 (28.4) 106 (27.9) P . value 0.7 0.5 IV Loop 22 (47.8) 139 (41.6) 161 (42.4) 20 (43.5) 149 (45.2) 169 (44.9) Arch 1 (2.2) 9 (2.7) 10 (2.6) 3 (6.5) 15 (4.5) 18 (4.8) Whorl 23 (50.0) 186 (55.7) 209 (55.0) 23 (50.0) 166 (50.3) 189 (50.3) P . value 0.8 0.9 V

Arch 4 (8.7) 24 (7.1) 28 (7.3) 6 (13.0) 27 (8.1) 33 (8.7) Whorl 10 (21.7) 91 (27.0) 101 (26.4) 11 (23.9) 95 (28.4) 106 (27.9) P . value 0.7 0.5 IV Loop 22 (47.8) 139 (41.6) 161 (42.4) 20 (43.5) 149 (45.2) 169 (44.9) Arch 1 (2.2) 9 (2.7) 10 (2.6) 3 (6.5) 15 (4.5) 18 (4.8) Whorl 23 (50.0) 186 (55.7) 209 (55.0) 23 (50.0) 166 (50.3) 189 (50.3) P . value 0.8 0.9 V Loop 30 (65.2) 246 (74.3) 276 (73.2) 29 (64.4) 251 (77.2) 280 (75.7) Arch 0 (0) 5 (1.5) 5 (1.3) 1 (2.2) 6 (1.8) 7 (1.9) Whorl 16 (34.8) 80 (24.2) 96 (25.5) 15 (33.3) 68 (20.9) 83 (22.4) P. value 0.1 0.2 Table 3 shows number and percentage of fingerprint pattern types in case and control groups. There was no significant difference in fingerprint patterns of both groups. Table 4 represents mean TRC in case and control groups. Significant differences in mean TRC between case and control groups were found in right digit IV (P=0.009), left digit III (P=0.02), left digit IV (P=0.03) and left digit V (P=0.03). The TRC asymmetry for all digits is shown in Table 5. There was no significant difference between both groups regarding TRC asymmetry. According to Table 6, significant differences were found in right hand atd angle (P=0.001), left hand atd angle (P=0.002), right hand a-b ridge (P=0.007) and left hand a-b ridge (P=0.001) between case and control group. No significant differences were found in atd angle asymmetry, a-b ridge count asymmetry or pattern dissimilarity score between both groups (Table 7). P-values less than 0.05 were considered statistically significant. Table 4 Mean TRC of CF patients and control group Digit CF patients Mean Rank Control group Mean Rank

According to Table 6, significant differences were found in right hand atd angle (P=0.001), left hand atd angle (P=0.002), right hand a-b ridge (P=0.007) and left hand a-b ridge (P=0.001) between case and control group. No significant differences were found in atd angle asymmetry, a-b ridge count asymmetry or pattern dissimilarity score between both groups (Table 7). P-values less than 0.05 were considered statistically significant. Table 4 Mean TRC of CF patients and control group Digit CF patients Mean Rank Control group Mean Rank P . value I R 156.85 197.35 0.2 L 181.49 191.74 0.5 II R 156.43 196.30 0.2 L 181.49 191.74 0.5 III R 167.84 194.15 0.1 L 153.87 195.54 0.02 IV R 150.29 195.48 0.009 L 155.51 192.54 0.03 V R 168.20 191.88 0.2 L 152.44 188.30 0.03 Table 5 Total ridge count asymmetry for five digits in CF patients and control group Digit CF Patients Mean (SD) Control group Mean (SD) P. value I 193.49 190.09 0.8 II 206.29 187.17 0.3 III 178.92 190.40 0.5 IV 175.41 188.63 0.4 V 174.02 183.10 0.6 CF: Cystic Fibrosis; SD: Standard deviation Discussion In this study, we observed significant differences in dermatoglyphic patterns including the mean TRC of the right digit IV, left digit III, left digit IV, left digit V and atd angle and a-b ridge in right and left hands of children with CF compared to control group.

V 174.02 183.10 0.6 CF: Cystic Fibrosis; SD: Standard deviation Discussion In this study, we observed significant differences in dermatoglyphic patterns including the mean TRC of the right digit IV, left digit III, left digit IV, left digit V and atd angle and a-b ridge in right and left hands of children with CF compared to control group. Kobylisky et al reported significant differences in fingerprint pattern types [17]. They showed that arches fingerprint patterns were higher in CF females in contrast to higher loop patterns in CF males. Our results did not establish significant differences considering these patterns in both groups. Based on Weizman et al, whorl patterns were more frequent than loop patterns as opposed to control group in celiac patients[18,19]. In another study, whorl pattern values were also higher in celiac children and there was a correlation between dermatoglyphic patterns of celiac patients and their parents[21]. Table 6 Mean (SD) for atd angles, a-b ridge counts in CF cases and control group Group CF cases Control group P -value atd – R 234.19 179.94 0.001** atd – L 231.91 178.48 0.002* a-b-R 148.75 194.59 0.007* a-b-L 137.59 194.87 0.001* SD: Standard deviation; CF: Cystic Fibrosis

Kobylisky et al reported significant differences in fingerprint pattern types [17]. They showed that arches fingerprint patterns were higher in CF females in contrast to higher loop patterns in CF males. Our results did not establish significant differences considering these patterns in both groups. Based on Weizman et al, whorl patterns were more frequent than loop patterns as opposed to control group in celiac patients[18,19]. In another study, whorl pattern values were also higher in celiac children and there was a correlation between dermatoglyphic patterns of celiac patients and their parents[21]. Table 6 Mean (SD) for atd angles, a-b ridge counts in CF cases and control group Group CF cases Control group P -value atd – R 234.19 179.94 0.001** atd – L 231.91 178.48 0.002* a-b-R 148.75 194.59 0.007* a-b-L 137.59 194.87 0.001* SD: Standard deviation; CF: Cystic Fibrosis In 1986, Gottlieb et al observed that the arch pattern values were significantly higher in congenital syndrome of early onset constipation and abdominal pain[20]. Mathew et al found increased loop patterns in children with oral cleft[1]. In this study, we observed significant differences in the mean TRC of the right digit IV, left digit III, left digit IV, left digit V of CF cases compared to the control group. Eslami et al observed significant differences in the mean TRC of the right digit IV, right digit V and left digit II in patients with cleft lip (CLP) compared to control group[3].

In 1986, Gottlieb et al observed that the arch pattern values were significantly higher in congenital syndrome of early onset constipation and abdominal pain[20]. Mathew et al found increased loop patterns in children with oral cleft[1]. In this study, we observed significant differences in the mean TRC of the right digit IV, left digit III, left digit IV, left digit V of CF cases compared to the control group. Eslami et al observed significant differences in the mean TRC of the right digit IV, right digit V and left digit II in patients with cleft lip (CLP) compared to control group[3]. This finding was in contrast to Kobylisky et al study[17] which reported no significant difference in the mean TRC. Kobylisky et al also showed that mean TRC values were lower in the CF group. Their results are in line with ours[17]. In 2002, Neiswanger et al observed no significant differences in TRC asymmetry in patients with cleft lip who had a negative family history[21]. Table 7 Mean atd angle asymmetry, a-b ridge count asymmetry and pattern dissimilarity score in Cystic Fibrosis cases and control group Group CF cases Control group P -value atd asymmetry 196.80 180.53 0.333 a-b asymmetry 183.54 186.34 0.8700 Pattern dissimilarity score 192.93 182.75 0.529 CF: Cystic Fibrosis Their results were similar to those of our study. Comparing the a-b ridge count, we found that a-b ridge count values were lower in CF patients. It was similar to Kobylisky et al study[17]. Rezaeinezhad et al showed that mean a-b ridge count values in patients with type 1 diabetes mellitus were higher in control group[10].

Their results were similar to those of our study. Comparing the a-b ridge count, we found that a-b ridge count values were lower in CF patients. It was similar to Kobylisky et al study[17]. Rezaeinezhad et al showed that mean a-b ridge count values in patients with type 1 diabetes mellitus were higher in control group[10]. In 1973, Taussing et al observed increased atd angle in CF children, which was in agreement with our results[22]. On the contrary, Kobylisky et al reported that the value of atd angles were significantly lower in CF females and significantly higher in CF males[17]. According to the study of Mathew et al, atd angles showed an increase in children with oral cleft[1]. Esalmi et al observed that atd angles were not significantly different in CLP patients and control group. In 2013, Eslami et al observed no significant differences in a-b ridge asymmetry and pattern dissimilarity score in CLP patients, which was in agreement with our results [3]. This study had its limitations. The limitations were confined to the small number of patients that may impose a negative effect on the final results. The authors recommend that further researches should be done in parents of CF children to assess the child and parent dermatoglyphic traits relation. Moreover, evaluating the distribution of fingerprint minutiae and palmar sweat glands in CF children would provide more comprehensive information of dermatoglyphic patterns in CF children.

end that further researches should be done in parents of CF children to assess the child and parent dermatoglyphic traits relation. Moreover, evaluating the distribution of fingerprint minutiae and palmar sweat glands in CF children would provide more comprehensive information of dermatoglyphic patterns in CF children. Conclusion Dermatoglyphic characteristics were significantly different in CF children and control group. These traits could be used as a supplementary diagnostic method in CF children. Acknowledgment We would like to thank Fatemeh Chaji for her invaluable assistance. This study was supported by a grant from the Vice Chancellor for Research of the Mashhad University of Medical Sciences with the approval number of 910821. Conflict of Interest: None Authors’ Contribution A. Ezzati: Concept, Acquisition of Data, Data Analysis, Manuscript Preparation, Critical Revision of the Manuscript F. Batoeh: Prepared the design of the study and drafted and Approval of the Article S.A. Jafari: Performed the study, acquired the data M.A. Kiani : performed the study, acquired the data N. Mahdavi-Shahri: Performed the study, acquired the data H. Ahanchian: Performed the study, acquired the data S. Tehraniam: Manuscript Preparation, Critical Revision of the Manuscript A. Jahanbin: Concept, acquired the data and Critical Revision of the Manuscript H.R. Kianifar: Concept, prepared design of the manuscript, data analysis, and interpretation All authors approved final version of the paper.

Introduction Birth asphyxia is a leading cause of mortality and morbidity in neonates in developing countries, with an incidence of 100-250/1000 live births compared to 5-10/1000 live births in the developed world[1]. It remains a significant cause of loss of life and adverse developmental outcome[2]. The major causes of neonatal deaths globally were estimated to be infections (35%), preterm births (28%) and birth asphyxia (23%)[3]. As large number of deliveries in the developing world takes place at home, there is no reliable data to precisely estimate the disease burden in countries like Pakistan. However, this figure is likely to be very high as two thirds of world’s neonatal mortality occurs in 10 developing countries[4]. In Pakistan, over 5 million children are born each year. Of them 0.45 million die before first birthday and nearly half of these deaths occur during the neonatal period[5]. In 2001, birth asphyxia was responsible for 35%, 14% and 11% of neonatal mortality in Lahore, Karachi and Khyber Pukhtunkhwa respectively[6].

s[4]. In Pakistan, over 5 million children are born each year. Of them 0.45 million die before first birthday and nearly half of these deaths occur during the neonatal period[5]. In 2001, birth asphyxia was responsible for 35%, 14% and 11% of neonatal mortality in Lahore, Karachi and Khyber Pukhtunkhwa respectively[6]. Various risk factors are associated with birth asphyxia. Though the topic has been extensively studied and reviewed worldwide, limited local data is available; Common intrapartum risk factors in a local data include non cephalic presentation, prolonged rupture of membranes, meconium staining, maternal anemia, vaginal bleeding, maternal fever at time of delivery, mode of delivery after prolonged second stage of labor like normal vertex delivery, cesarean section, and multiple births[5,6]. It is estimated that for every 1 death due to birth asphyxia 4 infants survive with long-term sequel[6]. The American Academy of Pediatrics and a task force on cerebral Palsy has suggested criterion to define birth asphyxia[7]: (1) Profound metabolic or mixed metabolic acidemia of pH <7.00 in an umbilical artery blood sample taken at birth. (2) Persistence of an Apgar score of 0-3 for ≥5 minutes. (3) Neonatal neurologic manifestations e.g. seizures, coma or hypotonia following an asphyxia insult. (4) Evidence of multi-organ involvement e.g. cardiovascular, gastrointestinal, or renal compromise.

(1) Profound metabolic or mixed metabolic acidemia of pH <7.00 in an umbilical artery blood sample taken at birth. (2) Persistence of an Apgar score of 0-3 for ≥5 minutes. (3) Neonatal neurologic manifestations e.g. seizures, coma or hypotonia following an asphyxia insult. (4) Evidence of multi-organ involvement e.g. cardiovascular, gastrointestinal, or renal compromise. There is nothing more tragic for a normally developed fetus to sustain cerebral anoxia during the last hours of perinatal life due to perinatal risk factors thus resulting in death or if escaped, has to live with major handicap. This study was planned with the aim to find out the leading perinatal risk factors in term babies causing birth asphyxia in our setup. Subjects and Methods This descriptive–cross sectional study was carried out at the neonatal intensive care unit (NICU) of Combined Military Hospital, Multan Pakistan, a tertiary care hospital from 1st December 2012 to 1st December 2013. Total of 196 asphyxiated cases that fulfilled the inclusion and exclusion criteria were included in the study by taking p 4.8% and d 3%[1] through consecutive non-probability sampling technique. Inclusion Criteria: • Neonates suffering from perinatal asphyxia diagnosed in the presence of at least 2 of the following factors, and admitted to neonatal intensive care unit within 6 hours of birth. • First cry delayed for 5 minutes. • Apgar score at 5 minutes of age <5 and didn’t improve to more than 7/10 at 20 minutes of age. • Post asphyxial seizures within first 48 hours after birth. Exclusion Criteria:

• Neonates suffering from perinatal asphyxia diagnosed in the presence of at least 2 of the following factors, and admitted to neonatal intensive care unit within 6 hours of birth. • First cry delayed for 5 minutes. • Apgar score at 5 minutes of age <5 and didn’t improve to more than 7/10 at 20 minutes of age. • Post asphyxial seizures within first 48 hours after birth. Exclusion Criteria: • Neonates with major congenital malformations of central nervous system, cardiovascular system, respiratory system or dysmorphic babies. • babies delivered by lower segment cesarean section (LSCS) to mothers who were given general anesthesia • Term intra-uterine growth retardation (IUGR) babies with birth weight less than 1.5 kg. • Preterm deliveries. • Other causes of central nervous system encephalopathy (infectious, metabolic). • All outdoor deliveries.

• Neonates with major congenital malformations of central nervous system, cardiovascular system, respiratory system or dysmorphic babies. • babies delivered by lower segment cesarean section (LSCS) to mothers who were given general anesthesia • Term intra-uterine growth retardation (IUGR) babies with birth weight less than 1.5 kg. • Preterm deliveries. • Other causes of central nervous system encephalopathy (infectious, metabolic). • All outdoor deliveries. Approval from hospital ethical committee was sought. Informed written consent was taken from all the patients’ parents participating in this study. All neonates admitted to neonatal intensive care unit (NICU) and fulfilling the inclusion criteria were taken as subjects of the study. Every baby was assigned a serial number. Detailed history was taken from mother and birth attendant (gynaecologist/ gynae nurse) on a predesigned questionnaire regarding selected perinatal factors (mode of delivery (spontaneous vertex delivery, cesarean section, instrumental delivery), abnormal presentation, rupture of membranes more than 18 hours, prolonged second stage of labor more than 30 minutes, maternal fever and anemia at the time of delivery, and meconium staining). Babies were examined in detail for the signs of birth asphyxia (tone, posture, reflexes, seizures etc). Data obtained was analyzed using SPSS version 15.0. Descriptive statistics were used to calculate mean, and standard deviation was used for gestational age, parity, maternal age, gravidity, and weight of the newborns. Frequencies and percentages were calculated for variables like mode of delivery, instrumental delivery, maternal complications like fever and anemia at delivery, prolonged rupture of membranes, meconium staining and multiple births. Stratification with respect to maternal age, gestational age, and weight of newborns, parity and gravidity was done and post stratification chi-square test was applied. P-value <0.05 was taken as significant.

tions like fever and anemia at delivery, prolonged rupture of membranes, meconium staining and multiple births. Stratification with respect to maternal age, gestational age, and weight of newborns, parity and gravidity was done and post stratification chi-square test was applied. P-value <0.05 was taken as significant. Findings A total of 196 cases fulfilling the inclusion/exclusion criteria were enrolled in the study, 125 (64%) were males and 71 (36%) females. Maternal age of the patients showed 54.59% (n=107) between 18-27 years and 45.41% (n=89) between 28-37 years, mean and standard deviation was calculated as 27.04 (±4.97) years. Majority (52.55%) of babies (n=103) were born between 37-39 weeks while 47.45 % (n=93) between 40-41 weeks of gestation, mean and standard deviation was calculated as 39.19 (+1.24) weeks. Majority (57.14 %, n=112) of mothers were between 1-3 and ≥4 paras and were recorded in 42.86 % (n=84) cases, mean and standard deviation 2.87 (+1.12). Most of the mothers were between 1-3 gravida, i.e. 64.80% (n=127) while 35.20 % (n=69) had ≥4 gravidities, mean and standard deviation was 3.45 (+0.87). Mean weight of the newborns was calculated as 2621.37 (+74.21) grams.

and ≥4 paras and were recorded in 42.86 % (n=84) cases, mean and standard deviation 2.87 (+1.12). Most of the mothers were between 1-3 gravida, i.e. 64.80% (n=127) while 35.20 % (n=69) had ≥4 gravidities, mean and standard deviation was 3.45 (+0.87). Mean weight of the newborns was calculated as 2621.37 (+74.21) grams. Frequency of perinatal factors leading to birth asphyxia among term newborns were evaluated; prolonged second stage of labor reported in 141 (72%) cases, mode of delivery was spontaneous vertex delivery in 44.39% (n=87), cesarean section in 32.14% (n=63), 23.47% (n= 46) instrumental delivery. Fifty-seven patients (29.08%) had prolonged rupture of membranes (PROM), 7.65% (n=15) had meconium staining, 5.61% (n=11) had multiple births, 21.94% (n=43) had maternal fever, 58.84% (n=113) had anemia at delivery. Stratification with regards to maternal age was recorded which shows anemia at delivery was significantly higher in 28-37 years of age, P value was calculated as 0.0001 while rest of risk factors were insignificant (Table 1). Stratification with regards to gestational age was recorded which shows that each factor was significantly higher in gestational age between 40-41 weeks of gestation as compared to 37-39 weeks (Table 2). In Table 3, stratification with regards to parity was computed where instrumental delivery, prolonged rupture of membranes and maternal fever was insignificant in both groups while others i.e. cesarean section, spontaneous vertex delivery, meconium staining and anemia at delivery had significant difference between 1-3 and ≥4 paras.

tratification with regards to parity was computed where instrumental delivery, prolonged rupture of membranes and maternal fever was insignificant in both groups while others i.e. cesarean section, spontaneous vertex delivery, meconium staining and anemia at delivery had significant difference between 1-3 and ≥4 paras. Discussion This study was planned to determine the various perinatal risk factors associated with birth asphyxia in term newborns in neonatal intensive care unit. As 23% of all neonatal deaths are attributable to birth asphyxia[2], it is important to be aware of factors that may predispose a newborn to a hypoxic insult at birth with the aim of formulating preventive strategies. Table 1 Stratification for maternal age (n=196) Factors Maternal age (years), n (%) P. value 18-27 years 28-37 years Instrumental delivery (n=46) 21 (45.65%) 25 (54.35%) 0.2 Spontaneous vertex delivery (n=87) 49 (56.32%) 38 (43.68%) 0.7 Cesarean section (n=63) 34 (53.97%) 29 (46.03%) 0.9 Prolonged rupture of membranes (n=57) 26 (45.61%) 31 (54.39%) 0.1 Meconium staining (n=15) 6 (40%) 9 (60%) 0.2 Maternal Fever (n=43) 26 (60.47%) 27 (62.79%) 0.3 Anemia at delivery (n=113) 45 (40.46%) 68 (59.54%) 0.0001 Table 2 Stratification for gestational age (n=196) Factors Gestational age (years), n (%) P. value 37-39 weeks (n=103) 40-41 weeks (n=93) Instrumental delivery (n=46) 15 (32.61) 31 (67.39) 0.002 Spontaneous vertex delivery (n=87) 33 (37.93) 54 (62.07) 0.001 Cesarean section (n=63) 43 (68.25) 20 (31.75) 0.002 Prolonged rupture of membranes (n=57) 19 (33.33) 38 (66.67) 0.001 Meconium staining (n=15) 3 (20) 12 (80) 0.008

Gestational age (years), n (%) P. value 37-39 weeks (n=103) 40-41 weeks (n=93) Instrumental delivery (n=46) 15 (32.61) 31 (67.39) 0.002 Spontaneous vertex delivery (n=87) 33 (37.93) 54 (62.07) 0.001 Cesarean section (n=63) 43 (68.25) 20 (31.75) 0.002 Prolonged rupture of membranes (n=57) 19 (33.33) 38 (66.67) 0.001 Meconium staining (n=15) 3 (20) 12 (80) 0.008 Maternal fever (n=43) 12 (27.91) 31 (72.09) 0.001 Anemia at delivery (n=113) 33 (28.24) 80 (71.76) 0.001 These factors may be antepartum in 50% of cases, intrapartum in 40% and postpartum in remaining 10%. Given the reduced availability of skilled care during delivery in developing countries like Pakistan, intrapartum causes may have greater contribution[6]. In this study full term neonates (>1.5kg and >37 weeks of gestation) of hospital deliveries admitted in the neonatal intensive care unit were included, male to female distribution was 3:2. Out of 196 cases 64% were males, Ibrahim[13] et al in their study identified weight of more than 2.5 kg and full term male neonates as risk factors leading to birth asphyxia in an analysis of 235 cases of birth asphyxia, as in our study.

e neonatal intensive care unit were included, male to female distribution was 3:2. Out of 196 cases 64% were males, Ibrahim[13] et al in their study identified weight of more than 2.5 kg and full term male neonates as risk factors leading to birth asphyxia in an analysis of 235 cases of birth asphyxia, as in our study. Prolonged second stage of labor (>30 mins) remained the most important determinant of birth asphyxia as in other studies as well as mode of delivery[8,12], Spontaneous vertex delivery (SVD) was the most common mode of delivery in our study associated with birth asphyxia in 87 (44.39%), followed by emergency cesarean section (CS) in 63 (32.14%), and instrumental delivery (included both forceps and vacuum delivery) in 46 (23.47%). This is similar to other studies like Chishty[12] et al where proportion of birth asphyxia was predominantly greater in spontaneous vertex delivery (SVD) in in-hospital cases, but Zulfiqar[14] et al in their study described 66% of cases associated with birth asphyxia delivered by cesarean section and 34% with spontaneous vertex delivery in in-hospital study.

ishty[12] et al where proportion of birth asphyxia was predominantly greater in spontaneous vertex delivery (SVD) in in-hospital cases, but Zulfiqar[14] et al in their study described 66% of cases associated with birth asphyxia delivered by cesarean section and 34% with spontaneous vertex delivery in in-hospital study. We had only 3 cases of birth asphyxia followed by elective cesarean section, which is significantly lower number. Different studies also describe this inverse association between elective cesarean section and birth asphyxia5. In recent years, the rate of cesarean section has risen to a record level of 46% in China and to levels of 25% and above in many Asian, European and Latin American countries[9]. The rate has increased significantly in the United States, to 33 percent of all births in 2011, up from 21 percent in 1996, and in the rate in 2009 varied widely between hospitals (ranging from 6.9% to 69.9% of births)[10]. Higher number of CS in in-cases shows skills of detecting perinatal asphyxia in a tertiary care hospital and going for early intervention. Table 3 Stratification for parity (n=196) Factors Parity n (%) P -value 1-3 (n=112) ≥4 (n=84) Instrumental delivery (n=46) 11 (23.91) 35 (76.09) 0.001 Spontaneous vertex delivery (n=87) 38 (43.68) 49 (56.32) 0.001 Cesarean section (n=63) 41 (65.08) 22 (34.92) 0.122 Prolonged rupture of membranes (n=57) 33 (57.89) 24 (42.11) 0.892 Meconium staining (n=15) 11 (73.33) 4 (26.67) 0.147 Maternal Fever(n=43) 19 (44.19) 24 (55.81) 0.052

Instrumental delivery (n=46) 11 (23.91) 35 (76.09) 0.001 Spontaneous vertex delivery (n=87) 38 (43.68) 49 (56.32) 0.001 Cesarean section (n=63) 41 (65.08) 22 (34.92) 0.122 Prolonged rupture of membranes (n=57) 33 (57.89) 24 (42.11) 0.892 Meconium staining (n=15) 11 (73.33) 4 (26.67) 0.147 Maternal Fever(n=43) 19 (44.19) 24 (55.81) 0.052 Anemia at delivery(n=113) 45 (38.17) 68 (61.83) 0.00 Emergency CS has also been identified as a risk factor having significant influence on birth asphyxia. Studies done by Milsom[15] and Seyal[16] noted similar association, that deliveries by emergency cesarean section and use of instruments have direct association with birth asphyxia. Maternal age at delivery did not appear to be significant in our study as 54% of mothers were between 18 to 27 years of age and 46% were between 28 to 37 years of age. This was similar to the results reported from Sweden15. But maternal age more than 35 years is a risk factor associated with birth asphyxia in many studies. A possible explanation could be as vast majority of our patient group came from a low income, illiterate segment of population, many women were unaware of their exact age and the age stated was an approximation with a tendency towards younger age. In our study 112 (57%) cases of birth asphyxia occurred in para 1-3, whereas 84 (43%) in para more than 4, a study by Sayel[16] described similar relation.

Maternal age at delivery did not appear to be significant in our study as 54% of mothers were between 18 to 27 years of age and 46% were between 28 to 37 years of age. This was similar to the results reported from Sweden15. But maternal age more than 35 years is a risk factor associated with birth asphyxia in many studies. A possible explanation could be as vast majority of our patient group came from a low income, illiterate segment of population, many women were unaware of their exact age and the age stated was an approximation with a tendency towards younger age. In our study 112 (57%) cases of birth asphyxia occurred in para 1-3, whereas 84 (43%) in para more than 4, a study by Sayel[16] described similar relation. In our study, PROM (>18 hours) was seen in 29% and pyrexia during delivery (>100 Fahrenheit) was seen in 22%. Both these factors are implicated with birth asphyxia as seen by Majeed[5] et al in their study “risk factors of birth asphyxia” where they found relation of 24% and 20% respectively for prolonged rupture of membranes and maternal pyrexia. Meconium aspiration was seen in 7.6% cases and multiple births in 5.6% which is comparable to a local study[5] where 9.6% and 4.8% of both these factors respectively are described.

In our study, PROM (>18 hours) was seen in 29% and pyrexia during delivery (>100 Fahrenheit) was seen in 22%. Both these factors are implicated with birth asphyxia as seen by Majeed[5] et al in their study “risk factors of birth asphyxia” where they found relation of 24% and 20% respectively for prolonged rupture of membranes and maternal pyrexia. Meconium aspiration was seen in 7.6% cases and multiple births in 5.6% which is comparable to a local study[5] where 9.6% and 4.8% of both these factors respectively are described. Gestational age of neonates did not appear to be significant risk factor in this study, as we excluded all preterm births less than 37 weeks and were only interested for the perinatal risk factors associated with birth asphyxia in term neonates. However, premature infants are reported in literature as being more prone to ischemic injuries of the white matter. These babies are more likely to have several other potentially fatal problems compared to term infants, since we only included term neonates, that could be explanation that no relation could be found. Maternal anemia at time of delivery was seen in 58% cases of birth asphyxia, maternal anemia is a significant risk factor for asphyxia, probably because of intrapartum hypoxia, and this observation is similar to Majeed et al[5] where 60% of mothers were found anemic at time of delivery.

Gestational age of neonates did not appear to be significant risk factor in this study, as we excluded all preterm births less than 37 weeks and were only interested for the perinatal risk factors associated with birth asphyxia in term neonates. However, premature infants are reported in literature as being more prone to ischemic injuries of the white matter. These babies are more likely to have several other potentially fatal problems compared to term infants, since we only included term neonates, that could be explanation that no relation could be found. Maternal anemia at time of delivery was seen in 58% cases of birth asphyxia, maternal anemia is a significant risk factor for asphyxia, probably because of intrapartum hypoxia, and this observation is similar to Majeed et al[5] where 60% of mothers were found anemic at time of delivery. Prolonged second stage of labor and birth asphyxia have strong correlation, reported in 72% of cases, improving awareness with easy access to a health services at delivery may play a role in reducing the incidence of birth asphyxia, as an average Pakistani woman is small sized therefore the chances of cephalo pelvic disproportion and prolonged labor leading to birth asphyxia is a possibility. Chishty[12] et al noted similar relation.

ness with easy access to a health services at delivery may play a role in reducing the incidence of birth asphyxia, as an average Pakistani woman is small sized therefore the chances of cephalo pelvic disproportion and prolonged labor leading to birth asphyxia is a possibility. Chishty[12] et al noted similar relation. A study in India showed that traditional birth attendants were able to recognize signs of asphyxia, but were unable to deal with it[11]. Training program targeting at traditional birth attendants may be of benefit in decreasing birth asphyxia in our country where only 31% of deliveries are attended by skilled health personnel. There are certain limitations of this study as it is hospital based study on term neonates where majority of births were attended by qualified personnel, this doesn’t reflect exact epidemiology and associated risk factors prevalent in the community, where more than 70% of births are attended by traditional birth attendants. Strong point of the study is that it highlighted major pitfalls in our set up or any tertiary care hospital for instance and there is always room for improvement and emphasis must be on the importance of development of preventive strategies to reduce the burden of birth asphyxia. That will only be possible by imparting health education, better perinatal and obstetric care facilities at a tertiary care hospital. However, observations made in this study can help in planning larger population-based studies to confirm and target the risk factors of perinatal asphyxia.

the burden of birth asphyxia. That will only be possible by imparting health education, better perinatal and obstetric care facilities at a tertiary care hospital. However, observations made in this study can help in planning larger population-based studies to confirm and target the risk factors of perinatal asphyxia. Conclusion There is nothing more tragic for a normally developed fetus then to sustain cerebral anoxia during the last hours of perinatal life. This results in death in most cases and if escaped, have to live with a major handicap or lifelong disability. Since it is a preventable problem and long term neurological sequels are almost untreatable once asphyxia set in, there is no denying the fact that preventive strategies must be built to reduce the burden of birth asphyxia. Early identification of high-risk cases with improved antenatal and perinatal care can decrease such high mortality. We need to bring major reforms in our health delivery system. A properly trained person in neonatal resuscitation, preferably a pediatrician, should be available during high risk deliveries. Neonatal life support courses should be made mandatory for personnel involved in newborn deliveries and care. Early antenatal recognition and practice of referral to tertiary care hospital of high risk pregnancies will give better outcomes. Acknowledgment This study was part of a post graduate dissertation of Dr. Asad Nauman Kiyani. Conflict of Interest: None Authors’ Contribution A. Khushdil: Concept / design, critical revision of the manuscript A.N. Kiyani: Acquisition of data, manuscript preparation

We need to bring major reforms in our health delivery system. A properly trained person in neonatal resuscitation, preferably a pediatrician, should be available during high risk deliveries. Neonatal life support courses should be made mandatory for personnel involved in newborn deliveries and care. Early antenatal recognition and practice of referral to tertiary care hospital of high risk pregnancies will give better outcomes. Acknowledgment This study was part of a post graduate dissertation of Dr. Asad Nauman Kiyani. Conflict of Interest: None Authors’ Contribution A. Khushdil: Concept / design, critical revision of the manuscript A.N. Kiyani: Acquisition of data, manuscript preparation A. Ehsan: Acquisition of data, data interpretation A. Ullah: Acquisition of data, data interpretation M.A. Khan: Concept / design, manuscript preparation All authors approved final version of the paper.

Introduction Low birth weight (LBW) is defined as a birth weight of less than 2500 g and is a major public health problem[1] as well as a major risk factor for neonatal and postnatal morbidity[2]. Based on WHO statistics the rate of LBW is 17% worldwide (6% in industrialized countries and 21% in developing countries)[3]. Vazirinejad et al found that at a public-sector referral hospital in Iran during a six month period, 9.6% of neonates were LBW[4]. As incidences are substantially higher, the magnitude of the problem is even larger in developing countries[5]. LBW neonates are categorized according to birth weight as follows: 1) moderately low birth weight (MLBW): between 1500 - 2499 g; 2) very low birth weight (VLBW): less than 1500 g; and extremely low birth weight (ELBW) less than 1000 g[6]. The latter groups are at special risk for developmental defects[7], but MLBW infants may suffer from these problems as well[8]. As the population of MLBW infants is 5 times larger than that of smaller infants[8], assessment and early detection of motor deficits in order to referring them for interventional program, can lead to the reduction of the later developmental problems and associated costs[9].

s may suffer from these problems as well[8]. As the population of MLBW infants is 5 times larger than that of smaller infants[8], assessment and early detection of motor deficits in order to referring them for interventional program, can lead to the reduction of the later developmental problems and associated costs[9]. Recent studies regarding developmental status of LBWs have received limited attention as to the importance of the development of MLBWs[8]. Huddy et al found that these children were at low risk for later neurodevelopmental problems but are prone to academic problems[10]. Middle et al found that MLBW children have higher rates of academic performance and educational problems[11]. Eglan et al and Breslau et al have reported increased rates of learning and behavioral problems among these children[12,13]. Some of these studies do not include a normal birth weight (NBW) control group for comparison. In this study, we evaluated the motor developmental status of a group of MLBW preterm infants at the corrected age of 18±2 months via the Peabody Developmental Motor Scale II (PDMS-2) test and compared the results with that of NBW infants.

Recent studies regarding developmental status of LBWs have received limited attention as to the importance of the development of MLBWs[8]. Huddy et al found that these children were at low risk for later neurodevelopmental problems but are prone to academic problems[10]. Middle et al found that MLBW children have higher rates of academic performance and educational problems[11]. Eglan et al and Breslau et al have reported increased rates of learning and behavioral problems among these children[12,13]. Some of these studies do not include a normal birth weight (NBW) control group for comparison. In this study, we evaluated the motor developmental status of a group of MLBW preterm infants at the corrected age of 18±2 months via the Peabody Developmental Motor Scale II (PDMS-2) test and compared the results with that of NBW infants. Subjects and Methods Upon approval of the ethical committee of human research from Tehran University of Medical Sciences, this study was carried out at Shahid Akbar-Abadi Hospital in Tehran. The sample size was calculated by computing the average and standard deviation of PDMS-2 scores for 20 children (10 LBW and 10 NBW). A total of 52 patients were required to achieve a statistical power of 0.90 with a type I error of 0.05 in consideration of the standard deviation of 11.918.

bar-Abadi Hospital in Tehran. The sample size was calculated by computing the average and standard deviation of PDMS-2 scores for 20 children (10 LBW and 10 NBW). A total of 52 patients were required to achieve a statistical power of 0.90 with a type I error of 0.05 in consideration of the standard deviation of 11.918. After reviewing hospital records, infants with a corrected age of 18±2 months with a history of preterm birth and MLBWs, from June–November 2008, were included. Infants with birth weights between 1500–2499 g were considered MLBW and with a gestational age of less than 37 weeks were considered preterm. Term infants (with a gestational age ≥37 weeks) with LBW due to intra uterine growth retardation or being small for gestational age, were excluded. In addition, multiple pregnancies, infants with low Apgar scores or severe asphyxia, abnormal brain imaging, congenital malformations, chromosomal and genetic syndromes, and children with history of rehabilitation therapy for more than 2 months or with drug treatments that affected motor function and those who were not residents of Tehran were also excluded. We contacted parents and requested to bring their infant to our clinic and asked for informed consent for the study. The control group consisted of healthy 18±2 month-old infants with birth weights between 2500–4000 g who presented at the clinic for routine checkup. Children with a history of admission to NICU, gestational age of <37 or >42 weeks, resulting from multiple pregnancy, those with musculoskeletal, neurologic, genetic, or any other disorder that negatively influenced development were also excluded. Children in the control group indicated normal development in previous well-child visits by physicians. All parents signed a consent form prior to enrollment in the study.

ultiple pregnancy, those with musculoskeletal, neurologic, genetic, or any other disorder that negatively influenced development were also excluded. Children in the control group indicated normal development in previous well-child visits by physicians. All parents signed a consent form prior to enrollment in the study. All children were referred to an occupational therapist that was blinded to their birth weight. Their gross and fine motor skills were assessed by the PDMS-2 test, i.e., a specifically designed motor scale that identifies most motor skill dysfunctions. It is a standardized and norm-referenced test of gross and fine motor skills from birth to 5 years of age with a determined reliability and validity[14]. It consists of a Gross Motor and a Fine Motor Scale, each is divided into skill subtests that detect typical motor tasks for each age. Gross motor development is the ability to use the large muscle systems to react to environmental changes, assume a stable posture, move from place to place, and catch, throw, and kick balls. While fine motor development is the ability to use fingers and hands to gasp objects, stack blocks, draw figures, and manipulate objects. Test item performance is summarized and analyzed using motor quotients derived by adding the subtest standard scores and converting the sum to a quotient that has a mean of 100 and a standard deviation of 15. They include gross motor quotient (reflexes or object manipulation, stationary and locomotion subtests), Fine motor quotient (FMQ, gasping and visual-motor integration subtests), and total motor quotient (TMQ) that is comprised of the quotient scores of the gross and fine motor. The total motor quotient (GMQ) is probably the best estimate of overall motor abilities.

bject manipulation, stationary and locomotion subtests), Fine motor quotient (FMQ, gasping and visual-motor integration subtests), and total motor quotient (TMQ) that is comprised of the quotient scores of the gross and fine motor. The total motor quotient (GMQ) is probably the best estimate of overall motor abilities. Quotient scores are interpreted as follows: very superior (131–165), superior (121–130), above average (110–120), average (90–109), below average (80–89), poor (70–79), and very poor (35–69). Evaluation of the reliability of the tests was performed in a test-retest pilot study. Two occupational therapists independently performed the tests in 10 randomly selected normal children. All children were re-examined by both testers at an interval of one-week. High inter-tester reliability was achieved, as intra-class correlation coefficients (ICC) were 1.00, 0.97, and 0.99 for GMQ, FMQ, and TMQ, respectively (P<0.001). The inter-tester reliability also was confirmed. ICCs were 0.94, 0.97, and 1.00 for GMQ, FMQ, and TMQ, respectively (P<0.001). The test is an objective assessment tool that does not compromise the results of the study. Nevertheless, we have validated it for the population under investigation that exists as unpublished data. Corrected age for LBW children was calculated by subtracting the gestational age from 37 weeks and the result was subtracted from chronological age (37 -gestational age=A, corrected age=chronological age -A).

Evaluation of the reliability of the tests was performed in a test-retest pilot study. Two occupational therapists independently performed the tests in 10 randomly selected normal children. All children were re-examined by both testers at an interval of one-week. High inter-tester reliability was achieved, as intra-class correlation coefficients (ICC) were 1.00, 0.97, and 0.99 for GMQ, FMQ, and TMQ, respectively (P<0.001). The inter-tester reliability also was confirmed. ICCs were 0.94, 0.97, and 1.00 for GMQ, FMQ, and TMQ, respectively (P<0.001). The test is an objective assessment tool that does not compromise the results of the study. Nevertheless, we have validated it for the population under investigation that exists as unpublished data. Corrected age for LBW children was calculated by subtracting the gestational age from 37 weeks and the result was subtracted from chronological age (37 -gestational age=A, corrected age=chronological age -A). Statistical analysis was performed using SPSS 17.0 software. Normal distribution of data was tested using the Kolmogorov–Smirnov test. Categorical data was analyzed using the Chi-square test. Continuous variables were compared between the two groups using independent sample t-tests or Mann–Whitney U tests when the distribution of data was abnormal. P. value <0.05 was considered significant.

ibution of data was tested using the Kolmogorov–Smirnov test. Categorical data was analyzed using the Chi-square test. Continuous variables were compared between the two groups using independent sample t-tests or Mann–Whitney U tests when the distribution of data was abnormal. P. value <0.05 was considered significant. Findings A total of 88 infants including 58 LBW and 30 NBW with a mean birth weight of 1900 (±382.4) and 3150 (±473.5) g respectively, were studied. 14 (46.7%) infants in NBW and 30 (51.7%) in LBW group were males, which was not statistically different (P=0.6). In the LBW group, there was no significant difference between the mean birth weights of male and female children (1970±423.6 vs 1820 (±430.2) g, respectively; P=0.2). The mean age of children in LBW was 18.2 (±0.0) and in NBW 18.0±0.7 months. The mean gestational age of male children in the LBW group was 33.5±2.7 weeks vs 32.3±2.7 weeks in female children (P=0.08). The mean duration of hospital stay in LBW group was 7.5 (4–11 days).

Findings A total of 88 infants including 58 LBW and 30 NBW with a mean birth weight of 1900 (±382.4) and 3150 (±473.5) g respectively, were studied. 14 (46.7%) infants in NBW and 30 (51.7%) in LBW group were males, which was not statistically different (P=0.6). In the LBW group, there was no significant difference between the mean birth weights of male and female children (1970±423.6 vs 1820 (±430.2) g, respectively; P=0.2). The mean age of children in LBW was 18.2 (±0.0) and in NBW 18.0±0.7 months. The mean gestational age of male children in the LBW group was 33.5±2.7 weeks vs 32.3±2.7 weeks in female children (P=0.08). The mean duration of hospital stay in LBW group was 7.5 (4–11 days). Table 1 demonstrates a comparison of PDMS-2 subtests standard scores for each group. As the data shows, LBW children achieved significantly lower scores in grasping and visual-motor integration skills. There was no significant difference found for stationary, locomotion, and object manipulation skills scores between LBW and NBW children. There was no statistically significant association between gender and motor quotients scores in LBW group (Table 2) and between LBW and NBW groups. According to motor quotients, in the LBW group, GMQ and FMQ were below average in 6 (10.3%) and 10 (17%) children, respectively (Table 2). Table 3 shows that there are no statistically significant difference between groups regarding GMQ (P=0.1). However, LBW children achieved significantly lower scores in FMQ (P=0.001) and in TMQ (P=0.001).

tients, in the LBW group, GMQ and FMQ were below average in 6 (10.3%) and 10 (17%) children, respectively (Table 2). Table 3 shows that there are no statistically significant difference between groups regarding GMQ (P=0.1). However, LBW children achieved significantly lower scores in FMQ (P=0.001) and in TMQ (P=0.001). Discussion LBW is a major public health problem that negatively influences infant development and the quality of life, and poses financial burdens on health care systems[15]. Table 1 Comparison of Peabody Developmental Motor Scale II (PDMS-2) subtests standard scores in low and normal birth weight children Variable Low birth weight Mean (SD) Normal birth weight Mean (SD) P -value Stationary 10.5 (1.5) 10.6 (1.3) 0.84 Locomotion 9.9 (1.4) 10.4 (1.0) 0.11 Object Manipulation 9.6 (1.5) 10.1 (0.9) 0.09 Grasping 9.05 (1.0) 9.9 (0.9) 0.001 Visual-Motor Integration 8.7 (1.2) 10 (1.0) 0.001 SD: Standard Deviation Table 2 Distribution of motor quotients scores of low birth weight children based on gender Parameter Below average (80–89) Average (90–109) Above average (110–120) Gross Motor Quotient Male 4 (13.3%) 21 (70.0%) 5 (16.7%) Female 2 (7.1%) 24 (85.7%) 2 (7.1%) Total 6 (10.3%) 45 (77.6%) 7 (12.1%) Fine Motor Quotient Male 8 (26.7%) 22 (73.3%) 0 (0%) Female 2 (7.1%) 25 (89.3%) 1 (3.6%) Total 10 (17.2%) 47 (81.0%) 1 (1.7%) Total Motor Quotient Male 6 (20.0%) 23 (76.7%) 1 (3.3%) Female 4 (14.3%) 24 (85.7%) 0 (0%)

Above average (110–120) Gross Motor Quotient Male 4 (13.3%) 21 (70.0%) 5 (16.7%) Female 2 (7.1%) 24 (85.7%) 2 (7.1%) Total 6 (10.3%) 45 (77.6%) 7 (12.1%) Fine Motor Quotient Male 8 (26.7%) 22 (73.3%) 0 (0%) Female 2 (7.1%) 25 (89.3%) 1 (3.6%) Total 10 (17.2%) 47 (81.0%) 1 (1.7%) Total Motor Quotient Male 6 (20.0%) 23 (76.7%) 1 (3.3%) Female 4 (14.3%) 24 (85.7%) 0 (0%) Total 10 (17.2%) 47 (81.0%) 1 (1.7%) Several studies have shown that LBW children are more likely to have neurological problems[16-18] that may persist into school age and adolescence periods[17]. Motor deficits in LBW children can influence the ability to learn and limit active participation in daily life at school and at home[19].

%) 47 (81.0%) 1 (1.7%) Several studies have shown that LBW children are more likely to have neurological problems[16-18] that may persist into school age and adolescence periods[17]. Motor deficits in LBW children can influence the ability to learn and limit active participation in daily life at school and at home[19]. Our findings indicate that LBW preterm infants at 18±2 month-old corrected age have impaired motor abilities compared with NBW infants especially for fine motor skills. This is in agreement with the results obtained by Cristian Alves da Silva et al in Brazil, who found that LBW preterm infants have delays in neuropsychomotor development and the lowest scores are for language, and hand-eye and fine motor coordination[9]. Our results are also comparable with Goyen T-A, who showed a significant proportion of LBW infants had fine motor deficits at 18 months of age that continued until 5 years of age[19]. Halpern et al reported that LBW children had a three times greater risk of developmental delay compared with NBW (P<0.001)[20]. Halpern et al also found the prevalence of infants with delay diminishes when incomes and birth weights increase[21]. In that study, children of poorer families were more prone to developmental delay and birth weight was a strong factor. In addition, Wilcox concluded that low birth weight is strongly associated with later developmental deficits[22].

evalence of infants with delay diminishes when incomes and birth weights increase[21]. In that study, children of poorer families were more prone to developmental delay and birth weight was a strong factor. In addition, Wilcox concluded that low birth weight is strongly associated with later developmental deficits[22]. We did not find any significant difference between LBW and NBW infants regarding gross motor ability. Other studies have shown that developmental problems of LBW children range from mild deficits in cognition and neuromotor functioning in the majority to cerebral palsy in a small minority[2,16,17]. We compared motor development between MLBW and NBW infants. Datar and Jacknowitz compared mental and motor development of VLBW and MLBW infants during the first two years of life with NBW, LBWs had a small defect in mental and motor development[23]. Middle et al also reported higher rates of neuro-motor problems in MLBW when compared with NBW children[11]. There were differences between LBW and NBW infants for FMQ and TMQ but not for GMQ in our study. This discrepancy may be related to the visual-motor integration subtest. Goyen et al showed a significant correlation between visual-motor and fine motor skills and concluded that previous reports of visual-motor problems in school-age VLBW children could be due to fine motor defects[19]. Differences observed for TMQ in our study could be a consequence of the magnitude of FMQ. Table 3 Comparison of motor quotient scores of low and normal birth weight children. Parameter Group Number Standard Score (Mean±SD) P. value

We compared motor development between MLBW and NBW infants. Datar and Jacknowitz compared mental and motor development of VLBW and MLBW infants during the first two years of life with NBW, LBWs had a small defect in mental and motor development[23]. Middle et al also reported higher rates of neuro-motor problems in MLBW when compared with NBW children[11]. There were differences between LBW and NBW infants for FMQ and TMQ but not for GMQ in our study. This discrepancy may be related to the visual-motor integration subtest. Goyen et al showed a significant correlation between visual-motor and fine motor skills and concluded that previous reports of visual-motor problems in school-age VLBW children could be due to fine motor defects[19]. Differences observed for TMQ in our study could be a consequence of the magnitude of FMQ. Table 3 Comparison of motor quotient scores of low and normal birth weight children. Parameter Group Number Standard Score (Mean±SD) P. value Gross Motor Quotient NBW 30 102.47 (5.52) 0.121 LBW 58 100.12 (7.18) Fine Motor Quotient NBW 30 99.6 (5.02) 0.001 LBW 58 93.33 (5.42) Total Motor Quotient NBW 30 101.53 (5.03) 0.001 LBW 58 97.02 (5.89) NBW: Normal Birth Weight, LBW: Low Birth Weight, SD: Standard Deviation Our results showed no differences in motor skills of LBW children regarding gender, which is consistent with those of other studies[3,24,25].

Fine Motor Quotient NBW 30 99.6 (5.02) 0.001 LBW 58 93.33 (5.42) Total Motor Quotient NBW 30 101.53 (5.03) 0.001 LBW 58 97.02 (5.89) NBW: Normal Birth Weight, LBW: Low Birth Weight, SD: Standard Deviation Our results showed no differences in motor skills of LBW children regarding gender, which is consistent with those of other studies[3,24,25]. It has been suggested that early intervention for children who are suspected of motor developmental delay can positively influence the outcome. In a systematic review of 34 studies, Blauw-Hospers and Hadders-Alga have evaluated the effect of intervention from birth to 18 months on outcomes for children who were at increased risk for developmental motor disorders and showed that infants benefit from intervention program[26]. Other studies have shown persistent deficits in LBW children with negative effect on academic performance[9]. Providing early intervention program is a reasonable goal to reduce motor deficit, its consequences, and to improve outcomes. As the problems of VLBW children are more significant and most recent studies have concentrated on this group, physicians may neglect the importance of developmental outcomes for MLBW children. The findings of our study are particularly important because they point to the need to assess the motor developmental status of MLBW infants with follow-ups throughout childhood.

t and most recent studies have concentrated on this group, physicians may neglect the importance of developmental outcomes for MLBW children. The findings of our study are particularly important because they point to the need to assess the motor developmental status of MLBW infants with follow-ups throughout childhood. Our study has some limitations. First, we have included 18±2 month-old toddlers and therefore, this study cannot answer whether LBW children with delayed fine motor abilities are able to catch up with peers. Second, we used only the PDMS-2 test to assess motor abilities. Although the PDMS-2 is a valid scoring system that is extensively used, some authors have suggested some limitations for it[27]. Conclusion MLBW infants may be at risk for developmental delay, which makes developmental assessments mandatory at an early age for this population. Physicians should take extra care with these infants and proceed to a thorough and systematic monitoring of developmental status. Together with frequent visits, these procedures enhance the developmental delay diagnosis and, hence, lead to earlier recognition and intervention that may reduce long-term problems associated with the developmental delay. Further studies are recommended to investigate motor abilities and the socio-behavioral and cognitive function in late childhood and adolescence of LBW children and to detect the effect of environmental factors on their developmental status.

Conclusion MLBW infants may be at risk for developmental delay, which makes developmental assessments mandatory at an early age for this population. Physicians should take extra care with these infants and proceed to a thorough and systematic monitoring of developmental status. Together with frequent visits, these procedures enhance the developmental delay diagnosis and, hence, lead to earlier recognition and intervention that may reduce long-term problems associated with the developmental delay. Further studies are recommended to investigate motor abilities and the socio-behavioral and cognitive function in late childhood and adolescence of LBW children and to detect the effect of environmental factors on their developmental status. Acknowledgment We would like to thank our colleagues and the staff at the medical record unit at Shahid Akbar Abadi Hospital and to Mrs. Shahrzad Soltanzadeh for her kind cooperation. Conflict of Interest: None Authors’ Contribution A. Tavasoli: Design of the study, drafted, and prepared the manuscript. F. Aliabadi: Design of the manuscript, data analysis, and interpretation. R. Eftekhari: data collection and data analysis All authors approved final version of the paper.

Dear Editor, Referring to synovitis, acne, pustulosis, hyperostosis and osteitis; SAPHO syndrome is defined as a chronic, relapsing rheumatologic disease of uncertain etiology characterized by distinct osteoarticular and cutaneous manifestations. There have been recent reports of chronic recurrent multifocal osteomyelitis (CRMO) occurring in adults and SAPHO syndrome occurring in children whereas just the vice-versa is normally expected[1,2]. Herein, we would like to emphasize a rare form of SAPHO syndrome in terms of age and the localization of the disease. A 12-year-old female patient was admitted to our hospital with the complaint of right hip and low back pain. She had tenderness on the right iliac crest. She had normal hemogram with an erythrocyte sedimentation rate (ESR) of 51 mm/hour. Serological tests for Salmonella and Brucella were negative. Having a BCG vaccination scar, PPD was negative and no abnormalities were found in chest radiography of the patient. Bone marrow aspiration and bone scintigraphy revealed no signs of malignancy. Anti-streptolysin O, C-reactive protein, anti-nuclear anitobody, anti double stranded DNA and HLA B27 were negative. Serum immunoglobulin levels were within the normal range. Magnetic resonance imaging (MRI) (Fig. 1), showed increased density of right acetabular area and surrounding soft tissue besides bone marrow edema. She received 15 mg/kg/day naproxen sodium administered.

obody, anti double stranded DNA and HLA B27 were negative. Serum immunoglobulin levels were within the normal range. Magnetic resonance imaging (MRI) (Fig. 1), showed increased density of right acetabular area and surrounding soft tissue besides bone marrow edema. She received 15 mg/kg/day naproxen sodium administered. In a month’s time, regression of complaints occurred while occasional emergence of fever and pain on the left hip was identified. Repeated pelvic MRI (Fig. 2) revealed involvement of head of femur on the left side, L5 and S1 vertebrates characterized with hyper-density and edema. Fig. 1 Coronal fat suppressed T2 weighted image shows edema in right acetabulum and surrounding soft tissues

In a month’s time, regression of complaints occurred while occasional emergence of fever and pain on the left hip was identified. Repeated pelvic MRI (Fig. 2) revealed involvement of head of femur on the left side, L5 and S1 vertebrates characterized with hyper-density and edema. Fig. 1 Coronal fat suppressed T2 weighted image shows edema in right acetabulum and surrounding soft tissues Naproxen sodium treatment was continued for almost next 4 months. She stopped having this treatment on her own accord, and then applied to our hospital with vesiculopustular skin lesions on the palmar, plantar and retroauricular surfaces. Histopathologic findings revealed subcorneal intraepidermal vesicles as well as intracorneal plasma insudation. Lymphocyte exocytosis, infiltrating vesicles and epidermal cells were also visualized. Patient received oral analgesic, non-steroidal anti-inflammatory drug (Ibuprofen) and topical steroid treatment. Remarkable increase in the intensity of edema signals was notable in the evaluation of serial radiological findings (Fig 3). Naproxen sodium administration was continued and after one month there was a clinical regression in the skeletal system findings and skin lesions. Being in the 18th month, she is still under follow up with no problems occurring to date. Classification of periodic fever syndromes under the auto-inflammatory disorders has been followed by the definition of auto-inflammatory diseases with particular involvement of bone tissue.

Naproxen sodium treatment was continued for almost next 4 months. She stopped having this treatment on her own accord, and then applied to our hospital with vesiculopustular skin lesions on the palmar, plantar and retroauricular surfaces. Histopathologic findings revealed subcorneal intraepidermal vesicles as well as intracorneal plasma insudation. Lymphocyte exocytosis, infiltrating vesicles and epidermal cells were also visualized. Patient received oral analgesic, non-steroidal anti-inflammatory drug (Ibuprofen) and topical steroid treatment. Remarkable increase in the intensity of edema signals was notable in the evaluation of serial radiological findings (Fig 3). Naproxen sodium administration was continued and after one month there was a clinical regression in the skeletal system findings and skin lesions. Being in the 18th month, she is still under follow up with no problems occurring to date. Classification of periodic fever syndromes under the auto-inflammatory disorders has been followed by the definition of auto-inflammatory diseases with particular involvement of bone tissue. These are mainly, CRMO, SAPHO syndrome, Majeed syndrome, deficiency of interleukin-1 receptor antagonist (DIRA) and cherubism. SAPHO syndrome is a clinical entity characterized by inflammation in bone, joint and skin. Inflammation manifests in the form of sterile osteomyelitis and hyperostosis in the bone, acne or pustulosis in the skin and synovitis in the joints.