Browse the corpus

1. Introduction In the world, obstetric hemorrhage is the most common cause of maternal death, causing 24% of, or an estimated 127,000, maternal deaths annually [1]. It has also been reported that massive (2,000 mL or more) and life-threatening obstetric hemorrhage occurs in 3–5% [2] and 0.1% [3] of deliveries, respectively, and blood product transfusion is required in 0.3–1% [2, 4]. Also in Japan, obstetric hemorrhage account causes 25% of maternal deaths [5] and massive obstetric hemorrhage (MOH) occurs in 1.1% of deliveries [6]. In the terminal stage of pregnancy, where the coagulation system is enhanced and the fibrinolysis system is inhibited [7, 8], MOH may be apt to induce consumptive loss of coagulation factors, which causes further hemorrhage, forming a vicious circle such as disseminated intravascular coagulation (DIC) [9]. Accurate evaluation of blood loss is important to determine whether transfusion should be performed, but it is difficult in obstetric hemorrhage [10–12]. In addition, high-level capacity of pregnant women to tolerate obstetric hemorrhage [13, 14] masks changes in their vital signs, resulting in a delay in the detection and treatment of hypovolemia, which causes further hemorrhage and hemorrhagic shock. Therefore, the comprehensive evaluation of not only blood loss but also the cause of hemorrhage, a patient's medical condition, age, vital signs, and blood biochemical data is required to determine whether transfusion is necessary [15].

nd treatment of hypovolemia, which causes further hemorrhage and hemorrhagic shock. Therefore, the comprehensive evaluation of not only blood loss but also the cause of hemorrhage, a patient's medical condition, age, vital signs, and blood biochemical data is required to determine whether transfusion is necessary [15]. Understanding the above-mentioned specificity of obstetric hemorrhage is also required for appropriate blood product support, which can effectively improve its pathophysiological condition, reduce the risk of DIC, and avoid the aggravation of hemorrhagic shock [16]. However, in Japan, “Principles for blood transfusion therapy” and “Principles of the use of blood products” [17] proposed by the Japanese Ministry of Health, Labour and Welfare give little consideration to the pathophysiological mechanism or no standard for the appropriate dosage of blood product transfusion specific to obstetric hemorrhage. Therefore, we retrospectively reviewed blood product administration in patients with obstetric hemorrhage at our tertiary obstetric center for a 5-year period and discuss appropriate dosage of blood product transfusion for obstetric hemorrhage.

riate dosage of blood product transfusion specific to obstetric hemorrhage. Therefore, we retrospectively reviewed blood product administration in patients with obstetric hemorrhage at our tertiary obstetric center for a 5-year period and discuss appropriate dosage of blood product transfusion for obstetric hemorrhage. 2. Material and Methods 2.1. Subjects Between January 1, 2004, and December 31, 2008, 243 obstetric patients underwent blood product transfusion in our tertiary perinatal institution, Saitama Medical Center/Saitama Medical University, which is only general medical institution for tertiary perinatal care in Saitama prefecture with population of 7.2 million. Their data were manually abstracted by our research staffs from our medical records, anonymized in an unlinkable fashion prior to our investigation, which exempted us from institutional review board approval according to “Ethical principles for etiological studies” [18] proposed by the Japanese Ministry of Health, Labour and Welfare. Blood products involved in this study are red cells concentrates (RCC), fresh-frozen plasma (FFP), and platelet concentrates (PC). Two units of RCC (approximately 140 mL/unit), 3 units of FFP (approximately 80 mL/unit), and 2 units of PC (approximately 20 mL/unit) are derived from 400 mL of whole blood, respectively, in this study.

s involved in this study are red cells concentrates (RCC), fresh-frozen plasma (FFP), and platelet concentrates (PC). Two units of RCC (approximately 140 mL/unit), 3 units of FFP (approximately 80 mL/unit), and 2 units of PC (approximately 20 mL/unit) are derived from 400 mL of whole blood, respectively, in this study. 2.2. Our Management Principles for Blood Product Transfusion (Table 1) Since blood loss in vaginal delivery or Caesarean section is difficult to evaluate accurately [11, 12] and hemoglobin (Hb) concentration necessary to maintain appropriate hemodynamics and oxygen supply is ≥7 g/dL [19, 20], Hb concentration <7 g/dL was determined to be an indication for blood product transfusion in principle. In addition to this principle, the patient's age, medical condition, state of hemorrhage, and blood test data were taken into consideration [8, 15]. Since the transfusion for patients with an Hb concentration ≥7 g/dL and stable vital signs may lead to excessive transfusion, RCC transfusion was performed with a goal Hb concentration of 7-8 g/dL [21]. FFP was concomitantly transfused until the coagulation function normalizes [9, 22]. We did not have any rule in advance to define the proportion of FFP to RCC in the present study. Cryoprecipitate, fibrinogen concentrate, or recombinant activated factor VII was not administered in general, since they are not approved or paid by public medical insurance in Japan.

N = 12 N = 24 Intraoperative characteristics Ovarian suspension 7 (58.3) 22 (91.7) 0.029 Rectosigmoid suspension 8 (66.7) 3 (12.5) 0.002 Systematic ureteral dissection 5 (41.7) 21 (87.5) 0.007 Division of uterine vein 0 (0.0) 3 (12.5) NS Excision of pelvic peritoneum 6 (50.0) 24 (100.0) <0.001 Partial colpectomy 3 (25.0) 0 (0.0) 0.031 Rectal wall suturing 3 (25.0) 0 (0.0) 0.031 Mean operation length (mins) 188 179 NS Mean blood loss (mls) 45 70 <0.001 Postoperative characteristics at 6/12 followup Complete resolution of CPP 7 (58.3) 18 (75.0) NS Vaginal scarring/induration 5 (41.7) 6 (25.0) NS Table 4 Intraoperative and postoperative characteristics according to size of the DIE nodule. Patient characteristics DIE nodule size ≤ 2 cm N (%) DIE nodule size > 2 cm N (%) Fisher's exact test P value N = 27 N = 9 Intraoperative characteristics Ovarian suspension 20 (74.1) 9 (100.0) NS Rectosigmoid suspension 5 (18.5) 6 (66.7) 0.012 Systematic ureteral dissection 18 (66.7) 8 (88.9) NS Division of uterine vein 0 (0.0) 3 (33.3) 0.012 Excision of pelvic peritoneum 22 (81.5) 8 (88.9) NS Partial colpectomy 0 (0.0) 3 (33.3) 0.012 Rectal wall suturing 0 (0.0) 3 (33.3) 0.012 Mean operation length (mins) 124 216 <0.0001 Mean blood loss (mls) 40 85 <0.001 Postoperative characteristics at 6/12 followup Complete resolution of CPP 20 (74.0) 5 (55.5) NS Vaginal scarring/induration 5 (18.5 ) 6 (84.4) 0.012

2.2. Our Management Principles for Blood Product Transfusion (Table 1) Since blood loss in vaginal delivery or Caesarean section is difficult to evaluate accurately [11, 12] and hemoglobin (Hb) concentration necessary to maintain appropriate hemodynamics and oxygen supply is ≥7 g/dL [19, 20], Hb concentration <7 g/dL was determined to be an indication for blood product transfusion in principle. In addition to this principle, the patient's age, medical condition, state of hemorrhage, and blood test data were taken into consideration [8, 15]. Since the transfusion for patients with an Hb concentration ≥7 g/dL and stable vital signs may lead to excessive transfusion, RCC transfusion was performed with a goal Hb concentration of 7-8 g/dL [21]. FFP was concomitantly transfused until the coagulation function normalizes [9, 22]. We did not have any rule in advance to define the proportion of FFP to RCC in the present study. Cryoprecipitate, fibrinogen concentrate, or recombinant activated factor VII was not administered in general, since they are not approved or paid by public medical insurance in Japan. 2.3. Evaluated Items The following items were retrospectively evaluated: underlying disorders which required blood product transfusion, types of blood product and their transfused volume, and data of hemoglobin (Hb) concentration, percent prothrombin activity (%PT; normal range: 84–117% in our institution), activated partial thromboplastin time (aPTT; 25–36 sec), and fibrinogen concentration (150–400 mg/dL) within 30 minutes before blood transfusion. Blood test data were excluded for further statistical analyses when they were obtained after blood product was transfused.

ty (%PT; normal range: 84–117% in our institution), activated partial thromboplastin time (aPTT; 25–36 sec), and fibrinogen concentration (150–400 mg/dL) within 30 minutes before blood transfusion. Blood test data were excluded for further statistical analyses when they were obtained after blood product was transfused. Prothrombin activity was assayed by STA-R Evolution (Roche Diagnostics), a fully automated coagulation analyzer. Clotting times were converted to percent normal plasma prothrombin activity from a log-log standard curve prepared with dilutions of control pooled plasma [23]. 2.4. Statistical Analyses The presence or absence of correlations was analyzed by JMP (SAS Institute) software employing Spearman's rank correlation coefficient, and equality of parameters among groups was analyzed employing Kruskal-Wallis one-way analysis of variance by ranks because each parameter did not show normal distribution. In each test, P < 0.05 was regarded as significant. 3. Results 3.1. Obstetric Patients Who Underwent Blood Product Transfusion (Table 2) We have experienced 243 obstetric patients who underwent blood product transfusion, consisting of 164 (67%) delivered by Cesarean section and 79 (33%) delivered vaginally. Eighty-two (34%) women were transported to our institution in their puerperium for our specialized management of obstetric hemorrhage. For 8 patients who had blood transfusion prior to or during transfer, we included the data on transfusion using their medical record of transfer source institution.

delivered vaginally. Eighty-two (34%) women were transported to our institution in their puerperium for our specialized management of obstetric hemorrhage. For 8 patients who had blood transfusion prior to or during transfer, we included the data on transfusion using their medical record of transfer source institution. We had one patient who died with amniotic fluid embolism resulting in severe coagulopathy and multiple organ failure and one vegetative patient with severe HELLP syndrome resulting in hypoxic encephalopathy. Two hundred and twenty (91%) patients underwent blood transfusion for obstetric hemorrhage, while 17 (7%) patients for hematological disorders during pregnancy such as idiopathic thrombocytopenic purpura (ITP), myelodysplastic syndrome (MDS), and leukemia. Four other patients underwent blood transfusion for major disorders during pregnancy such as cerebral infarction, gastric cancer, colon cancer, and head injury. One pregnant woman with cardiac dysfunction and one with no routine checkup but severe anemia had blood transfusion in their emergency Cesarean delivery. 3.2. Obstetric Indications of Blood Transfusion (Table 3) Table 3 shows the 220 cases with obstetric hemorrhage that required blood product transfusion. Most of the cases underwent blood transfusion in their peripartum, except for a few cases who had massive hemorrhage several days after their delivery.

Two hundred and twenty (91%) patients underwent blood transfusion for obstetric hemorrhage, while 17 (7%) patients for hematological disorders during pregnancy such as idiopathic thrombocytopenic purpura (ITP), myelodysplastic syndrome (MDS), and leukemia. Four other patients underwent blood transfusion for major disorders during pregnancy such as cerebral infarction, gastric cancer, colon cancer, and head injury. One pregnant woman with cardiac dysfunction and one with no routine checkup but severe anemia had blood transfusion in their emergency Cesarean delivery. 3.2. Obstetric Indications of Blood Transfusion (Table 3) Table 3 shows the 220 cases with obstetric hemorrhage that required blood product transfusion. Most of the cases underwent blood transfusion in their peripartum, except for a few cases who had massive hemorrhage several days after their delivery. Some of the uterine atony cases resulted from retained placental tissue or overdistension due to multiple pregnancy. Genital tract trauma was noted in the vaginal wall, cervix, and/or uterine body (uterine rupture was detected in 6 cases). Thirteen cases of placenta previa were complicated by placenta acreta, increta, or percreta. 3.3. Blood Product Transfusion for Obstetric Indications

1. Introduction Abnormal fetal umbilical artery Doppler (UAD) studies represent a problem that is complex in both antenatal prevention and management and postnatal management [1]. In particular, absent and reversed end-diastolic flow of the fetal umbilical arteries are associated with poor neonatal outcomes, ranging from premature delivery and stillbirth to postnatal neurodevelopmental impairment [2] and diseases such as obesity and hypertension later in life [2]. Reversed end-diastolic flow (REDF) is the most advanced stage of abnormal umbilical artery Doppler flow and represents obliteration of nearly 70% of the placental function [3]. REDF also represents a higher risk of NICU admission, need for respiratory support, and perinatal mortality, regardless of age at delivery [4].

Some of the uterine atony cases resulted from retained placental tissue or overdistension due to multiple pregnancy. Genital tract trauma was noted in the vaginal wall, cervix, and/or uterine body (uterine rupture was detected in 6 cases). Thirteen cases of placenta previa were complicated by placenta acreta, increta, or percreta. 3.3. Blood Product Transfusion for Obstetric Indications Table 4 shows the number of cases and median volumes for each blood product transfused for the 220 cases with obstetric hemorrhage shown in Table 3. No cryoprecipitate, fibrinogen concentrate, or recombinant factor VII was administered. Autologous whole blood was transfused for 24 patients with placenta previa with or without placenta acreta, increta, or percreta. These patients who had autologous blood transfusion were excluded for the analysis. In 196 patients with obstetric hemorrhage who underwent only allogenic transfusion, there was a significant positive correlation (P < 0.001) between the volume of RCC transfused and that of FFP (Figure 1). This significant positive correlation was shown irrespective of underlying obstetric disorders (Table 5).

Table 4 shows the number of cases and median volumes for each blood product transfused for the 220 cases with obstetric hemorrhage shown in Table 3. No cryoprecipitate, fibrinogen concentrate, or recombinant factor VII was administered. Autologous whole blood was transfused for 24 patients with placenta previa with or without placenta acreta, increta, or percreta. These patients who had autologous blood transfusion were excluded for the analysis. In 196 patients with obstetric hemorrhage who underwent only allogenic transfusion, there was a significant positive correlation (P < 0.001) between the volume of RCC transfused and that of FFP (Figure 1). This significant positive correlation was shown irrespective of underlying obstetric disorders (Table 5). A ratio of total transfused units of FFP to RCC was 2.1 (3,550/1,665) in the 196 patients with allogenic transfusion alone. The median of FFP/RCC ratio for each patient was 2.0 in total (Table 5), and 2.1 and 2.0 in subgroups with moderate (6–9 units) and massive (10 units or more) RCC transfusion, respectively (Table 6). The median of FFP/RCC ratio was not significantly different between underlying obstetric disorders, but 2.0 or more except for uterine atony (Table 5).

each patient was 2.0 in total (Table 5), and 2.1 and 2.0 in subgroups with moderate (6–9 units) and massive (10 units or more) RCC transfusion, respectively (Table 6). The median of FFP/RCC ratio was not significantly different between underlying obstetric disorders, but 2.0 or more except for uterine atony (Table 5). 3.4. Comparison of Blood Test Data among Underlying Obstetric Disorders Next, we compared Hb concentration, percent prothrombin activity (%PT), aPTT, and fibrinogen concentration immediately before blood transfusion among underlying obstetric disorders which required blood product transfusion. From the 220 obstetric disorder cases, 8 patients who had our blood tests only after transfusion were excluded for the analysis. One patient with amniotic fluid embolism was also excluded. As shown in Figure 2, aPTT was not significantly different among obstetric disorders. Hb concentration was significantly higher in HELLP syndrome compared with other disorders. %PT in placental abruption was not significantly different from those in uterine atony/inversion or genital tract trauma, but significantly lower compared with placenta previa and HELLP syndrome. Fibrinogen concentration in placental abruption was significantly lower, and that in HELLP syndrome was significantly higher compared to other disorders.

s not significantly different from those in uterine atony/inversion or genital tract trauma, but significantly lower compared with placenta previa and HELLP syndrome. Fibrinogen concentration in placental abruption was significantly lower, and that in HELLP syndrome was significantly higher compared to other disorders. 3.5. Correlation between Data of Blood Tests and the Volumes of Blood Products Transfused To clarify the characteristics of hemorrhage in total or each underlying obstetric disorders, we evaluated the possible correlation between data from blood tests and the volumes of blood products transfused. As mentioned previously, 32 patients who had our blood tests only after transfusion or autologous blood transfusion were excluded for the analysis. One maternal death with amniotic fluid embolism was also excluded since severe coagulopathy resulting in MOF was not controlled by massive blood product transfusion. 3.5.1. Correlation between the Volume of RCC Transfusion and Hb Concentration or %PT In the 187 obstetric disorder patients, there was a significant negative correlation (P < 0.001) between the volume of RCC transfused and Hb concentration immediately before transfusion (Figure 3). This significant negative correlation was shown irrespective of underlying obstetric disorders except for placenta previa (data not shown).

ric disorder patients, there was a significant negative correlation (P < 0.001) between the volume of RCC transfused and Hb concentration immediately before transfusion (Figure 3). This significant negative correlation was shown irrespective of underlying obstetric disorders except for placenta previa (data not shown). Similarly, there was a significant negative correlation (P < 0.001) between the volume of RCC transfused and %PT (Figure 3). This significant negative correlation was shown irrespective of underlying obstetric disorders except for placenta previa and HELLP syndrome (data not shown). 3.5.2. Correlation between the Volume of FFP Transfusion and %PT or Fibrinogen Concentration Also in FFP transfusion, there was a significant negative correlation (P < 0.001) between the volume of FFP transfused and %PT immediately before transfusion (Figure 4). This significant negative correlation was shown irrespective of underlying obstetric disorders except for placenta previa (data not shown). Similarly, there was a significant negative correlation (P < 0.001) between the volume of FFP transfused and fibrinogen concentration (Figure 4). This significant negative correlation was shown irrespective of underlying obstetric disorders except for placenta previa and HELLP syndrome (data not shown).

3.5.2. Correlation between the Volume of FFP Transfusion and %PT or Fibrinogen Concentration Also in FFP transfusion, there was a significant negative correlation (P < 0.001) between the volume of FFP transfused and %PT immediately before transfusion (Figure 4). This significant negative correlation was shown irrespective of underlying obstetric disorders except for placenta previa (data not shown). Similarly, there was a significant negative correlation (P < 0.001) between the volume of FFP transfused and fibrinogen concentration (Figure 4). This significant negative correlation was shown irrespective of underlying obstetric disorders except for placenta previa and HELLP syndrome (data not shown). 4. Discussion From 2004 to 2008, the incidence of blood product transfusion for obstetric patients in our tertiary perinatal institution was 4.6% (243/5,311), which was much higher than previously reported incidences, 0.3–1% [2–4]. In these patients who had blood product transfusion, the percentage of patients with 6 or more units of RCC was 57% (138/243), which was also higher than previously reported, 32–42% [2, 4, 24]. This may be because our institution is only general medical institution for tertiary perinatal care in Saitama prefecture with population of 7.2 million and has accepted obstetric patients with life-threatening conditions.

nits of RCC was 57% (138/243), which was also higher than previously reported, 32–42% [2, 4, 24]. This may be because our institution is only general medical institution for tertiary perinatal care in Saitama prefecture with population of 7.2 million and has accepted obstetric patients with life-threatening conditions. Of the 220 patients with obstetric hemorrhage who underwent blood product transfusion (Table 3), FFP was transfused for 92.3% (203/220) (Table 4), suggesting importance of coagulation factors in blood transfusion for obstetric hemorrhage. In the patients with obstetric hemorrhage who underwent only allogenic transfusion, there was a significant positive correlation between the volume of RCC transfused and that of FFP (Figure 1) irrespective of underlying obstetric disorders (Table 5). A ratio of total transfused units of FFP to RCC and the median of FFP/RCC ratio in each patient was 2.1 and 2.0 (Table 5), respectively. Moreover, the median FFP/RCC ratio was 2.0 and 2.1 in subgroups with moderate and massive RCC transfusion, respectively (Table 6). Since two units of RCC and 3 units of FFP are derived from 400 mL of whole blood, respectively, in this study, the RCC : FFP ratio of 1 : 2.0–2.1 in units is equivalent to 1 : 1.3–1.4 when these volumes are converted to whole blood. This RCC : FFP ratio of 1 : 1.3–1.4 is almost consistent with the report of Borgman et al. who recommended the transfusion of RCC and plasma at a ratio of 1 : 1.4 for massive hemorrhage [25].

study, the RCC : FFP ratio of 1 : 2.0–2.1 in units is equivalent to 1 : 1.3–1.4 when these volumes are converted to whole blood. This RCC : FFP ratio of 1 : 1.3–1.4 is almost consistent with the report of Borgman et al. who recommended the transfusion of RCC and plasma at a ratio of 1 : 1.4 for massive hemorrhage [25]. To evaluate possible differences among obstetric hemorrhagic disorders in their pathophysiological condition which may affect the volumes of RCC and FFP transfused, we compared Hb concentration, %PT, aPTT, and fibrinogen concentration immediately before blood transfusion. Placental abruption causes dissection of blood at the decidual-placental interface, resulting in entry of placental tissue factor into the circulation to promote thrombin generation which eventually leads to disseminated intravascular coagulation (DIC) [26]. As shown in Figure 2, fibrinogen concentration is significantly lower in placental abruption compared with other obstetric disorders, suggesting severer coagulopathy than other obstetric disorders.

he circulation to promote thrombin generation which eventually leads to disseminated intravascular coagulation (DIC) [26]. As shown in Figure 2, fibrinogen concentration is significantly lower in placental abruption compared with other obstetric disorders, suggesting severer coagulopathy than other obstetric disorders. As mentioned previously, accurate evaluation of blood loss is difficult in obstetric hemorrhage [10–12], so comprehensive evaluation of not only blood loss but also the cause of hemorrhage and blood test data is required to determine the timing and volume of blood transfusion [15]. Moreover, our present data lack some other important variables prior to blood transfusion, such as rates and volumes of IV fluid resuscitation and use of surgical or pharmacologic interventions. Nonetheless, as shown in Figure 3, there was a significant negative correlation between the volume of RCC transfused and Hb concentration immediately before transfusion. This significant negative correlation was shown in all obstetric disorders except for placenta previa. This may be because for some patients with placenta previa the interval between onset of hemorrhage and blood test was so short that data did not reflect blood loss, and the transfusion volume was determined by intraoperative blood loss count.

Reversed end-diastolic flow (REDF) is the most advanced stage of abnormal umbilical artery Doppler flow and represents obliteration of nearly 70% of the placental function [3]. REDF also represents a higher risk of NICU admission, need for respiratory support, and perinatal mortality, regardless of age at delivery [4]. As placental function declines, the changes noted in fetal venous Doppler studies represent major changes in the fetal circulation in response to hypoxia. The increase in placental resistance leads to an obliteration of small muscular placental arteries, which leads to a decrease in the diastolic flow in the umbilical artery Doppler. The fetus responds with an increase in red blood cell mass and shunting of blood to nonessential vascular beds in order to increase oxygen utilization [5, 6]. This results in preferential cardiac and cerebral blood flow, with reduced blood flow to the rest of the body [7, 8]. As this process continues, the fetal right ventricular afterload increases in the setting of myocardial impairment, and changes in the fetal ductus venosus can often be visualized as a late and ominous finding [9].

correlation was shown in all obstetric disorders except for placenta previa. This may be because for some patients with placenta previa the interval between onset of hemorrhage and blood test was so short that data did not reflect blood loss, and the transfusion volume was determined by intraoperative blood loss count. Similarly, there was a significant negative correlation between the volume of FFP transfused and %PT or blood fibrinogen concentration (Figure 4), showing that patients with severer coagulation factor depletion required a larger volume of FFP for its supplementation. Interestingly, there was a significant negative correlation also between the volume of RCC transfused and %PT as data of coagulation system tests (Figure 3), suggesting that coagulation dysfunction due to coagulation factor depletion may result in even more blood loss and increase in transfusion volume of not only FFP but also RCC. These results are consistent with studies showing an increase in blood loss due to secondary atonic bleeding unless coagulation factors are rapidly supplemented [27], further dilution of coagulation factors and blood loss resulting from RCC and extracellular fluid supplementation alone without coagulation factors to pregnant and puerperal women with MOH [28], an increase in blood loss along with a decrease in coagulation factors, particularly with blood fibrinogen level less than 200 mg/dL [29], and the indispensability of coagulation factor supplementation when the blood fibrinogen level is ≤100 mg/dL [30]. Appropriate supplementation of coagulation factors normalizes the coagulation function in the early stage and reduces not only blood loss but also the volume of blood product transfusion [22, 27, 28, 31].

and the indispensability of coagulation factor supplementation when the blood fibrinogen level is ≤100 mg/dL [30]. Appropriate supplementation of coagulation factors normalizes the coagulation function in the early stage and reduces not only blood loss but also the volume of blood product transfusion [22, 27, 28, 31]. According to Figures 3 and 4, it is also noted that considerable number of patients with Hb, %PT, and fibrinogen values in the normal range were transfused with RCC and/or FFP. This may be because above-mentioned comprehensive evaluation of blood loss, cause of hemorrhage, vital signs, underlying disorders, and so forth has led to the clinical decision for blood product transfusion before the deterioration of their blood test values. On the other hand, Table 6 shows that 24 (12%) patients received FFP without RCC. In these patients consisting of 5 with placental abruptions, 7 with uterine atonies, 1 with genital tract trauma, and 11 with HELLP syndromes, RCC transfusion was not required since coagulation factors were promptly supplemented with FFP and their Hb levels could be maintained above 7 g/dL. Although some retrospective analyses reported that a percentage of patients were inappropriately transfused [24, 32], we believe that prompt decision making is inevitable to avoid secondary atonic bleeding and DIC especially in life-threatening obstetric hemorrhage.

and their Hb levels could be maintained above 7 g/dL. Although some retrospective analyses reported that a percentage of patients were inappropriately transfused [24, 32], we believe that prompt decision making is inevitable to avoid secondary atonic bleeding and DIC especially in life-threatening obstetric hemorrhage. In conclusion, for massive obstetric hemorrhage where appropriate supplementation of coagulation factors is essential, the transfusion of RCC : FFP = 1 : 1.3–1.4 in terms of whole blood is desirable according to our retrospective analysis as well as previous report [25]. Acknowledgments The authors thank all the doctors in their department for their great contributions to patient management and this study, and doctors in their district for their prompt referral and initial management. Figure 1 Correlation between the volume of RCC and FFP in obstetric hemorrhage patients with allogenic transfusion (n = 196). A significant positive correlation was observed between the volume of RCC and that of FFP, as also shown in Table 5. Patients who underwent autologous transfusion were excluded. An outlier with amniotic fluid embolism who had 50 units of RCC and 116 units of FFP was also excluded.

with allogenic transfusion (n = 196). A significant positive correlation was observed between the volume of RCC and that of FFP, as also shown in Table 5. Patients who underwent autologous transfusion were excluded. An outlier with amniotic fluid embolism who had 50 units of RCC and 116 units of FFP was also excluded. Figure 2 Hemoglobin (Hb) concentration, percent prothrombin activity (%PT), activated partial thromboplastin time (aPTT), and fibrinogen concentration immediately before blood transfusion in each obstetric hemorrhagic disorder (n = 211). Box plot graphs represent the median value as well as the upper and lower quartiles. The line across the middle of the box identifies the median sample value. The whiskers extend from the ends of the box to the outermost data point. The thick lines denote the mean values. Different superscript letters (a, b, c) denote significant difference at P < 0.05. Patients were excluded as mentioned in the text. Figure 3 Correlation between the volume of RCC transfused and Hb concentration or %PT in obstetric hemorrhage patients (n = 187). A significant negative correlation was observed between the volume of RCC transfused and Hb (a), or RCC and %PT (b). Patients were excluded as mentioned in the text. Figure 4 Correlation between the volume of FFP transfused and %PT or fibrinogen concentration in obstetric hemorrhage patients (n = 187). A significant negative correlation was observed between the volume of FFP transfused and %PT (a), or FFP and fibrinogen (b). Patients were excluded as mentioned in the text.

Figure 3 Correlation between the volume of RCC transfused and Hb concentration or %PT in obstetric hemorrhage patients (n = 187). A significant negative correlation was observed between the volume of RCC transfused and Hb (a), or RCC and %PT (b). Patients were excluded as mentioned in the text. Figure 4 Correlation between the volume of FFP transfused and %PT or fibrinogen concentration in obstetric hemorrhage patients (n = 187). A significant negative correlation was observed between the volume of FFP transfused and %PT (a), or FFP and fibrinogen (b). Patients were excluded as mentioned in the text. Table 1 Our transfusion management principles. (1) While evaluating the bleeding state, consumed coagulation factors are rapidly supplemented until the coagulation function normalizes [9, 22] (2) With the above as a goal, 4–6 units of FFP are transfused at a time, and the coagulation function is evaluated after each transfusion (3) Stabilization of the vital signs (4) RCC transfusion is performed to achieve an Hb level of 7-8 g/dL as a goal (5) PC transfusion is performed to achieve a platelet count above 50,000/mm3 as a goal (6) Cryoprecipitate, as well as specific coagulation factor preparations, is not administered in general Table 2 Demographics of obstetric patients with blood transfusion (n = 243). Age in years, mean ± SD 32.1 ± 4.7 Gestational age in weeks, mean ± SD 35.3 ± 4.9 Primipara, n (%) 95 (39.1) Multiple pregnancy, n (%) 14 (5.8) Cesarean delivery, n (%) 164 (67.5) Assisted vaginal delivery, n (%) 26 (10.7) Table 3 Obstetric hemorrhagic disorders with blood transfusion (n = 220).

Table 2 Demographics of obstetric patients with blood transfusion (n = 243). Age in years, mean ± SD 32.1 ± 4.7 Gestational age in weeks, mean ± SD 35.3 ± 4.9 Primipara, n (%) 95 (39.1) Multiple pregnancy, n (%) 14 (5.8) Cesarean delivery, n (%) 164 (67.5) Assisted vaginal delivery, n (%) 26 (10.7) Table 3 Obstetric hemorrhagic disorders with blood transfusion (n = 220). n (%) Uterine atony 57 (25.9) Genital tract trauma including uterine rupture/injury 51 (23.2) Placental abruption 48 (21.8) Placenta previa without acreta/increta/percreta 30 (13.6) Placenta previa with acreta/increta/percreta 13 (5.9) Uterine inversion 5 (2.3) HELLP syndrome 15 (6.8) Amniotic fluid embolism 1 (0.5) Table 4 Blood products transfused for obstetric hemorrhage (n = 220). Blood product n (%) Median units (range) Red cell concentrate 188 (85.5) 8 (2–50) Fresh frozen plasma 203 (92.3) 14 (2–116) Platelet concentrate 62 (28.2) 20 (10–80) Autologous whole blood 24 (10.9) 3 (1–8) Table 5 Significant positive correlation between RCC and FFP and an FFP/RCC ratio in each obstetric hemorrhagic disorder. n Spearman's rank correlation coefficient (ρ) P FFP/RCC* Uterine atony/inversion 62 0.7843 <0.0001 1.5 (1.2–2.5) Genital tract trauma 51 0.7841 <0.0001 2.3 (1.5–2.9) Placental abruption 48 0.7818 <0.0001 2.3 (1.5–3.0) Placenta previa 19 0.7765 <0.0001 2.0 (1.1–2.1) HELLP syndrome 15 0.5290 0.0426 2.0 (1.9–2.8) Total 196 0.7769 <0.0001 2.0 (1.4–2.5) *Data shown are median (interquartile range).

n Spearman's rank correlation coefficient (ρ) P FFP/RCC* Uterine atony/inversion 62 0.7843 <0.0001 1.5 (1.2–2.5) Genital tract trauma 51 0.7841 <0.0001 2.3 (1.5–2.9) Placental abruption 48 0.7818 <0.0001 2.3 (1.5–3.0) Placenta previa 19 0.7765 <0.0001 2.0 (1.1–2.1) HELLP syndrome 15 0.5290 0.0426 2.0 (1.9–2.8) Total 196 0.7769 <0.0001 2.0 (1.4–2.5) *Data shown are median (interquartile range). Table 6 FFP transfusion volume and an FFP/RCC ratio in obstetric hemorrhage patients with minimal, moderate, and massive RCC transfusion. RCC transfusion n FFP (units)* FFP/RCC* None 24 5 (4–8) — Minimal (2–4 units) 49 6 (4–8) 1.5 (1.0–3.0) Moderate (6–9 units) 53 14 (10–20) 2.0 (1.3–2.7) Massive (10- units) 70 30 (20–42) 2.1 (1.5–2.5) *Data shown are median (interquartile range).

1. Introduction Placental abruption or premature separation of the normally implanted placenta is a serious and life-threatening obstetric complication for both mother and fetus [1–3]. Although the cause of placental abruption remains elusive, the presence of inflammation and infection has been suggested to be the primary cause of placental abruption [2, 4–8]. Some previous studies have observed an association between intrauterine infection, especially chorioamnionitis (CAM), and placental abruption [2, 4–8]. In addition, evidence from prior studies suggests that women exposed to prolonged preterm premature rupture of membranes (p-PROM) are at increased risk of placental abruption [9–11], because recent evidence has linked neutrophil infiltration into the deciduas with preterm placental abruption [2, 7]. In our earlier studies [12, 13], for example, the incidence of preterm delivery, p-PROM, and low birth weight in the cases of placental abruption with chorioamnionitis were higher than in cases without chorioamnionitis: however there were no significant differences in the incidence of other outcomes between the cases of placental abruption with and without histological chorioamnionitis. However, there have been few examinations concerning the influence of precedent p-PROM on the severity of placental abruption at preterm only. In this study, we examined the obstetric and neonatal outcomes of preterm singleton pregnancies complicated by placental abruption following p-PROM compared with those without p-PROM.

1. Introduction Placental abruption or premature separation of the normally implanted placenta is a serious and life-threatening obstetric complication for both mother and fetus [1–3]. Although the cause of placental abruption remains elusive, the presence of inflammation and infection has been suggested to be the primary cause of placental abruption [2, 4–8]. Some previous studies have observed an association between intrauterine infection, especially chorioamnionitis (CAM), and placental abruption [2, 4–8]. In addition, evidence from prior studies suggests that women exposed to prolonged preterm premature rupture of membranes (p-PROM) are at increased risk of placental abruption [9–11], because recent evidence has linked neutrophil infiltration into the deciduas with preterm placental abruption [2, 7]. In our earlier studies [12, 13], for example, the incidence of preterm delivery, p-PROM, and low birth weight in the cases of placental abruption with chorioamnionitis were higher than in cases without chorioamnionitis: however there were no significant differences in the incidence of other outcomes between the cases of placental abruption with and without histological chorioamnionitis. However, there have been few examinations concerning the influence of precedent p-PROM on the severity of placental abruption at preterm only. In this study, we examined the obstetric and neonatal outcomes of preterm singleton pregnancies complicated by placental abruption following p-PROM compared with those without p-PROM. 2. Patients and Methods The protocol for this paper was approved by the Ethics Committee of the Japanese Red Cross Katsushika Maternity Hospital. In addition, informed consent concerning analysis from a retrospective database was obtained from each subject at their first hospital visit.

In this study, we examined the obstetric and neonatal outcomes of preterm singleton pregnancies complicated by placental abruption following p-PROM compared with those without p-PROM. 2. Patients and Methods The protocol for this paper was approved by the Ethics Committee of the Japanese Red Cross Katsushika Maternity Hospital. In addition, informed consent concerning analysis from a retrospective database was obtained from each subject at their first hospital visit. All subjects in this study had received care at Japanese Red Cross Katsushika Maternity Hospital between April 2002 and March 2011. We reviewed the obstetric records of 95 singleton deliveries complicated by placental abruption, defined as complete or partial separation of a normally implanted placenta indicated by evidence of retro-placental bleeding at 22–36 weeks' gestation. (In our hospital, there were 65 cases complicated by placental abruption after 37 weeks' gestation during the 9-year period.) We excluded the cases referred from other hospitals after the onset of placental abruption. In this paper, we examined the incidence of hypertensive disorders such as gestational hypertension and preeclampsia, emergency Cesarean delivery, disseminated intravascular coagulation (DIC), maternal blood loss requiring hemotransfusion, small-for-gestational-age infants, fetal demise, nonreassuring fetal status (NRFS), Apgar score <4 at 1 and 5 minutes and umbilical artery pH < 7. Infants who were small-for-gestational-age were defined as those with sex- and age-adjusted birth weights below the tenth percentile according to the neonatal birth weight standards for gestational age in Japanese [14]. In addition, microscopic histological analyses of the placentas were performed to diagnose chorioamnionitis (CAM). The severity of CAM, that is, inflammation of the placental surface, was determined by the degree of maternal polymorphonuclear lymphocyte infiltration into either the subchorionic space (intervillositis: stage I), the intervillous space (chorionitis: stage II) or the amniotic cavity (CAM in a narrow sense: stage III) according to Blanc's criteria [15].

mmation of the placental surface, was determined by the degree of maternal polymorphonuclear lymphocyte infiltration into either the subchorionic space (intervillositis: stage I), the intervillous space (chorionitis: stage II) or the amniotic cavity (CAM in a narrow sense: stage III) according to Blanc's criteria [15]. 2.1. Analysis Data are presented as number (%) or mean ± SD. For statistical analysis, the χ 2 test with Yates' correction for categorical variables was used. While the Student's t-test for continuous variables was used. Odds ratios (ORs) and 95% confidence intervals (CIs) were also calculated. Differences with P < 0.05 were considered significant. 3. Results Of the 95 cases of placental abruption in deliveries at 22–36 weeks' gestation, 64 cases (67.4%) occurred without p-PROM and 31 cases (32.6%) occurred following p-PROM. During the 9-year period, there were 17,667 singleton deliveries after 22 weeks' gestation in our hospital. Of these, 655 cases were complicated by p-PROM (3.7%). In our hospital, therefore, the incidence of placental abruption in singleton pregnancies with p-PROM was 4.7% and the crude OR of placental abruption for women following p-PROM was 6.50 (95% CI: 4.4–9.7, P < 0.01).

veries after 22 weeks' gestation in our hospital. Of these, 655 cases were complicated by p-PROM (3.7%). In our hospital, therefore, the incidence of placental abruption in singleton pregnancies with p-PROM was 4.7% and the crude OR of placental abruption for women following p-PROM was 6.50 (95% CI: 4.4–9.7, P < 0.01). Table 1 shows the perinatal outcomes of preterm singleton pregnancies complicated by placental abruption with and without p-PROM. The incidence of histological CAM stage III in the patients following p-PROM was significantly higher than that in the patients without p-PROM (crude OR: 5.22, 95% CI: 1.4–19, P = 0.02). On the other hand, the rate of emergency Cesarean deliveries due to NRFS in the patients following p-PROM was significantly lower than that in the patients without p-PROM (emergency Cesarean delivery, crude OR: 0.13, 95% CI: 0.05–0.33, P < 0.01; NRFS, crude OR: 0.34, 95% CI: 0.13–0.87, P = 0.04). However, there were no significant differences in the maternal and neonatal outcomes between the patients with and without p-PROM. 4. Discussion The major findings of the current study are: (1) the preterm singleton pregnancies complicated by placental abruption following p-PROM was strongly associated with the presence of histological CAM more than those without p-PROM, and (2) the perinatal outcomes of preterm placental abruption following p-PROM were not different from those without p-PROM at preterm.

(1) the preterm singleton pregnancies complicated by placental abruption following p-PROM was strongly associated with the presence of histological CAM more than those without p-PROM, and (2) the perinatal outcomes of preterm placental abruption following p-PROM were not different from those without p-PROM at preterm. Some previous studies have reported that p-PROM is a common obstetric complication, occurring in approximately 1-2% of pregnancies, and it is one of important risk factors for placental abruption [9–11]. Our current results also support these previous studies.

(1) the preterm singleton pregnancies complicated by placental abruption following p-PROM was strongly associated with the presence of histological CAM more than those without p-PROM, and (2) the perinatal outcomes of preterm placental abruption following p-PROM were not different from those without p-PROM at preterm. Some previous studies have reported that p-PROM is a common obstetric complication, occurring in approximately 1-2% of pregnancies, and it is one of important risk factors for placental abruption [9–11]. Our current results also support these previous studies. Histological CAM, defined as inflammation of the extraplacental membrane, has been consistently linked with prematurity and low birth weight of neonates [16]. In addition, the relationship between histological CAM and infection (positive culture) of the chorioamnion has been reported to be strongest among preterm deliveries [17, 18]. In cases with histological CAM, because the prematurely delivered placentas have been observed to be often accompanied by an acute marginal hemorrhage that undermines the edge of the placenta and that originates from deciduitis [2]. This hemorrhage process can cause premature labor and/or p-PROM and has been reported to differ from the typical placental abruption due to other causes such as preeclampsia [5]. Vintzileos et al. [8] also suggested that true placental abruption following the presence of CAM usually occurs after PROM. On the other hand, Nelson et al. [9] speculated that an acute reduction in the uterine volume and intrauterine surface as a consequence of p-PROM could ultimately lead to disruption of the site of placental attachment in the decidual spongiosa layer, thereby predisposing to abruption. In this study, unfortunately, we could not examine the intervals between p-PROM and onset of placental abruption. However, the progress of placental abruption in the cases following p-PROM may not tend to be acute; because the rate of cases with NRFS requiring cesarean operation in women following p-PROM was lower than those without p-PROM. Therefore, our results support the previous studies suggesting the association among CAM, p-PROM, and placental abruption [10, 11].

tion in the cases following p-PROM may not tend to be acute; because the rate of cases with NRFS requiring cesarean operation in women following p-PROM was lower than those without p-PROM. Therefore, our results support the previous studies suggesting the association among CAM, p-PROM, and placental abruption [10, 11]. In this study, the perinatal outcomes of preterm placental abruption following p-PROM were not different from those without p-PROM at preterm, although the rate of emergency Cesarean delivery due to NRFS in cases following p-PROM was lower than in those without p-PROM. One reason may be the small sample size in this study. The other possible reason is that a more rapid Cesarean delivery might tend to be carried out in cases without PROM due to more typical symptoms, (bleeding, abdominal pain, and NRFS) and more easy diagnosis of placental abruption compared with cases following PROM [19]. Because rupture of membranes (amniotomy) has been supposed to decrease bleeding from the implantation site of the placenta and reduce entry of thromboplastin into the maternal circulation [19]. In our series, therefore, the influence of placental abruption on maternal and fetal conditions might be larger in cases without PROM than those following PROM; however, a rapid delivery of the fetus by Cesarean section might prevent serious complications in many cases without PROM. In conclusion, although p-PROM may be one of important risk factors for placental abruption associated with CAM, it may not influence the perinatal outcomes in preterm placental abruption.

In this study, the perinatal outcomes of preterm placental abruption following p-PROM were not different from those without p-PROM at preterm, although the rate of emergency Cesarean delivery due to NRFS in cases following p-PROM was lower than in those without p-PROM. One reason may be the small sample size in this study. The other possible reason is that a more rapid Cesarean delivery might tend to be carried out in cases without PROM due to more typical symptoms, (bleeding, abdominal pain, and NRFS) and more easy diagnosis of placental abruption compared with cases following PROM [19]. Because rupture of membranes (amniotomy) has been supposed to decrease bleeding from the implantation site of the placenta and reduce entry of thromboplastin into the maternal circulation [19]. In our series, therefore, the influence of placental abruption on maternal and fetal conditions might be larger in cases without PROM than those following PROM; however, a rapid delivery of the fetus by Cesarean section might prevent serious complications in many cases without PROM. In conclusion, although p-PROM may be one of important risk factors for placental abruption associated with CAM, it may not influence the perinatal outcomes in preterm placental abruption. Table 1 Obstetric complications and perinatal outcomes of preterm singleton pregnancies complicated by placental abruption with and without preterm premature rupture of membranes (p-PROM). p-PROM P value* (−) (+) N 60 42 Maternal age (years) 31.9 ± 4.4 30.5 ± 4.6 0.16 Parity 0.8 ± 1.1 0.7 ± 0.7 0.59 Multiparous 32 (50.0) 18 (58.1) 0.61

Table 1 Obstetric complications and perinatal outcomes of preterm singleton pregnancies complicated by placental abruption with and without preterm premature rupture of membranes (p-PROM). p-PROM P value* (−) (+) N 60 42 Maternal age (years) 31.9 ± 4.4 30.5 ± 4.6 0.16 Parity 0.8 ± 1.1 0.7 ± 0.7 0.59 Multiparous 32 (50.0) 18 (58.1) 0.61 Gestational age at delivery Average (weeks) 33.3 ± 2.6 32.5 ± 3.5 0.26 <32 weeks 12 (18.8) 10 (32.3) 0.23 Hypertensive disorders 6 (9.4) 0 (0.0) 0.19 Emergency Cesarean delivery 52 (81.3) 11 (35.5) <0.01 DIC 12 (18.8) 1 (3.2) 0.08 Blood loss requiring transfusion 13 (20.3) 3 (9.7) 0.34 Histological chorioamnionitis** Stage II 14 (21.9) 9 (29.0) 0.61 Stage III 4 (6.3) 8 (25.8) 0.02 Neonatal birth weight Average (g) 2,006 ± 544 1,891 ± 642 0.39 Small for gestational age 11 (17.2) 6 (19.4) 0.98 Fetal demise 9 (14.1) 1 (3.2) 0.21 Surviving fetuses/neonates n 55 (85.9) 30 (96.8) 0.21 NRFS 41 (74.5) 15 (50.0) 0.04 Apgar 1′ < 4 14 (25.5) 2 (6.7) 0.07 Apgar 5′ < 4 4 (7.3) 1 (3.3) 0.77 Umbilical artery pH < 7 9 (16.4) 1 (3.3) 0.14 Values are expressed as n (%) or mean ± SD. *P values by Student's t-test or χ 2 test. **Microscopic histological analysis of the placenta. p-PROM: preterm premature rupture of membranes. DIC: disseminated intravascular coagulation. NRFS: nonreassuring fetal status.

1. Introduction Deep infiltrating endometriosis (DIE) is a particular form of endometriosis that extends >5 mm under the peritoneal surface [1]. These lesions develop in the form of retroperitoneal nodules that consist histologically of endometrial epithelium and stroma, surrounded by muscular hyperplasia and fibrosis [2]. DIE nodules are rich in nerve fibers [3] and are commonly associated with severe cyclic or acyclic pelvic pain such as dysmenorrhea, deep dyspareunia, and nonmenstrual pain and organ-specific symptoms related to bladder or intestinal dysfunction (dyschezia, constipation, diarrhea, rectal bleeding, frequency of micturition, and hematuria) [4–6]. Radical surgical exeresis of DIE lesions is the mainstay of treatment for this form of endometriosis. Medical therapies may temporarily alleviate painful symptoms, but recurrence rates after their discontinuation are high [7, 8]. Furthermore, performing inadequate primary surgery not only results in disease progression with persistence or aggravation of painful symptoms but also renders any future procedure difficult and potentially dangerous [9, 10].

alleviate painful symptoms, but recurrence rates after their discontinuation are high [7, 8]. Furthermore, performing inadequate primary surgery not only results in disease progression with persistence or aggravation of painful symptoms but also renders any future procedure difficult and potentially dangerous [9, 10]. DIE nodules represent a real operative challenge due to common involvement of vital retroperitoneal structures (ureter, bowel, vessels, and nerves). Laparoscopy appears to be the ideal tool to perform such surgery, offering the advantages of magnification, accurate hemostasis, precise dissection, and careful handling of delicate tissue. Nevertheless, laparoscopic management of retroperitoneal endometriosis should not be undertaken by inexperienced operators and thorough knowledge of pelvic retroperitoneal anatomy is a prerequisite for radical and uncomplicated removal of DIE nodules. Posterior nodules represent the commonest form of deep infiltrating endometriosis [11, 12]. Their radical exeresis may require extensive retroperitoneal surgery which may include resection of the uterosacral ligaments (USL), partial colpectomy, and resection of rectal disease. A multidisciplinary surgical approach may be necessary involving a urologist or a bowel surgeon with expertise in advanced laparoscopic surgery to successfully conclude the procedure [13, 14].

al surgery which may include resection of the uterosacral ligaments (USL), partial colpectomy, and resection of rectal disease. A multidisciplinary surgical approach may be necessary involving a urologist or a bowel surgeon with expertise in advanced laparoscopic surgery to successfully conclude the procedure [13, 14]. Although laparoscopic excision of large DIE nodules may become a quite unpredictable operation in terms of following distinct and timely operative steps, an effort should be made during one's learning curve to develop a standardized way of approaching such lesions. We conducted this study in an effort to standardize our laparoscopic technique of excision of posterior DIE according to the size, location, and geometry of the lesion, attempting at the same time to develop a rough guide for relatively inexperienced surgeons embarking on the surgical management of posterior DIE nodules.

conducted this study in an effort to standardize our laparoscopic technique of excision of posterior DIE according to the size, location, and geometry of the lesion, attempting at the same time to develop a rough guide for relatively inexperienced surgeons embarking on the surgical management of posterior DIE nodules. 2. Materials and Methods From September 2008 to July 2011 we recruited for this study 40 consecutive patients with a prospective diagnosis of DIE based on their symptoms, clinical examination, and preoperative workup. Institutional review board approval was obtained for this study, and a detailed informed consent was signed by all women. All patients were initially referred to our institution for chronic pelvic pain (CPP), infertility, or the presence of endometriotic ovarian cysts and, in the process of their evaluation, were found with a pelvic nodule suggestive of DIE. All women completed a detailed pain, sexual function, and quality of life (QoL) questionnaire [15] and underwent a thorough pelvic examination (i.e., vaginal and rectal). A transvaginal (TVS) and/or transrectal (TRS) ultrasound scan was performed to assess the adnexa and uterus and to estimate the size and exact position of the nodule in relation to the wall of the rectosigmoid and the pelvic sidewall. Magnetic resonance imaging (MRI) of the pelvis was also performed in cases with dubious findings and in cases with large lesions, to obtain more detailed information on the exact geometry of the nodule. For posterolateral lesions an upper abdominal scan was performed to investigate the possibility of ureteral involvement resulting in hydronephrosis by the disease process. Cases with considerable disturbances of bowel function were submitted to rectosigmoidoscopy. Preoperative CA-125 and CA-19-9 levels were also measured in all cases.

al lesions an upper abdominal scan was performed to investigate the possibility of ureteral involvement resulting in hydronephrosis by the disease process. Cases with considerable disturbances of bowel function were submitted to rectosigmoidoscopy. Preoperative CA-125 and CA-19-9 levels were also measured in all cases. All patients were scheduled for a laparoscopic procedure outside their days of menstruation. No medical ovarian suppression such as GnRH analogs was administered preoperatively. Peritoneal entry and the pneumoperitoneum were established following a standardized transumbilical blind technique using the Veress needle. In cases with a history of previous gynecological or lower abdominal surgery, initial entry was achieved through the left subcostal area, followed by placement of a 10 mm trocar for the optic through the umbilicus. Three 5 mm accessory trocars for the laparoscopic instruments were used, placed in a standardized fashion above the pubic hairline [14].

ious gynecological or lower abdominal surgery, initial entry was achieved through the left subcostal area, followed by placement of a 10 mm trocar for the optic through the umbilicus. Three 5 mm accessory trocars for the laparoscopic instruments were used, placed in a standardized fashion above the pubic hairline [14]. During the diagnostic phase of the procedure, pelvic endometriosis was staged according to the revised American Fertility Society (rAFS) classification [16] and DIE according to the ENZIAN classification [17]. The laparoscopic procedure to treat ovarian and superficial endometriosis and to perform pelvic adhesiolysis followed certain operative rules in a timely fashion and was common for all types of DIE nodules. Excision of DIE nodules followed completion of these initial operative steps. Both posterolateral and central nodules were approached step by step, recording each operative step according to its absolute necessity for the procedure to advance smoothly. Recorded technical parameters included the necessity for ovarian and/or sigmoid suspension, systematic ureteral dissection, removal of parts of the pelvic peritoneum, division of large uterine vascular branches, partial colpectomy, and rectosigmoid reinforcement suturing. Other recorded parameters included intraoperative blood loss, length of the procedure, and hospitalization times. All excised specimens were sent for histology. A prerequisite for a case to be included in this study was the histologic confirmation of the presence of endometriosis (epithelium and stroma) in the excised DIE nodule.

ded parameters included intraoperative blood loss, length of the procedure, and hospitalization times. All excised specimens were sent for histology. A prerequisite for a case to be included in this study was the histologic confirmation of the presence of endometriosis (epithelium and stroma) in the excised DIE nodule. 2.1. Operative Technique In all cases the operation began with adhesiolysis commencing at the level of the pelvic brim, following a direction from top to bottom and laterally to medially. Congenital sigmoid adhesions were routinely divided in order to facilitate rectosigmoid suspension, should this appeared necessary during the procedure. Both adnexae were mobilized with division of their adhesions with the ovarian fossa, and careful hemostasis was performed to avoid contamination of the operative field. In cases with endometriomas the cyst was evacuated and the cavity thoroughly washed for the same reason. Ovarian surgery, when necessary, was left to follow excision of the DIE. Following these common preparatory steps the DIE nodule-harboring area to be excised was selected. Selection was facilitated through careful intraoperative digital transvaginal and/or transrectal palpation performed by the surgeon. Suspension of either the adnexa or the rectosigmoid was decided at this point according to quality of the operative field obtained without the need to systematically occupy an instrument as a retractor. The ovarian and rectosigmoid suspension techniques used have already been described by others [14].

d by the surgeon. Suspension of either the adnexa or the rectosigmoid was decided at this point according to quality of the operative field obtained without the need to systematically occupy an instrument as a retractor. The ovarian and rectosigmoid suspension techniques used have already been described by others [14]. In cases with posterolateral nodules the retroperitoneal space was entered through a small peritoneal incision at a healthy area as close as possible to the nodule. The incision was enlarged along the periphery of the diseased peritoneum of the ovarian fossa, and the peritoneal area to be excised was selected. At this point it was also decided whether systematic ureteral dissection (Figure 1) appeared necessary. The decision depended on the following parameters: the degree of lateral fibrosis, the degree of peritoneal involvement, the degree of uterosacral ligament involvement, and the difficulty to recognize relevant retroperitoneal structures. Ureteral dissection was advanced distally towards the ureter's point of crossing with the uterine artery. Similarly, the decision to bypass, fully dissect, or sacrifice large uterine vascular branches (Figure 2) was made at that point, according to degree of their involvement by the DIE nodule. The uterosacral ligament was subsequently divided at the point of its insertion to the uterine cervix. The 2nd assistant's vaginal finger served as a guide to limit the incision to a healthy plane in relation to the vaginal fornix. At this point the decision to perform or not partial colpectomy was made. To achieve radical excision of the nodule both medial and lateral ipsilateral rectovaginal spaces were developed in all cases with posterolateral nodules. In cases with rectal wall involvement the shaving technique was always attempted to excise the nodule. Following its' removal a decision was made whether or not to reinforce the bowel wall with horizontal interrupted absorbable sutures.

ateral rectovaginal spaces were developed in all cases with posterolateral nodules. In cases with rectal wall involvement the shaving technique was always attempted to excise the nodule. Following its' removal a decision was made whether or not to reinforce the bowel wall with horizontal interrupted absorbable sutures. In cases with predominantly central lesions, the technique was modified according to presence of unilateral or bilateral involvement of the uterosacral ligaments. In the case of purely central nodules, opening of the retroperitoneal space was achieved from the least involved side by incising the peritoneum medially to the ipsilateral uterosacral ligament. The ipsilateral medial rectovaginal space was thoroughly dissected below the level of the lowest limit of the nodule. Similarly, the contralateral space was developed, as far down as possible. At this point a sponge was inserted in the posterior vaginal fornix and another in the rectum, to assist in the identification of the plane between these two organs. In cases without a recognizable plane, possibly suggestive of a nodule predominantly attached to the rectosigmoid, the procedure was advanced by incising the peritoneum lying below the uterine torus laterally to medially. At this point a decision was made to perform or not partial colpectomy depending on the degree of vaginal induration caused by the nodule. Although preoperative findings during examination under anesthesia were taken into account, colpectomy was by no means a predetermined operative step. Following colpotomy, the vaginal lesion was circumscribed by distally incising the vaginal mucosa, and the lesion was left attached to the rectosigmoid. The same applied to cases managed without colpotomy. In such patients multiple intraoperative vaginal examinations assisted in identifying the correct plane for vaginal dissection. Following complete mobilization of the nodule from the vagina with or without colpotomy the nodule was gradually detached from the rectosigmoid using the shaving technique (Figures 3(a) and 3(b)). The decision to reinforce the bowel wall with horizontal sutures depended on the degree of muscular wall involvement.

tion. Following complete mobilization of the nodule from the vagina with or without colpotomy the nodule was gradually detached from the rectosigmoid using the shaving technique (Figures 3(a) and 3(b)). The decision to reinforce the bowel wall with horizontal sutures depended on the degree of muscular wall involvement. In cases with central nodules bearing a lateral component which involved significantly the uterosacral ligament(s) the technique was modified according to geometry of the lateral extension. Similar operative steps and timely decisions were made as those described above, in order to optimally approach the DIE nodule. Occasionally, the order of decision making was changed and the operative steps were adapted to serve better the individual patient. Discoid rectal excision or rectosigmoidectomy, where necessary, was undertaken by a general surgeon with relevant expertise after the preparation of the operative field as described above by the laparoscopic gynecologic team. A trained urologist was also involved in cases where ureteral surgery other than ureterolysis was necessary. Follow-up examinations were scheduled at six monthly intervals for two years. During each appointment patients complete the pain, sexual function, and QoL questionnaire and receive a vaginal and rectal examination. TVS and/or TRS are performed in case of abnormal findings.

Discoid rectal excision or rectosigmoidectomy, where necessary, was undertaken by a general surgeon with relevant expertise after the preparation of the operative field as described above by the laparoscopic gynecologic team. A trained urologist was also involved in cases where ureteral surgery other than ureterolysis was necessary. Follow-up examinations were scheduled at six monthly intervals for two years. During each appointment patients complete the pain, sexual function, and QoL questionnaire and receive a vaginal and rectal examination. TVS and/or TRS are performed in case of abnormal findings. 2.2. Statistical Analysis Fisher's exact test was used to compare differences between frequencies of adnexal and rectosigmoid suspension, systematic ureteral dissection, removal of parts of the pelvic peritoneum, division of large uterine vascular branches, partial colpectomy, rectosigmoid reinforcement suturing, and rates of intraoperative and postoperative complications, according to location (predominantly central versus predominantly lateral) and size (≤2 cm versus >2 cm) of the DIE nodule. Student's 2-independent samples t-test was used to compare operative parameters expressed in numerical values such as intraoperative blood loss, operation times, and duration of hospitalization between groups. Significance level was set to P < 0.05.

redominantly lateral) and size (≤2 cm versus >2 cm) of the DIE nodule. Student's 2-independent samples t-test was used to compare operative parameters expressed in numerical values such as intraoperative blood loss, operation times, and duration of hospitalization between groups. Significance level was set to P < 0.05. 3. Results Patient characteristics are presented in Table 1. Four patients were excluded from further analysis. Of these, one was found with complete obliteration of the Pouch of Douglas by adhesions, but without real deeper involvement. Three further patients were laparoconverted. The first case was a 46-year-old patient, with a poorly mobile voluminous adenomyotic uterus combined with a 4 cm central DIE nodule, who was managed with modified radical hysterectomy. The second case had left ureteral obstruction with hydronephrosis due to a 3 cm nodule in whom excision of the involved segment and end-to-end ureteral reanastomosis was performed. The last case was a 29-year-old patient with a 3 cm hemorrhagic lesion invading the vaginal mucosa that were surprisingly found at frozen section to correspond to a primary serous papillary carcinoma of the peritoneum.

nodule in whom excision of the involved segment and end-to-end ureteral reanastomosis was performed. The last case was a 29-year-old patient with a 3 cm hemorrhagic lesion invading the vaginal mucosa that were surprisingly found at frozen section to correspond to a primary serous papillary carcinoma of the peritoneum. In Table 2 our cases are presented according to their ENZIAN classification taking into account the two most involved compartments. Figure 4 presents distribution of our cases according to size of the DIE nodule. For purposes of more meaningful analysis of our technical considerations, we grouped together (group 1, Figure 5(a)) nodules spreading along compartments a and c (predominantly central nodules). Spread along the horizontal plane b constituted the group with predominantly lateral nodules (group 2, Figure 5(b)). Suspension of the adnexa to facilitate exposure of the ovarian fossa and ipsilateral uterosacral ligament was more frequently necessary in cases of group 2 (91.7% versus 58.3%, resp., P = 0.029). On the contrary, suspension of the rectosigmoid was considered necessary for the procedure to advance smoothly in the majority of group 1 patients (66.7% versus 12.5%, resp., P = 0.002). The peritoneum of the ovarian fossa was the initial point of entry to the retroperitoneal space in all cases with lateral nodules compared with only 2 cases of the group with central nodules in whom the peritoneum overlying the lateral rectovaginal space was opened (100.0% versus 16.6%, resp., P < 0.001).

, P = 0.002). The peritoneum of the ovarian fossa was the initial point of entry to the retroperitoneal space in all cases with lateral nodules compared with only 2 cases of the group with central nodules in whom the peritoneum overlying the lateral rectovaginal space was opened (100.0% versus 16.6%, resp., P < 0.001). The ureter was systematically dissected down to its' crossing with the uterine artery in the majority of group 2 cases (85.7% versus 41.7%, resp., P = 0.007). Similarly, the diseased and fibrotic part of the peritoneum of the ovarian fossa was removed in all patients with lateral nodules, compared with less than half of patients with central nodules (100.0% versus 50.0%, resp., P < 0.001). During the process of liberating the nodule from its retroperitoneal attachments it was considered necessary to sacrifice the uterine vein in 3 of group 2 cases compared with none of group 1 (12.5%, versus 0.0%, resp., NS). On the contrary, colpectomy was considered necessary in 3 cases with central nodules compared with none of those with lateral lesions (25.0% versus 0.0%, resp., P = 0.031).

ents it was considered necessary to sacrifice the uterine vein in 3 of group 2 cases compared with none of group 1 (12.5%, versus 0.0%, resp., NS). On the contrary, colpectomy was considered necessary in 3 cases with central nodules compared with none of those with lateral lesions (25.0% versus 0.0%, resp., P = 0.031). The DIE nodule was firmly attached to the wall of the rectosigmoid in 5/12 (41.7%) of group 1 and in 7/24 (33.3%) of group 2 cases, respectively (Table 2). The shaving technique was performed in all but one patient. This last case corresponded to an E4c lesion according to the ENZIAN classification and was managed with rectosigmoidectomy of the affected segment and end-to-end reanastomosis. Bowel wall reinforcement sutures were considered necessary in 3 of group 1 cases compared to none of group 2 (37.5% versus 0.0%, resp., P = 0.011). Table 3 summarizes the above findings.

according to the ENZIAN classification and was managed with rectosigmoidectomy of the affected segment and end-to-end reanastomosis. Bowel wall reinforcement sutures were considered necessary in 3 of group 1 cases compared to none of group 2 (37.5% versus 0.0%, resp., P = 0.011). Table 3 summarizes the above findings. When grouping patients according to nodule size (group A: ≤2 cm, group B: >2 cm), almost all of the examined operative parameters were more frequently applied to patients of group B: adnexal suspension (100.0% versus 74.1%, resp., NS), suspension of the rectosigmoid (66.7% versus 18.5%, respectively, P = 0.012), systematic ureteral dissection (88.9% versus 66.7%, resp., NS), division of uterine vein (33.3% versus 0.0%, resp., P = 0.012), colpectomy (33.3% versus 0.0%, resp., P = 0.012), and placement of bowel wall reinforcement sutures (33.3% versus 0.0%, resp., P = 0.012). On the contrary, excision of pelvic peritoneum was more frequent in group A cases, but not significantly so compared to group B (81.5% versus 77.8%, resp., NS) (Table 4).

= 0.012), colpectomy (33.3% versus 0.0%, resp., P = 0.012), and placement of bowel wall reinforcement sutures (33.3% versus 0.0%, resp., P = 0.012). On the contrary, excision of pelvic peritoneum was more frequent in group A cases, but not significantly so compared to group B (81.5% versus 77.8%, resp., NS) (Table 4). Operation times were significantly longer in cases with larger (>2 cm) nodules. Similarly, blood loss was significantly more in cases with larger (>2 cm) and posterolateral nodules (Table 4). We had no intraoperative or immediate postoperative complications. Hospitalization times were not significantly different between groups. All patients included in this study have completed so far at least six months of followup. Complete resolution of CPP (all types) occurred in 75.0% of patients with posterolateral nodules versus 58.3% of those with central nodules and in 74.1% of patients with nodules ≤2 cm versus 55.5% of nodules >2 cm. These differences did not reach statistical significance. During patients' gynecological examination at 6-month followup we recorded presence of any vaginal scarring or induration. Patients with nodule size >2 cm were more likely to present with some degree of scarring compared to those with nodule size ≤2 cm (18.5% versus 66.7%, P = 0.012) (Table 4).

tatistical significance. During patients' gynecological examination at 6-month followup we recorded presence of any vaginal scarring or induration. Patients with nodule size >2 cm were more likely to present with some degree of scarring compared to those with nodule size ≤2 cm (18.5% versus 66.7%, P = 0.012) (Table 4). 4. Discussion Posterior deep infiltrating endometriosis nodules (DIE) represent an operative challenge due to common involvement of vital retroperitoneal structures. Careful dissection is necessary to restore pelvic anatomy and preserve function. Such surgery despite the fact that it may at times become of unpredictable complexity should follow certain rules and well defined operative steps, in order to proceed without serious complications. The above apply even more to the less experienced surgeon who builds up his/her learning curve in the treatment of such lesions. We conducted this study in an effort to develop a rough guide for the novice operator of DIE, taking into consideration the importance of size, location, and geometry of the nodule, in the process of selecting the appropriate technique to approach each type of lesion. We also feel that by identifying steps absolutely necessary for dissection of the retroperitoneum to advance smoothly, an inexperienced surgeon would benefit, by correcting weaknesses both theoretical and operative and by concentrating on practicing certain operative steps in less complicated cases.

ach type of lesion. We also feel that by identifying steps absolutely necessary for dissection of the retroperitoneum to advance smoothly, an inexperienced surgeon would benefit, by correcting weaknesses both theoretical and operative and by concentrating on practicing certain operative steps in less complicated cases. The ENZIAN staging system of DIE was published in 2005. It classifies DIE nodules according to their size and spread along three compartments of the pelvis, vertical, horizontal, and posterior [17]. The truth is that, excluding German speaking countries, the ENZIAN score has been poorly accepted by gynecologists due to the complexity of its documentation and to the absence of significant factors such as pain or infertility incorporated in this system [18, 19]. Nevertheless, we found this system an invaluable aid for the intraoperative development of a strategy to approach these DIE lesions. A simpler yet not descriptive surgical classification is that proposed by Chapron et al., which correlates the type of lesion to the type of procedure necessary to treat each case [12]. In our study we tried to design our surgery by creating two groups of patients: cases with nodules expanding in a predominant fashion laterally and cases with predominantly central nodules, irrespective of the type of their spread along the vertical and posterior axes. In our opinion the key point in attacking both types of posterior nodules is the invariable development of lateral and medial rectovaginal spaces in all DIE cases, a step which the novice should master during his/her learning curve.

odules, irrespective of the type of their spread along the vertical and posterior axes. In our opinion the key point in attacking both types of posterior nodules is the invariable development of lateral and medial rectovaginal spaces in all DIE cases, a step which the novice should master during his/her learning curve. Systematic ureteral dissection is a vital step especially when dealing with predominantly lateral (>1 cm, E2b/2bb–E4b/4bb) and larger (>2 cm, all compartments) nodules. Lateral retroperitoneal dissection frequently encounters large vascular structures which represent branches of the internal iliac artery and vein. It is absolutely essential to have a thorough and working knowledge of their anatomy, as it may become necessary to perform extensive vascular dissection with or without division of vessels incorporated in the DIE nodule. The surgeon involved should therefore possess such expertise or be prepared to call for a senior and more experienced assistant.

have a thorough and working knowledge of their anatomy, as it may become necessary to perform extensive vascular dissection with or without division of vessels incorporated in the DIE nodule. The surgeon involved should therefore possess such expertise or be prepared to call for a senior and more experienced assistant. The technique to manage central nodules is different and has been described in detail [14, 20–23]. We prefer the approach of attacking the vaginal part of the lesion first, leaving the nodule attached to the rectosigmoid, before applying the shaving technique for its excision [14]. The surgeon dealing with central and larger lesions (>2 cm) should have mastered laparoscopic suturing as the application of such skills may become necessary to reinforce or repair the bowel wall, when the shaving technique or discoid excision is performed. In this group of patients we did not have to perform discoid excision. When such or more advanced bowel surgery appears likely, a general surgeon with appropriate skills should be informed in advance, as in many countries such as in Greece, no legal cover exists so far for gynecologists performing intestinal tract procedures. The same applies to ureteral surgery other than ureterolysis.

. When such or more advanced bowel surgery appears likely, a general surgeon with appropriate skills should be informed in advance, as in many countries such as in Greece, no legal cover exists so far for gynecologists performing intestinal tract procedures. The same applies to ureteral surgery other than ureterolysis. Very large lesions (>3 cm) represent a different operative platform. In such cases all compartments are frequently involved in a variable manner and designing of surgery in a comprehensive and predictable way, as that described above, may not be of practical value [19]. In such cases, a full preoperative workup should be performed and a multidisciplinary approach is necessary from the start of the procedure to achieve radical excision of the nodule and avoid complications.

g of surgery in a comprehensive and predictable way, as that described above, may not be of practical value [19]. In such cases, a full preoperative workup should be performed and a multidisciplinary approach is necessary from the start of the procedure to achieve radical excision of the nodule and avoid complications. Performing routine colpectomy in cases with nodules involving the vaginal wall but sparing the vaginal mucosa appears to be a debatable issue [9, 14, 23, 24]. Recurrence rates of up to 25% have been reported [23], despite the fact that histologically proven endometriosis in the excised vaginal specimen may not exceed 10% [9]. Our technique of removing such nodules without opening the vagina involves a combination of monopolar and bipolar energy. We observed a significant percentage of limited and painless vaginal scaring at 6-month followup in patients treated with the above technique. This finding was more common in patients with central and larger (>2 cm) nodules. Interestingly, in several cases this scarring had subsided considerably at the 12-month followup examination. Whether postoperative asymptomatic vaginal scarring in the noncolpectomized group is associated with an increased risk of recurrence or represents development of fibrosis due to extensive use of electrical energy that tends to subside with time deserves further investigation and longer followup.

h followup examination. Whether postoperative asymptomatic vaginal scarring in the noncolpectomized group is associated with an increased risk of recurrence or represents development of fibrosis due to extensive use of electrical energy that tends to subside with time deserves further investigation and longer followup. In conclusion, surgical steps to excise DIE nodules with laparoscopy may be codified according to lesion size, location, and geometry. Although the ENZIAN scoring system is an important and comprehensive tool to assist in designing surgery, we feel that by simplifying grouping into predominantly lateral and predominantly central lesions, two basic techniques with distinct differences between them arise at least for smaller lesions (≤2 cm). The relatively inexperienced surgeon should try to develop his/her learning curve for each technique by adhering to strict and timely operative steps bearing in mind that for larger or multicompartmental lesions intraoperative adaptation and assistance by more experienced surgeons are of paramount importance to safely conclude the procedure. Conflict of Interests The authors declare that there is no conflict of interests regarding the publication of this paper. Figure 1 Complete dissection of the ureter, with sparing of the uterine vessels in a case with a posterolateral E2b DIE nodule and grossly involved pelvic peritoneum. Figure 2 Division of the uterine vein in a case with an E3b DIE nodule.

Conflict of Interests The authors declare that there is no conflict of interests regarding the publication of this paper. Figure 1 Complete dissection of the ureter, with sparing of the uterine vessels in a case with a posterolateral E2b DIE nodule and grossly involved pelvic peritoneum. Figure 2 Division of the uterine vein in a case with an E3b DIE nodule. Figure 3 (a) Shaving technique to remove an E2a–E2c legion with opening the vagina (arrow). (b) Shaving technique to remove an E3c legion from the rectosigmoid. Figure 4 Distribution of our cases according to DIE nodule size. Figure 5 Grouping of our patients according to location of the DIE nodules. Table 1 Characteristics of our 36 cases with posterior DIE nodules. Patient characteristics Mean (range) Age (years) 29.2 (19–36) BMI (kg/m2) 23.3 (20.2–27.6) Gravidity 0.7 (0–3) Parity 0.3 (0–3) N % Previous surgery for endometriosis 4 11.0 Chronic pelvic pain 36 100.0 Dysmenorrhea 32 88.8 Deep dyspareunia 36 100.0 Nonmenstrual pain 23 63.9 Infertility 7 19.4 Ovarian endometriomas 29 80.5 r-AFS classification Stage I 1 2.8 Stage II 4 11.1 Stage III 13 36.1 Stage IV 18 50.0 Table 2 Distribution of our cases, according to the two main compartments involved, by the DIE nodule. Numbers in brackets represent bilateral involvement (N = 36). ENZIAN system Less involved compartment Total a b c Compartment with principal involvement a 1* 6 (2) 2 9 b 4 13* (8) 7 (3) 24 c 2 — 1* 3 Total 7 19 10 36 *These cases represent involvement of a single compartment only. Table 3 Intraoperative and postoperative characteristics according to location of the DIE nodule.

Previous surgery for endometriosis 4 11.0 Chronic pelvic pain 36 100.0 Dysmenorrhea 32 88.8 Deep dyspareunia 36 100.0 Nonmenstrual pain 23 63.9 Infertility 7 19.4 Ovarian endometriomas 29 80.5 r-AFS classification Stage I 1 2.8 Stage II 4 11.1 Stage III 13 36.1 Stage IV 18 50.0 Table 2 Distribution of our cases, according to the two main compartments involved, by the DIE nodule. Numbers in brackets represent bilateral involvement (N = 36). ENZIAN system Less involved compartment Total a b c Compartment with principal involvement a 1* 6 (2) 2 9 b 4 13* (8) 7 (3) 24 c 2 — 1* 3 Total 7 19 10 36 *These cases represent involvement of a single compartment only. Table 3 Intraoperative and postoperative characteristics according to location of the DIE nodule. Patient characteristics Central nodules N (%) Lateral nodules N (%) Fisher's exact test P value N = 12 N = 24 Intraoperative characteristics Ovarian suspension 7 (58.3) 22 (91.7) 0.029 Rectosigmoid suspension 8 (66.7) 3 (12.5) 0.002 Systematic ureteral dissection 5 (41.7) 21 (87.5) 0.007 Division of uterine vein 0 (0.0) 3 (12.5) NS Excision of pelvic peritoneum 6 (50.0) 24 (100.0) <0.001 Partial colpectomy 3 (25.0) 0 (0.0) 0.031 Rectal wall suturing 3 (25.0) 0 (0.0) 0.031 Mean operation length (mins) 188 179 NS Mean blood loss (mls) 45 70 <0.001 Postoperative characteristics at 6/12 followup Complete resolution of CPP 7 (58.3) 18 (75.0) NS Vaginal scarring/induration 5 (41.7) 6 (25.0) NS Table 4 Intraoperative and postoperative characteristics according to size of the DIE nodule. Patient characteristics DIE nodule size ≤ 2 cm

in preferential cardiac and cerebral blood flow, with reduced blood flow to the rest of the body [7, 8]. As this process continues, the fetal right ventricular afterload increases in the setting of myocardial impairment, and changes in the fetal ductus venosus can often be visualized as a late and ominous finding [9]. Many studies have attempted to elucidate the factors that most strongly predict perinatal outcomes after delivery in the setting of abnormal UAD; however to date gestational age has always been most predictive [10]. More recently postnatal hemodynamic evaluation of preterm neonates has become part of the routine assessment in many European and Australian centers. Postnatal functional echocardiography (fECHO) uses measures of systemic blood flow (SBF) that have been shown to be more predictive than traditional measures of perfusion such as heart rate and blood pressure monitoring for poor outcomes. Infants identified to have low SBF, as indicated either by low superior vena cava flow (SVC) or low right ventricular output (RVO), have a higher risk of mortality and morbidity such as intraventricular hemorrhage [11, 12]. Given the high rate of poor neonatal outcomes in the setting of abnormal fetal UAD and low SBF, we sought to identify which antenatal factors could predict low SBF in pregnancies complicated by abnormal UAD.

icular output (RVO), have a higher risk of mortality and morbidity such as intraventricular hemorrhage [11, 12]. Given the high rate of poor neonatal outcomes in the setting of abnormal fetal UAD and low SBF, we sought to identify which antenatal factors could predict low SBF in pregnancies complicated by abnormal UAD. 2. Materials and Methods This is a retrospective review of fetuses who are delivered prematurely in the setting of abnormal UAD who received a fECHO in the first 72 hours. IRB approval for the study was obtained from our institution. A list of all fetuses with abnormal Doppler studies that were cared for and delivered at the University of California, San Diego, between August 2008 and April of 2012 was collected into a database.

of abnormal UAD who received a fECHO in the first 72 hours. IRB approval for the study was obtained from our institution. A list of all fetuses with abnormal Doppler studies that were cared for and delivered at the University of California, San Diego, between August 2008 and April of 2012 was collected into a database. Antenatal variables identified and collected from the electronic charts were gestational age at delivery, gravity and parity, ethnicity, chorionicity, maternal age at delivery, gestational age at the time of initial abnormal Doppler studies, number of days from initial identification of abnormal UAD until delivery, administration of maternal steroids, estimated fetal weight percentile prior to delivery, last measured amniotic fluid index (AFI), maternal BMI, maternal disease (including diabetes, hypertension, preeclampsia, and abruption), indication for delivery, and mode of delivery. Prenatal ultrasound data collected for each delivery included gestational age at first abnormal Doppler flow (defined as absent or reversed end-diastolic flow in the umbilical artery), the number of days of abnormal Doppler flow prior to delivery, and the presence of any other Doppler flow abnormalities at the time of delivery (such as abnormal ductus venosus flow or middle cerebral artery abnormalities).

bnormal Doppler flow (defined as absent or reversed end-diastolic flow in the umbilical artery), the number of days of abnormal Doppler flow prior to delivery, and the presence of any other Doppler flow abnormalities at the time of delivery (such as abnormal ductus venosus flow or middle cerebral artery abnormalities). Postnatal clinical variables collected were birth weight and birth weight percentile, APGAR scores, gender, presence of congenital anomalies, number of hospital days, death prior to discharge, presence of respiratory distress syndrome (RDS), presence of intraventricular hemorrhage (IVH), and placental pathology. Postnatal fECHO measurements collected were SVC flow and RVO. A low SVC flow was defined as <50 mL/kg/min and a low RVO was defined as <150 mL/kg/min.

mber of hospital days, death prior to discharge, presence of respiratory distress syndrome (RDS), presence of intraventricular hemorrhage (IVH), and placental pathology. Postnatal fECHO measurements collected were SVC flow and RVO. A low SVC flow was defined as <50 mL/kg/min and a low RVO was defined as <150 mL/kg/min. Patients were included in the study if they had both abnormal antenatal UAD studies and a postnatal echocardiogram within the first 72 hours of life. Pregnancies were excluded from the study if the fetuses were known to have congenital anomalies or any heart defect other than a patent ductus arteriosus, or a small ventral septal defect. Indication for evaluation with Doppler studies was at the discretion of the provider; however, common indications included suspected growth abnormalities, abnormal fluid levels, or previously documented Doppler abnormalities. All pregnant patients were scanned with a General Electric E8 ultrasound (GE Medical Systems, Milwaukee, WI, USA) by either a perinatologist or sonographer with advanced fetal sonography training, and umbilical artery Doppler velocimetry waveforms were obtained in the midportion of the cord during periods of fetal inactivity without breathing being present (see Figure 1).

tric E8 ultrasound (GE Medical Systems, Milwaukee, WI, USA) by either a perinatologist or sonographer with advanced fetal sonography training, and umbilical artery Doppler velocimetry waveforms were obtained in the midportion of the cord during periods of fetal inactivity without breathing being present (see Figure 1). Postnatal functional echocardiograms were performed when a trained provider in echocardiography was available and/or there was a clinical indication. Prematurity, hypotension, clinical instability, and evaluation for patent ductus arteriosus (PDA) were common clinical indications for fECHO in the first 72 hours of life. fECHO was performed and interpreted at the bedside by neonatologists trained in echocardiography using the General Electric Vivid E9 cardiovascular ultrasound system (GE Medical Systems, Milwaukee, WI, USA) with either the 7S or 10S phased array transducer probe. Original recorded measures for SBF without knowledge of the antenatal Dopplers were used for purposes of minimizing any bias for the study.

ocardiography using the General Electric Vivid E9 cardiovascular ultrasound system (GE Medical Systems, Milwaukee, WI, USA) with either the 7S or 10S phased array transducer probe. Original recorded measures for SBF without knowledge of the antenatal Dopplers were used for purposes of minimizing any bias for the study. SVC flow was calculated from the vessel diameter obtained in the parasternal long axis window in a sagittal plane and from the velocity obtained in the subcostal window. The average SVC diameter was obtained by measuring the maximum and minimum diameters at the junction of the SVC and right atrium over three cardiac cycles and all 6 measurements averaged. The complete velocity time integral from 10 consecutive cardiac cycles displaying laminar flow was obtained and averaged. SVC flow was calculated by measuring the average velocity time integral and multiplying it by the average cross-sectional area of the superior vena cava (mm) and the heart rate (beats per minute). Right ventricular output (RVO) was obtained by imaging the pulmonary artery from the parasternal long axis window in the sagittal plane to obtain both the vessel diameter and the velocity. The complete velocity time integral from 5 consecutive cardiac cycles displaying laminar flow was obtained and averaged. RVO was calculated by multiplying the velocity time integral by the cross-sectional area of the pulmonary artery (cm) and the heart rate (beats per minute) (see Figure 2).

diameter and the velocity. The complete velocity time integral from 5 consecutive cardiac cycles displaying laminar flow was obtained and averaged. RVO was calculated by multiplying the velocity time integral by the cross-sectional area of the pulmonary artery (cm) and the heart rate (beats per minute) (see Figure 2). Descriptive statistics were performed using Student's t-test and Mann-Whitney U tests (when nonparametric data was present), along with chi square analysis for categorical outcomes. Multivariate logistic regression was used to determine independent variables associated with low SBF including maternal age, gestational age or birth percentile at delivery, and length of stay in the NICU. 3. Results 63 subjects were identified with abnormal uterine artery Doppler studies; 20 subjects had both abnormal UAD and fECHO performed within the first 72 hours of life. Six infants had abnormal fECHO defined as either low RVO (<150 mL/kg/min) or low SVC flow (<50 mL/kg/min). The maternal demographics were overall similar between the two groups with the exception of age, which was lower in the abnormal fECHO group (Table 1). The gestational age at delivery was similar between the two groups. Those with abnormal fECHO had fewer days of abnormal UAD prior to delivery and trended towards a greater length of NICU stay (P value). The indications for deliveries were similar between the two groups as were the amniotic fluid indices at time of delivery and modes of delivery (Table 2).

milar between the two groups. Those with abnormal fECHO had fewer days of abnormal UAD prior to delivery and trended towards a greater length of NICU stay (P value). The indications for deliveries were similar between the two groups as were the amniotic fluid indices at time of delivery and modes of delivery (Table 2). Infants with abnormal fECHO had higher birth weight percentiles than those with normal fECHO and universally developed RDS. Due to the small frequency of more morbid neonatal outcomes (such as NEC, IVH, and pulmonary hemorrhage), the risk of these outcomes was not calculated (Table 3). The individual outcomes of infants with low systemic blood flow are shown in Table 4. The use of multivariate logistic regression did not significantly change the statistical significance of any of the above variables.

such as NEC, IVH, and pulmonary hemorrhage), the risk of these outcomes was not calculated (Table 3). The individual outcomes of infants with low systemic blood flow are shown in Table 4. The use of multivariate logistic regression did not significantly change the statistical significance of any of the above variables. 4. Discussion The timing of when to deliver a fetus with abnormal UAD has long been challenging. The goal of delivering as mature a fetus as possible has to be balanced with the desire to minimize poor neural outcomes due to significant hypoxemia, or even death. Baschat advocated prolongation of pregnancy to 34 weeks whenever possible, due to the significant morbidities associated with preterm delivery [10]. While some suggest delivering only when either an abnormal BPP is noted or ductus venosus a-wave reversal occurs, other studies have only recommended delivery prolongation to 28 weeks with an attempt to deliver prior to development of cardiac decompensation [5]. The challenge with many of these studies is the correlation between prenatal cardiac function and postnatal hemodynamics.

s noted or ductus venosus a-wave reversal occurs, other studies have only recommended delivery prolongation to 28 weeks with an attempt to deliver prior to development of cardiac decompensation [5]. The challenge with many of these studies is the correlation between prenatal cardiac function and postnatal hemodynamics. This is the first study to describe an association between abnormal UAD and low SBF as an attempt to identify the highest risk infants. Of interest, lower postnatal SBF (abnormal SVC or RVO) was associated with a shorter duration of time from the first abnormal UAD until delivery. The factors indicating the need for imminent delivery, such as the severity of the UAD or the fetal tracings at the time of delivery, were similar between the two groups. Infants who had lower SBF were more immature, suggesting that delaying delivery to allow for more maturity was likely outweighed by other acute factors driving the decision to deliver. It is possible that the short duration abnormal Doppler studies prior to delivery were indicative of a more acute and severe underlying process, which gave insufficient time to allow a normal fetal adaptive response.

o allow for more maturity was likely outweighed by other acute factors driving the decision to deliver. It is possible that the short duration abnormal Doppler studies prior to delivery were indicative of a more acute and severe underlying process, which gave insufficient time to allow a normal fetal adaptive response. While low SBF has been shown to correlate with adverse outcomes such as death and IVH [12, 13], infants in our study with abnormal UAD as well as low SBF were at much higher risk of needing surfactant and mechanical ventilation due to RDS. While RDS is primarily directly related to the degree of prematurity, there was no significant difference in gestational age between groups that could explain the difference in rates of RDS. This similarly could be related to either acutely impaired transitional hemodynamics causing abnormal pulmonary blood flow or a short duration of fetal stress limiting the time allowed for a fetal adaptive response. While this study was not large enough to evaluate the risk of more severe neonatal morbidities (such as NEC, pulmonary, or intraventricular hemorrhage), prior studies have already demonstrated these associations [12–14].

d flow or a short duration of fetal stress limiting the time allowed for a fetal adaptive response. While this study was not large enough to evaluate the risk of more severe neonatal morbidities (such as NEC, pulmonary, or intraventricular hemorrhage), prior studies have already demonstrated these associations [12–14]. There are several limitations to our study. Our study had a small sample size, due in part to the limited number of abnormal scans plus a neonatal provider who had performed a postnatal echocardiogram. Often, infants who had abnormal UAD who were more mature did not warrant an echocardiogram because of their stability. We also did not include infants who had normal UAD as a third control group. Unfortunately, these infants could have a number of confounding variables for both antenatal causes of delivery and reasons for postnatal low SBF. Future prospective studies should control for premature infants without abnormal UAD and similarly perform time scans to minimize changes that may occur with adaptation. Our study suggests that if antenatal measures of systemic blood flow such as SVC flow could be performed at the time of Doppler measurements of umbilical flow, this could help determine the degree of fetal impairment. These measures need further prospective evaluation. In conclusion, our findings suggest that other measures of SBF may be a useful tool in the assessment of fetuses with abnormal UAD and may be helpful in identifying the most at risk infants in this subset of patients.

There are several limitations to our study. Our study had a small sample size, due in part to the limited number of abnormal scans plus a neonatal provider who had performed a postnatal echocardiogram. Often, infants who had abnormal UAD who were more mature did not warrant an echocardiogram because of their stability. We also did not include infants who had normal UAD as a third control group. Unfortunately, these infants could have a number of confounding variables for both antenatal causes of delivery and reasons for postnatal low SBF. Future prospective studies should control for premature infants without abnormal UAD and similarly perform time scans to minimize changes that may occur with adaptation. Our study suggests that if antenatal measures of systemic blood flow such as SVC flow could be performed at the time of Doppler measurements of umbilical flow, this could help determine the degree of fetal impairment. These measures need further prospective evaluation. In conclusion, our findings suggest that other measures of SBF may be a useful tool in the assessment of fetuses with abnormal UAD and may be helpful in identifying the most at risk infants in this subset of patients. Conflict of Interests The authors declare that there is no conflict of interests regarding the publication of this paper. Figure 1 Umbilical artery Doppler studies. Examples of (a) normal, (b) absent, and (c) reversed end-diastolic flow. Figure 2 fECHO—normal SVC and RO measurements. Examples of (a) normal RO flow, (b) low (abnormal) RO flow, (c) normal SVC flow, and (d) low (abnormal) SVC flow.

Conflict of Interests The authors declare that there is no conflict of interests regarding the publication of this paper. Figure 1 Umbilical artery Doppler studies. Examples of (a) normal, (b) absent, and (c) reversed end-diastolic flow. Figure 2 fECHO—normal SVC and RO measurements. Examples of (a) normal RO flow, (b) low (abnormal) RO flow, (c) normal SVC flow, and (d) low (abnormal) SVC flow. Table 1 Maternal demographics. Normal fECHO Abnormal fECHO P value (n = 14) (n = 6) Age 33.4 ± 5.5 27.2 ± 2.5 0.003 BMI 27.6 ± 4.3 31 ± 2.4 0.08 Prior term deliveries   0.142 Prior preterm deliveries   0.143 Race   0.607 Caucasian 4 3 Hispanic 7 3 Asian/Pacific Islander 1 0 Other/unknown 2 0 Preeclampsia 5 2 0.919 Abnormal serum analytes 7 1 0.347 Table 2 Antenatal variables. Normal fECHO Abnormal fECHO P value (n = 14) (n = 6) Gestational age at first abnormal Doppler study (weeks) 26.8 ± 4.3 27.3 ± 3.5 0.794 Gestational age at delivery (weeks) 29.9 ± 2.9 27.9 ± 3.3 0.184 Duration of abnormal UAD prior to delivery (days) 23 ± 28 4.5 ± 4 0.03 Indication for delivery   0.239 Labor/nonspecified 0 1 Fetal distress 4 3 IUGR/oligohydramnios 8 2 Preeclampsia 2 0 Cesarean delivery 12 6 0.329 Table 3 Neonatal outcomes. Normal fECHO Abnormal fECHO P value (n = 14) (n = 6) RVO flow (mL/kg/min) 241 ± 59 118 ± 52 <0.001 SVC flow (mL/kg/min) 91 ± 27 38 ± 12 <0.001 Male infant 8 (57%) 3 (50%) 0.636 1 minute Apgar scores <7 9 (64%) 5 (83%) 0.394 5 minute Apgar scores <7 2 (14%) 0 0.329 Length of NICU stay (days) 63 ± 38 97 ± 52 0.173 Birth weight percentile 5 ± 5 25 ± 24 0.009 Small for gestational age (%) 14 (100%) 2 (33%) <0.001

Normal fECHO Abnormal fECHO P value (n = 14) (n = 6) RVO flow (mL/kg/min) 241 ± 59 118 ± 52 <0.001 SVC flow (mL/kg/min) 91 ± 27 38 ± 12 <0.001 Male infant 8 (57%) 3 (50%) 0.636 1 minute Apgar scores <7 9 (64%) 5 (83%) 0.394 5 minute Apgar scores <7 2 (14%) 0 0.329 Length of NICU stay (days) 63 ± 38 97 ± 52 0.173 Birth weight percentile 5 ± 5 25 ± 24 0.009 Small for gestational age (%) 14 (100%) 2 (33%) <0.001 RDS (%) 5 (36%) 6 (100%) 0.008 IVH 2 (14%) 2 (33%) 0.329 Table 4 Neonates with abnormal fECHO. Subj. number Number of days UAD prior to delivery (days) GA at delivery (weeks) Weight percentile at delivery SVC flow (mL/kg/min) RVO (mL/kg/min) NICU length of stay (days) ARDS Intubated IVH Demise 1 9 25 4/7 2% 47 213 147 Yes Yes No No 2 10 25 1/7 25% 29 79 NA1 Yes Yes Grade IV Yes 3 1 25 5/7 16% nc2 64 152 Yes Yes Grade I No 4 4 32 6/7 11% 47 127 29 Yes No No No 5 3 27 70% 21 105 87 Yes No No No 6 0 31 25% 44 123 69 Yes No No No 1Demise on day of life 2. 2Not calculable.

1. Introduction The development of ovarian follicles begins during fetal life with the transformation of primordial germ cells into oocytes enclosed in structures called follicles [1, 2]. Some of these follicles are recruited to start a long progress of growth and differentiation, during which the proteins required for oocyte maturation are progressively synthesized and accumulated [3, 4]. All events related to follicular development are regulated by appropriate signals originating from the growing oocyte itself and from the somatic cells that surround it [5, 6] and also by complex interactions between gonadotropin hormones, sex steroids, and diverse growth factors [7, 8]. The sex steroids produced by follicular cells are known to play major roles in the regulation of ovarian function. When present in the systemic circulation, these steroids actively participate in the regulation of pituitary gonadotropin secretion, and when present in the ovarian microenvironment, they act as important paracrine factors for the maintenance of follicular development [9]. Although much of the information about the role of sex steroids in ovarian functioning has been obtained in studies directed at the action of estrogens [10–12] and progestogens [13–15], increasing attention is being devoted to the action of androgen hormones because the activation of the androgen receptor located in follicular cells [16, 17] modulates the expression and activity of important genes for the maintenance of ovarian follicle development [17–19].

estrogens [10–12] and progestogens [13–15], increasing attention is being devoted to the action of androgen hormones because the activation of the androgen receptor located in follicular cells [16, 17] modulates the expression and activity of important genes for the maintenance of ovarian follicle development [17–19]. Additional evidence of the action of androgens in the regulation of folliculogenesis has arisen from in vitro studies showing that various androgens, including testosterone, androstenedione, and dihydrotestosterone, can stimulate the growth and development of ovarian follicles in mammals [20–22]. The reduction of reproductive function and the development of premature ovarian failure in mice with the nonselective deletion of the androgen receptor gene [23–25] support the hypothesis of the involvement of androgens in the regulation of follicular development. Reinforcing these findings, mice carrying this deletion show the impaired in vitro development of preantral follicles [26]. Additionally, a more pronounced expression of the androgen receptors has been reported to occur in preantral follicles [27, 28], suggesting the major action of androgens during the initial stages of folliculogenesis.

ndings, mice carrying this deletion show the impaired in vitro development of preantral follicles [26]. Additionally, a more pronounced expression of the androgen receptors has been reported to occur in preantral follicles [27, 28], suggesting the major action of androgens during the initial stages of folliculogenesis. From a clinical point of view, polycystic ovary syndrome (PCOS) is a nosologic entity that affects approximately 5–10% of women of reproductive age and is characterized by increased ovarian androgen production and chronic anovulation. The excessively androgenic microenvironment of the ovary is believed to have a negative impact on follicular development, which, in addition to LH hypersecretion, promotes follicle stagnation in the early stages of development (initial antral), inhibiting the development of a dominant and ovulatory follicle and leading to chronic anovulation and infertility. Among the various therapeutic alternatives for infertile patients with this diagnosis is the in vitro maturation of follicles obtained from their ovaries without previous induction. The results of this technique are limited in terms of the reproductive outcome, likely due to the impaired quality of the oocytes developed in hyperandrogenic environments [29]. Another therapeutic possibility in these cases is in vitro fertilization procedure, which shows that patients with PCOS, despite a larger number of oocytes, have similar pregnancy rates. This suggests a poorer utilization of the oocytes obtained, again indicating impaired oocyte quality [30].

rogenic environments [29]. Another therapeutic possibility in these cases is in vitro fertilization procedure, which shows that patients with PCOS, despite a larger number of oocytes, have similar pregnancy rates. This suggests a poorer utilization of the oocytes obtained, again indicating impaired oocyte quality [30]. Therefore, there is evidence in the literature of the participation of androgens in follicular development both as essential adjuvants and as harmful agents when present in excessive amounts. This indicates the relevance of understanding the role of androgens in the regulation of folliculogenesis, as well as the possibility of recreating in vitro conditions capable of guaranteeing the full growth of ovarian follicles to mimic as much as possible the in vivo intrafollicular environment [31]. On this basis, the objective of the current review was to present more relevant data regarding the involvement of androgens in the regulation of folliculogenesis and to provide information for the design of future culture strategies that can be used to promote the in vitro development of ovarian follicles.

onment [31]. On this basis, the objective of the current review was to present more relevant data regarding the involvement of androgens in the regulation of folliculogenesis and to provide information for the design of future culture strategies that can be used to promote the in vitro development of ovarian follicles. 2. Androgen Synthesis in the Ovary The androgens androstenedione, testosterone, and dihydrotestosterone are primarily synthesized from cholesterol and are produced by the ovary in a sequential manner together with other sex steroids, progestogens, and estrogens, with each steroid serving as a substrate for the subsequent one in a cascade of events known as steroidogenesis [32–34]. The classical two-cells-two-hormones model describes the role of follicular cells (theca and granulose) and of gonadotropins (follicle stimulating hormone (FSH) and luteinizing hormone (LH)) in steroid synthesis and secretion in the ovary, with emphasis on the cooperation of the two cell types that are necessary for androgen production. In general, the androstenedione synthesized from progestogens is converted to testosterone by the action of the enzyme 17β-hydroxysteroid dehydrogenase in theca cells under the LH stimulus, and the produced androgen is passively transported to the granulose cells where it is converted in estrogen by the action of aromatase under the FSH stimulus [35] (Figure 1).

from progestogens is converted to testosterone by the action of the enzyme 17β-hydroxysteroid dehydrogenase in theca cells under the LH stimulus, and the produced androgen is passively transported to the granulose cells where it is converted in estrogen by the action of aromatase under the FSH stimulus [35] (Figure 1). Due of this conversion of testosterone in estradiol in the granulosa, many of the actions of androgens on the growth and differentiation of ovarian follicles can be indirectly mediated by the action of androgens as precursors in the biosynthesis of estrogens. Although the actions of estrogens on the ovary are well known in terms of the pattern of expression and function of estrogen receptors [36], little is known about the direct involvement of androgens in terms of their interaction with the specific receptor in the regulation and maintenance of folliculogenesis.

strogens. Although the actions of estrogens on the ovary are well known in terms of the pattern of expression and function of estrogen receptors [36], little is known about the direct involvement of androgens in terms of their interaction with the specific receptor in the regulation and maintenance of folliculogenesis. 3. Expression of the Androgen Receptor in the Ovary The cellular actions of androgens require the binding and activation of the specific ligand receptor called the androgen receptor (AR) [37]. Both the protein and messenger RNA of AR have been detected in the ovary of various mammalian species, such as rodents [38, 39], cattle [40, 41], sheep [42], swine [43, 44], non-human primates [16, 17], and humans [45, 46]. Although most of these studies have indicated that granulosa cells are the predominant sites of AR expression, theca and ovarian stromal cells also express AR [20, 47, 48]. In oocytes, AR expression exhibits an evolutive profile that is highly expressed in amphibians [49], moderately expressed in rodents [20], little expressed in ruminants [42, 44], and incipient or absent in non-human primates and in humans [17, 50–52].

ssion, theca and ovarian stromal cells also express AR [20, 47, 48]. In oocytes, AR expression exhibits an evolutive profile that is highly expressed in amphibians [49], moderately expressed in rodents [20], little expressed in ruminants [42, 44], and incipient or absent in non-human primates and in humans [17, 50–52]. In rodents and primates, AR expression appears to be regulated along follicular development, being increased in ovaries containing a larger number of preantral and antral follicles of small diameter and reduced in ovaries containing periovulatory follicles [53, 54]. The analysis of the expression in isolated follicles shows that, in these species, follicles in the early stages of development express a larger number of AR than those in more advanced stages (Table 1, ∗) [20, 55]. Additionally, a differential gradient of AR expression is noted in mature follicles, which are little expressed in mural granulosa cells and are highly expressed in cumulus cells [56]. The profile of mRNA and the protein of AR in different follicular classes are presented in Table 1.

anced stages (Table 1, ∗) [20, 55]. Additionally, a differential gradient of AR expression is noted in mature follicles, which are little expressed in mural granulosa cells and are highly expressed in cumulus cells [56]. The profile of mRNA and the protein of AR in different follicular classes are presented in Table 1. 4. Mechanisms of Action of the Androgens Like all steroid hormones, the androgens primarily exert their functions by binding and activating specific nuclear receptors that trigger the intracellular events responsible for the beginning of the transcription of target genes [57, 58]. Additionally, the androgens can also exert their effects by interacting with receptors located on the cell membrane to perform rapid, non-genomic actions involved in the activation of various transcription factors [59–61]. Thus, the activated ARs transcriptionally regulate the expression of a selected group of genes via direct or indirect association with the regulatory regions of (enhancer/promoter) upstream elements [37, 62, 63]. Although several autocrine and/or paracrine factors involved in the regulation of the development of ovarian follicles have been described [6, 64, 65], only some genes responsible for the transcription of these factors have been tested as AR targets. Particularly important among them are the genes related to FSH receptors, insulin-like growth factor-1 (IGF-1), and aromatase enzyme [18, 19, 66, 67]. The main findings regarding the direct action of androgen hormones on the in vivo and in vitro control of follicular development in mammals are based on the transcriptional actions of AR in follicular cells. As reported for AR expression in the oocytes, the physiological role of androgens in oocyte maturation appears to have been lost over evolution, being phylogenetically replaced by the action of gonadotropins [68].

l of follicular development in mammals are based on the transcriptional actions of AR in follicular cells. As reported for AR expression in the oocytes, the physiological role of androgens in oocyte maturation appears to have been lost over evolution, being phylogenetically replaced by the action of gonadotropins [68]. 5. Role of Androgens in Early Follicular Development 5.1. In Vivo Effects Although the effects of chronic exposure to high androgen concentrations during the prenatal or postnatal period in different mammalian species are extensively known and are correlated with irregularities of the reproductive cycle and changes in the ovarian morphology in patients with a diagnosis of PCOS [69], few studies have been designed to assess the effects of short-term exposure to low androgen concentrations. The main results obtained in studies evaluating the effects of the in vivo administration of androgen hormones on early follicular development in different species are listed in Table 2 and described below.

[69], few studies have been designed to assess the effects of short-term exposure to low androgen concentrations. The main results obtained in studies evaluating the effects of the in vivo administration of androgen hormones on early follicular development in different species are listed in Table 2 and described below. Subcutaneous implants for the controlled release of low androgen doses have been used as an efficient tool for the study of the effects of short-term androgen exposure using experimental animal models. In non-human primates, subcutaneous implants containing different doses of testosterone promoted an expressive increase in follicular recruitment, growth, and survival [70]. These effects appear to be mediated by a local amplification of the action of both IGF-1 and FSH because exposure to testosterone induced an increase in IGF-1, IGF-1 receptor, and FSH receptor mRNA in the ovaries of these animals [18, 19, 66]. The increase of follicular recruitment was positively correlated with increases in IGF-1 and IGF-1 receptor mRNA in the oocytes of primordial follicles, suggesting an indirect action of androgens via the IGF-1 system on follicular activation [19]. All of the effects on both ovarian morphology and the IGF-1 and FSH system induced via exposure to testosterone were fully replicated when subcutaneous implants containing the nonaromatizable androgen dihydrotestosterone were used, showing that the effects of androgens are mediated by AR and not by conversion to estrogens. Similarly, the in vitro treatment of granulosa cells obtained from the antral follicles of swine with dihydrotestosterone increases the action of IGF-1 as a stimulus to cell proliferation, as well as the effect of growth and differentiation factor 9 (GDF9) in the presence of IGF-1 [71, 72], both of which are effects that are blocked by the presence of the AR antagonist hydroxyflutamide. The positive correlation between the expression of the AR gene and the proliferation of granulosa cells and follicular growth further supports the hypothesis of the involvement of androgens in follicular growth via AR [17] (Figure 2).

h are effects that are blocked by the presence of the AR antagonist hydroxyflutamide. The positive correlation between the expression of the AR gene and the proliferation of granulosa cells and follicular growth further supports the hypothesis of the involvement of androgens in follicular growth via AR [17] (Figure 2). In other mammalian species, short-term exposure to low androgen doses was also related to increased follicular recruitment and growth. In swine, the intramuscular administration of dihydrotestosterone during the first 3 days of the early follicular phase and during the last 3 days of the late follicular phase of the reproductive cycle significantly increased the ovulatory rate of these animals [73]. The administration of 10x diluted doses administered from the 13th day of the estrous cycle to the next estrus promoted a significant increase of mRNA expression of the FSH receptor in periovulatory follicles, suggesting that the increased ovulatory response detected in androgenized animals is in fact related to the increased sensitivity of ovarian follicles to the gonadotropic action induced by the androgen. In rodents, the administration of subcutaneous implants containing dihydrotestosterone promoted the increased expression of FSH receptor mRNA in preantral follicles [74]. Additionally, the potential for the in vitro development of preantral follicles isolated from the ovaries of androgenized animals was superior to that of nonandrogenized follicles, showing that in this species the androgens also promote follicular growth via increased gonadotropic sensitization.

eantral follicles [74]. Additionally, the potential for the in vitro development of preantral follicles isolated from the ovaries of androgenized animals was superior to that of nonandrogenized follicles, showing that in this species the androgens also promote follicular growth via increased gonadotropic sensitization. Taken together, these findings suggest that the in vivo action of androgens via AR may regulate both the expression and the action of one or more of the ovarian growth factors necessary for the regulation of follicular recruitment and growth. It should be noted that no in vivo studies have been conducted on the human species because of the limitation represented by the side effects that this practice could cause in women exposed to systemic androgens. A satisfactory model for these evaluations in humans is represented by the study of oocytes obtained from patients with PCOS. In 1991, Cha et al. [75] reported the first pregnancy resulting from in vitro maturation (IVM); in 1994, Trounson [76] first reported a successful pregnancy process using oocytes aspirated from nonstimulated patients with PCOS. Since then, various studies have performed IVM without stimulation in patients with PCOS with the objective of obtaining pregnancy rates similar to those of fertile women [75, 77–79] and to assess the efficacy of IVM and fertilization in nonstimulated patients.

g oocytes aspirated from nonstimulated patients with PCOS. Since then, various studies have performed IVM without stimulation in patients with PCOS with the objective of obtaining pregnancy rates similar to those of fertile women [75, 77–79] and to assess the efficacy of IVM and fertilization in nonstimulated patients. In the studies cited above, immature oocytes were aspirated from infertile patients between days 6 and 14 of the cycle with the aid of transvaginal ultrasound, and those of normal morphology were placed in culture for 24–48 hours; only those with extrusion of the first polar body were submitted to intracytoplasmic sperm injection (ICSI), and transfer was performed 2 to 3 days after ICSI [75, 77–79]. In the cited reports, fertilization rates ranged from 62% to 75.3%, and cleavage rates ranged from 81.4% to 95%, which are in agreement with data reported by Trounson et al. [76]. The pregnancy rates reported by Zhao et al. [78] (40%), Holzer et al. [77] (50%), and Zhao et al. [79] (40%) are similar to those of patients with unknown causes of infertility, although Cha et al. [75] and Bos-Mikich et al. [80] reported lower pregnancy rates in PCOS patients (27.1% and 32%, resp.). According to Zhao et al. [79], ovarian stimulation is unnecessary; they obtained pregnancy rates similar to those of patients with stimulated cycles (40%). A similar result was obtained by Ge et al. [81], who showed that the addition of hCG does not contribute to oocyte maturation. The advantages of nonstimulation are many and include the prevention of ovary hyperstimulation syndrome induced by gonadotropin use, reduction of costs, shorter and continuous treatment cycles, and the prevention of a series of other long-term complications such as hormone-dependent neoplasias [75, 79] (Table 3).

tion. The advantages of nonstimulation are many and include the prevention of ovary hyperstimulation syndrome induced by gonadotropin use, reduction of costs, shorter and continuous treatment cycles, and the prevention of a series of other long-term complications such as hormone-dependent neoplasias [75, 79] (Table 3). According to Das et al. [82], the anti-Mullerian hormone (AMH) plays a role in PCOS because its values are increased in affected patients compared with control ovulatory women (466 ng/mL and 78 ng/mL, resp., p = 11 and 8, resp.). It was also observed that early antral and preantral follicles express AMH, which is absent in primordial and in atresic follicles [82]. These authors speculated about the influence of high androgen levels on the elevation of AMH values in patients with PCOS. The relationship between GDF9 and bone morphogenetic protein (BMP15) factor has been studied in the oocytes and granulosa cells of patients with PCOS because these factors play a crucial role in follicle development, ovulation, oocyte maturation, and embryo development. It has been reported that GDF9 and BMP15 are not expressed in patients with PCOS, with a consequent later impairment of cytoplasm maturation and poor oocyte quality, whereas they are expressed in normal ovulatory women [83, 84].

crucial role in follicle development, ovulation, oocyte maturation, and embryo development. It has been reported that GDF9 and BMP15 are not expressed in patients with PCOS, with a consequent later impairment of cytoplasm maturation and poor oocyte quality, whereas they are expressed in normal ovulatory women [83, 84]. 5.2. In Vitro Effects The development of in vitro culture systems able to guarantee the growth and differentiation of isolated ovarian follicles represents a valuable tool for the study of the direct effects of androgens on folliculogenesis. Various in vitro systems have been employed for the culture of preantral follicles in various mammalian species, such as cattle [85], goats [86], non-human primates [66], and humans [87]. However, the production of healthy offspring from oocytes of preantral follicles matured in vitro has been reported only in mice thus far [88]. The main results obtained in studies aiming to evaluate the in vitro effects of androgens on early follicular development in different species are listed in Table 4 and described below.

the production of healthy offspring from oocytes of preantral follicles matured in vitro has been reported only in mice thus far [88]. The main results obtained in studies aiming to evaluate the in vitro effects of androgens on early follicular development in different species are listed in Table 4 and described below. The in vitro treatment of mouse ovarian follicles with AR antagonists (hydroxyflutamide and bicalutamide) reduced follicular growth during the preantral phase, as well as the meiotic maturation of the enclosed oocyte [20], suggesting the importance of androgen action in follicle maturation. The inability of preantral follicles to develop in vitro to preovulatory stages in the presence of antiandrogen antibodies [21] supports the hypothesis that the actions mediated by ARs are important for the early stages of follicular growth. In the same study, the addition of the AR antagonist casodex inhibited the positive effect of FSH on follicular growth, which was completely reversed when dihydroxytestosterone was added, revealing a joint action of androgens and FSH on follicular development. Additionally, the increased survival and growth of preantral follicles in the presence of androstenedione [89] represents further evidence of the positive action of androgens on follicular development. In the same study [21], the addition of antiestrogen antibodies and of estradiol receptor antagonists (ICI 182, 780) did not modify the positive effects of androstenedione on follicular growth, confirming the direct action of androgens on the development of preantral follicle maintenance.

ens on follicular development. In the same study [21], the addition of antiestrogen antibodies and of estradiol receptor antagonists (ICI 182, 780) did not modify the positive effects of androstenedione on follicular growth, confirming the direct action of androgens on the development of preantral follicle maintenance. The supplementation of the culture medium with other androgen hormones, that is, dihydrotestosterone, testosterone, DHEA, and DHEA sulfate, at different concentrations (10−11 to 10−7 M) also promoted the growth of preantral follicles in a dose-dependent manner [22]. In this study, the AR antagonist hydroxyflutamide but not the aromatase inhibitor fadrozole hydrochloride inhibited the growth response, indicating that estrogens converted from androgens during culture were not responsible for follicular growth.

omoted the growth of preantral follicles in a dose-dependent manner [22]. In this study, the AR antagonist hydroxyflutamide but not the aromatase inhibitor fadrozole hydrochloride inhibited the growth response, indicating that estrogens converted from androgens during culture were not responsible for follicular growth. Androgen hormones are able to promote follicular growth not only during the culture of isolated follicles but also during in situ culture. The addition of testosterone (10−10 to 10−7 M) to the culture medium of fragments of ovarian cortex from bovine fetuses increased the transition from primary to secondary follicles in a dose-dependent manner [85]. In the same study, the addition of estradiol did not promote the same effect; also, in the presence of the AR antagonist flutamide, the positive effect of testosterone on follicular development was completely abolished, indicating that the observed effect was due to the direct action of androgens via AR. The addition of androgens (10−7 to 10−10 M) to the culture medium of fragments of human ovarian cortex tissue significantly inhibited cell apoptosis [87]. Similarly, the addition of estradiol to the culture medium was unable to reproduce the effect of androgen; this effect was also abolished in the presence of the AR antagonist casodex. These findings suggest a positive effect of androgen hormones on the maintenance of the viability of ovarian tissue during culture, which is exclusively due to a direct androgen action mediated by AR.

nable to reproduce the effect of androgen; this effect was also abolished in the presence of the AR antagonist casodex. These findings suggest a positive effect of androgen hormones on the maintenance of the viability of ovarian tissue during culture, which is exclusively due to a direct androgen action mediated by AR. In general, the results reported here suggest that, at least under in vitro conditions, androgen hormones can promote the growth of ovarian follicles during early stages of development. As observed in vivo, high androgen doses can also have a negative influence on follicular development under in vitro conditions. The in vitro exposure of mouse preantral follicles to androgen concentrations higher than 10−5 M induced the precocious luteinization of granulosa cells [90] and also significantly reduced follicular growth and viability [91]. Additionally, under these conditions, the in vitro estradiol and progesterone secretion by developing mouse follicles is exacerbated and is associated with reduced oocyte quality and abnormal chromosome distribution on the metaphase plate [92].

s [90] and also significantly reduced follicular growth and viability [91]. Additionally, under these conditions, the in vitro estradiol and progesterone secretion by developing mouse follicles is exacerbated and is associated with reduced oocyte quality and abnormal chromosome distribution on the metaphase plate [92]. 6. Conclusion In general, the results compiled in the present review indicate that during the early and intermediate stages of follicular maturation, when AR expression is more pronounced, the androgens locally produced by the developing follicles facilitate the transcription of genes involved in the control of follicle transition from the reserve pool to the growth pool and of genes involved in the promotion of subsequent follicle development. Additionally, because androgens increase the activities of FSH, especially those related to cell proliferation and differentiation, the fall in AR expression in mature follicles reduces the action of androgens; this is possibly a critical event during the processes of follicular selection and atresia. Under in vitro conditions, submicromolar androgen doses can have a positive influence on the development of preantral follicles, promoting both survival and growth, especially when combined with the addition of FSH. Conflict of Interests The authors declare that they have no conflict of interests regarding the publication of this paper.

6. Conclusion In general, the results compiled in the present review indicate that during the early and intermediate stages of follicular maturation, when AR expression is more pronounced, the androgens locally produced by the developing follicles facilitate the transcription of genes involved in the control of follicle transition from the reserve pool to the growth pool and of genes involved in the promotion of subsequent follicle development. Additionally, because androgens increase the activities of FSH, especially those related to cell proliferation and differentiation, the fall in AR expression in mature follicles reduces the action of androgens; this is possibly a critical event during the processes of follicular selection and atresia. Under in vitro conditions, submicromolar androgen doses can have a positive influence on the development of preantral follicles, promoting both survival and growth, especially when combined with the addition of FSH. Conflict of Interests The authors declare that they have no conflict of interests regarding the publication of this paper. Figure 1 Schematic view of the production of androgen hormones by ovarian follicular cells according to the two-cells-two-hormones model. In the theca cells, the androgens (androstenedione and testosterone) are produced in response to the luteinizing hormone (LH) stimulus. After diffusing towards the granulosa cells, the androgens are converted to estrogens (estradiol and estrone) by the enzyme aromatase under the action of the follicle stimulating hormone (FSH). P450scc, enzyme responsible for the cleavage of the lateral chain of cholesterol; 3β-HSD, 3β-hydroxysteroid dehydrogenase; 17β-HSD, 17β-hydroxysteroid dehydrogenase.

ells, the androgens are converted to estrogens (estradiol and estrone) by the enzyme aromatase under the action of the follicle stimulating hormone (FSH). P450scc, enzyme responsible for the cleavage of the lateral chain of cholesterol; 3β-HSD, 3β-hydroxysteroid dehydrogenase; 17β-HSD, 17β-hydroxysteroid dehydrogenase. Figure 2 Schematic presentation of the action of androgen hormones on early follicular development. The androgens produced by the growing follicles can promote the transition from (1) primordial to primary follicles and (2) from primary to secondary follicles through the amplification of the actions of the IGF-1 system and can also amplify the actions of follicle stimulating hormone (FSH) on the promotion of the subsequent growth of (3) preantral and (4) antral follicles. Table 1 Profile of mRNA expression and of AR protein in the different follicular classes. Lenie and Smitz, 2009 [20] Juengel et al., 2006 [42] Cárdenas and Pope, 2002 [43] Salvetti et al., 2012 [41] Weil et al., 1998 [17] Chadha et al., 1994 [45] Animal Rodent Sheep Swine Cattle Non-human primate Human

Figure 2 Schematic presentation of the action of androgen hormones on early follicular development. The androgens produced by the growing follicles can promote the transition from (1) primordial to primary follicles and (2) from primary to secondary follicles through the amplification of the actions of the IGF-1 system and can also amplify the actions of follicle stimulating hormone (FSH) on the promotion of the subsequent growth of (3) preantral and (4) antral follicles. Table 1 Profile of mRNA expression and of AR protein in the different follicular classes. Lenie and Smitz, 2009 [20] Juengel et al., 2006 [42] Cárdenas and Pope, 2002 [43] Salvetti et al., 2012 [41] Weil et al., 1998 [17] Chadha et al., 1994 [45] Animal Rodent Sheep Swine Cattle Non-human primate Human Final Product mRNA Protein mRNA Protein mRNA Protein mRNA Protein mRNA Protein mRNA Protein Primordial X + + + + X X − − X − X Primary X* +* +* +* +* X* X* −* −* X* −* X* Secondary X* +* +* −* +* X* X* +* +* X* −* X* Initial antral X* +* +* −* +* X* X* ++* +* X* +/−* X* Final antral X + − − + X X ++ − X +/− X Oocyte X +++ + +/− − X X X − X X X Theca X + + − − X X + − X +/− X Granulosa X + ++ + − X X + + X +/− X Legend: (X): not done; (−): No expression; (−/+): Little expression; (+): Low expression; (++): Moderate expression; (+++): Abundant expression. Table 2 Effects of the in vivo administration of androgen hormones on early follicular development. Animal Methodology used Main results Reference

Final Product mRNA Protein mRNA Protein mRNA Protein mRNA Protein mRNA Protein mRNA Protein Primordial X + + + + X X − − X − X Primary X* +* +* +* +* X* X* −* −* X* −* X* Secondary X* +* +* −* +* X* X* +* +* X* −* X* Initial antral X* +* +* −* +* X* X* ++* +* X* +/−* X* Final antral X + − − + X X ++ − X +/− X Oocyte X +++ + +/− − X X X − X X X Theca X + + − − X X + − X +/− X Granulosa X + ++ + − X X + + X +/− X Legend: (X): not done; (−): No expression; (−/+): Little expression; (+): Low expression; (++): Moderate expression; (+++): Abundant expression. Table 2 Effects of the in vivo administration of androgen hormones on early follicular development. Animal Methodology used Main results Reference Non-human primate Subcutaneous implants containing testosterone (4 mg/kg animal weight/day) for 3 days, (400 μg/kg animal weight/day) for 10 days, (20 g/kg animal weight/day) for 5 days, or dihydrotestosterone (145 μg/kg animal weight/day) for 5 days. Increased follicular retrieval, growth, and survival. Increased theca and granulosa cell proliferation. Vendola et al., 1998 [70] Subcutaneous implants containing testosterone (20 μg/kg animal weight/day) or dihydrotestosterone (145 μg/kg animal weight/day) for 5 days. Increased expression of mRNA for IGF-I and for IGF-I receptor in granulosa and theca cells of growing follicles. Vendola et al., 1999 [18] Subcutaneous implants containing testosterone (20 μg/kg animal weight/day) or dihydrotestosterone (145 μg/kg animal weight/day) for 5 days; testosterone (400 μg/kg animal weight/day) for 3 or 10 days. Increased follicular retrieval and expression of IGF-I mRNA and of IGF-I receptor in oocytes of primordial follicles. Vendola et al., 1999 [19] Subcutaneous implants containing testosterone (4 mg/kg animal weight/day) for 3 days or testosterone (0.4 mg/kg animal weight/day) for 10 days. Increased expression of FSH receptor mRNA in granulosa cells of growing follicles. Weil et al., 1999 [66]

receptor in oocytes of primordial follicles. Vendola et al., 1999 [19] Subcutaneous implants containing testosterone (4 mg/kg animal weight/day) for 3 days or testosterone (0.4 mg/kg animal weight/day) for 10 days. Increased expression of FSH receptor mRNA in granulosa cells of growing follicles. Weil et al., 1999 [66] Swine Intramuscular administration of dihydrotestosterone (60 μg/kg animal weight) for the first 3 days of the early follicular phase or for the last 3 days of the late follicular phase; dihydrotestosterone (6 μg/kg animal weight) administered from the 13th day of the estrous cycle to the next estrus. Increased ovulatory rate and increased expression of FSH receptor mRNA in periovulatory follicles. Cárdenas et al., 2002 [73] Rodent Administration of a subcutaneous implant containing dihydrotestosterone (83 μg/kg animal weight/day) for 90 days. Increased follicular retrieval in preantral follicles. Increased potential for in vitro development of preantral follicles. Xue et al., 2012 [74] Table 3 Reproductive results after the in  vitro maturation in nonstimulated cycles of patients with polycystic ovary syndrome. Maturation (%) (N) Fertilization (%) (N) Cleavage (%) (N) Pregnancy (%) (N) Zhao et al., 2006 [78] 73.7% (632/857) 75.3% (476/632) 91.2% (434/476) 40% (19/47) Cha et al., 2000 [75] 62.2% (708/1.139) 68% (481/708) 88.1% (266/302) 27.1% (85/23)

Rodent Administration of a subcutaneous implant containing dihydrotestosterone (83 μg/kg animal weight/day) for 90 days. Increased follicular retrieval in preantral follicles. Increased potential for in vitro development of preantral follicles. Xue et al., 2012 [74] Table 3 Reproductive results after the in  vitro maturation in nonstimulated cycles of patients with polycystic ovary syndrome. Maturation (%) (N) Fertilization (%) (N) Cleavage (%) (N) Pregnancy (%) (N) Zhao et al., 2006 [78] 73.7% (632/857) 75.3% (476/632) 91.2% (434/476) 40% (19/47) Cha et al., 2000 [75] 62.2% (708/1.139) 68% (481/708) 88.1% (266/302) 27.1% (85/23) Holzer et al., 2007 [77] 68.3% (104/154) 73.3% (71/104) 81.4% (62/71) 50% (6/12) Zhao et al., 2009 [79] 68.8% (1.753/2.548) 70.28% (1.232/1.753) 90.2% (1.111/1.232) 40% (56/140) Bos-Mikich et al. 2011 [80] 63% (350/556) 62% (218/350) 95% (208/218) 32% (11/34) Table 4 Effects of the in vitro administration of androgen hormones on early follicular development. Animal Methodology used Main results Reference Cattle Culture of ovarian cortex fragments for 10 days in the presence of (i) 10−10to 10−7 M testosterone, (ii) 10−7 M estradiol, and (iii) testosterone in combination with flutamide (AR antagonist). Testosterone (10−7 and 10−6 M) promoted an increased transition from primary to secondary follicles, which was inhibited in the presence of flutamide and was not replicated in the presence of estradiol. Yang and Fortune, 2006 [85]

) 10−7 M estradiol, and (iii) testosterone in combination with flutamide (AR antagonist). Testosterone (10−7 and 10−6 M) promoted an increased transition from primary to secondary follicles, which was inhibited in the presence of flutamide and was not replicated in the presence of estradiol. Yang and Fortune, 2006 [85] Human Culture of ovarian cortex fragments for 24 hours in the presence of FSH in combination with (i) 10−10 to 10−7 M testosterone, (ii) 10−10 to 10−7 M dihydrotestosterone, (iii) 10−10 to 10−8 M estradiol, and (iiii) dihydrotestosterone in combination with casodex (AR antagonist). Androgens promoted a reduction of ovarian tissue apoptosis which was inhibited in the presence of casodex and was not replicated in the presence of estradiol. Otala et al., 2004 [87]

10−7 M dihydrotestosterone, (iii) 10−10 to 10−8 M estradiol, and (iiii) dihydrotestosterone in combination with casodex (AR antagonist). Androgens promoted a reduction of ovarian tissue apoptosis which was inhibited in the presence of casodex and was not replicated in the presence of estradiol. Otala et al., 2004 [87] Rodent Culture of isolated preantral follicles for 6 days in the presence of (i) anti-androgen antibody in combination or not with 1 μg/mL androstenedione, (ii) casodex (AR antagonist) in combination with FSH, and (iii) 1 μg/mL dihydroxytestosterone in combination with FSH. Treatment with an anti-androgen antibody inhibited follicular growth and differentiation; an effect that was reversed by the addition of androstenedione. Treatment with casodex inhibited the positive effect of FSH on follicular growth which was reversed by the addition of dihydroxytestosterone. Murray et al., 1998 [21] Culture of isolated preantral follicles for 6 days in the presence of FSH in combination or not with 1 μg/mL androstenedione. Treatment with FSH promoted follicular growth and differentiation which were improved when FSH was combined with androstenedione. Spears et al., 1998 [89] Culture of isolated preantral follicles for 4 days in the presence of (i) dihydrotestosterone or testosterone or dihydroxytestosterone or dihydroxytestosterone sulfate (10−11 to 10−7 M) (ii) in combination or not with hydroxyflutamide (AR antagonist) or FSH. Treatment with the different androgens promoted a dose-dependent follicular growth which was inhibited by the presence of the AR antagonist. The combination with FSH improved the effect of androgens on follicular growth. Wang et al., 2001 [22] Culture of isolated preantral follicles for 13 days in the presence of (i) 50 μM hydroxyflutamide (AR antagonist) in combination with FSH and (ii) 50 μM bicalutamide (AR antagonist) in combination with FSH. Follicular growth, production of inhibiting B, and steroids and oocyte maturation were impaired by the addition of both AR antagonists. Lenie and Smitz 2009 [20] Culture of isolated preantral follicles for 13 days in the presence of (i) 20 or 200 nM androstenedione and (ii) testosterone (20 or 200 nM or 2 μM) both in the presence of FSH. Treatment with both androgens at concentrations of more than 200 nM impaired oocyte maturation. Romero and Smitz, 2010 [92] Culture of isolated preantral follicles for 10 days in the presence of 10−5 M androstenedione.

0 or 200 nM androstenedione and (ii) testosterone (20 or 200 nM or 2 μM) both in the presence of FSH. Treatment with both androgens at concentrations of more than 200 nM impaired oocyte maturation. Romero and Smitz, 2010 [92] Culture of isolated preantral follicles for 10 days in the presence of 10−5 M androstenedione. Treatment with androstenedione induced changes in the morphology of granulosa cells compatible with luteinization. Okutsu et al., 2010 [90] Culture of isolated preantral follicles for 12 days in the presence of androstenedione (10−11, 10−9, 10−5 M) in combination with FSH. Treatment with androstenedione at the dose of 10−5 M promoted follicular growth, whereas higher doses impaired follicular development. Treatment with androstenedione impaired oocyte maturation regardless of the dose used. Tarumi et al., 2012 [91]

1. Introduction Pain management during labour is an essential part of good obstetric care. Though this severe pain during labour is not life threatening, it can have neuropsychological consequences. Postnatal depression may be more common when labour analgesia is not used [1]. Pain during labor has also been correlated with the development of posttraumatic stress disorder [2]. It has been proven beyond a doubt that epidural analgesia, when compared with other methods, provides superior analgesia for labor. However there can be situations where either it is not available or it is not feasible. Parenteral opioids, thus, are still popular for pain relief in labor in many countries throughout the world. A recent Cochrane review emphasized that as parenteral opioid drugs are very widely used for labor analgesia, it is important that more research is carried out so that women can make informed choices about these forms of pain relief [3]. Use of parenteral opioids for labour pain relief has been quite common, pethidine being the most commonly used drug in this group worldwide [4]. Studies on pethidine have, however, raised concerns about its adverse effects on the parturient and the newborn. It has been implicated to result in neonatal behavioral and feeding problems up to six weeks after delivery [5]. These concerns about pethidine have promoted a search for alternative opioids like pentazocine and tramadol.

hidine have, however, raised concerns about its adverse effects on the parturient and the newborn. It has been implicated to result in neonatal behavioral and feeding problems up to six weeks after delivery [5]. These concerns about pethidine have promoted a search for alternative opioids like pentazocine and tramadol. Tramadol is a synthetic analogue of codeine and is a centrally acting agent. It has a relatively low affinity for opiate receptors. Studies have shown that tramadol is an effective analgesic without the maternal and neonatal respiratory depression common to other opioids and it does not delay gastric emptying. Pentazocine, a benzomorphan derivative, is a synthetic opioid with both agonist and weak antagonistic action. It produces analgesia with little or no respiratory depression. When used for the relief of labor pain many studies have demonstrated equal analgesic effect with a better safety profile compared with pethidine. These two may be useful in producing effective pain relief in labor without the adverse effects seen in pethidine. Tramadol or pentazocine can be used as labor analgesics with minimum cost and less training as compared to the proven epidural analgesia that requires trained staff and equipment and has higher cost. It also avoids the side effects associated with epidural analgesia like hypotension, foetal heart rate changes, impaired motor ability, shivering, urinary retention, delayed pushing, and a prolonged second stage of labour.

d to the proven epidural analgesia that requires trained staff and equipment and has higher cost. It also avoids the side effects associated with epidural analgesia like hypotension, foetal heart rate changes, impaired motor ability, shivering, urinary retention, delayed pushing, and a prolonged second stage of labour. In this study we aimed to evaluate and compare the efficacy of intramuscular tramadol and pentazocine in the first stage of labour. 2. Material and Methods This prospective study was carried out in the labour ward at a tertiary care centre for one year. The study protocol was approved by the institutional ethics committee. All enrolled women provided written informed consent for participation. A total of 65 women with normal singleton, cephalic-presenting foetuses, between 37- and 41-week period of gestation, and who requested analgesia in labour were recruited for the study. Only those women who were having at least 2 regular contractions every 10 min with cervical dilatation of at least 3 cm were included. Patients with previous uterine scar, multiple gestation, malpresentation, absent membranes, antepartum haemorrhage, cephalopelvic disproportion, and pregnancies complicated with medical complications were excluded.

ing at least 2 regular contractions every 10 min with cervical dilatation of at least 3 cm were included. Patients with previous uterine scar, multiple gestation, malpresentation, absent membranes, antepartum haemorrhage, cephalopelvic disproportion, and pregnancies complicated with medical complications were excluded. A total of 80 women, 40 in each arm were recruited using computer generated random number table, as a convenient sample. Written consent was taken from these women once they crossed 37 weeks of gestation (i.e., once they reached term). All these women were informed about the study; however they were blinded as to which arm of study they belonged to. Among the women recruited for the pentazocine group nine women went for caesarean before the drug could be administered (five for nonreassuring labour admission test and four for failed induction), four women refused to participate in the study, and two within one hour of administration of pentazocine requested epidural analgesia. Thus, unfortunately, 15 women had to be excluded from the final analysis.

re the drug could be administered (five for nonreassuring labour admission test and four for failed induction), four women refused to participate in the study, and two within one hour of administration of pentazocine requested epidural analgesia. Thus, unfortunately, 15 women had to be excluded from the final analysis. The “pentazocine group” received 30 mg pentazocine (Ranbaxy) and the “tramadol group” received tramadol (Zydac Aliloc) 1 mg/ kg intramuscularly at request for labour analgesia. Clinical data and analgesic efficacy were assessed. Pain was assessed by a 10 cm long visual analogue scale with 0 representing no pain and 10 as the worst pain. Pain was then graded into mild (scores of 0–3), moderate (scores of 4–6), and severe (scores of 7–10). Pain was assessed before the administration of the drug (0 h), at 1 h, 2 h, and 4 h following drug administration, and at full dilatation. Simultaneously maternal pulse rate, blood pressure, respiratory rate, and foetal heart rate were recorded before administering the drug (0 h), at 1 h, 2 h, and 4 h, and at full dilatation. Side effects like sedation, vomiting, drowsiness, tachycardia, and foetal distress were noted following the administration of the drug. Maternal sedation was assessed on a three-point scale as 0 = alert, 1 = drowsy, and 2 = asleep. Intrapartum monitoring was done according to the standard labour ward protocol using the partogram. The time interval between drug administration and delivery was recorded. Neonatal evaluation was done by the neonatologist using APGAR score. The neonatologist was informed about the type of analgesia given to the mother. Naloxone usage for any presumed opioid induced respiratory depression was recorded.

the partogram. The time interval between drug administration and delivery was recorded. Neonatal evaluation was done by the neonatologist using APGAR score. The neonatologist was informed about the type of analgesia given to the mother. Naloxone usage for any presumed opioid induced respiratory depression was recorded. 2.1. Statistical Analysis Statistical analysis of the data was done using SPSS 11.0. Results were expressed as mean ± standard deviation (SD). Qualitative analysis was done using Student's t-test. For quantitative analysis Chi-square test was used. Nonparametric data were compared with Mann-Whitney U test. A P value of <0.05 was considered significant. 3. Results Of the total 65 patients who requested labour analgesia, 40 women were in the tramadol group and 25 in the pentazocine group. All of the patients in this study received only a single dose of either tramadol or pentazocine. Maternal characteristics like age, height, weight, blood pressure, parity, gestational age, and cervical dilatation at initiation of analgesia were comparable (Table 1).

3. Results Of the total 65 patients who requested labour analgesia, 40 women were in the tramadol group and 25 in the pentazocine group. All of the patients in this study received only a single dose of either tramadol or pentazocine. Maternal characteristics like age, height, weight, blood pressure, parity, gestational age, and cervical dilatation at initiation of analgesia were comparable (Table 1). Median pain score before administering the drug (at 0 h) was 8 in both tramadol and pentazocine groups. This score decreased to a median pain score of 7 in the tramadol group and 6 in the pentazocine group one hour after drug administration. Difference in pain relief between the two drugs was not significant (P > 0.05). Twenty-three women in the tramadol group and three in the pentazocine group delivered within four hours of the analgesic administration. In the patients who had not delivered the median pain score was 8 at 4 h in both groups. Thus, the analgesic effect of tramadol and pentazocine was comparable. At the end of the first stage the median pain score was 10 in both groups (Table 2).

tazocine group delivered within four hours of the analgesic administration. In the patients who had not delivered the median pain score was 8 at 4 h in both groups. Thus, the analgesic effect of tramadol and pentazocine was comparable. At the end of the first stage the median pain score was 10 in both groups (Table 2). On characterizing pain into mild, moderate, and severe, at 1 h after drug administration mild pain was experienced by one (4%) patient in the pentazocine group and none in the tramadol group. In the tramadol group, the majority of women (55%) rated pain as severe, whereas in the pentazocine group, the majority of women (60%) rated pain as moderately severe. However this difference was not statistically significant. At 4 h after drug administration mild pain was experienced by only one (4.5%) patient in the pentazocine group. The majority of women in both groups rated pain as severe. However only 88.2% women in the tramadol group experienced severe pain compared to 77.2% in the pentazocine group and this difference was statistically significant (P < 0.05) (Figure 1). The mode of delivery did not differ statistically between the two groups. Birth weight was also found to be comparable (2.86 ± 0.41 kg in the tramadol group and 2.94 ± 0.365 kg in the pentazocine group). Mean injection to delivery interval was shorter in the tramadol group as compared to the pentazocine group and the difference was statistically significant. Mean Apgar score of the neonates at 1 and 5 min in both groups were also comparable (Table 3).

in the tramadol group and 2.94 ± 0.365 kg in the pentazocine group). Mean injection to delivery interval was shorter in the tramadol group as compared to the pentazocine group and the difference was statistically significant. Mean Apgar score of the neonates at 1 and 5 min in both groups were also comparable (Table 3). The side effect profile of these two opioids was found to be not significant in our study. Only one lady had nausea and vomiting, while one felt drowsy. Both these women belonged to the pentazocine group. 4. Discussion In our study we found that the analgesic effect of the two drugs was not significantly different. Neither of these analgesics was effective towards the end of the first stage. However in the tramadol group, the majority of women (55%) rated pain as severe, whereas in the pentazocine group, the majority of women (60%) rated pain as moderately severe. There were not many side effects with either of the drugs in the given dosage. We also noticed that mean injection to delivery interval was shorter in the tramadol group as compared to the pentazocine group and the difference was statistically significant. Similarly, another study from India also noted that injection to delivery interval was significantly shorter in the tramadol group [6]. The same results were obtained from one more recent study wherein the investigators found that tramadol significantly shortens the duration of labour [7].

as statistically significant. Similarly, another study from India also noted that injection to delivery interval was significantly shorter in the tramadol group [6]. The same results were obtained from one more recent study wherein the investigators found that tramadol significantly shortens the duration of labour [7]. A randomized controlled trial done in Nigeria also showed results similar to us. They concluded that both pentazocine and tramadol are safe for the relief of labor pain. Pentazocine, however, provides better pain relief than tramadol in labor, but it is associated with higher incidence of drowsiness. There was no significant difference between the two drugs in outcome of labor and neonatal side effects [8]. Jain et al. compared intramuscular opioids with epidural analgesia in labour and concluded that, in developing nations where availability of facilities is the main limiting factor, intramuscular opioids can be considered suitable alternatives [9]. The limitations of this study are that opioids were administered only once in this study, the effect of which lasts for approximately 4 hours. Since the duration of labour is usually longer, the analgesic effect does not last long enough to provide adequate analgesia for the entire duration of labour. The sample size is small, so it is difficult to extrapolate these results into the general population.

udy, the effect of which lasts for approximately 4 hours. Since the duration of labour is usually longer, the analgesic effect does not last long enough to provide adequate analgesia for the entire duration of labour. The sample size is small, so it is difficult to extrapolate these results into the general population. The positive point in our study methodology was that none of the other studies comparing tramadol and pentazocine have used VAS for assessing the analgesic effect of these drugs. VAS is a measurement instrument used for the quantification of subjective characteristics that cannot be directly measured like pain perception. It has been suggested that this scaling system has superior metrical characteristics as compared to any other classification of pain (like mild, moderate, and severe). The other important finding was that all the previous studies have used 100 mg of tramadol, whereas we tried to tailor it with 1 mg/kg body weight and got similar efficacy. This concept might help reduce the adverse effect of the drug while still maintaining the desired analgesic effect.

ke mild, moderate, and severe). The other important finding was that all the previous studies have used 100 mg of tramadol, whereas we tried to tailor it with 1 mg/kg body weight and got similar efficacy. This concept might help reduce the adverse effect of the drug while still maintaining the desired analgesic effect. 5. Conclusion Tramadol and pentazocine both offer similar pain relief during labor and are safe for the mother and the baby. The analgesic effect of pentazocine was sustained up till 4 h as compared to tramadol where the analgesic effect wears off early. However tramadol seems to shorten the overall duration of labour. Pentazocine or tramadol can be given for labour pain relief as an alternative to epidural analgesia in resource poor setting. Further studies with multidose regimen of these analgesics are worth looking into. Conflict of Interests The authors declare that there is no conflict of interests regarding the publication of this paper. Figure 1 Pain characteristics at 1 h and 4 h after drug administration. *Pain perception at the end of 4 h was severe in 88.2% in the tramadol group while 77.2% perceived it as severe in the pentazocine group. This difference was statistically significant (P = 0.017). Table 1 Maternal characteristics among the two groups. Characteristics Tramadol

Figure 1 Pain characteristics at 1 h and 4 h after drug administration. *Pain perception at the end of 4 h was severe in 88.2% in the tramadol group while 77.2% perceived it as severe in the pentazocine group. This difference was statistically significant (P = 0.017). Table 1 Maternal characteristics among the two groups. Characteristics Tramadol n = 40 Pentazocine n = 25 P value Age (mean ± SD) 27.15 ± 2.99 26.68 ± 3.23 0.552 Height cm (mean ± SD) 155.63 ± 6.50 155.04 ± 5.35 0.708 Weight kg (mean ± SD) 64.93 ± 6.71 64.80 ± 6.55 0.941 Primigravida (%) Multigravida 25 (62.5) 15 (37.5) 19 (76) 06 (24) 0.326 Gestational age weeks (mean ± SD) 37.73 ± 1.62 38.24 ± 1.36 0.190 Cervical dilatation at initiation of analgesia cm (mean ± SD) 3.11 ± 0.737 3.12 ± 0.726 0.130 Systolic BP before analgesia mm Hg (mean ± SD) 123.2 ± 6.12 123.2 ± 7.21 1.000 Table 2 Comparison of pain score (VAS) among the two groups. Tramadol Pentazocine P value Median Range   Median Range Before injection (0 h) n = 40 8 4–10 n = 25 8 5–10 0.281 1 hour n = 40 7 4–10 n = 25 7 2–10 0.073 4 hours n = 17 8 5–10 n = 22 8 2–10 0.732 End of first stage n = 36 10 6–10 n = 25 10 6–10 0.914 Table 3 Labour characteristics and neonatal outcome among the two groups. Labour Tramadol n = 40 (%) Pentazocine

Tramadol Pentazocine P value Median Range   Median Range Before injection (0 h) n = 40 8 4–10 n = 25 8 5–10 0.281 1 hour n = 40 7 4–10 n = 25 7 2–10 0.073 4 hours n = 17 8 5–10 n = 22 8 2–10 0.732 End of first stage n = 36 10 6–10 n = 25 10 6–10 0.914 Table 3 Labour characteristics and neonatal outcome among the two groups. Labour Tramadol n = 40 (%) Pentazocine n = 25 (%) P value Induced 22 (55) 8 (32) 0.388 Normal vaginal delivery 26 (65) 23 (92) 0.080 Forceps delivery 5 (12.5) 1 (4) 0.080 Vacuum delivery 3 (7.5) 1 (4) 0.080 Caesarean section 6 (15) 0 Birth weight in kg (mean ± SD) 2.86 ± 0.41 2.94 ± 0.365 0.428 Mean APGAR (1 min) 8.93 9.00 Mean APGAR (5 min) 9.98 10.00 Injection delivery interval in min (mean ± SD) 265.91 ± 103.35 430.28 ± 117.29 0.0001* (*P value < 0.05; ie: statistically significant).

1. Introduction Premenstrual syndrome (PMS) is one of the most common problems in women at their reproductive age [1, 2]. PMS is defined as the recurrent mood and physical symptoms which is usually in the luteal phase, and it remits in the follicular phase of the menstrual cycle [3–6]. There is a high prevalence of PMS; about 80% of women reported mild premenstrual symptoms, 20%–50% reported moderate symptoms, and about 5% of women had severe symptoms [7, 8]. Despite the high incidence of premenstrual syndrome, causes of it have not been clear and several etiologies have been suggested (e.g., hormonal change, neurotransmitters, prostaglandins, diet, drugs, and lifestyle) [7, 9–14]. Although the exact etiology of PMS is not known, a wide range of therapeutic interventions have been tested to treat premenstrual symptoms (e.g., lifestyle changes, pharmacological intervention, and nonpharmacological treatments) [15–22]. Due to the side effects of chemical drugs, except severe cases, chemical drugs consumption is not recommended. Today, complementary and herbal medicine are commonly used in the treatment of many chronic conditions such as PMS, menopausal symptoms, and dysmenorrhea [17, 20, 23–27].

onpharmacological treatments) [15–22]. Due to the side effects of chemical drugs, except severe cases, chemical drugs consumption is not recommended. Today, complementary and herbal medicine are commonly used in the treatment of many chronic conditions such as PMS, menopausal symptoms, and dysmenorrhea [17, 20, 23–27]. Ginger is commonly used in the herbal medicine and traditionally it used to treat dysmenorrhea [28, 29]. Zingiber officinale is the scientific name of plant Ginger [30]. Ginger rhizome is used in traditional medicine [31]. Studies showed beneficial effects of ginger on vomiting, nausea, motion sickness, arthritis, migraine, headache, and so forth [32]. In addition, studies have shown that the ginger can modulate prostaglandins system [33–35]. With regard to the role of prostaglandins in PMS and effect of ginger on modulation of prostaglandins system, the aim of this study was evaluating the effects of ginger on the severity of the PMS symptoms. 2. Materials and Methods This clinical trial was a randomized, double-blinded, and placebo-controlled study. The study was done on 70 female students in dormitories of Tehran University of Medical Sciences in the year of 2013. Data were collected during 7 months. The ethics committee of Tehran University of Medical Sciences approved the study (No. 97/130/D/92) and it was registered at Iranian Registry of Clinical Trial (IRCT No. 201301012751N7).

n 70 female students in dormitories of Tehran University of Medical Sciences in the year of 2013. Data were collected during 7 months. The ethics committee of Tehran University of Medical Sciences approved the study (No. 97/130/D/92) and it was registered at Iranian Registry of Clinical Trial (IRCT No. 201301012751N7). Inclusion criteria were as follows: being healthy premenopausal women aged between 18 and 35 years, having regular menstrual cycles of 21–35 days, being single, lacking sensitivity to ginger, not taking any medication, not drinking alcohol, not smoking, and not having stressful events in the last 3 months.

n 70 female students in dormitories of Tehran University of Medical Sciences in the year of 2013. Data were collected during 7 months. The ethics committee of Tehran University of Medical Sciences approved the study (No. 97/130/D/92) and it was registered at Iranian Registry of Clinical Trial (IRCT No. 201301012751N7). Inclusion criteria were as follows: being healthy premenopausal women aged between 18 and 35 years, having regular menstrual cycles of 21–35 days, being single, lacking sensitivity to ginger, not taking any medication, not drinking alcohol, not smoking, and not having stressful events in the last 3 months. The participants recorded symptoms with daily record questionnaires (this form contains a table with 19 symptoms of premenstrual syndrome questionnaire based on the DSM-IV (the fourth edition of the Diagnostic and Statistical Manual of Mental Disorders of the American Psychiatric Association), Self-Rating Scale) for two cycles before the intervention. This questionnaire determines the severity of PMS using 3 items, including mood symptoms (restlessness; irritability; anxiety, depression, or sadness; crying; feeling of isolation), physical symptoms (headache, breast tenderness, backache, abdominal pain, weight gain, swelling of extremities, muscle stiffness, gastrointestinal symptoms, and nausea), and behavioral characteristics (fatigue, lack of energy, insomnia, difficulty in concentrating, increased or decreased appetite). Then the severity of premenstrual syndrome was evaluated for all participants ((0) absence of symptoms, (1) mild symptoms that may not interfere with everyday activities, (2) moderate symptoms that interfere with daily activities, and (3) severe symptoms that impede doing daily activities). Each participant with at least five symptoms was finally diagnosed as a person with PMS. After identifying patients with PMS, they were randomly divided into two groups (n = 35). The first group received ginger capsule and the second group received placebo capsule. Doses of ginger and placebo were 250 mg/12 hours and given from 7 days before and until 3 days after onset of menstrual bleeding. The subjects completed the daily record questionnaire at their first, second, and third menstrual cycles. After interventions, again the severity of PMS was evaluated. Participants and researcher were blind about the kind of drug throughout the study. Exclusion criteria were incidence of side effects drugs or drugs allergy, use of other drugs, drug use disorder, identifying any diseases during the study, getting married during the study, menstrual irregularities, and irregular bleeding event during the study.

r were blind about the kind of drug throughout the study. Exclusion criteria were incidence of side effects drugs or drugs allergy, use of other drugs, drug use disorder, identifying any diseases during the study, getting married during the study, menstrual irregularities, and irregular bleeding event during the study. Statistical analysis was performed using software SPSS-18. Intergroup analyses were performed to compare the total score of PMS, mood, and behavioral and physical symptoms before and after the intervention. Intergroup analysis was performed by independent samples t-test. P < 0.05 was considered significant. 3. Results Out of the 70 students who participated in the study 2 persons in the ginger group and 2 persons in the placebo group did not complete the study (Figure 1). In the ginger group two samples were excluded, one because of incidence of side effect and second one due to misuse of ginger. In the placebo group reason for exclusion for each of the 2 people was drug use disorder. No significant differences between two groups were seen with regard to demographic characteristics and menstrual history data (Table 1).

3. Results Out of the 70 students who participated in the study 2 persons in the ginger group and 2 persons in the placebo group did not complete the study (Figure 1). In the ginger group two samples were excluded, one because of incidence of side effect and second one due to misuse of ginger. In the placebo group reason for exclusion for each of the 2 people was drug use disorder. No significant differences between two groups were seen with regard to demographic characteristics and menstrual history data (Table 1). There were no significant differences regarding the total score of PMS, severity of mood, and physical and behavioral symptoms between the two groups before the intervention (resp., P = 0.71, P = 0.69, P = 0.56, and P = 0.76) (Table 2). Statistical analysis demonstrated that there were significant differences in total score of PMS, severity of mood, and physical and behavioral symptoms between the two groups after the intervention, and ginger could reduce severity symptom of PMS (Table 2).

(resp., P = 0.71, P = 0.69, P = 0.56, and P = 0.76) (Table 2). Statistical analysis demonstrated that there were significant differences in total score of PMS, severity of mood, and physical and behavioral symptoms between the two groups after the intervention, and ginger could reduce severity symptom of PMS (Table 2). 4. Discussion The findings of this study showed that ginger could significantly reduce the total score of PMS, severity of mood, and physical and behavioral symptoms of the first month intervention. So according to the availability and the safety of ginger, the ginger can be an appropriate treatment in reducing symptoms of premenstrual syndrome. One of the proposed mechanisms that lead to premenstrual syndrome is change in prostaglandin system [7, 16, 36]. Ginger through the inhibition of the metabolism of cyclooxygenase and lipoxygenase prevents the production of prostaglandins [35]. To our knowledge, this study is the first report about the effect of ginger on premenstrual syndrome. Rahnama et al. [28] and Ozgoli et al. [29] showed that the use of ginger is effective in reducing symptoms of dysmenorrheal [28, 29]. Primary dysmenorrhea is caused by excessive production of prostaglandins from the endometrial tissue, and 80% of cases of dysmenorrhea with prostaglandin inhibitors can be improved [29]. Because some ginger compounds are prostaglandin inhibitors and are effective on dysmenorrhea, so perhaps through this mechanism they could be effective on other problems of menstruation cycle [28, 29]. Some of symptoms of PMS like pain are common with dysmenorrhea (such as backache and abdominal pain). In our study these symptoms of PMS have been reduced.

staglandin inhibitors and are effective on dysmenorrhea, so perhaps through this mechanism they could be effective on other problems of menstruation cycle [28, 29]. Some of symptoms of PMS like pain are common with dysmenorrhea (such as backache and abdominal pain). In our study these symptoms of PMS have been reduced. In women with PMS, severe headache outbreaks during the luteal phase and an increase in headache resistance to painkillers and anti-inflammatory drugs lead to increase in depression, irritability, anxiety, anger, and food intolerance during the luteal phase [37–39]. Cady et al. [40] reported that ginger administration caused a significant deceleration in severity of headache in patients with migraine [40]. One of the symptoms of PMS is incidence of headache and exacerbation of migraine. In our study ginger was found to be effective in relieving headaches. Levine et al. reported that the ginger reduced the delayed nausea of chemotherapy and reduced use of antiemetic medications [41]. Ginger was effective in treating nausea and vomiting during pregnancy [31, 42]. Study of K. S. Naeine and S. S. Naeine [43] showed that the use of 1 gram of powdered ginger, 1 hour before taking contraceptive pills, induced reduction in contraceptive pills nausea [43]. Nausea is one of symptoms of PMS; in our study the incidence of gastrointestinal disturbances and nausea after taking ginger reduced.

dy of K. S. Naeine and S. S. Naeine [43] showed that the use of 1 gram of powdered ginger, 1 hour before taking contraceptive pills, induced reduction in contraceptive pills nausea [43]. Nausea is one of symptoms of PMS; in our study the incidence of gastrointestinal disturbances and nausea after taking ginger reduced. Studies showed that ginger has anti-inflammatory properties and is effective in treatment of pain in patients with osteoarthritis, muscle pain [30, 33, 44]. Joint and muscle pain are also symptoms of PMS; in the present study, ginger was effective in relieving severity of these symptoms of PMS. The only side effect observed in the present study was nausea in the group of ginger. In addition, the use of ginger in traditional medicine to treat colds, fever, sore throat, nausea, stomach upset, muscle aches, and arthritis and for cancer prevention is effective [32]. Therefore, the use of ginger in the treatment of PMS can also benefit from other advantages of ginger.

nausea in the group of ginger. In addition, the use of ginger in traditional medicine to treat colds, fever, sore throat, nausea, stomach upset, muscle aches, and arthritis and for cancer prevention is effective [32]. Therefore, the use of ginger in the treatment of PMS can also benefit from other advantages of ginger. 5. Conclusions The findings of the present study demonstrated the likelihood of usefulness of ginger in treating PMS while no specific side effects have been seen, and consumption of it is associated with health benefits. Ginger is readily available because of its low cost and, so, can be widely used in the treatment of premenstrual syndrome. According to increased tendency in the use of traditional medicine and herbal medicine, especially for people who have no desire to use chemical drugs, with more side effects, use of ginger, with very low side effects, is very useful. Furthermore, we cannot find study that evaluates effect of ginger on PMS; we suggest doing more studies with larger number of patients concerning the efficacy and safety of different doses of ginger. Acknowledgments This work was supported by the Tehran University of Medical Sciences. The authors thank the Department of Student and Research Assistance of Tehran University of Medical Sciences, and Golestan 8 and Vesal dormitories' administrators, and students residing in Golestan 8 and Vesal dormitories for their nice assistance in data collection. (This paper is relevant to the midwifery master degree thesis of Samira khayat.)

ment of Student and Research Assistance of Tehran University of Medical Sciences, and Golestan 8 and Vesal dormitories' administrators, and students residing in Golestan 8 and Vesal dormitories for their nice assistance in data collection. (This paper is relevant to the midwifery master degree thesis of Samira khayat.) Conflict of Interests The authors declare that there is no conflict of interests regarding the publication of this paper. Figure 1 Trial profile. Table 1 The demographic characteristics and menstrual history of the participants. Characteristics Group Test results Control Ginger Mean ± SD Mean ± SD Age (year) 25.52 ± 3.13 24.76 ± 3.17 P = 0.33 Weight (kg) 59.61 ± 9.69 57.55 ± 7.4 P = 0.24 Height (cm) 160.9 ± 6.66 161.4 ± 5.72 P = 0.92 BMI (kg/m2) 23.33 ± 3.44 22.13 ± 2.95 P = 0.1 Age at menarche (year) 13.48 ± 1.25 13.21 ± 1.31 P = 0.39 Duration of cycle (day) 29 ± 1.42 28.7 ± 1.6 P = 0.46 Duration of menstruation (day) 5.3 ± 1.46 5.8 ± 1.36 P = 0.1 Table 2 Comparison of PMS scores in ginger and placebo groups before and after the intervention.

0.92 BMI (kg/m2) 23.33 ± 3.44 22.13 ± 2.95 P = 0.1 Age at menarche (year) 13.48 ± 1.25 13.21 ± 1.31 P = 0.39 Duration of cycle (day) 29 ± 1.42 28.7 ± 1.6 P = 0.46 Duration of menstruation (day) 5.3 ± 1.46 5.8 ± 1.36 P = 0.1 Table 2 Comparison of PMS scores in ginger and placebo groups before and after the intervention. Symptoms Group Test results Control Ginger Mean ± SD Mean ± SD Unpaired test Total severity of PMS Before intervention 106.7 ± 44.65 110.2 ± 30.77 P = 0.71 One month after intervention 105.7 ± 45.76 51.18 ± 32.76 P < 0.0001 Two months after intervention 107.2 ± 50.68 49.48 ± 33.12 P < 0.0001 Three months after intervention 106 ± 48.74 47.06 ± 33.41 P < 0.0001 Mood symptoms Before intervention 37.42 ± 17.37 38.97 ± 14.16 P = 0.69 One month after intervention 37.18 ± 16.13 16.39 ± 10.54 P < 0.0001 Two months after intervention 37.61 ± 18.58 14.61 ± 8.84 P < 0.0001 Three months after intervention 38.37 ± 20.31 13.45 ± 10.65 P < 0.0001 Physical symptoms Before intervention 42.64 ± 23.83 45.76 ± 19.76 P = 0.56 One month after intervention 43.48 ± 24.25 21.85 ± 22.54 P = 0.0004 Two months after intervention 43.42 ± 24.13 21.12 ± 20.31 P < 0.0001 Three months after intervention 42.06 ± 22.76 22.76 ± 19.62 P = 0.0005 Behavioral symptoms Before intervention 26.64 ± 16.20 25.42 ± 16.05 P = 0.76 One month after intervention 25.06 ± 17.86 12.94 ± 12.35 P = 0.002 Two months after intervention 26.15 ± 19.08 13.76 ± 13.79 P = 0.003 Three months after intervention 25.61 ± 18.47 10.85 ± 13.05 P = 0.0004

1. Introduction Agenesis or premature closure of the ductus arteriosus is an uncommon anomaly. The defect, when present, is usually associated with Tetrology of Fallot (TOF), absent pulmonary valve, truncus arteriosus, or maternal use of prostaglandin synthetase inhibitors for the arrest of premature labor. Isolated premature closure of the ductus with or without congenital heart disease can result in right sided heart failure and fetal hypoxemia resulting in hydrops fetalis. We present a case involving the prenatal diagnosis of agenesis of the ductus arteriosus and Tetrology of Fallot complicating a twin pregnancy.

1. Introduction Agenesis or premature closure of the ductus arteriosus is an uncommon anomaly. The defect, when present, is usually associated with Tetrology of Fallot (TOF), absent pulmonary valve, truncus arteriosus, or maternal use of prostaglandin synthetase inhibitors for the arrest of premature labor. Isolated premature closure of the ductus with or without congenital heart disease can result in right sided heart failure and fetal hypoxemia resulting in hydrops fetalis. We present a case involving the prenatal diagnosis of agenesis of the ductus arteriosus and Tetrology of Fallot complicating a twin pregnancy. 2. Case Report A 34-year-old African American woman Gravida-4, Para-3, presented for prenatal care at 15 weeks of gestation by her last menstrual period. She had no significant pastobstetric, medical, or family history. Comprehensive sonographic evaluation at 18 weeks gestation revealed a diamniotic dichorionic twin gestation, cleft lip in Twin B, and large echogenic intracardiac foci in the left ventricles of both twins. A fetal echocardiographic study revealed the following. Twin A was noted to have an echogenic focus in the left ventricle with evidence of ventricular septal defect, overriding aorta, and a small dysplastic pulmonary valve with pulmonary regurgitation. Twin B was also noted to have an echogenic focus in the left ventricle with evidence of ventricular septal defect, overriding aorta, and a small pulmonary valve characteristic of TOF. The patient underwent genetic consultation, but declined amniocentesis for kayotypic analysis. Subsequent echocardiographic evaluation performed at 27 weeks gestation revealed the following. Twin B was noted to have decreased right ventricular function, bradycardia with absent end diastolic flow in the umbilical artery and hydrops. Twin A was also noted to have frequent episodes of bradycardia. Due to the findings of fetal hydrops, and nonreassuring antenatal testing, a cesarean section was performed after administration of steroid therapy.

right ventricular function, bradycardia with absent end diastolic flow in the umbilical artery and hydrops. Twin A was also noted to have frequent episodes of bradycardia. Due to the findings of fetal hydrops, and nonreassuring antenatal testing, a cesarean section was performed after administration of steroid therapy. At birth, Twin B weighed 820 grams and was noted to have a cleft lip and penile hypospadias; Apgars at one and five minutes were of 4 and 6, respectively. The infant was subsequently electively intubated and ventilated. Initial arterial blood gas revealed mild metabolic acidosis. Cardiac evaluation revealed the patient to be in sinus rhythm and echocardiography confirmed the diagnosis of Tetralogy of Fallot with mild right ventricular outflow tract obstruction; no ductal shunt was appreciated (Figure 1). On day two of life, the neonate was noted to have decreasing oxygen saturation associated with hypotension and metabolic acidosis. Echocardiogram revealed increasing infundibular stenosis and right ventricular outflow tract gradient above 40 mm Hg. Ductal flow was still not demonstrated which was unusual given the gestational age and development of hypoxemia. Dopamine at 10 microgram/kg/min and Prostaglandin E-1 (PGE-1) at 0.05 microgram/kg/min was initiated with minimal clinical improvement. Cardiothoracic consultation from an adjacent tertiary care center was requested, however given the patient's weight and unstable condition surgical intervention was not considered an option. There was progressive deterioration in the form of oliguric renal failure with anasarca and severe metabolic acidosis and the infant expired on day 7 of life. Karyotyping revealed 46 XY genotype with no evidence of micro- or macrodeletions of long arm of chromosome-22. Autopsy confirmed the ante-mortem cardiac findings of TOF and the pulmonary artery was markedly diminished in size. The ductus arteriosus could not be demonstrated (Figure 2).

infant expired on day 7 of life. Karyotyping revealed 46 XY genotype with no evidence of micro- or macrodeletions of long arm of chromosome-22. Autopsy confirmed the ante-mortem cardiac findings of TOF and the pulmonary artery was markedly diminished in size. The ductus arteriosus could not be demonstrated (Figure 2). Twin A at birth weighed 790 grams, had one and five minute Apgars of 4 and 6, respectively and was also electively intubated and ventilated. Arterial blood gas did not reveal any metabolic acidosis. Echocardiography confirmed Tetrology of Fallot and pulmonary regurgitation. The main pulmonary artery annulus was small and measured 2.5 mm and the distal branch of pulmonary arteries measured 4.5 mm (Figure 3). No flow was appreciable across the ductus arteriosus. Initially oxygen saturations were within low normal range, however after four days of life, the neonate began to have decreasing values. Echocardiogram revealed increasing subinfundibular and valvar stenosis with gradients increasing to 60–80 mm hg over time. PGE-1 drip at 0.1 mcg/kg/min was initiated with no significant improvement in the saturations and no ductal flow was demonstrated on the echocardiogram. As with Twin B, cardiothoracic surgical intervention was sought but declined given the high risk. The patient's clinic course progressively worsened with progressive metabolic acidosis, cardiac decompensation, and sepsis. The infant expired on day 30 of life. Normal 46 XY karyotype was noted. The parents declined autopsy.

with Twin B, cardiothoracic surgical intervention was sought but declined given the high risk. The patient's clinic course progressively worsened with progressive metabolic acidosis, cardiac decompensation, and sepsis. The infant expired on day 30 of life. Normal 46 XY karyotype was noted. The parents declined autopsy. 3. Discussion Absence of the ductus arteriosis was first described in 1671 by Stensen in a fetus with TOF [1, 2]. Most such cases of TOF with absent ductus have the associated absent pulmonary valve syndrome and aneurysmally dilated pulmonary arteries. [3, 4]. The usual presentation with this condition is increasing respiratory distress due to compression of the bronchi. [3]. Another form of presentation is heart failure or hydrops diagnosed in utero [5, 6]. Both our patients had TOF variants but presented with progressive cyanosis not responding to PGE 1 due to absent ductus arteriosus. Twin A had TOF with a dysplastic pulmonary valve with moderate stenosis and regurgitation but over time developed increasing infundibular stenosis, cyanosis and right ventricular failure rather than progressive respiratory distress as is typically described in the TOF/absent pulmonary valve syndrome. Twin B had typical features of TOF but presented with right ventricular dysfunction and early hydrops requiring urgent delivery. Absent ductus arteriosus was suspected only after birth, when there was no response to prostaglandin. Both patients became progressively cyanotic over time and failed to improve with increasing doses of PGE-1.

features of TOF but presented with right ventricular dysfunction and early hydrops requiring urgent delivery. Absent ductus arteriosus was suspected only after birth, when there was no response to prostaglandin. Both patients became progressively cyanotic over time and failed to improve with increasing doses of PGE-1. This case brings to the literature the association of TOF and absent or premature closure of the ductus not associated with usual pulmonary regurgitation and aneurysmal pulmonary arteries. This association should be considered and looked for specifically during in utero studies as premature delivery may not be a viable option as was in our case. Although we were not able to do so in our case, it is possible to make the diagnosis of ductal closure in utero by Doppler flow studies as described by Huhta et al. [7]. Moreover, if congenital heart disease is associated with myocardial dysfunction especially of the right ventricle and hydrops, absent ductus arteriosus should be considered, as it is not usual for congenital heart disease to be associated with cardiac dysfunction unless there is an associated arrhythmia, heart block, or myocarditis. The case emphasizes the need to consider absent or premature ductal closure in a fetus with hydrops of unknown etiology and especially associated with any variant of TOF. Figure 1 Echocardiogram of Twin B showing overriding aorta and right ventricular hypertrophy. Figure 2 Autopsy of Twin B heart and lungs showing small sized pulmonary artery and right ventricular hypertrophy.

This case brings to the literature the association of TOF and absent or premature closure of the ductus not associated with usual pulmonary regurgitation and aneurysmal pulmonary arteries. This association should be considered and looked for specifically during in utero studies as premature delivery may not be a viable option as was in our case. Although we were not able to do so in our case, it is possible to make the diagnosis of ductal closure in utero by Doppler flow studies as described by Huhta et al. [7]. Moreover, if congenital heart disease is associated with myocardial dysfunction especially of the right ventricle and hydrops, absent ductus arteriosus should be considered, as it is not usual for congenital heart disease to be associated with cardiac dysfunction unless there is an associated arrhythmia, heart block, or myocarditis. The case emphasizes the need to consider absent or premature ductal closure in a fetus with hydrops of unknown etiology and especially associated with any variant of TOF. Figure 1 Echocardiogram of Twin B showing overriding aorta and right ventricular hypertrophy. Figure 2 Autopsy of Twin B heart and lungs showing small sized pulmonary artery and right ventricular hypertrophy. Figure 3 Echocardiogram of Twin A showing small pulmonary annulus and dilated distal branch of pulmonary artery.

1. Introduction The presence of adnexal masses during pregnancy ranges from 1 to 81 to 1 in 2,500 pregnancies, but only 3% of these masses are malignant. Krukenberg tumor is an ovarian metastasis of a gastric tumor and accounts for 1%—2% of all ovarian tumors [1]. These tumors during pregnancy are even rarer, as the incidence of gastric cancer in women of reproductive age group is only 0.4%-0.5% [2]. Thus they are a diagnostic and treatment challenge to the physician. We present a case report of a woman 38 weeks pregnant with massive bilateral Krukenberg tumors, which had an unfortunate outcome despite of treatment. 2. Case Report A 30-year-old woman, gravida 3 para 2, from a very remote community was referred at 38 weeks to our tertiary care center due to the presence a bilateral ovarian masses and shortness of breath. The patient complained of history of 1 year before the pregnancy with mild bloating and early satiety that was attributed to gastritis and gastroesophageal reflux, but despite the symptoms she never saught for medical attention. A pelvic sonogram suggested that the ovarian tumors were malignant. Decision to proceed with exploratory laparotomy and cesarean section was taken. Circulating levels were as follows: testosterone (0.93 μg/L, normal range: <0.62 μg/L), progesterone (22.29 nmol/L, normal range: 0.64–2.58 nmol/L), and 17-OH-progesterone (4.96 μg/L, normal range: 0.40–1.02 μg/L), and the tumor markers levels were 43.4 U/mL (CA 19–9), 1.7 ng/mL (carcinoembrionic antigen) 12.8 U/mL (CA72–4) and 33.9 U/mL (CA 125), respectively.

erone (0.93 μg/L, normal range: <0.62 μg/L), progesterone (22.29 nmol/L, normal range: 0.64–2.58 nmol/L), and 17-OH-progesterone (4.96 μg/L, normal range: 0.40–1.02 μg/L), and the tumor markers levels were 43.4 U/mL (CA 19–9), 1.7 ng/mL (carcinoembrionic antigen) 12.8 U/mL (CA72–4) and 33.9 U/mL (CA 125), respectively. Intraoperatively she was found to have two bilateral adnexal masses measuring 15 cm in its greatest diameter for the right ovary and 22 cm for the left (Figure 1) and also a 10 cm tumor in the major gastric curvature that resembled linitis plastica. She underwent an elective cesarean delivery and debulking surgery including bilateral oophorectomy omentectomy, and gastric tumor biopsy. Histopathology of the gastric biopsy was consistent with the presence of poorly differentiated tumor cells corroborating the diagnosis of linitis plastica. Since debulking does not improve the prognosis we decided against debulking. In the postoperative period the patient was started on chemotherapy with 6 cycles of chemotherapy using 5-fluorouracil (2000 mg/m2) and oxaliplatin (50 mg/m2) in a weekly schedule. Despite all our efforts the patient succumbed to disease in 3 months. 3. Discussion Krukenberg tumor refers to gastrointestinal cancer metastatic to the ovaries, accounting for 1%-2% of all ovarian tumors.

In the postoperative period the patient was started on chemotherapy with 6 cycles of chemotherapy using 5-fluorouracil (2000 mg/m2) and oxaliplatin (50 mg/m2) in a weekly schedule. Despite all our efforts the patient succumbed to disease in 3 months. 3. Discussion Krukenberg tumor refers to gastrointestinal cancer metastatic to the ovaries, accounting for 1%-2% of all ovarian tumors. The eponym of this condition was given due by Krukenberg initially described in 1896 and the criteria given were (1) the presence of a tumor in the ovary, (2) evidence of intracellular mucin secretion by the formation of signet cells, and (3) diffuse infiltration of stroma giving a sarcoma-like appearance [2]. A search in English, Spanish, and French literature revealed four reports of bilateral ovarian Krukenberg tumors, with our case being the fifth in the literature. The cornerstone of the management of these tumors is the diagnosis of the gastrointestinal primary tumor, and also the prognosis worsens when the primary tumor is identified after the metastasis to the ovary is discovered [3]. Due to the rare nature of these tumors, there is no current standardization for the diagnosis and the treatment.

The cornerstone of the management of these tumors is the diagnosis of the gastrointestinal primary tumor, and also the prognosis worsens when the primary tumor is identified after the metastasis to the ovary is discovered [3]. Due to the rare nature of these tumors, there is no current standardization for the diagnosis and the treatment. Also due to the rarity of the condition it is not appropriate to comment on the effect of tumor on pregnancy and vice versa. There are only 3 cases of pregnancy-related Krukenberg the tumors in the literature; of these also 2 were detected postpartum, and this is the only tumor that is not associated to virilization. Some authors are of the opinion that there might be an increased risk of acute abdomen secondary to tumor torsion or rupture [4]. The role of debulking surgery and chemotherapy with platinum-based chemotherapy can be reasonable, and even relatively safe to be administered during pregnancy. But despite the interventions, usually the discovery of the presence of masses of the size that our patient exhibited the prognosis is poor as it generally represents an advanced stage disease, However, possible early detection with debulking surgery, possible hysterectomy with/without delivery, and platinum-based chemotherapy may improve the survival of these patients [5, 6]. Figure 1 Presence of a bilateral ovarian tumor and a full-term gravid uterus.

The differential diagnosis for a dorsal midline mass presenting in fetuses and newborns includes a wide range of pathological conditions, consisting of spinal dysraphisms, teratoma, hamartoma, disturbances in regression of fetal tail, and pseudotail formation [1]. Posterior meningocele is the least common form of spina bifida. It appears as a sac or cyst which contains cerebrospinal fluid. The majority of cases have no underlying bony defect or communication to the meninges. The spinal cord and nerves are not involved and their function is normal. Its incidence is 1 : 1000 live births [2, 3]. Apart from spina bifida, causes of meningocele include teratoma and other tumors of the sacrococcyx and of the presacral space and Currarino syndrome. Usually a meningocele has no negative long-term effects, although there are reports of tethered cord. We present a case of unique posterior meningocele diagnosed at the early second trimester anatomical scan using 2D and 3D ultrasound. A healthy 29-year-old woman presented to our ultrasound unit at her first pregnancy for a routine anatomical scan at 15-week gestation.

Apart from spina bifida, causes of meningocele include teratoma and other tumors of the sacrococcyx and of the presacral space and Currarino syndrome. Usually a meningocele has no negative long-term effects, although there are reports of tethered cord. We present a case of unique posterior meningocele diagnosed at the early second trimester anatomical scan using 2D and 3D ultrasound. A healthy 29-year-old woman presented to our ultrasound unit at her first pregnancy for a routine anatomical scan at 15-week gestation. The anatomical scan revealed a small cystic lesion with very tiny stalk close to the caudal aspect of the spine, “floating" in the amniotic fluid, with no flow when Doppler was applied. This lesion was suspected to be meningocele, and based on its appearance we have termed it as the “Lasso sign". No opened spina bifida was noted. 3D ultrasound using sectional planes and surface rendering was performed, and it demonstrated the same lesion and confirmed it to be meningocele alone. The anatomical scan of the brain including the posterior fossa was normal. During the present pregnancy she performed nuchal translucency of 0.8 mm. The triple test screen was normal and consisted of alpha-fetoprotein level of 0.95 MOM. The patient did not perform MRI due to claustrophobia.

The anatomical scan revealed a small cystic lesion with very tiny stalk close to the caudal aspect of the spine, “floating" in the amniotic fluid, with no flow when Doppler was applied. This lesion was suspected to be meningocele, and based on its appearance we have termed it as the “Lasso sign". No opened spina bifida was noted. 3D ultrasound using sectional planes and surface rendering was performed, and it demonstrated the same lesion and confirmed it to be meningocele alone. The anatomical scan of the brain including the posterior fossa was normal. During the present pregnancy she performed nuchal translucency of 0.8 mm. The triple test screen was normal and consisted of alpha-fetoprotein level of 0.95 MOM. The patient did not perform MRI due to claustrophobia. One of the complications of meningocele is tethered cord, caused by pathological fixation of the spinal cord, resulting in traction on the neural tissue which, in turn, leads to ischemia and progressive neurological deterioration. The prenatal diagnosis of tethered cord is based on ultrasound evaluations of the spinal cord and the conus medullaris at the level of L2-L3 at the second and third trimester [4, 5]. Repeated sonograms in our case demonstrated the same lesion at L3-L4 (see Figure 1 and supplementary movie Lasso Movie 1 in Supplementary Material available online at doi:10.5402/2011/175916) without sonographic signs of tethered cord.

the conus medullaris at the level of L2-L3 at the second and third trimester [4, 5]. Repeated sonograms in our case demonstrated the same lesion at L3-L4 (see Figure 1 and supplementary movie Lasso Movie 1 in Supplementary Material available online at doi:10.5402/2011/175916) without sonographic signs of tethered cord. At 40 + 2 weeks of gestation the patient underwent a cesarean section because of arrest of dilatation. A 3384gr female infant was delivered with Apgar scores of 9 and 10 at 1 minute and 5 minutes, respectively.

the conus medullaris at the level of L2-L3 at the second and third trimester [4, 5]. Repeated sonograms in our case demonstrated the same lesion at L3-L4 (see Figure 1 and supplementary movie Lasso Movie 1 in Supplementary Material available online at doi:10.5402/2011/175916) without sonographic signs of tethered cord. At 40 + 2 weeks of gestation the patient underwent a cesarean section because of arrest of dilatation. A 3384gr female infant was delivered with Apgar scores of 9 and 10 at 1 minute and 5 minutes, respectively. The neonate's physical exam revealed an atretic string of about 2 cm on the lower aspect of her back in the midline, confirmed by a neurosurgeon to be meningocele. Additional evaluation included ultrasound of the brain which was normal, ultrasound of the sacrum which ruled out tethered cord, and X-ray of the spine which demonstrated normal structure of the vertebrae. Movements of four limbs and muscle tone were normal. There was no leakage of urine, and the tonus of the anal sphincter was good. The string dried out and fell 12 hours after birth. Pathological evaluation was not performed; therefore, the diagnosis was based on the prenatal ultrasound performed by gynecologists and pediatric neurosurgeons and by the clinical examination after birth by the pediatric neurosurgeon. Photos of the back of the infant taken at the age of 2 days were unremarkable, apart from a tiny scar at the middle of her lower back that disappeared after a few days. Followup examination including neurological examination at the age of two months and four months was unremarkable. MRI performed at the age of four months was normal.

ack of the infant taken at the age of 2 days were unremarkable, apart from a tiny scar at the middle of her lower back that disappeared after a few days. Followup examination including neurological examination at the age of two months and four months was unremarkable. MRI performed at the age of four months was normal. In summary, detection of the “Lasso” sign may indicate the diagnosis of posterior meningocele. Thorough evaluation of the fetal brain and spinal cord to exclude Chiari II malformation and spina bifida is mandatory. The use of colour Doppler to rule out the possibility of umbilical cord as part of the examination is necessary. Level of alpha-fetoprotein should be examined. Prenatal followup should include repeated sonographic evaluation of the brain and spinal cord with emphasis on the conus medullaris to rule out tethered cord. 3D ultrasound may aid both in the diagnosis and followup. MRI may also be offered in order to aid in the diagnosis. In general, posterior meningocele has no long-term effect. “Lasso" appearing meningocele should be included in the differential diagnosis of lower spine posterior cystic lesions. Figure 1 (a) Transverse 2D views of the lower spine revealing posterior meningocele at 15 weeks. (b) Sagittal 2D view of the posterior meningocele at 22 + 3 weeks. (c) Sagittal 3D view of the posterior meningocele at 22 + 3 weeks.

1. Introduction Neural tube defects (NTDs), including spina bifida, anencephaly, and encephalocele, are among the most common birth defects and cause morbidity and mortality among fetuses and babies [1]. The incidence of NTDs in the United States is approximately one per 1,000 pregnancies. Approximately one third of 4,000 pregnancies with NTDs end with spontaneous or induced abortion, so there are about 2,500 live births with NTD annually in the US [2]. There is a significant geographical variation in the rate of NTDs [3]. In the years 1999-2000 there were at least 394 cases of isolated NTD in Israel, with a nearly fourfold incidence among non-Jews compared to Jews [4]. The embryonic basis of NTD is not fully understood, but a common abnormality is lack of closure of the neural tube [3]. The normal development and closure of the neural tube are completed by the 25th day after fertilization. The mortality rate among newborns with spina bifida is 10% in the United States, but nearly 100% in China [3]. Among surviving babies there is a high rate of neurological complications such as paralysis, hydrocephalus, Arnold Chiari type II and syringomyelia [3]. The known risk factors for the development of NTD are low socioeconomical class, maternal diabetes, antiepileptic drugs, fever in the early stage of pregnancy, and obesity [3].

viving babies there is a high rate of neurological complications such as paralysis, hydrocephalus, Arnold Chiari type II and syringomyelia [3]. The known risk factors for the development of NTD are low socioeconomical class, maternal diabetes, antiepileptic drugs, fever in the early stage of pregnancy, and obesity [3]. In families with a history of NTD, there is a 2%-3% risk of NTD in future pregnancies. The association between folic acid supplementation and reduced rates of NTD is well established [5]. A study among 130,142 Chinese women between 1993–1995 demonstrated a reduction in the risk for NTD of up to 85% in women who took 0.4 mg/day of folic acid prior to conception and during the first trimester [6] and other studies have provided additional evidence of this beneficial effect [1]. Folic acid is absorbed in the proximal part of the small intestine and stored in the liver and CSF. About 4-5 microgram is excreted daily in the urine. The mean dietary intake of folic acid is about 0.1-0.2 mg a day. Folic acid is present in food products such as liver, kidney, nuts, yeast, and green vegetables. It is reduced in the body to tetrahydrofolate, a coenzyme for many metabolic reactions including DNA synthesis and amino acid conversion. Folate deficiency can result in megaloblastic anemia. The recommended daily dose of folic acid to prevent megaloblastic acid in pregnancy is 0.2–0.5 mg/day. Folic acid supplementation is well tolerated, although gastrointestinal side effects may occur. Folic acid also plays an important role in the prevention of arteriosclerosis [5].

cy can result in megaloblastic anemia. The recommended daily dose of folic acid to prevent megaloblastic acid in pregnancy is 0.2–0.5 mg/day. Folic acid supplementation is well tolerated, although gastrointestinal side effects may occur. Folic acid also plays an important role in the prevention of arteriosclerosis [5]. The Israel Ministry of Heath recommends a daily intake of folic acid of 0.4 mg/day three months before conception and throughout the first trimester. Women who previously conceived an infant with NTD should take 0.4 mg/day of folic acid throughout fertility and 4-5 mg/day beginning one month before conception and throughout the first trimester [7]. In the present study we assessed the consumption of folic acid for the prevention of NTDs among Jewish and Muslim Bedouin women in southern Israel and analyzed possible correlations with the recommendations of family physicians and gynecologists. The Jewish sector in Beer-Sheva is classified as low-middle class. The Bedouin sector in southern Israel is of a very low socioeconomic status, with many living in small stone houses, wooden or tin huts, or even tents. A considerable percentage of this population is supported by welfare and has only a few years of formal education [8].

in Beer-Sheva is classified as low-middle class. The Bedouin sector in southern Israel is of a very low socioeconomic status, with many living in small stone houses, wooden or tin huts, or even tents. A considerable percentage of this population is supported by welfare and has only a few years of formal education [8]. 2. Methods 2.1. Study Group The study population included 65 pregnant Jewish women from the city of Beer-Sheva, 64 pregnant Bedouin women from the Negev desert region of southern Israel, 39 gynecologists (specialists and residents) from the Soroka University Medical Center in Beer-Sheva and 60 family physicians (specialists and residents) from the Negev region. The Helsinki Committee of the Soroka Medical Center in Beer-Sheva and the Israel Ministry of Health approved the study. The interviews in the Jewish sector were held in three clinics in Beer-Sheva that represent populations of different socioeconomic status, as determined by the Israel Ministry of Health. The pregnant women were interviewed personally by the first author (EF) upon arrival for routine check-up at the “Mother and Child Health Clinics” after consenting to be interviewed and after it was clear that they had not been interviewed previously. Women visiting the clinic for the first time and women with a history of congenital anomaly in a previous pregnancy were excluded.

thor (EF) upon arrival for routine check-up at the “Mother and Child Health Clinics” after consenting to be interviewed and after it was clear that they had not been interviewed previously. Women visiting the clinic for the first time and women with a history of congenital anomaly in a previous pregnancy were excluded. The interviews in the Bedouin sector were held at a clinic in a Bedouin village and in a Beer-Sheva clinic that serves Bedouins living in rural areas outside established villages. In those clinics we used a questionnaire that was translated into Arabic with the aid of an interpreter. The doctors completed the questionnaires at work or while attending continuing medical education courses at the university. 2.2. The Study Tool An anonymous questionnaire that included questions on demographic background, knowledge and use of folic acid was administered to the pregnant women. A second anonymous questionnaire that included questions on demographic background, knowledge, attitudes, and recommendations regarding folic acid use was given to the doctors. 2.3. Statistical Methods The study data were organized with the Excel software program and processed using the SSPS statistical package. Group differences were compared by the χ2 test and statistical significance was set at P < .05.

2.2. The Study Tool An anonymous questionnaire that included questions on demographic background, knowledge and use of folic acid was administered to the pregnant women. A second anonymous questionnaire that included questions on demographic background, knowledge, attitudes, and recommendations regarding folic acid use was given to the doctors. 2.3. Statistical Methods The study data were organized with the Excel software program and processed using the SSPS statistical package. Group differences were compared by the χ2 test and statistical significance was set at P < .05. 3. Results 3.1. Pregnant Women The mean maternal age among 65 Jewish women was 28.6 ± 4.9 years (range 20–39) and 27.9 ± 5.6 among Bedouin women (range 17–40 years). Four Bedouin women and nine Jewish women refused to participate. The mean number of children per woman was significantly higher in the Bedouin sector. The women from the Jewish sector were significantly more educated than the women from the Bedouin sector. Table 1 presents the demographic background of the Jewish and Bedouin women.

women and nine Jewish women refused to participate. The mean number of children per woman was significantly higher in the Bedouin sector. The women from the Jewish sector were significantly more educated than the women from the Bedouin sector. Table 1 presents the demographic background of the Jewish and Bedouin women. Fifty-one Jewish women (78.5%) reported taking folic acid during pregnancy, but only seven (10.8%) began prior to conception. Sixty Bedouin women (93.75%) reported taking folic acid during pregnancy, but only four (6.25%) began prior to conception (P < .05) (Table 2). Only five Jewish women (7.7%) and two Bedouin women (3.1%) began taking folic acid three months before conception as recommended by the Israel Ministry of Health. Sixty-one Jewish women (94%) and all of the Bedouin women said that they had received these recommendations or knew about them. Seven of the ten Jewish women and three of the four Bedouin women who refrained from taking folic acid cited as the reason previous side effects such as nausea, weakness and dizziness that they attributed to it. Twenty-eight Jewish women (43.1%) 18 Bedouin women (28.1%) believed that folic acid could cause side effects. Twenty three Jewish women (35.4%) and four Bedouin women (6.3%) knew that folic acid could prevent NTDs. Twenty of the Bedouin women knew that taking folic acid could “help” the fetus, but could not define specific health benefits.

omen (43.1%) 18 Bedouin women (28.1%) believed that folic acid could cause side effects. Twenty three Jewish women (35.4%) and four Bedouin women (6.3%) knew that folic acid could prevent NTDs. Twenty of the Bedouin women knew that taking folic acid could “help” the fetus, but could not define specific health benefits. Only two Jewish women (3.1%) and 10 Bedouin women (15.6%) had received and read the Ministry of Health publication on prevention of NTDs, which is available at the Mother and Child Clinics. 3.2. Doctors The study was conducted among 60 family physicians (54% women, mean age 38.9 ± 7.6 years, range 29–56) and 39 gynecologists (26% women, mean age 39.5 ± 8.5 years, range 29–59). Table 3 presents the demographic background of the doctors. More gynecologists than family physicians were department or unit heads, while more family physicians were still in various stages of professional training (residents).

ars, range 29–56) and 39 gynecologists (26% women, mean age 39.5 ± 8.5 years, range 29–59). Table 3 presents the demographic background of the doctors. More gynecologists than family physicians were department or unit heads, while more family physicians were still in various stages of professional training (residents). Thirty-three gynecologists (87.9%) and 36 family physicians (60%) claim to always recommend folic acid on preconception examination. All the gynecologists and 55 family physicians (92%) reported that they almost always or always recommend folic acid supplementation at this stage (P < .05). Thirty-three gynecologists (84%) and 44 family physicians (73%) always recommend folic acid supplementation during pregnancy. Table 4 presents the physician recommendations before and during pregnancy. Only 35 of 56 family physicians (62.5%) and 28 of 39 gynecologists (71.8%) recommended the correct daily dose of 0.4 mg. Eight family physicians (32.1%) and nine gynecologists (21.3%) recommended a high daily dose of 5 mg. Seventeen of 37 gynecologists (45.9%) and 32 of 57 family physicians (56.1%) believe that more than 30% of the women actually begin to take folic acid prior to conception.

Thirty-three gynecologists (87.9%) and 36 family physicians (60%) claim to always recommend folic acid on preconception examination. All the gynecologists and 55 family physicians (92%) reported that they almost always or always recommend folic acid supplementation at this stage (P < .05). Thirty-three gynecologists (84%) and 44 family physicians (73%) always recommend folic acid supplementation during pregnancy. Table 4 presents the physician recommendations before and during pregnancy. Only 35 of 56 family physicians (62.5%) and 28 of 39 gynecologists (71.8%) recommended the correct daily dose of 0.4 mg. Eight family physicians (32.1%) and nine gynecologists (21.3%) recommended a high daily dose of 5 mg. Seventeen of 37 gynecologists (45.9%) and 32 of 57 family physicians (56.1%) believe that more than 30% of the women actually begin to take folic acid prior to conception. 4. Discussion Folic acid supplementation before and during pregnancy is an inexpensive, simple strategy that can significantly reduce the incidence of NTD, a very severe malformation. The average lifetime cost of a baby with NTD is estimated at about $294,000 [3]. There is evidence that folic acid can also prevent other types of congenital anomalies [1]. Folic acid treatment before and during pregnancy is recommended by the Israel and the United States Ministries of Health and by the American Academy of Pediatrics [2, 9].

st of a baby with NTD is estimated at about $294,000 [3]. There is evidence that folic acid can also prevent other types of congenital anomalies [1]. Folic acid treatment before and during pregnancy is recommended by the Israel and the United States Ministries of Health and by the American Academy of Pediatrics [2, 9]. The results of the present study demonstrate that despite the clear recommendations, only 10.8% of Jewish women and 6.25% of Bedouin women take folic acid prior to conception, although 78.5% of the Jewish women and 93.75% of the Bedouin women said they took folic acid at some time during pregnancy. The issue of folic acid supplementation prior to conception has been studied in many countries. In British Columbia [10] 49.4% of 1,004 took folic acid prior to conception. In Puerto Rico [11] 31.5% of 479 women took folic acid appropriately. In Ireland [12] 16% of 300 women took folic acid prior to conception. In a previous Israeli study conducted in 1999 [13] only 2.8% of 920 women took folic acid before conception. In the present study we found that although more Bedouin women take folic acid during pregnancy than Jewish women, fewer Bedouin women begin taking it prior to conception. Since a relatively high rate of Jewish and Bedouin women take folic acid during pregnancy, even if inappropriately, the potential exists for better adherence to recommendations with a concomitant reduction in the rate of NTDs.

during pregnancy than Jewish women, fewer Bedouin women begin taking it prior to conception. Since a relatively high rate of Jewish and Bedouin women take folic acid during pregnancy, even if inappropriately, the potential exists for better adherence to recommendations with a concomitant reduction in the rate of NTDs. The rate of NTDs is almost fourfold among Bedouins and other non-Jewish populations compared to Jews [4]. The reasons for this finding include consanguineous marriage among Bedouins, differences in nutrition, and religious prohibition of abortions. Special intervention plans should be developed for each population based on its cultural and socioeconomic characteristics. A minority of women who did not take folic acid during pregnancy despite recommendations cited fear of side effects that they attributed to its use during previous pregnancies. Even the high dose of folic acid (5 mg) prescribed by 32.1% of the family physicians and 23.1% of gynecologists is unlikely to explain the reported side effects, since these side effects are rare even at this higher dose. It is possible that some of the women used this explanation as an excuse because they were embarrassed that they did not take folic acid.

prescribed by 32.1% of the family physicians and 23.1% of gynecologists is unlikely to explain the reported side effects, since these side effects are rare even at this higher dose. It is possible that some of the women used this explanation as an excuse because they were embarrassed that they did not take folic acid. There is a significant difference between doctors' reports and the actual degree to which women took folic acid prior to conception. All the gynecologists and 92% of the family physicians in the study said that they always/almost always recommend taking folic acid prior to conception, but only 10.8% of the Jewish women and 6.25% of the Bedouin women reported taking folic acid at this stage. On the other hand, there is correspondence between the doctors' reports of recommendations and the pregnant women's reports of folic acid use during pregnancy. Despite the study's small sample size the data demonstrate that only a small minority of women use folic acid appropriately. Since folic acid supplementation is the only way to provide 4-5 mg of folic acid daily it is important to increase the public's awareness of this requirement. An intervention plan should include three basic strategies [14]: diet, folic acid supplementation, and food fortification. According to a survey conducted in the year 2000 the mean daily dietary intake of folic acid is 213.8 ± 181.8 micrograms among Jewish women and 302.0 ± 226.9 micrograms among Bedouin women (unpublished data). This difference may be due to the relatively high intake of fresh vegetables by Bedouins.

d food fortification. According to a survey conducted in the year 2000 the mean daily dietary intake of folic acid is 213.8 ± 181.8 micrograms among Jewish women and 302.0 ± 226.9 micrograms among Bedouin women (unpublished data). This difference may be due to the relatively high intake of fresh vegetables by Bedouins. Our study demonstrates that only a small minority of the women received and read the Ministry of Health's publication on this issue. Other studies have reported that popular newspapers and the electronic media are more effective than printed material disseminated by health authorities [9]. 5. Conclusions It is important to train health system workers in the community to recommend folic acid before, and not only after, conception. It is important for physicians to encourage a preconception examination in which counseling and recommendations for folic acid supplementation are provided. Special attention should be given to family physicians since, according to the results of the present study, only 60% always recommend folic acid prior to conception. Table 1 The demographic characteristics of the jewish and bedouin women. Variable Jews (n = 65) Bedouins (n = 64) P Age [N (%)] 17–20 2 (3) 7 (11) NS 21–30 41 (63) 36 (57) >30 22 (34) 20 (32) Mean ± SD 28.6 ± 4.9 27.9 ± 5.6 Present pregnancy First to third 48 (74) 32 (50) <.05 Fourth or more 17 (26) 32 (50) Mean ± SD 2.6 ± 1.6 4.0 ± 3.0 Children 0 26 (40) 14 (22) <.05 1–3 34 (52) 29 (46) 4+ 5 (8) 20 (32) Mean ± SD 1.2 ± 1.3 2.8 ± 2.7

Variable Jews (n = 65) Bedouins (n = 64) P Age [N (%)] 17–20 2 (3) 7 (11) NS 21–30 41 (63) 36 (57) >30 22 (34) 20 (32) Mean ± SD 28.6 ± 4.9 27.9 ± 5.6 Present pregnancy First to third 48 (74) 32 (50) <.05 Fourth or more 17 (26) 32 (50) Mean ± SD 2.6 ± 1.6 4.0 ± 3.0 Children 0 26 (40) 14 (22) <.05 1–3 34 (52) 29 (46) 4+ 5 (8) 20 (32) Mean ± SD 1.2 ± 1.3 2.8 ± 2.7 Education None 1 (1.5) 14 (22) <.0001 Elementary 1 (1.5) 20 (31) High school 63 (97) 30 (47) Table 2 Comparison of consumption of folic acid, by sector and time during pregnancy. Relation to pregnancy Bedouins N (%) Jews N (%) P Before and during 4 (6.25) 7 (10.8) <.05 Only during 56 (87.5) 44 (67.7) Never 4 (6.25) 14 (21.5) Table 3 Comparison of gynecologists and family physicians. Variable Gynecologists (n = 39) Family physicians (n = 60) P Sex Male 28 (74) 26 (46) <.05 Female 10 (26) 31 (54) Age (yrs) <40 20 (62.5) 30 (64) NS ≥40 12 (37.5) 17 (36) Mean ± SD 39.5 ± 8.5 38.8 ± 7.6 Years as physician <10 19 (49) 30 (55) NS ≥10 20 (51) 24 (45) Mean ± SD 13.7 ± 11.3 11.2 ± 7.9 Position Department head 6 (15) 1 (2) <.01 Other specialists 17 (43) 20 (33) Resident 16 (42) 39 (65) Table 4 Comparison of gynecologists and family physicians as to recommendations to take folic acid prior to conception and during pregnancy. Extent of recommendation Gynecologists N (%) Family physicians N (%) P Prior to conception Always 33 (87) 36 (60) <.05 Almost always 5 (13) 19 (32) Rarely — 5 (8) During pregnancy Always 33 (84) 44 (73) NS Almost always 5 (13) 14 (23) Rarely 1 (3) 2 (4)

1. Background Herniation of the uterine tube is rare due to its anatomical position. If it happens, its status should be properly checked as the treatment depends on its viability. 2. Case Presentation A 39-years-old patient was admitted to the surgical ward with a painful lump in the right groin which had been present for about three years. The lump had recently increased in size. She was investigated by the gynaecologists for primary infertility as she had failed to conceive for many years, but no cause was found. Clinical examination revealed a 5 cm × 5 cm tender, irreducible, nonpulsatile mass in the right groin just under the inguinal ligament. There was no cough impulse. A diagnosis of a femoral hernia was made. Exploration of the mass confirmed the diagnosis of the femoral hernia, but sac contained the right fallopian tube (Figure 1). The uterine tube was easily reduced back into the pelvic cavity as it looked healthy and the femoral hernia was repaired. The patient made good recovery, and she has been referred back to the gynaecology clinic for followup.

med the diagnosis of the femoral hernia, but sac contained the right fallopian tube (Figure 1). The uterine tube was easily reduced back into the pelvic cavity as it looked healthy and the femoral hernia was repaired. The patient made good recovery, and she has been referred back to the gynaecology clinic for followup. 3. Discussion The diagnosis of a femoral hernia is usually clinical. Most patients present as an emergency with symptoms and signs of intestinal obstruction. A typical femoral hernia presents as a tender, nonreducible swelling with no cough impulse and is situated below and lateral to the pubic tubercle. On exploration, it often contains only omentum, or may contain a knuckle of bowel known as Richter's hernia. The differential diagnoses include inguinal hernia, lipoma, saphena varix, enlarged lymph nodes, femoral artery aneurysm, sarcoma, obturator hernia, psoas abscess, psoas bursa, and in males, ectopic testis.

cle. On exploration, it often contains only omentum, or may contain a knuckle of bowel known as Richter's hernia. The differential diagnoses include inguinal hernia, lipoma, saphena varix, enlarged lymph nodes, femoral artery aneurysm, sarcoma, obturator hernia, psoas abscess, psoas bursa, and in males, ectopic testis. Inguinal herniation of the adnexa is rare and is generally found in children with associated congenital abnormalities of the genital tract. In his analysis of 1,950 patients with inguinal hernias, Gurer et al. [1] found that most of his patients were adults with no genital abnormalities. The incidence of inguinal hernias containing ovary and fallopian tube in his study was 2.9%. Different contents in femoral hernias have been reported in the literature, such as appendix, small intestine, omentum, bladder, Meckel's diverticulum, ectopic testis, and stomach [2]. Herniation of the fallopian tube in a femoral hernia however is very rare due to its normal anatomical position as it lies at a lower level than the femoral ring [3]. In infants, the fallopian tube is near the internal inguinal ring and can more readily pass through this structure than later in life [4], due to the closure of the canal of Nuck. Due to its deep pelvic position, it is very difficult for the uterine tube to herniate in adults unless there is a sudden anomalous movement when the body is forcibly inclined towards the inguinal region, for example, or a pathological condition of the internal genital organs such as a uterine fibroid [4].

nal of Nuck. Due to its deep pelvic position, it is very difficult for the uterine tube to herniate in adults unless there is a sudden anomalous movement when the body is forcibly inclined towards the inguinal region, for example, or a pathological condition of the internal genital organs such as a uterine fibroid [4]. Patients with herniation of the fallopian tube might present with unusual symptoms which vary from intermittent attacks of abdominal pains, irreducibility and strangulation, increased hernia size during muscular effort, to those which resemble symptoms of ileus. Sometimes, a cord-like body in the sac leading to the hernial opening might be palpated. A vaginal bimanual examination help diagnose such cases by findings a deviation of the uterus towards the side of the hernia; for instance, the uterus being drawn over and inclined towards the side on which the hernia was situated, or the uterus is in a position of marked anteflexion and seemingly fixed [4]. If the tube is healthy on exploring the hernial sac, it should be reduced into the pelvic cavity. Otherwise, it should be removed if its viability is in doubt, gangrenous, or profound pathological changes have occurred [4]. In children, the peritoneal attachment of the fallopian tube and the ovary to the posterior wall has to be divided before reducing them if they are situated in the canal of Nuck as they are more or less fixed at this position [5]. 4. Conclusion We present a rare case of a femoral hernia containing a fallopian tube which clinicians should be aware of when managing women with femoral masses.

If the tube is healthy on exploring the hernial sac, it should be reduced into the pelvic cavity. Otherwise, it should be removed if its viability is in doubt, gangrenous, or profound pathological changes have occurred [4]. In children, the peritoneal attachment of the fallopian tube and the ovary to the posterior wall has to be divided before reducing them if they are situated in the canal of Nuck as they are more or less fixed at this position [5]. 4. Conclusion We present a rare case of a femoral hernia containing a fallopian tube which clinicians should be aware of when managing women with femoral masses. Conflict of Interests The authors declare that they have no competing interests. Consent Informed written consent was obtained from patient's relatives for publication of the manuscript and figures. Acknowledgment The authors would like to thank Dr. David Nunn, consultant gynaecologist for editing this paper. Figure 1 Fallopian tube (black arrow) in the explored femoral hernia (red arrow).

1. Introduction Müllerian duct malformations delineate a miscellaneous group of congenital anomalies that result from arrested development, abnormal formation, or incomplete fusion of the mesonephric ducts [1]. The incidence of uterine anomalies in a fertile population is reported to be around 3.2% [2]. The unicornuate uterus results from normal differentiation of the Müllerian duct, but a rudimentary functional horn may be found [3]. Patients with an asymmetric uterus and a rudimentary horn constitute 5% to 10% of those with uterovaginal anomalies [4]. Approximately 75% of such horns do not communicate with the normal hemiuterus [5]. Vaginal obstruction is associated with perivaginal mass, pain, and endometriosis, but cyclic menstrual flow may be present because of the normally functioning opposite side [4]. This anomaly is usually associated with ipsilateral renal agenesis (67%) or ipsilateral pelvic kidney [6].

ate with the normal hemiuterus [5]. Vaginal obstruction is associated with perivaginal mass, pain, and endometriosis, but cyclic menstrual flow may be present because of the normally functioning opposite side [4]. This anomaly is usually associated with ipsilateral renal agenesis (67%) or ipsilateral pelvic kidney [6]. 2. Case A 16-year-old nulliparous woman presented with severe dysmenorrhoea since menarche in November 2006 which was only minimally relieved with oral contraceptives and nonsteroidal anti-inflammatory drugs (NSAIDs). In April 2007, the patient experienced an episode of severe pain in the left lower quadrant of the abdomen. Pelvic ultrasound revealed a large irregular complex mass in the left hemipelvis with multiple cystic and solid components. The uterus and right ovary were thought to be normal. The left ovary could not be identified. The patient underwent diagnostic laparoscopy with hysteroscopy. Hysteroscopy showed a right unicornuate uterus and revealed a patent right cornus with no sign of ostium on the left side. Laparoscopy showed a right unicornuate uterus with a normal adnexa, a left non-communicating rudimentary horn (4 × 3 × 2 cm) with an enlarged and thickened tube, and a left ovarian endometrioma of 6 cm. A large endometrial cyst was washed out with irrigation fluid, and a biopsy was taken. After washing, the interior wall of the cyst was carefully examined to confirm the absence of intracystics lesions suspected to be malignant. The interior wall of the cyst was then destroyed using bipolar coagulation. Additionally, fibrous adhesions involving the ascending colon and small intestine were destroyed. Lysis of omental adhesions allowed identification of multiple areas of endometriosis in the posterior cul-de-sac, on the right and left uterosacral ligaments. There were no external genital abnormalities. A subsequent urogram revealed absence of the left kidney (Figure 1). Medical treatment for endometriosis using 6 months of gonadotropin-releasing hormone (GnRHa) was done. After extensive discussion with the patient laparoscopic removal of the left horn was indicated. The second laparoscopic examination was performed on September 2007. A four-puncture laparoscopy was performed with a 10-mm infraumbilical port, a 10 mm suprapubic port, and two 5-mm suprapubic ports laterally in the right abdominal side and in midline. Laparoscopy revealed a normal right hemiuterus, tube, and ovary, and a left rudimentary uterine horn (Figure 2).

med on September 2007. A four-puncture laparoscopy was performed with a 10-mm infraumbilical port, a 10 mm suprapubic port, and two 5-mm suprapubic ports laterally in the right abdominal side and in midline. Laparoscopy revealed a normal right hemiuterus, tube, and ovary, and a left rudimentary uterine horn (Figure 2). A left salpingectomy was started at the fimbriated end using bipolar coagulation and laparoscopic scissors. The left tube was used to pull up the rudimentary horn. The left uterus was dissected apart from the bladder using scissors and bipolar coagulation and was removed using a morcellator (Karl Storz, Germany). She went home 1 day after surgery and began a regimen of oral contraception with 75 μg of desogestrel. She continues free of pelvic pain one year after surgery.

ry horn. The left uterus was dissected apart from the bladder using scissors and bipolar coagulation and was removed using a morcellator (Karl Storz, Germany). She went home 1 day after surgery and began a regimen of oral contraception with 75 μg of desogestrel. She continues free of pelvic pain one year after surgery. 3. Comment Around 75%–90% of cases of unicornuate uterus with rudimentary horn are non-communicanting [7]. The management of the present case illustrates the value of simultaneous laparoscopic and hysteroscopy evaluation of known uterine abnormalities. The literature suggests the need to remove the rudimentary horn of a unicornuate uterus and supports the laparoscopic approach if such a decision is taken [8–11]. A high incidence of associated endometriosis has been documented in cases of obstructive müllerian anomalies [8–11]. In the present case, the procedure was effective in resolving the pelvic pain and dysmenorrhea and avoided the risk of endometriosis. A GnRH agonist was used preoperatively in this case to reduce the vascularization and the inflammation that are often present around endometriotic lesions, facilitating surgical procedures [12]. In conclusion, operative laparoscopy is an excellent alternative to laparotomy for the management of unicornuate uterus with non-communicanting rudimentary horn. Commonly accepted benefits of minimally invasive surgery are enhanced visualization of the cul-de-sac, less adhesion formations, smaller incisions, reduced postoperative pain, and shortened hospital stay. Conflict of Interests None to declare.

In conclusion, operative laparoscopy is an excellent alternative to laparotomy for the management of unicornuate uterus with non-communicanting rudimentary horn. Commonly accepted benefits of minimally invasive surgery are enhanced visualization of the cul-de-sac, less adhesion formations, smaller incisions, reduced postoperative pain, and shortened hospital stay. Conflict of Interests None to declare. Figure 1 Urogram revealed absence of the left kidney. Figure 2 Normal right hemiuterus, tube, and ovary, and a left rudimentary uterine horn (arrow).

11/22) of UPSC patients with advanced disease (Stage III~IV), compared to 14.3% (2/14) of patients with early disease (Stage I~II). Patients with overexpression of Her-2/neu were significantly more likely to have advanced-stage disease when compared to Her-2/neu negative patients (OR = 6.0, 95%CI: 1.08–33.32, P = .03). CISH was performed in all UPSC cases. Four out of the 36 (11.1%) cases were found with Her-2/neu gene amplification, in which 3 amplified in high level and 1 in low level (Figure 1). The correlation between IHC and CISH results were assessed (Table 2). All IHC 3+ (100%, 3/3) cases had Her-2/neu gene amplification. And all IHC negative cases had normal Her-2/neu gene copies (1–5 copies). Therefore, complete concordance between IHC and CISH was observed in IHC 3+ and negative cases (P = .001, Spearman Correlation). Only one out of 10 cases scored as 2+ by IHC (10%, 1/10) was found with gene amplification by CISH. 3.3. Survival Analyses The median follow-up time for all of the patients was 31 months (range, < 1 to 115 months). During the follow-up time, 17 patients died (47.2%), and 15 of these deaths were related to this disease. Nine patients were observed to have disease progression during followup, with 4 in pelvis, 2 in liver, 1 in brain, 1 in both liver and pelvis, and 1 in both liver and brain. The 1-year, 3-year, and 5-year overall survival for all patients was 73.1%, 51.9%, and 43.9%, respectively (Figure 2,).

1. Introduction Urinary tract infections (UTI) are the most common bacterial infections of pregnancy [1]. Asymptomatic bacteriuria (ASB), occurring in 2–11% of pregnancies, is a major predisposition to the development of pyelonephritis, which is associated with obstetrical complications, such as preterm labor and low birth weight infants. Bacteriuria is defined as the presence of >105 colonies of a single pathogen per milliliter of urine. It may be either an asymptomatic bacteriuria (ASB of pregnancy) or symptomatic acute cystitis and acute pyelonephritis [2]. When left untreated, ASB may lead to symptomatic uriner tract infections (UTI) [3]. Untreated, ASB is found to be associated with subsequent acute pyelonephritis in 20–50% of cases [4]. The female urethra is relatively short and is anatomically proximal to the vagina, which is colonized with organisms from the gastrointestinal tract. Normal physiological changes in pregnancy place women at risk for pyelonephritis. There is also relative obstruction of the ureters because the enlarging uterus physically blocks them, and the hormonal milieu of pregnancy leads to relaxation of the smooth muscle of the ureters and the bladder. Furthermore, the glycosuria and aminoaciduria of pregnancy provide an excellent medium for bacterial proliferation [5–7]. The relatively high prevalence of ASB during pregnancy, the significant consequences for women and pregnancy, and the ability to avoid undesired outcomes with treatment justify screening and treatment of ASB in pregnancy [8].

The female urethra is relatively short and is anatomically proximal to the vagina, which is colonized with organisms from the gastrointestinal tract. Normal physiological changes in pregnancy place women at risk for pyelonephritis. There is also relative obstruction of the ureters because the enlarging uterus physically blocks them, and the hormonal milieu of pregnancy leads to relaxation of the smooth muscle of the ureters and the bladder. Furthermore, the glycosuria and aminoaciduria of pregnancy provide an excellent medium for bacterial proliferation [5–7]. The relatively high prevalence of ASB during pregnancy, the significant consequences for women and pregnancy, and the ability to avoid undesired outcomes with treatment justify screening and treatment of ASB in pregnancy [8]. The frequency of isolated pathogens and antimicrobial resistance patterns can vary in different geographical regions; therefore, the most common causative agents should be investigated and communities should be aware of their local antimicrobial resistances. The objective of this prospective study was to identify the prevalence of ASB, the most common causative agents and the antibacterial susceptibilities of the isolated microorganisms in a Research Hospital in Ankara, Turkey.

should be investigated and communities should be aware of their local antimicrobial resistances. The objective of this prospective study was to identify the prevalence of ASB, the most common causative agents and the antibacterial susceptibilities of the isolated microorganisms in a Research Hospital in Ankara, Turkey. 2. Materials and Method Between December 2009 and May 2010, 2011 of 2132 pregnant women applying for their first antenatal visit to the Zekai Tahir Burak Women's Health Education and Research Hospital antenatal outpatient clinic were recruited into the study. The remaining 121 women refused to take part in this study, and informed consent for the participating patients was taken. Patients with fever, urinary symptoms (such as dysuria, hesitancy, urgency, frequent voiding, incontinence and incomplete voiding), current antibiotic therapy, and a history of urolithiasis and diabetes were excluded. Urine cultures of these symptomatic patients were collected and antimicrobial therapy was given accordingly. Along with the routine antenatal laboratory tests, patients were instructed to collect mid-stream urine samples. Collected samples were immediately inoculated on 5% sheep blood agar and eosin-methylene blue agar plates with a 0.01-mL loop. After 24–48 hours of aerobic inoculation at 37°C, the plates were interpreted. Colony counts of a single microorganism of >105 colony forming units (cfu)/mL were interpreted as bacteriuria. The presence of multiple organisms or of skin flora was considered to result from contamination.

agar plates with a 0.01-mL loop. After 24–48 hours of aerobic inoculation at 37°C, the plates were interpreted. Colony counts of a single microorganism of >105 colony forming units (cfu)/mL were interpreted as bacteriuria. The presence of multiple organisms or of skin flora was considered to result from contamination. Antimicrobial susceptibility testing by disc diffusion was employed according to the Clinical and Laboratory Standards Institute Guidelines. Identification and antimicrobial susceptibility testing of microorganisms was performed according to the Clinical and Laboratory Standards Institute Guidelines with conventional microbiological methods and confirmed by The BD Phoenix Automated Microbiology System (BD Diagnostics, Sparks, MD). The Phoenix is designed for the rapid identification (ID) and antimicrobial susceptibility testing (AST) of clinically significant human bacterial pathogens. The Phoenix identification method uses modified conventional, fluorogenic, and chromogenic substrates. Research-use-only combination panels (UMIC/ID) for both the identification and susceptibility testing were used for this comparison. Software versions V3.34A and V3.54A were used for this study. The ID side contains 45 wells with dried biochemical substrates and 2 fluorescent control wells. The ID broth was inoculated with bacterial colonies from a pure culture that was adjusted to a 0.5 McFarland standard by using a CrystalSpec nephelometer (BD Diagnostics) according to the manufacturer's recommendations. A 25-μL aliquot of this suspension was removed for AST, and the remaining suspension was then poured into the ID side of the Phoenix panel. The specimen was logged and loaded into the instrument within the specified timeline of 30 min. Quality control was performed according to the manufacturer's recommendations. The AST side of the combination panel contains up to 84 wells with dried antimicrobial panels and growth control well. The assay is a broth-based microdilution test. The system uses a redox indicator for the detection of organism growth in the presence of an antimicrobial agent. The previously described 25 μL of the standardized ID broth suspension was transferred to the AST broth, yielding a final concentration of approximately 5 × 105 CFU/mL. Quality control was performed according to the manufacturer's recommendations.

detection of organism growth in the presence of an antimicrobial agent. The previously described 25 μL of the standardized ID broth suspension was transferred to the AST broth, yielding a final concentration of approximately 5 × 105 CFU/mL. Quality control was performed according to the manufacturer's recommendations. The differences in the demographic features between the two groups were analyzed by the SPSS v14.0 Student's t-test. 3. Results Demographic features of the women screened for ASB are displayed in Table 1. Age, gravid, and parity are similar between the two groups (P > .05). Hemoglobin levels, however, display a significant difference between the ASB and nonbacteriuria groups (P = .01). Of the 2011 pregnant women included in the study, 171 (8.5%) had ASB. E. coli, detected in 131 (76.6%) patients, was the most frequently isolated organism, followed by Klebsiella pneumonia, which was found in 25 (14.6%) of the cases. Table 2 shows the isolated microorganisms. E. coli, the most common isolate, was found to be only 57.2% and 68% sensitive to ampicillin and amoxicillin + clavulanate, respectively. Its sensitivity fosfomycin was very high, 99.2%. Sensitivities to ceftriaxone and ciprofloxacin were both 90%, and sensitivity to amikacin was 99%. A 100% sensitivity was found for the broad spectrum penicillins imipenem and meropenem. An 86% sensitivity was documented for cefuroxime, a commonly prescribed oral agent for UTIs in pregnancy.

mycin was very high, 99.2%. Sensitivities to ceftriaxone and ciprofloxacin were both 90%, and sensitivity to amikacin was 99%. A 100% sensitivity was found for the broad spectrum penicillins imipenem and meropenem. An 86% sensitivity was documented for cefuroxime, a commonly prescribed oral agent for UTIs in pregnancy. Klebsiella pneumonia, the second most frequent organism grown on the cultures, was only 4% sensitive to ampicillin, while its sensitivity to amoxicillin/clavulanic acid was 84%. We found a 88% sensitivity to fosfomycin. Sensitivity to cefuroxime was 84%, and a 100% sensitivity was found for cefepime, ceftriaxone, imipenem, and meropenem. Antibiotic sensitivities of the isolated gram (−) and gram (+) microorganism are shown separately in Tables 3 and 4. 4. Discussion Pregnant women with ASB are more likely to deliver premature or low birth weight infants and have a 20–30-fold increased risk of developing pyelonephritis compared to women without bacteriuria [9]. In this study, 171 of 2011 (8.5%) pregnant women had ASB, which is comparable with the results of studies by Uncu et al. [10] in 2002 and Hazhir [11] in 2007. E. coli was the most common isolate (76.6%), which is consistent with previous studies [12–14]. E. coli, the most frequent isolate, was 99.2% sensitive to fosfomycin. Fosfomycin is an oral agent administered in a single dose with relatively limited side effects. Klebsiella pneumonia, the second most common microorganism isolated in this study is, also 88% sensitive to fosfomycin.

In this study, 171 of 2011 (8.5%) pregnant women had ASB, which is comparable with the results of studies by Uncu et al. [10] in 2002 and Hazhir [11] in 2007. E. coli was the most common isolate (76.6%), which is consistent with previous studies [12–14]. E. coli, the most frequent isolate, was 99.2% sensitive to fosfomycin. Fosfomycin is an oral agent administered in a single dose with relatively limited side effects. Klebsiella pneumonia, the second most common microorganism isolated in this study is, also 88% sensitive to fosfomycin. Ampicillin and amoxicillin/clavulanate are two frequently prescribed oral antimicrobial agents for UTI in pregnant women. Our culture results yielded 57.2% and 68% sensitivities to these agents, respectively. Cefuroxime is another drug commonly prescribed for treating ASB, and sensitivity to this drug was 86%, which is comparable to that of cefazolin (87%), cefepime (91.6%), and ceftriaxone (90%). Sensitivities to imipenem and meropenem were both 100% (Table 3). In some studies, gram-positive microorganisms are found to be important causative agents of ASB. Enayat et al. reported that up to 16.8% of causative organisms are coagulase-negative staphylococci [13]. In our study, only 5.8% of the urinary isolates were gram-positive organisms, the most frequent one being Enterococcus faecalis (4%).

e related to this disease. Nine patients were observed to have disease progression during followup, with 4 in pelvis, 2 in liver, 1 in brain, 1 in both liver and pelvis, and 1 in both liver and brain. The 1-year, 3-year, and 5-year overall survival for all patients was 73.1%, 51.9%, and 43.9%, respectively (Figure 2,). The results of univariate survival analysis of Her-2/neu protein expression and other clinicopathological factors are shown in Table 3. The 5-year overall survival rate for Her-2/neu IHC positive and negative cases was 12.9% and 68.6%, respectively. Poorer outcome (Kaplan–Meier) was observed for patients with Her-2/neu overexpression positive than negative patients (P = .0023) (Figure 3). The 5-year overall survival rate for early and advanced stage cases was 60.4% and 16%, respectively. Advanced stages (P = .0006) and deep myometrial invasion (P = .0138) were also significantly associtated with a shorter OS (Figures 4 and 5).

gram-positive microorganisms are found to be important causative agents of ASB. Enayat et al. reported that up to 16.8% of causative organisms are coagulase-negative staphylococci [13]. In our study, only 5.8% of the urinary isolates were gram-positive organisms, the most frequent one being Enterococcus faecalis (4%). Contributing risk factors for developing bacteriuria during pregnancy are age, parity, sexual intercourse, diabetes mellitus, sickle cell disease, trait, anatomical abnormalities of the urinary tract, previous history of UTI, and socioeconomic status [15–18]. In this study, four variables, age, parity, and hemoglobin levels, were evaluated. Among these variables, hemoglobin levels were associated with ASB (P = .01). Because an association between bacteriuria and anemia has not been confirmed, we concluded that this result may be attributed to the confounding effects of low socioeconomic status. Two of the most important points to consider in the choice of antimicrobial agents for use in a population of pregnant women within a certain geographical region are the frequently isolated urinary pathogens and the patterns of antimicrobial resistance.

Contributing risk factors for developing bacteriuria during pregnancy are age, parity, sexual intercourse, diabetes mellitus, sickle cell disease, trait, anatomical abnormalities of the urinary tract, previous history of UTI, and socioeconomic status [15–18]. In this study, four variables, age, parity, and hemoglobin levels, were evaluated. Among these variables, hemoglobin levels were associated with ASB (P = .01). Because an association between bacteriuria and anemia has not been confirmed, we concluded that this result may be attributed to the confounding effects of low socioeconomic status. Two of the most important points to consider in the choice of antimicrobial agents for use in a population of pregnant women within a certain geographical region are the frequently isolated urinary pathogens and the patterns of antimicrobial resistance. 5. Conclusion Every symptomatic or asymptomatic woman should have a clean-catch urine culture, and detected cases should be treated according to the antimicrobial susceptibility tests. A clean-catch culture is the most sensitive test for the detection of ASB, but it is expensive and requires trained laboratory staff. When these facilities are not available, knowledge of the most frequent isolates and local antimicrobial resistance patterns will aid physicians in the successful empirical treatment of the infection.

culture is the most sensitive test for the detection of ASB, but it is expensive and requires trained laboratory staff. When these facilities are not available, knowledge of the most frequent isolates and local antimicrobial resistance patterns will aid physicians in the successful empirical treatment of the infection. In this certain geographical region, we found E. coli as the most common causative agent of ASB in the obstetric population and it is very sensitive to fosfomycin. We recommend fosfomycin for ASB in pregnant women due to its high sensitivity, ease of administration, and safety for use in pregnancy. Table 1 Demographic characteristics of the pregnant women screened for asymptomatic bacteriuria. Characteristics Non-Bacteriuria Bacteriuria P Value Age (years) Mean (SD) 24.6 (5.1) 23.9 (5.0) >.05 Range 17–43 17–44 Pregnancy Mean (SD) 1.96 (1.0) 2.01 (1.1) >.05 Range 0–6 0–7 Parity Mean (SD) 0.86 (0.92) 0.91 (0.98) >.05 Range 0–5 0–6 Hemoglobin (g/dL) Mean (SD) 12.8 (6.8) 11.1 (5.3) .01 Range 8.3–14.9 7.6–13.7 SD = standard deviation. Table 2 Bacteriologic isolates from pregnant women with asymptomatic bacteriuria. Microorganism (s) N (%) E. coli 131 (76.6) Klebsiella pneumoniae 25 (14.6) Enterococcus faecalis 7 (4) Klebsiella oxytoca 2 (1.2) Proteus spp. 2 (1.2) Enterobacter sakazakii 1 (0.6) Enterococcus spp. 1 (0.6) Staphylococcus aureus 1 (0.6) Staphylococcus saprophyticus 1 (0.6) Total 171 (100) Table 3 Gram negative microorganisms. Amo/clav Am Cf Cef Ceft Cefu Fos Fm Sxt Microorganism (s) Antibiotics sensitivity (%)

E. coli 131 (76.6) Klebsiella pneumoniae 25 (14.6) Enterococcus faecalis 7 (4) Klebsiella oxytoca 2 (1.2) Proteus spp. 2 (1.2) Enterobacter sakazakii 1 (0.6) Enterococcus spp. 1 (0.6) Staphylococcus aureus 1 (0.6) Staphylococcus saprophyticus 1 (0.6) Total 171 (100) Table 3 Gram negative microorganisms. Amo/clav Am Cf Cef Ceft Cefu Fos Fm Sxt Microorganism (s) Antibiotics sensitivity (%) E. coli 89 75 115 120 118 113 130 125 108 (68) (57.2) (87) (91.6) (90) (86) (99.2) (95.4) (82) Klebsiella pneumoniae 21 1 23 25 25 21 22 9 25 (84) (4) (92) (100) (100) (84) (88) (36) (100) Klebsiella oxytoca 0 0 0 0 0 0 2 1 2 (100) (50) (100) Proteus spp. 2 1 2 2 2 2 2 2 0 (100) (50) (100) (100) (100) (100) (100) (100) Enterobacter sakazakii 0 0 0 1 1 1 1 1 1 (100) (100) (100) (100) (100) (100) Amo/clav=amoxicillin/clavulanic acid, Am=ampicillin, Cf=Cefazoline, Cef=Cefepime, Ceft=Ceftriaxone, Cefu=Cefuroxime, Fos=Fosfomycin, Fm=Nitrofurantoin, Sxt=co trimoxazole. Table 4 Gram positive microorganisms. Am Fos penG Sxt Clin Eryt Fm Microorganism (s) Antibiotics sensitivity (%) Enterococcus faecalis 7 7 6 ND ND ND ND (100) (100) (85.7) Enterococcus spp. 1 ND 1 ND ND ND ND (100) (100) Staphylococcus aureus ND ND 0 ND 1 1 1 (100) (100) (100) Staphylococcus saprophyticus 0 ND 0 1 ND ND ND (100) Am=ampicillin, Fos=Fosfomycin, PenG=Penicillin G, Sxt=co-trimoxazole, Clin=Clindamycin, Eryt=Erythromycin, Fm=Nitrofurantoin, ND=not done.

-2/neu overexpression positive than negative patients (P = .0023) (Figure 3). The 5-year overall survival rate for early and advanced stage cases was 60.4% and 16%, respectively. Advanced stages (P = .0006) and deep myometrial invasion (P = .0138) were also significantly associtated with a shorter OS (Figures 4 and 5). 4. Discussion UPSC is a rare type of endometrial cancer, whose clinical features are different from endometrioid endometrial adenocarcinoma. The median age of UPSC was 64 years in this study, similar to previous report, but older than the median age of women with endometrioid endometrial cancers in China [25]. Unlike endometrioid endometrial cancers, UPSC was not associated with obesity. The median BWI was 23.8 kg/m2 in this study. And most important of all, compared to patients with endometrioid endometrial adenocarcinomas, patients with UPSC tend to have deep myometrial invasion, high-stage diseases, and worse overall survival, as we had discussed previously in [25]. In our study, 61% were late stages. More than half of patients (58.3%) had deep myometrial invasion. The 1-year, 3-year, and 5-year overall survival was 73.1%, 51.9%, and 43.9%, respectively.

1. Introduction Primary malignant melanoma of the cervix constitutes a rare disease [1–4], representing less than 2% of cases of malignant melanoma that affects the female genital tract [1]. It was not until 1960, after the description by Cid [5] of melanocytic cells in the cervix, when the concept was accepted of primary melanoma in uterine cervix. It is considered as a very aggressive neoplasia [6], whose diagnosis is untimely due to the unusualness of the disease and the scarce suspicion of the clinician. At present, there is no consensus on the therapeutic approach to follow [1, 2, 7]; in the reports found, initial treatment is referred of radical hysterectomy accompanied or not by radical pelvic lymphadectomy or superior vaginectomy [1, 2, 6–11]. Although there are no significant statistics, prognosis in general is poor [2, 9, 11] and unpredictable [1] and is even worse when early visceral metastases are discovered [11]. In the current literature, there are no more than 60 cases reported of this entity. Our purpose is to report the case of a patient with this neoplasia who was treated with ultraradical surgery followed by radiotherapy and who has had long-term followup.

Although there are no significant statistics, prognosis in general is poor [2, 9, 11] and unpredictable [1] and is even worse when early visceral metastases are discovered [11]. In the current literature, there are no more than 60 cases reported of this entity. Our purpose is to report the case of a patient with this neoplasia who was treated with ultraradical surgery followed by radiotherapy and who has had long-term followup. 2. Case Report A female patient 34 years of age, Mexican mestizo, nubile. History of 2 months of abnormal genital bleeding, for which she was evaluated by abdominal Ultrasound (US) with a report of uterine myomatosis. Carrying out a gynecological examination is decided upon, in which a blackish lesion is observed in the area of the uterine cervix. Thus, a biopsy of the cervix was taken, which revealed melanoma. The patient was therefore referred to the National Cancerology Institute (INCan) in Mexico City in January 2001.

yomatosis. Carrying out a gynecological examination is decided upon, in which a blackish lesion is observed in the area of the uterine cervix. Thus, a biopsy of the cervix was taken, which revealed melanoma. The patient was therefore referred to the National Cancerology Institute (INCan) in Mexico City in January 2001. On initial physical exploration at this Institute, cervix-dependent tumor was noted that extended to the right pelvic wall, of petraeus consistency, and another at the level of the anterior wall of the vagina. A new biopsy was taken, reporting malignant melanoma. Chest X-ray, cystoscopy, and colonoscopy were performed, and reported as normal, in addition to complete revision of skin and eyes, discarding other sites of melanocytic lesions. Additionally, a Computed tomography (CT) was requested, which reported tumor in the pelvic floor toward the right annex of approximately 8 × 6 cm, which displaced the bladder and ureters (Figure 1). Due to the extension of the lesion, the patient was considered to be a candidate for exploratory laparotomy, and total pelvic exenteration was planned, which was performed in March 2001. Reconstruction of the digestive and urinary tracts was carried out with a terminal colostomy and Bricker ileal conduit operation, respectively. The final histopathological report of the surgical specimen obtained demonstrated malignant melanoma of the cervix of 7.5 × 5 cm, surgical margins were reported as negative. Intrahospital evolution was satisfactory, and the patient was discharged on day 10. Adjuvant radiotherapy was administered, 21 Gy in three sessions, concluding in June 2001. The patient has been under routine surveillance for 8 years, and has only referred four occasional urinary tract infections, which have been resolved medically. To the moment of this report, there has been no evidence of local or distant recurring disease.

administered, 21 Gy in three sessions, concluding in June 2001. The patient has been under routine surveillance for 8 years, and has only referred four occasional urinary tract infections, which have been resolved medically. To the moment of this report, there has been no evidence of local or distant recurring disease. 3. Pathology For histopathological studies, the product of the total pelvic exenteration was received, in which a neoplastic lesion of the cervix measured 7.5 × 5.5 cm with involvement of the upper third of the vagina into anterior vaginal septum. The neoplasia was polypoid and light brown in color with necrosis. Microscopically, the neoplasia was formed by nodules of broad cytoplasmic cells with poorly defined borders with pleomorphic nuclei with prominent nucleolae. These areas displayed transition with areas with fascicules of elongated cells of moderate cytoplasm and similar elongated nuclei to those described previously (Figure 2). Immunohistochemical stains were positive for S-100 protein, Melan A, and HMB 45 in neoplastic cells. CKAE1/AE3 and CEA were negative. No binding activity was found. A total of 15 pelvic lymph nodes were removed with the specimen, one of them was positive for disease on H&E, all the surgical margins were negative. 4. Discussion Malignant melanomas are generally found in areas of skin exposed to the sun, but can also be present in nonexposed sites, such as genital tract and esophagus, among others [8].

Immunohistochemical stains were positive for S-100 protein, Melan A, and HMB 45 in neoplastic cells. CKAE1/AE3 and CEA were negative. No binding activity was found. A total of 15 pelvic lymph nodes were removed with the specimen, one of them was positive for disease on H&E, all the surgical margins were negative. 4. Discussion Malignant melanomas are generally found in areas of skin exposed to the sun, but can also be present in nonexposed sites, such as genital tract and esophagus, among others [8]. Cervical melanoma arises from melanocytic cells of the cervix; in fact, the cervical epithelium is capable of forming the complete spectrum of melanocytic lesions, from benign lentigines to blue nevi to melanoma [10]. The usual form of presentation of primary melanoma of the cervix on physical examination is a polypoid exophytic mass, red, brown, grey, black, or blue in color [6], or a colorless in the case of amelanotic melanomas, which constitute up to 55% of cases at this anatomic site [12], presented with vaginal bleeding [1, 6, 8, 9]. Age range varies from 20 to 78 years [6], being more common between 60 and 70 years [1]. Due to that the cervix is an unusual site for this type of neoplasm, the International Federation of Gynecology and Obstetrics (FIGO) staging system for cervical cancer is used [11], rather than the Clark and Breslow scales, because the FIGO staging system correlates better with the prognosis.

ween 60 and 70 years [1]. Due to that the cervix is an unusual site for this type of neoplasm, the International Federation of Gynecology and Obstetrics (FIGO) staging system for cervical cancer is used [11], rather than the Clark and Breslow scales, because the FIGO staging system correlates better with the prognosis. Diagnosis of primary melanoma of the cervix entertains a high probability of being confused with another entity, due to the rarity of the disease [9]. Differential diagnosis between a primary cervical melanoma and a metastatic tumor is important because the latter can be part of a metastatic disease spreading to the cervix [1]. Norris and Taylor criteria are used [13] to distinguish whether it is a primary malignant melanoma of the cervix [6, 8, 12, 13]: (a) presence of melanin in the cervical epithelium; (b) absence of melanoma in another site of the body; (c) presence of binding activity in the cervical epithelium near the lesion; (d) if metastatic disease is found, it should be according to the cervical carcinoma pattern.

gnant melanoma of the cervix [6, 8, 12, 13]: (a) presence of melanin in the cervical epithelium; (b) absence of melanoma in another site of the body; (c) presence of binding activity in the cervical epithelium near the lesion; (d) if metastatic disease is found, it should be according to the cervical carcinoma pattern. At the present moment there is no standard treatment for this disease, while there is no doubt that the surgical approach is the most usual and radical hysterectomy with or without pelvic lymphadectomy and/or superior vaginectomy is reported most frequently [1, 2, 7, 8], some authors entertain doubts concerning survival if pelvic lymphadectomy is performed [2]. Although there is no enough information about the real role of negative margins in primary melanoma of the cervix, the primary surgery should have the purpose of obtaining negative margins [6]; some authors recommend 2-cm margins as minimum [7, 9]. The role of radiotherapy (RT) has not been well established, but it has been demonstrated that RT reduces the tumor size [1, 8, 11]. The use of adjuvant pelvic RT is considered in the case of not obtaining a satisfactory surgical resection margins, when the parametrium is involved, or when lymph nodes are found to be involved [6]. Despite the low level of radiosensitivity exhibited by melanoma, the use of external or intracavitary RT is recommended after surgery, or palliatively when the tumor is inoperable because of its own conditions or those of the patient [1, 6, 8, 10].

ametrium is involved, or when lymph nodes are found to be involved [6]. Despite the low level of radiosensitivity exhibited by melanoma, the use of external or intracavitary RT is recommended after surgery, or palliatively when the tumor is inoperable because of its own conditions or those of the patient [1, 6, 8, 10]. No chemotherapy regimes have been reported that substantially may reduce the possibility of recurrence. Dacarbazin is utilized in advanced disease, and it has been observed that up to 20% of patients may have response [6, 7]. It has been proposed that the combination of cisplatin, bleomycin, and vinblastine can provoke a better response than the use of solely dacarbazine [1], while in other cases, the greater effectiveness of combining dacarbazine with vincristine and carmustine has been reported as well as immunotherapy utilizing local BCG or the transfusion of activated lymphocytes [10]. Average survival reported in the world literature of these patients ranges from 6 months to 14 years. The majority of them report that they succumb to the disease in the first 3 years after diagnosis [8]. Five-year survival after radical hysterectomy as only treatment is very low: less than 40% in stage I and 14% in stage II [1, 14]. The limited experience in the management of these cases, the unpredictable biological behavior of the disease, and the great variety of treatments employed render the choice of an optimal therapy for primary malignant melanoma of the cervix difficult [4, 7].

Average survival reported in the world literature of these patients ranges from 6 months to 14 years. The majority of them report that they succumb to the disease in the first 3 years after diagnosis [8]. Five-year survival after radical hysterectomy as only treatment is very low: less than 40% in stage I and 14% in stage II [1, 14]. The limited experience in the management of these cases, the unpredictable biological behavior of the disease, and the great variety of treatments employed render the choice of an optimal therapy for primary malignant melanoma of the cervix difficult [4, 7]. In our case, due to the magnitude of the tumor, it was decided to employ total pelvic exenteration, showing extraordinary results, because the patient continues to be alive and disease-free at 8 years of surgery, which is in contrast importantly with that reported in the literature after treatment with radical hysterectomy. We propose consideration of this surgical approach and adjuvant RT for primary melanoma of the cervix as the initial treatment of choice. It is necessary to carry out long-term followup and biological research to obtain a consensus with regard to the diagnosis, treatment, and prognosis of the disease [11].

In our case, due to the magnitude of the tumor, it was decided to employ total pelvic exenteration, showing extraordinary results, because the patient continues to be alive and disease-free at 8 years of surgery, which is in contrast importantly with that reported in the literature after treatment with radical hysterectomy. We propose consideration of this surgical approach and adjuvant RT for primary melanoma of the cervix as the initial treatment of choice. It is necessary to carry out long-term followup and biological research to obtain a consensus with regard to the diagnosis, treatment, and prognosis of the disease [11]. 5. Conclusion Primary malignant melanoma of the cervix is a rare disease with a poor prognosis, especially if it is not detected in a timely fashion or if it is not treated correctly. To date, no consensus has been established concerning treatment of primary melanoma of the cervix, but it is recommended that this be surgical, procuring the establishment of 2-cm margins, accompanied by radio- or chemotherapy. The majority of the case reports found suggest radical hysterectomy as the treatment indicated for these patients; notwithstanding this, survival is less than three years in general when managed in this manner. Total pelvic exenteration for primary malignant melanoma of the cervix offers a feasible initial treatment for these unusual cases. It is necessary initiate collaborative studies and followup of patients to be able to achieve establishing a therapeutic approach that may offer the best results to these unfortunate patients.

l pelvic exenteration for primary malignant melanoma of the cervix offers a feasible initial treatment for these unusual cases. It is necessary initiate collaborative studies and followup of patients to be able to achieve establishing a therapeutic approach that may offer the best results to these unfortunate patients. Conflict of Interests The authors declare that they have no competing interests. All authors read and approved the final paper. Figure 1 Contrast enhanced CT scan of the pelvis showing a solid cervical mass without evidence of direct involvement of bladder or rectum. Figure 2 Microscopic picture showing a medium power view of the lesion. (HE 10x) in the box Melan A reaction diffuse positive.

1. Introduction This is a case of 18-year-old African American female presenting with profuse per vaginal bleeding three days following an induced second trimester curettage abortion. She was diagnosed to be a case of disseminated intravascular coagulation disorder due to retained products of conception. 2. Case Presentation This 18-year-old (gravida 2 parity 1 living issue 1) African American female was brought by the paramedics in the emergency department with history of profuse bleeding per vagina since morning. She had an induced abortion done by dilatation and curettage 3 days ago at 13 weeks of gestation (by LMP). It was uncomplicated, and she was asymptomatic till morning, when she woke up in a pool of blood followed by a lower abdominal menstrual-like pain. She denied any history of fever, any systemic illness, any foul smelling vaginal discharge, or any use of medications except prenatal vitamins. Her past medical and surgical history and her family history was not significant, and she denied smoking, use of alcohol or substance abuse. Physical examination showed that she was alert, conscious and cooperative, pale, and hypotensive (BP: 84/60 mm of Hg) with tachycardia (pulse: 112/min) and temperature 98.4 F. Her systemic examination was unremarkable except the tachycardia and tenderness in the suprapubic area. She has no evidence of any respiratory distress or calf tenderness. Per vaginal examination demonstrated enlarged uterus (16-week size) and some amount of tenderness in the fornix.

e: 112/min) and temperature 98.4 F. Her systemic examination was unremarkable except the tachycardia and tenderness in the suprapubic area. She has no evidence of any respiratory distress or calf tenderness. Per vaginal examination demonstrated enlarged uterus (16-week size) and some amount of tenderness in the fornix. Resuscitation with intravenous fluids was started immediately at the emergency department, and blood was sent for cross matching and lab works. Packed red blood cells (RBCs) was started as soon as it was available. Transvaginal sonogram done in the emergency department showed enlarged uterus with intrauterine retained products of conception and also a collection in the pouch of Douglas. Her hemoglobin was 5.5 gm/dl, WBC count 9000/mm3, and platelet count 17000/mm3. P-time was 100 seconds, aPTT−104 seconds, Fibrinogen <50, D−dimer−980 ng/ml, and her B-HCG in blood was raised. Chest X-ray, urine analysis, and her basic metabolic panel were within normal limits. After 2 units of packed RBCs, platelet transfusion was started followed by FFP. She was then taken to the operation room, and under aseptic conditions and general anesthesia, suction and evacuation of the retained products followed by dilatation and curettage were done. Hemostasis was checked and patient revived. She was further transfused packed RBCs, platelets, and FFP in the ICU.

llowed by FFP. She was then taken to the operation room, and under aseptic conditions and general anesthesia, suction and evacuation of the retained products followed by dilatation and curettage were done. Hemostasis was checked and patient revived. She was further transfused packed RBCs, platelets, and FFP in the ICU. Post operative period was uneventful with no evidence of any further bleeding from any sites. She was monitored for any evidence of bleeding and also her hemoglobin and coagulation profile rechecked. In total, she was transfused with 5 units of packed RBCs, 3 units of volume-reduced platelet concentrates, and 4 units of FFP. She had an uncomplicated recovery period and was discharged after 3 days with depot progesterone contraception. The histopathological report of the curettage material later showed products of conception. 3. Discussion Postabortion complications usually are a result of 3 major mechanisms: incomplete evacuation of the uterus and uterine atony, which leads to hemorrhagic complications, infection, Injury due to instruments used during the procedure. DIC should be suspected in all patients who present with severe postabortion bleeding, especially after mid trimester abortions. Incidence is approximately 200 cases per 100,000 abortions; this rate is even higher for saline instillation techniques (660 per 100,000 abortions) [1].

Injury due to instruments used during the procedure. DIC should be suspected in all patients who present with severe postabortion bleeding, especially after mid trimester abortions. Incidence is approximately 200 cases per 100,000 abortions; this rate is even higher for saline instillation techniques (660 per 100,000 abortions) [1]. Disseminated intravascular coagulation (DIC) is an acquired syndrome characterized by the intravascular activation of coagulation with loss of localization arising from different causes. It can originate from and cause damage to the microvasculature, which, if sufficiently severe, can produce organ dysfunction [2]. The resultant clinical condition is characterized by intravascular coagulation and hemorrhage. The affected person is often acutely ill and shocked with widespread hemorrhage (common bleeding sites are mouth, nose, and venepuncture sites), extensive bruising, renal failure, and gangrene. DIC usually presents as an acute, often catastrophic, acquired hemorrhagic tendency. Rarely it can also manifest as a low-grade disorder with predominantly thrombotic manifestations [3].

despread hemorrhage (common bleeding sites are mouth, nose, and venepuncture sites), extensive bruising, renal failure, and gangrene. DIC usually presents as an acute, often catastrophic, acquired hemorrhagic tendency. Rarely it can also manifest as a low-grade disorder with predominantly thrombotic manifestations [3]. The most common cause of abortion-related DIC is amniotic fluid embolism, which is when amniotic fluid gets into the mother's blood stream. This can be caused by lacerations of the uterus or by compromised blood vessels when the placenta detaches. Infection, either localized in the uterus or generalized (septicemia), can also trigger DIC. Retained products of conception which are most likely to cause DIC when they gets into the maternal bloodstream are fetal brain. There is much debate about the best method of dealing with the fetal head. Some believe that it is best to suction the fetal brain out of the head before crushing it. This eliminates fetal brain matter from the area when the sharp pieces of fetal skull are being removed. These sharp bony fragments are the fetal parts most likely to scratch the cervix and allow fetal tissue to get into the mother's bloodstream.

it is best to suction the fetal brain out of the head before crushing it. This eliminates fetal brain matter from the area when the sharp pieces of fetal skull are being removed. These sharp bony fragments are the fetal parts most likely to scratch the cervix and allow fetal tissue to get into the mother's bloodstream. Others argue that it is sometimes difficult to be sure that the structure you are grasping really is the fetal head and not some part of the mother's body. They recommend squeezing the structure and watching the mother's cervix to see if grey material (fetal brain) oozes out (calvarium show). The sight of “calvarium show" means that the grasped structure is indeed the head of the fetus and it is safe to extract it. Since DIC can quickly become fatal, it is important that medical professionals recognize the symptoms and diagnose it quickly. Definitive diagnosis depends on the result of [4] thrombocytopenia, Prolongation of prothrombin time and activated partial thromboplastin time, a low fibrinogen concentration, increased levels of fibrin degradation products. The only effective treatment is the reversal of the underlying cause. Anticoagulants are rarely used. Platelets may be transfused if counts are less than 5,000–10,000/mm3 and massive hemorrhage is occurring, and fresh frozen plasma may be administered in an attempt to replenish coagulation factors and antithrombotic factors.

1. Introduction Endometrial cancer is the fourth most common cancer among women in the United States. According to the latest data from the National Cancer Institute, approximately 40,100 new cases and 7,470 deaths from uterine cancer are expected in 2008 [1]. In Shanghai, China, the age-adjusted incidence of endometrial cancer increased by 4.41% per year between 1973 to 1975 and 1997 to 1999 [2–4]. Endometrial cancer has been divided into two subtypes (type I and II) based on different morphological appearance and clinicalpathological characteristics [5–7]. Type I, which is endometrioid histology with relatively low-grade features and good prognoses, occurs most often in postmenopausal women and is associated with unopposed estrogen exposure. In contrast, type II endometrial cancer comprises of nonendometrial histology with an aggressive clinical course and poor prognoses.

ype I, which is endometrioid histology with relatively low-grade features and good prognoses, occurs most often in postmenopausal women and is associated with unopposed estrogen exposure. In contrast, type II endometrial cancer comprises of nonendometrial histology with an aggressive clinical course and poor prognoses. Uterine papillary serous carcinoma (UPSC) is the most common subtype of type II endometrial cancer. It was first described by Hendrickson et al. [8] in 1982, and histologically similar to high-grade ovarian cancer. The microscopic criteria for diagnosis were described as neoplastic epithelium, characterized by serous differentiation with psammoma bodies present and predominantly papillary architecture [8]. Although UPSC represents only about 10% of endometrial cancers, they account for over 50% of recurrences and deaths caused by endometrial cancer. The 5-year overall survival (OS) for women with UPSC is only 46% for all stages [9]. Comprehensive surgical staging is recommended for all patients with UPSC, regardless of the depth of myometrial invasion of the tumor [9, 10]. Chemotherapy is recommended after the surgery, but unlike ovarian papillary serous carcinomas, UPSC is a chemoresistant disease from onset, with low response rates and a short duration of response [11, 12]. Given the overall poor prognosis of these patients, new treatment modalities are urgently needed for UPSC.

, 10]. Chemotherapy is recommended after the surgery, but unlike ovarian papillary serous carcinomas, UPSC is a chemoresistant disease from onset, with low response rates and a short duration of response [11, 12]. Given the overall poor prognosis of these patients, new treatment modalities are urgently needed for UPSC. Her-2/neu is one of the most studied molecular markers in anticancer therapy. Her-2/neu, also known as c-erbB2, is a member of the epidermal growth factor receptor transmembrane receptor tyrosine kinase family [13]. 25% to 30% of breast cancers were found to have Her-2/neu overexpression, which is considered as a negative prognostic factor [14, 15] as well as a predictive marker of resistance to tamoxifen therapy, benefit from doxorubicin-based adjuvant chemotherapy, and trastuzumab (anti-Her-2 monoclonal antibody) [16–18]. Recently, a striking overexpression of the Her-2/neu in UPSC has been identified by Santin et al, with the rates ranging from 18% to 80% in different studies [19–21]. In this study, we conducted a single institutional review of 10 years summary of this rare disease, which, to our knowledge, is the largest one so far analysing the clinicopathological features, prognostic factors, and Her-2/neu status of Chinese patients with UPSC. The purpose of this study is to better understand clinicopathological features of UPSC and the relationship with Her-2/neu abnormality.

his rare disease, which, to our knowledge, is the largest one so far analysing the clinicopathological features, prognostic factors, and Her-2/neu status of Chinese patients with UPSC. The purpose of this study is to better understand clinicopathological features of UPSC and the relationship with Her-2/neu abnormality. 2. Materials and Methods 2.1. Patient Population and Tumor Specimens All the slides of the patients with endometrial carcinoma who underwent surgery at Cancer hospital, Fudan University, from January 1996 to January 2006, were reviewed by two pathologists separately. If there was dispute in diagnosis between them, slides would be reviewed by another pathologist who made the final diagnosis. Altogether 457 patients with endometrial cancer were treated in our hospital during the last 10 years and 36 cases of UPSC were rediagnosed, which accounted for about 8%. Their clinical and surgical pathology reports were recorded. Patients were staged according to the 1988 FIGO surgical criteria [22]. Overall Survival (OS) was the primary end-point evaluated. It was calculated from the date of the surgical diagnosis. The information on survival was obtained in the medical record and death certificate. If this information was not available, the patient was censored after her last contact.

2. Materials and Methods 2.1. Patient Population and Tumor Specimens All the slides of the patients with endometrial carcinoma who underwent surgery at Cancer hospital, Fudan University, from January 1996 to January 2006, were reviewed by two pathologists separately. If there was dispute in diagnosis between them, slides would be reviewed by another pathologist who made the final diagnosis. Altogether 457 patients with endometrial cancer were treated in our hospital during the last 10 years and 36 cases of UPSC were rediagnosed, which accounted for about 8%. Their clinical and surgical pathology reports were recorded. Patients were staged according to the 1988 FIGO surgical criteria [22]. Overall Survival (OS) was the primary end-point evaluated. It was calculated from the date of the surgical diagnosis. The information on survival was obtained in the medical record and death certificate. If this information was not available, the patient was censored after her last contact. 2.2. Immunohistochemistry (IHC) Formalin-fixed, paraffin-embedded specimens were collected from tissue bank of Cancer hospital, Fudan University. For all cases, a routine hematoxylin and eosin slide was evaluated to ensure that the specimen evaluated had serous carcinoma. IHC was performed using c-erbB-2/Her-2/neu Ab (A0485, Dako Corp., Denmark). This antibody is a polyclonal rabbit anti-human c-erbB-2 oncoprotein. IHC staining was performed on 4-um-thick paraffin-embedded sections according to the manufacturer's instructions. Briefly, after pretreatment with 10 mM citrate buffer at pH 6.0 using a steamer, they were incubated with c-erbB-2/Her-2/neu Ab at 37°C for 2 hours, then incubated with immunoperoxidase anti-rabbit Ab (K4003, Dako Corp., Denmark) for half an hour and followed by substrate-chromogen solution( DAB). Negative controls were analyzed on slides incubated without the antibodies. Her-2/neu-expressing breast cancer was used as a positive control. The intensity of immunostaining was graded as follows: negative (0); incomplete membranous staining or complete membranous staining in less than 10% of the tumor cells (1+); moderate intensity, complete membranous staining in greater than 10% of the tumor cells (2+); or strong intensity, complete membranous staining in greater than 10% of the tumor cells (3+). Tumors with 2+ or 3+ staining were considered Her-2/neu overexpression [23, 24]. The sections were examined by light microscopy by two different pathologists without knowledge of the clinical outcome.

of the tumor cells (2+); or strong intensity, complete membranous staining in greater than 10% of the tumor cells (3+). Tumors with 2+ or 3+ staining were considered Her-2/neu overexpression [23, 24]. The sections were examined by light microscopy by two different pathologists without knowledge of the clinical outcome. 2.3. Chromogenic In Situ Hybridization (CISH) CISH was done using Zymed SPoT-Light HER-2 CISH Kit (84-0146) according to instructions of manufacture. Sections of 3 μm were cut from paraffin-embedded archival tissue from tissue bank. The sections were deparaffinized and pretreated in a microwave oven at 100°C for 15 minutes. After a brief rinsing with Tris-buffered saline (0.05 mol/L Tris/HCl saline, pH 7.4–7.6), the slides were digested with Enzyme for 5 to 10 minutes (the time for each slide was different), followed by rinsing with distilled water for 3 times, and dehydrating with graded ethanols. The digoxigenin-labelled Her-2/neu probe (Zymed) was applied onto the slides, covered with coverslips and denatured at 95°C for 5 minutes. The hybridization was performed overnight at 37°C. The slides were then washed with standard saline citrate (SSC) in room temperature briefly then with SSC at 75°C for 5 minutes. Immunodetection was performed by the labelled avidin-biotin complex peroxidase system according to the manufacturer's instructions. Finally, sections were lightly counterstained with haematoxylin. Breast cancer with known Her-2/neu-amplification was used as a positive control. Benign endometrium was used as a negative control. Her-2/neu gene signals were scored in histological sections, in at least 150 neoplastic cells, using a conventional microscope (Olympus). Tumours were classified, depending on the number of gene copies in the nuclei, as normal (1–5 copies) and amplified (>6 copies or when large gene copy clusters were seen in at least 50%). In addition, amplified tumours were classified as low-level (6–10 copies) or high level (>10 copies).

entional microscope (Olympus). Tumours were classified, depending on the number of gene copies in the nuclei, as normal (1–5 copies) and amplified (>6 copies or when large gene copy clusters were seen in at least 50%). In addition, amplified tumours were classified as low-level (6–10 copies) or high level (>10 copies). 2.4. Statistical Analysis X 2 tests were performed to assess prognostic factors. Kaplan-Meier survival curves were used to estimate OS. The log-rank test was used to compare survival curves. Cox's proportional hazards regression was used to model survival with Her-2/neu IHC overexpression and clinicopathological variables typically associated with prognosis. Spearman correlation was used to analyze the relation between the results of IHC and CISH. SPSS version 11.5 software was used for all analyses. 3. Results 3.1. Clinicopathological Features of Patients The median age of the patients was 63 years (mean 64 years, range 45~81 years). The median BWI (Body Weight Index) was 23.8 kg/m2 (mean 23.73, range 17.6~33.8). The main presenting symptoms were abnormal vaginal bleeding (86%, 31/36), while other 4 patients first presented with abdominal symptoms such as abdominal mass and ascites, and 1 with vaginal discharge. 16 patients (45.7%) were complicated with hypertension and 4 patients (11.4%) with diabetes. 8 cases or relatives had history of other cancers, including 1 with ovarian mucous adenocarcinoma, 2 with breast cancer, 1 with endometrial cancer, 3 with gastrointestinal cancer, and 1 with thyroid neoplasms.

aginal discharge. 16 patients (45.7%) were complicated with hypertension and 4 patients (11.4%) with diabetes. 8 cases or relatives had history of other cancers, including 1 with ovarian mucous adenocarcinoma, 2 with breast cancer, 1 with endometrial cancer, 3 with gastrointestinal cancer, and 1 with thyroid neoplasms. Of these 36 patients, 11 (31%) were stage I (1 IA, 6 IB, 4 IC), 3 (8%) were stage II (1 IIA, 2 IIB), 9 (25%) were stage III (4 IIIA, 1 IIIB, 4 IIIC), and 13 (36%) were stage IV (13 IVB). Altogether 39% were early stages (stage I/II) and 61% were late stages (stage III/IV). 21 tumors invaded more than half of myometrium, in which 11 invaded to serosa of the uterus. 15 tumors invaded less than half of myometrium. And only two patients had no myometrial invasion. 20 tumors (55.6%) demonstrated LVSI (Lymph-vascular space invasion). Twenty-seven tumors (75%) had overexpression of p53, while positive ER expression was only detected in 10 cases (27.8%), and PR expression in 7 cases (19.4%).

mors invaded less than half of myometrium. And only two patients had no myometrial invasion. 20 tumors (55.6%) demonstrated LVSI (Lymph-vascular space invasion). Twenty-seven tumors (75%) had overexpression of p53, while positive ER expression was only detected in 10 cases (27.8%), and PR expression in 7 cases (19.4%). All patients underwent exploratory surgery, hysterectomy, bilateral salpingo-oophorectomy, omentectomy, pelvic lymphadenectomy, and/or para-aortic lymphadenectomy. Patients with stage IVB disease also received cytoreductive surgery. 4 patients received chemotherapy before the surgery. After the surgery, 14 patients received platinum-based chemotherapy including cisplatin, cyclophosphamide and anthracycline, or carboplatin and paclitaxel, and so forth. 4 patients received whole pelvic radiotherapy with 2 extended to the para-aortic field. 4 patients received whole pelvic radiotherapy combined with platinum-based chemotherapy. 6 patients had oral tamoxifen or progestin, and 8 patients were observed after the surgery.

cline, or carboplatin and paclitaxel, and so forth. 4 patients received whole pelvic radiotherapy with 2 extended to the para-aortic field. 4 patients received whole pelvic radiotherapy combined with platinum-based chemotherapy. 6 patients had oral tamoxifen or progestin, and 8 patients were observed after the surgery. 3.2. IHC and CISH Of the 36 cases, 13 (36.1%) patients had Her-2/neu protein overexpression by IHC. In this 13 patients, 10 (27.8%) showed 2+ and 3 (8.3%) showed 3+. (Figure 1). The remaining 11 samples showed 1+ and 12 showed 0 for Her-2/neu protein. The clinicopathological features by Her-2/neu IHC status were summarized in Table 1. Overexpression of Her-2/neu was seen in 50.0% (11/22) of UPSC patients with advanced disease (Stage III~IV), compared to 14.3% (2/14) of patients with early disease (Stage I~II). Patients with overexpression of Her-2/neu were significantly more likely to have advanced-stage disease when compared to Her-2/neu negative patients (OR = 6.0, 95%CI: 1.08–33.32, P = .03).

tend to have deep myometrial invasion, high-stage diseases, and worse overall survival, as we had discussed previously in [25]. In our study, 61% were late stages. More than half of patients (58.3%) had deep myometrial invasion. The 1-year, 3-year, and 5-year overall survival was 73.1%, 51.9%, and 43.9%, respectively. Although racial differences in the uterine cancer have been previously published, most of these studies have focused on disparities between African Americans and whites. Santin et al. noted that black patients with UPSC tended to be younger, had higher Her-2/neu expression and short survival than white [26]. Slomovitz [9] summarized the clinical features of 129 cases with UPSC. In his study, 93% patients were Caucasian, and only 2% were Asian. Therefore, little is known about Asian patients with UPSC till now. Recent study had reported the racial disparities between Asians and whites. Zhang et al. [27] compared 2,144 Asians and 32,999 whites with corpus cancer in the United States, and found that Asians presented at a younger age, with more advanced stage disease and higher 5-year survival rates than whites. In our study, similar to whites, main presenting symptoms were abnormal vaginal bleeding and with median BWI of 23.8 kg/m2, patients with UPSC ware not associated with obesity. On the other hand, Asian patients with UPSC had younger age (64 years) and more late stage diseases (61%) than whites compared to the previous report [9], in which median age was 68 years for whites and 56% patients were in late stages. The 3-year and 5-year overall survival in our study was 51.9% and 43.9%, respectively, which was a little worse than whites, whose 3-year and 5-year overall survival was 62.6% and 45.9%, respectively [9].

pared to the previous report [9], in which median age was 68 years for whites and 56% patients were in late stages. The 3-year and 5-year overall survival in our study was 51.9% and 43.9%, respectively, which was a little worse than whites, whose 3-year and 5-year overall survival was 62.6% and 45.9%, respectively [9]. Recently, Her-2/neu overexpression was found to be a common event in UPSC. Grushko et al. [28] found that UPSC had more Her-2/neu overexpression than all other types of endometrial cancers (23 of 38, 61% versus 81 of 196, 41%, resp., P = .03). Santin et al. [20, 21] reported that 62%~80% of UPSC overexpressed Her-2/neu protein, and others reported overexpression rates varyng from 40% to 48% [29–31]. In contrast, Slomovitz et al. [19] reported overexpression of Her-2/neu in only 18% (12 of 68) of UPSC patients. Table 4 summmariued the results and methods of the studies on Her-2/neu status in UPSC. In our study, we demonstrated that 36% of patients with UPSC overexpressed Her-2/neu protein. The differences of the rate of Her-2/neu overexpression, might be due to different antibody, inherent intraobserver variability, and most important of all, racial disparity.

methods of the studies on Her-2/neu status in UPSC. In our study, we demonstrated that 36% of patients with UPSC overexpressed Her-2/neu protein. The differences of the rate of Her-2/neu overexpression, might be due to different antibody, inherent intraobserver variability, and most important of all, racial disparity. In our study, patients with Her-2/neu overexpression were significantly more likely to have advanced stage disease when compared to Her-2/neu negative patients (P = .03). Overexpression of Her-2/neu was seen in 50.0% (11/22) of UPSC patients with advanced disease, compared to 14.3% (2/14) of patients with early disease. Similar findings were reported by Slomovitz, Santin, and Díaz-Montes et al., in which overexpression of Her-2/neu in patients with advanced disease were 24%~81.8%, compared to 8%~28.6% of patients with early disease [19, 20, 30]. Díaz-Montes et al. also demonstrated that overexpression of Her-2/neu was associated with higher Ki-67 index, larger tumor sizes, and worse survival outcome.

s et al., in which overexpression of Her-2/neu in patients with advanced disease were 24%~81.8%, compared to 8%~28.6% of patients with early disease [19, 20, 30]. Díaz-Montes et al. also demonstrated that overexpression of Her-2/neu was associated with higher Ki-67 index, larger tumor sizes, and worse survival outcome. In survival analysis, we found that the overall survival was worse for patients with Her-2/neu protein positive tumors than Her-2/neu negative ones (P = .0023). The 5-year overall survival for Her-2/neu IHC positive and negative cases was 12.9% and 68.6%, respectively. Slomovitz et al. [19] also revealed that the 5-year overall survival in UPSC patients was 0% in Her-2/neu IHC positive cases versus 45% in Her-2/neu negative ones (P = .008). And Santin et al. [26] demonstrated that short survival was associated significantly with Her-2/neu overexpression compared with IHC expression 0~1+(P = .002). All of these findings suggested that Her-2/neu overexpression was a useful prognostic factor for this aggressive subtype of endometrial cancer. Besides Her-2/neu IHC overexpression, we also found that deep myometrial invasion (P = .0138) and late stages (P = .003) were associtated with a shorter OS.

ion 0~1+(P = .002). All of these findings suggested that Her-2/neu overexpression was a useful prognostic factor for this aggressive subtype of endometrial cancer. Besides Her-2/neu IHC overexpression, we also found that deep myometrial invasion (P = .0138) and late stages (P = .003) were associtated with a shorter OS. We used CISH to detect the Her-2/neu gene amplification in our study and found Her-2/neu gene amplification in four cases (11.1%). Santin et al. [32] revealed that Her-2/neu gene amplification was correlated with a poor survival outcome in patients with UPSC (P = .0084). Because of the small size of Her-2/neu amplification in our study, we did not find the effect of Her-2/neu amplification on overall survival.

amplification in four cases (11.1%). Santin et al. [32] revealed that Her-2/neu gene amplification was correlated with a poor survival outcome in patients with UPSC (P = .0084). Because of the small size of Her-2/neu amplification in our study, we did not find the effect of Her-2/neu amplification on overall survival. Her-2/neu gene amplification was found in all IHC 3+ cases (100%, 3/3). But in IHC 2+ cases, only one out of ten cases had Her-2/neu gene amplification. As we know, patients who have Her-2/neu gene amplification respond better to trastuzumab. Therefore, like breast cancer, the entry criteria of trastuzumab in UPSC should be confined to IHC 3+ cases, excluding the 2+ cases unless it is proved to have Her-2/neu amplification by CISH or FISH. The Gynecologic Oncology Group conducted a phase II study using trastuzumab as a single agent in patients with advanced stage or recurrent endometrial cancer with Her-2/neu IHC 2+ or 3+ staining, but found no activity of trastuzumab in heavily pretreated patients. 13% of cases demonstrated gene amplification and 37% demonstrated moderate or high immunostaining. Entry criteria were then revised to include only those patients whose tumors were FISH positive, but the study was finally closed due to slow enrollment [33].

und no activity of trastuzumab in heavily pretreated patients. 13% of cases demonstrated gene amplification and 37% demonstrated moderate or high immunostaining. Entry criteria were then revised to include only those patients whose tumors were FISH positive, but the study was finally closed due to slow enrollment [33]. Santin et al. [21] studied the effects of trastuzumab in UPSC cell lines and found that UPSC cell lines were highly sensitive to Herceptin mediated antibody-dependent cellular cytotoxicity (ADCC) and cell proliferation was inhibited. Villella et al. [34] tried to use trastuzumab therapy (4 mg/kg intravenously over 90 min, then maintenance dose of 2 mg/kg intravenously over 30 min weekly until progression of disease) in patients with UPSC who expressed Her-2/neu 3+ by IHC. Two patients with 3+ overexpression received trastuzumab treatment. One patient with IVB disease had complete response and the other with IIIC disease had stable disease for 3 months. Targeting Her-2/neu might be beneficial for a select group of patients with UPSC. In conclusion, our data revealed that Chinese patients with UPSC had characteristics of deep myometrial invasion, advanced stages and poor overall survival. Her-2/neu overexpression, rather than Her/neu gene amplification, is associated with advanced stage and poor survival outcome. Her-2/neu might be a therapeutic target in refractory UPSC patients, but it should be further evaluated by randomized clinical trials.

trial invasion, advanced stages and poor overall survival. Her-2/neu overexpression, rather than Her/neu gene amplification, is associated with advanced stage and poor survival outcome. Her-2/neu might be a therapeutic target in refractory UPSC patients, but it should be further evaluated by randomized clinical trials. Figure 1 (a) Hematoxylin and eosin stain and (b) immunohistochemical staining 3+ for Her-2/neu expression on paraffin-embedded uterine serous carcinoma specimens. ×200. (c) Her-2/neu gene amplification amplified in high level. ×400. Figure 2 Overall survival curve for all 36 patients with UPSC. Figure 3 Kaplan-Meier curve based on Her-2/neu protein expression (IHC positive versus IHC negative, P = .0023). Figure 4 Kaplan-Meier curve based on stage (stage I/II versus stage III/IV, P = .003). Figure 5 Kaplan-Meier curve based on myometrial invasion (≥1/2 versus <1/2, P = .0138). Table 1 Clinicopathological features by Her-2/neu IHC status.

Figure 3 Kaplan-Meier curve based on Her-2/neu protein expression (IHC positive versus IHC negative, P = .0023). Figure 4 Kaplan-Meier curve based on stage (stage I/II versus stage III/IV, P = .003). Figure 5 Kaplan-Meier curve based on myometrial invasion (≥1/2 versus <1/2, P = .0138). Table 1 Clinicopathological features by Her-2/neu IHC status. Clinicopathological features by Her-2 IHC status positive (n = 13) negative (n = 23) P value Age (years) Median (range) 64 (58–77) 61 (45–81) .22 Stage .03 I~II 2 (15.4%) 12 (52.2%) III~IV 11 (84.6%) 11 (47.8%) Myometrial invasion .78 <1/2 5 (38.5%) 10 (43.5%) ⩾1/2 8 (61.5%) 13 (56.5%) Histology .33 Pure 9 (69.2%) 12 (52.2%) Mixed 4 (30.8%) 11 (47.8%) LVSI .4 Absent 7 (53.8%) 9 (39.1%) Present 6 (46.2%) 14 (60.9%) Grade .45 II 3 (23.1%) 3 (13.0%) III 10 (76.9%) 20 (87.0%) p53 .08 Absent 1 (7.7%) 8 (34.8%) Present 12 (92.3%) 15 (65.2%) ER .69 Absent 10 (76.9%) 16 (69.6%) Present 3 (23.1%) 7 (30.4%) PR .66 Absent 11 (84.6%) 18 (78.3%) Present 2 (15.4%) 5 (21.7%) Table 2 Correlation of Her-2/neu status by IHC and CISH. Her-2 protein expression by IHC P value 3 + (n = 3) 2+ (n = 10) 1+ (n = 11) 0 (n = 12) Her-2 Amplification by CISH High-level 2 1 0 0 .001 Low-level 1 0 0 0 No 0 9 11 12 Table 3 Univariate analysis of Her-2/neu IHC status and clinicopathological factors with overall survival.

Clinicopathological features by Her-2 IHC status positive (n = 13) negative (n = 23) P value Age (years) Median (range) 64 (58–77) 61 (45–81) .22 Stage .03 I~II 2 (15.4%) 12 (52.2%) III~IV 11 (84.6%) 11 (47.8%) Myometrial invasion .78 <1/2 5 (38.5%) 10 (43.5%) ⩾1/2 8 (61.5%) 13 (56.5%) Histology .33 Pure 9 (69.2%) 12 (52.2%) Mixed 4 (30.8%) 11 (47.8%) LVSI .4 Absent 7 (53.8%) 9 (39.1%) Present 6 (46.2%) 14 (60.9%) Grade .45 II 3 (23.1%) 3 (13.0%) III 10 (76.9%) 20 (87.0%) p53 .08 Absent 1 (7.7%) 8 (34.8%) Present 12 (92.3%) 15 (65.2%) ER .69 Absent 10 (76.9%) 16 (69.6%) Present 3 (23.1%) 7 (30.4%) PR .66 Absent 11 (84.6%) 18 (78.3%) Present 2 (15.4%) 5 (21.7%) Table 2 Correlation of Her-2/neu status by IHC and CISH. Her-2 protein expression by IHC P value 3 + (n = 3) 2+ (n = 10) 1+ (n = 11) 0 (n = 12) Her-2 Amplification by CISH High-level 2 1 0 0 .001 Low-level 1 0 0 0 No 0 9 11 12 Table 3 Univariate analysis of Her-2/neu IHC status and clinicopathological factors with overall survival. No. of patients No. of Deaths Survival Rate (%) P value 1-year 3-years 5-years 36 17 73.1 51.9 43.9 Her-2/neu by IHC .0023 Positive 13 11 60 25.7 12.9 Negative 23 6 81 68.6 68.6 Stage .0006 I~II 14 3 92.6 84.5 60.4 III~IV 22 14 60 26.7 16 Myometrial invasion .0138 <1/2 15 4 85.7 77.6 55.4 ⩾1/2 21 13 64.1 33.1 19.9 Histology .948 Pure 21 10 74.4 50.6 30.3 Mixed 15 7 71.4 54 36 LVSI .4884 Present 20 10 63.2 51.1 30.7 Absent 16 7 86.2 52.8 37.7 Grade .8689 II 6 3 75 50 50 III 30 14 68.4 51.8 41.5 Table 4 Summary of related articles on Her-2/neu status in UPSC.

etrial invasion .0138 <1/2 15 4 85.7 77.6 55.4 ⩾1/2 21 13 64.1 33.1 19.9 Histology .948 Pure 21 10 74.4 50.6 30.3 Mixed 15 7 71.4 54 36 LVSI .4884 Present 20 10 63.2 51.1 30.7 Absent 16 7 86.2 52.8 37.7 Grade .8689 II 6 3 75 50 50 III 30 14 68.4 51.8 41.5 Table 4 Summary of related articles on Her-2/neu status in UPSC. Year Author Methods Number of Patients Positivity (%) 1994 Prat et al. IHC 10 40% (4/10) 2001 Halperin et al. IHC 22 45% (10/22) 2002 Santin et al. IHC 10 80% (8/10) 2004 Slomovitiz et al. IHC 68 18% (12/68) FISH 12 17% (2/12) 2005 Santin et al. IHC 26 62% (16/26) FISH 42% (11/26) 2006 Diaz-Montes IHC 25 48% (12/25) 2008 Grushko et al. IHC 38 61% (23/38) FISH 28 21% (6/28)

1. Introduction Imperforate hymen is reported at an approximate rate of 0,1% and occurs due to the incomplete canalization of the Mullerian system and the urogenital system [1]. The treatment of this condition that can be easily diagnosed through gynecological examination is surgical [2]. In this paper, we present an adolescent girl with a spontaneous ruptured imperforate hymen who is the first case in the literature.

1. Introduction Imperforate hymen is reported at an approximate rate of 0,1% and occurs due to the incomplete canalization of the Mullerian system and the urogenital system [1]. The treatment of this condition that can be easily diagnosed through gynecological examination is surgical [2]. In this paper, we present an adolescent girl with a spontaneous ruptured imperforate hymen who is the first case in the literature. 2. Case Report Gynecological examination of a 13-year-old female patient admitted to our clinic with a cyclic abdominal pain and amenorrhea revealed bulging out of a thinned hymen from the introitus with a blue reflection. Furthermore, on rectal examination, a mass lesion of approximately 10 cm in size was palpated and ultrasonography showed a 14 × 8 cm mass lesion compatible with hematocolpometra. The patient was hospitalized with a diagnosis of imperforate hymen, and hymenotomy was planned. She had severe abdominal and pelvic pain with a dark bloody discharge on the night just before the operation. Reexamination revealed an opening with irregular borders in the upper half of the hymenal ring. The ruptured area was assessed to be adequate to provide menstrual drainage, and the patient was decided to be followedup. She was also informed about the possibility of reclosure of the hymen. On her next visit two months later, she described a normal menstrual period. Although lower half of the hymen was still closed, it was open in upper half with deflorations at 3 and 9 o'clock positions (Figure 1). Hematocolpometra did not exist any more. In her last visit, one year later, her menstruations were still regular and adequate without any complaint.

escribed a normal menstrual period. Although lower half of the hymen was still closed, it was open in upper half with deflorations at 3 and 9 o'clock positions (Figure 1). Hematocolpometra did not exist any more. In her last visit, one year later, her menstruations were still regular and adequate without any complaint. 3. Discussion Cyclical pelvic pain 2.5 to 4 years after thelarche is the usual presentation in a teenager with imperforate hymen [3]. Sometimes it can be fairly asymptomatic, and rarely there is obstruction of the urinary tract from the hematocolpos [4]. With the improving effect of estrogenization on healing, hymenal resection is best performed in adolescence [3]. This case report is an example of spontaneous hymenal rupture and no similar case report was found in the literature. Bakos and Berglund [5] reported a case of acute abdomen due to bilateral hematosalpinx and unilateral rupture of the hematosalpinx owing to an imperforate hymen. In that report, rupture of the tuba was thought to be as a result of distention due to the increased pressure along with hematometrocolpos. Similarly, in our case, we think that the retrograde accumulation of menstrual blood leaded to hematometrocolpos, increased intravaginal pressure and perforated the hymen. Therefore, when there is an increase in the intensity of pain of an adolescent girl with hematocolpometra, spontaneous rupture of the imperforate hymen should be kept in mind and examination should be repeated.

ion of menstrual blood leaded to hematometrocolpos, increased intravaginal pressure and perforated the hymen. Therefore, when there is an increase in the intensity of pain of an adolescent girl with hematocolpometra, spontaneous rupture of the imperforate hymen should be kept in mind and examination should be repeated. It may be interesting to see what will happen when this girl gets married. There is a possibility that she can develop a rigid hymen that has to be surgically removed. As a conclusion, the time of spontaneous rupture cannot be predicted, and, therefore, waiting for that in an adolescent girl with imperforate hymen should not be recommended. However, hymenotomy may not be needed in cases with spontaneous rupture of imperforate hymen if adequate opening for menstrual discharge is warranted. Close followup and operating only the ones with reclosure or inadequate hymenal opening may be logical as far as a minimal invasive approach is concerned. Figure 1 The appearance of the hymen two months after spontaneous rupture, with deflorations at the 3 and 9 o'clock positions (arrows) and the lower half closed (asterisk).

curacy in results [7]. The finding of small groups of nonatypical epithelial cells in an inflammatory and proteinaceous background in FNA biopsy was similar to those of endometriosis in other sites. The absence of atypia helps to differentiate inguinal endometriosis from metastatic carcinoma to the inguinal region [7]. 3. Use of Hormonal Therapy Recurrences are rare after adequate surgical treatment. Hormonal therapy has shown to be a useful adjunct in the management of these cases. Batistta Candiani et al. [11] reported that in one out of the six patients with inguinal endometriosis treated by surgical excision, the inguinal mass recurred 14 years after the initial surgery [11]. This patient underwent a second exploration which revealed a multinodular subinguinal endometriotic lesion with ill-defined borders mixed with scar tissue and infiltrating the thigh muscles. Excision was incomplete and patient was given Danazol 400 mg/day for the residual endometriosis [11]. In terms of the usage of prophylactic hormonal therapy for endometriosis, Quagliarello et al. [8] (1985) cautioned that, due to its side effects, suppressive therapy with Danazol should be reserved only for patients with laparoscopic evidence of coexisting pelvic endometriosis [8].

1. Introduction Fetal abnormalities are the major cause of perinatal death and contribute to the increase of the morbidity after birth [1, 2]. In this context prenatal ultrasound has become the main method for detecting fetal anomalies [3, 4]. Accurate prenatal diagnosis of fetal anomalies is essential to the parental decision and the assessment of the recurrence risk. The genetic evaluation and fetal pathology examination after the TOP allow to improve the ability to achieve a diagnosis, to refine the prognosis, and to enhance the genetic counseling, as well as to audit the quality and accuracy of prenatal ultrasound [5–8]. The main goal of this study was to review the prenatal diagnosis experience in cases of TOP for fetal abnormalities in our department over a period of eight years. We have collected and compared the prenatal clinical findings with the genetic evaluation and the postmortem results. This was done in order to assess the contribution of genetic evaluation and autopsy in improving the diagnostic skills and the couple's counseling. 2. Material and Methods We audited 385 cases of TOP performed because of fetal abnormalities between January 1, 2000, and December 31, 2007, at the Prenatal Diagnosis Center of Maternidade Bissaya Barreto, Centro Hospitalar de Coimbra. All cases had data regarding the results of prenatal ultrasound and laboratory evaluation, observation of the fetus after TOP by clinical geneticist, who could request further exams (such as fetal X-rays cytogenetic, molecular, or enzymatic tests of fetal samples) and fetal pathology.

eto, Centro Hospitalar de Coimbra. All cases had data regarding the results of prenatal ultrasound and laboratory evaluation, observation of the fetus after TOP by clinical geneticist, who could request further exams (such as fetal X-rays cytogenetic, molecular, or enzymatic tests of fetal samples) and fetal pathology. All cases with fetal malformations were evaluated for the same team of sonographers, trained in fetal medicine. The ultrasound equipment used was a Toshiba Xario that has transabdominal and transvaginal transducers with frequencies ranging from 3.5 to 5 MHz. Cytogenetic (karyotype and FISH analysis for specific probes) and molecular tests were performed when indicated after amniocentesis or chorionic villus sampling biopsy. In many cases with cytogenetic abnormalities, invasive prenatal diagnosis was carried out due to advanced maternal age (age greater or equal to 35 years at the estimated due date). The postmortem studies were performed after informed consent and were done according to protocols described by Keeling and adapted by Brandão and Laurini [9, 10]. The ultrasound and genetic findings were available to the pathologists before the postmortem evaluation.

Cytogenetic (karyotype and FISH analysis for specific probes) and molecular tests were performed when indicated after amniocentesis or chorionic villus sampling biopsy. In many cases with cytogenetic abnormalities, invasive prenatal diagnosis was carried out due to advanced maternal age (age greater or equal to 35 years at the estimated due date). The postmortem studies were performed after informed consent and were done according to protocols described by Keeling and adapted by Brandão and Laurini [9, 10]. The ultrasound and genetic findings were available to the pathologists before the postmortem evaluation. All TOPs performed at our department were consecutively recorded. We reviewed the TOP due to fetal abnormalities and gathered all the data from the sonographic, genetic (fetal observation and laboratorial tests), and postmortem records. The results were compared for the presence or absence of agreement at the TOP and the final genetic counseling. Total agreement was considered when no relevant findings (regarding abnormalities that threaten life, require surgery or medical treatment, impair significantly organic function or appearance or were relevant to the etiological diagnosis) were added to ultrasound results after genetic evaluation or autopsy. Disagreement was stated when relevant findings were identified at the genetic and/or fetal pathology evaluation. Whenever disagreement was identified, we also evaluated whether it led to a change in the diagnosis and/or prognosis. All the information was used to improve the genetic counseling of these couples.

All TOPs performed at our department were consecutively recorded. We reviewed the TOP due to fetal abnormalities and gathered all the data from the sonographic, genetic (fetal observation and laboratorial tests), and postmortem records. The results were compared for the presence or absence of agreement at the TOP and the final genetic counseling. Total agreement was considered when no relevant findings (regarding abnormalities that threaten life, require surgery or medical treatment, impair significantly organic function or appearance or were relevant to the etiological diagnosis) were added to ultrasound results after genetic evaluation or autopsy. Disagreement was stated when relevant findings were identified at the genetic and/or fetal pathology evaluation. Whenever disagreement was identified, we also evaluated whether it led to a change in the diagnosis and/or prognosis. All the information was used to improve the genetic counseling of these couples. 3. Results A total of 385 cases were included in this study. Chromosome abnormalities were the most common indication for TOP (149/385; 39%). Numerical chromosome abnormalities were diagnosed in 122 fetuses in which trisomy 21 was the leading cause (67 cases) followed by trisomy 18 (21 cases), trisomy 13 (16 cases), X monosomy with hydrops (12 cases), and mosaics for other numerical abnormalities (6 cases). Euploidies occurred in 5 cases (4 triploidies and 1 tetraploidy). Structural unbalanced abnormalities were presented in 22 cases. In 52% (77/149) of the chromosome abnormalities, the invasive prenatal diagnosis was performed due to advanced maternal age.

cases), and mosaics for other numerical abnormalities (6 cases). Euploidies occurred in 5 cases (4 triploidies and 1 tetraploidy). Structural unbalanced abnormalities were presented in 22 cases. In 52% (77/149) of the chromosome abnormalities, the invasive prenatal diagnosis was performed due to advanced maternal age. Central nervous system malformations were diagnosed in 77 cases (20%). Neural tube defects (anencephaly, myelomeningocele and encephalocele) were found in 59 cases (15%). A central nervous tumor was diagnosed in two fetuses. One was an embryonic tumor and the other a giant dermoid cyst. Monogenic disorders were responsible for 11% of TOP (44/385 cases), which could be divided in two groups: those with previous familial history and those in which the fetus was the index case. On the first group, the molecular diagnosis was offered to 13 cases (3.4%). The analysis of an autosomal recessive disease was the most common request. In one couple with a previous child with Walker-Warburg syndrome, the diagnosis was exclusively based on ultrasound findings, because no mutation was found on the index case. Thirty-one cases (8%) belonged to the second group. On these disorders and after the fetal observation and postmortem findings, the genetic assessment allowed an accurate diagnosis in 84% (26/31 cases) of the cases. Their work was crucial in 16 of these cases, the pathological study by itself in five cases of them and a collaboration of both areas in other five cases.

. On these disorders and after the fetal observation and postmortem findings, the genetic assessment allowed an accurate diagnosis in 84% (26/31 cases) of the cases. Their work was crucial in 16 of these cases, the pathological study by itself in five cases of them and a collaboration of both areas in other five cases. Sequences (defined as a pattern of developmental anomalies consequent upon a primary defect, often with heterogeneous etiology) were presented in 37 cases (9.6%). The most common sequence was oligohydramnios sequence (Potter syndrome) in 17 cases. Amnion rupture sequence (amniotic bands) was diagnosed in six cases and early urethral obstruction sequence in also six cases; laterality sequences with left isomerism (polysplenia syndrome) and right isomerism (Ivemark syndrome) were identified in two cases. Other sequences were identified in six cases. Twenty cases (5.2%) had a recognized nonmonogenic nonchromosomal syndrome. In all of these cases, no specific etiologic diagnosis could be found; however the observation of the fetus by the geneticist and the autopsy added essential information for the genetic counseling. Sixteen cases (4%) had the prenatal diagnosis of an isolated congenital heart disease. The postmortem studies did not change the severity of these anomalies but were able to better characterize it in 11 cases.

Twenty cases (5.2%) had a recognized nonmonogenic nonchromosomal syndrome. In all of these cases, no specific etiologic diagnosis could be found; however the observation of the fetus by the geneticist and the autopsy added essential information for the genetic counseling. Sixteen cases (4%) had the prenatal diagnosis of an isolated congenital heart disease. The postmortem studies did not change the severity of these anomalies but were able to better characterize it in 11 cases. In five cases of nonimmune hydrops fetalis no etiologic diagnosis was established, even after assessment of structural abnormalities of fetal heart, chromosome abnormalities, hemoglobinopathies, infections, and metabolic diseases (such as mucolipidosis I and II, galactosialidosis, mucopolysaccharidosis type I, IV, and VII, Gaucher disease type 2, gangliosidosis, Niemann-Pick disease type IA and C, multiple sulfatase deficiency, and sialic acid storage disease). Infections were the cause of fetal abnormalities in five cases, cytomegalovirus in four cases, and parvovirus B19 in one case, as fetal hydrops. Teratogens were identified in five cases. The use of misoprostol was the most common cause, found in 4 cases, and there was one warfarin embryopathy. Other diagnoses were identified in 24 cases, such as acardiac fetus, umbilical cord abnormalities, placental abnormalities, cervical lymphangioma, severe decreased/absent fetal movement, and structural anomalies (bilateral micropthalmia, phocomelia of upper limbs, and spinal anomalies). The anomalies are summarized in Table 1.

Other diagnoses were identified in 24 cases, such as acardiac fetus, umbilical cord abnormalities, placental abnormalities, cervical lymphangioma, severe decreased/absent fetal movement, and structural anomalies (bilateral micropthalmia, phocomelia of upper limbs, and spinal anomalies). The anomalies are summarized in Table 1. We have compared all the results from the obstetric, genetic, and fetal pathology evaluations for the presence or absence of agreement. Total agreement on the three areas was found in 165 cases (42%). The central nervous system anomalies were the main group with total agreement—70% (54/77 cases)—specifically the neural tube defects 81% (48/59 cases). Concordance in oligohydramnios sequence was 35% (6/17 cases). Prenatal diagnosis of familial syndromes had a total agreement in 84% (11/13 cases) of the cases. On the chromosome abnormality group we had a total of concordance of only 37%. All data were concordant in 38% of the prenatal diagnoses for advanced maternal age (29/77 cases). Total concordance between obstetric data and genetic observation and/or laboratory analysis was present in 111 cases (29%). Total concordance between genetic findings and autopsy results was present in 116 cases (30%). In 100 cases (26%) there was total agreement between obstetric data and fetal pathology evaluation.

We have compared all the results from the obstetric, genetic, and fetal pathology evaluations for the presence or absence of agreement. Total agreement on the three areas was found in 165 cases (42%). The central nervous system anomalies were the main group with total agreement—70% (54/77 cases)—specifically the neural tube defects 81% (48/59 cases). Concordance in oligohydramnios sequence was 35% (6/17 cases). Prenatal diagnosis of familial syndromes had a total agreement in 84% (11/13 cases) of the cases. On the chromosome abnormality group we had a total of concordance of only 37%. All data were concordant in 38% of the prenatal diagnoses for advanced maternal age (29/77 cases). Total concordance between obstetric data and genetic observation and/or laboratory analysis was present in 111 cases (29%). Total concordance between genetic findings and autopsy results was present in 116 cases (30%). In 100 cases (26%) there was total agreement between obstetric data and fetal pathology evaluation. Etiological diagnoses were changed or established after the genetic and/or autopsy observations in 80 cases (21%). After the genetic assessment (laboratorial results and fetal observation) the etiological diagnosis was changed in 49% (39/80 cases) of the cases. The postmortem results contributed to a modification of the diagnosis in 31% (25/80 cases) of the cases, and the identification of fetal infection in amniotic fluid changed the diagnosis in 6% (5/80 cases). The association between the autopsy findings and the data from the genetic evaluation allowed to identify the etiological diagnosis or to change it in 14% of the cases (11/80 cases).

in 31% (25/80 cases) of the cases, and the identification of fetal infection in amniotic fluid changed the diagnosis in 6% (5/80 cases). The association between the autopsy findings and the data from the genetic evaluation allowed to identify the etiological diagnosis or to change it in 14% of the cases (11/80 cases). There was only one case in which the prenatal prognosis was changed: in a twin pregnancy the diagnosis of one fetus with structural abnormalities and the other with hydrops being not confirmed in the latter. 4. Discussion In the last years, several studies tried to correlate the prenatal diagnosis with the results provided by fetopathogical and genetics evaluation and to compare the impact of the new information to final diagnosis, prognosis, and genetic counseling. In our study, chromosome abnormalities were the most common diagnosis, representing 39% of cases. Advanced maternal age was the major reason of its realization (52%). Central nervous system anomalies (20%) and genetic syndromes (11%) were the most frequent diagnosis after chromosome abnormalities. Nonmonogenic nonchromosomal polimalformative syndromes without specific etiology were presented in 5.2% of cases. In this group the genetic and autopsy evaluations provided data that improved significantly the genetic counseling. Isolated congenital heart disease was found only in 4% and congenital infections in 1% of the cases.

ic nonchromosomal polimalformative syndromes without specific etiology were presented in 5.2% of cases. In this group the genetic and autopsy evaluations provided data that improved significantly the genetic counseling. Isolated congenital heart disease was found only in 4% and congenital infections in 1% of the cases. Vaknin et al. found that chromosome abnormalities were the most common cause of TOP in 32% of the cases, followed by central nervous system in 27%, genetic syndromes in 5%, and (22 cases) fetal infection of the cases in 8% [1]. Amini et al. referred to 13.7% of chromosome abnormalities, 34.8% of central nervous system anomalies, and 3.7% of isolated congenital heart disease [13]. Boyd et al. found an abnormal karyotype in 46%, central nervous system abnormalities in 21%, monogenic disease in 8%, and isolated heart defect in 3% of the cases [7]. These studies and ours allow us to suggest a fairly constant prevalence of central nervous system anomalies and isolated congenital heart disease. There is a significant difference between the prevalence of genetic syndromes; in our opinion this value is dependent on the extent of the etiological investigation performed. There is a wide difference in the prevalence of chromosome abnormalities, which we relate to differences in the design of each study. Genetic evaluation, laboratory tests, and fetus observation by a clinical geneticist are a reality in our department and proved to bring out valuable information. This was shown mainly in precise the diagnosis in many situations (49% of the cases in our study).

These studies and ours allow us to suggest a fairly constant prevalence of central nervous system anomalies and isolated congenital heart disease. There is a significant difference between the prevalence of genetic syndromes; in our opinion this value is dependent on the extent of the etiological investigation performed. There is a wide difference in the prevalence of chromosome abnormalities, which we relate to differences in the design of each study. Genetic evaluation, laboratory tests, and fetus observation by a clinical geneticist are a reality in our department and proved to bring out valuable information. This was shown mainly in precise the diagnosis in many situations (49% of the cases in our study). Fetal postmortem studies were not always performed in other series. Boyd et al. showed that autopsy rates fell from 84% to 67% in 2000 [7]. Vaknin et al. had only 11% (52/462 cases) of cases submitted to fetopathological studies [1], and in Hungary study performed by Kaiser et al. 60% of cases had autopsy studies [12].

Fetal postmortem studies were not always performed in other series. Boyd et al. showed that autopsy rates fell from 84% to 67% in 2000 [7]. Vaknin et al. had only 11% (52/462 cases) of cases submitted to fetopathological studies [1], and in Hungary study performed by Kaiser et al. 60% of cases had autopsy studies [12]. In our study, the importance of fetal pathology can be seen when we analyze its weight in the alteration of diagnosis by itself (31%) and in addition with the genetic evaluation (14%). So the collaboration of a clinical geneticist and a fetal pathologist in a prenatal diagnosis team in all tertiary centers is essential. Their participation improved the diagnostic skills in 45% of cases, in our department. This can be also supported by the study of Phadke and Gupta They found in their study that support from dysmorphologists and geneticists in fetal evaluation changed or improved the diagnosis in 33% of cases (13) and that is plays a crucial role in specific etiologic diagnosis and genetic counseling [11]. Full agreement between all data (ultrasound, genetics, and postmortem results) was obtained in 42% of cases in contrast with other studies. Phadke and Gupta referred to 72.5% of total concordance, Ramalho et al. 61.1%, Kaasen et al. 40%, Kaiser et al. 49%, and Boyd et al. 55% [4, 7, 8, 11, 12].

In our study, the importance of fetal pathology can be seen when we analyze its weight in the alteration of diagnosis by itself (31%) and in addition with the genetic evaluation (14%). So the collaboration of a clinical geneticist and a fetal pathologist in a prenatal diagnosis team in all tertiary centers is essential. Their participation improved the diagnostic skills in 45% of cases, in our department. This can be also supported by the study of Phadke and Gupta They found in their study that support from dysmorphologists and geneticists in fetal evaluation changed or improved the diagnosis in 33% of cases (13) and that is plays a crucial role in specific etiologic diagnosis and genetic counseling [11]. Full agreement between all data (ultrasound, genetics, and postmortem results) was obtained in 42% of cases in contrast with other studies. Phadke and Gupta referred to 72.5% of total concordance, Ramalho et al. 61.1%, Kaasen et al. 40%, Kaiser et al. 49%, and Boyd et al. 55% [4, 7, 8, 11, 12]. This discrepancy in the data can be attributed to the methodology used to comparison. In our study we considered all relevant data obtained by observation of the fetus by geneticist, in addition to the results of fetopathology, as able to change or improve the etiologic diagnosis. In these studies, except in that of Phadke et al., routine genetic evaluation isolated was not referred to as a source of information to diagnosis.

red all relevant data obtained by observation of the fetus by geneticist, in addition to the results of fetopathology, as able to change or improve the etiologic diagnosis. In these studies, except in that of Phadke et al., routine genetic evaluation isolated was not referred to as a source of information to diagnosis. In only one situation the data changed the prognosis. Kaiser et al. referred to 2.4% of cases, where indication to TOP could not be supported after fetopathological results, Sankar and Phadke had 2 cases with total disagreement in 206 cases analyzed, and Phadke and Gupta also had 2 cases in 91examineted [2, 11, 12]. The data are summarized in Table 2. The diagnostic accuracy was improved in 80 cases (21%) which corroborate the need of a multidisciplinary team in prenatal centers, as already noted by other authors [2, 3, 6, 11–13]. In conclusion, our study in concordance with others showed that when fetal anomalies have been detected by ultrasound, the specific diagnosis can be made or refined with association of genetic evaluation and autopsy studies. Altogether they affect favorably the genetic counseling and represent an important aspect to control the quality of information given to a couple previously to a TOP. Table 1 Anomalies found in 385 cases after TOP. Anomalies Number of cases Chromosome abnormalities 149 Numerical 122 Trisomy 13 16 Trisomy 18 21 Trisomy 21 67 Monosomy X 12 Other 6 Structural unbalanced 22 Euploidies 5 Nervous system anomalies 77 Neural tube defects 59 Other 18

In conclusion, our study in concordance with others showed that when fetal anomalies have been detected by ultrasound, the specific diagnosis can be made or refined with association of genetic evaluation and autopsy studies. Altogether they affect favorably the genetic counseling and represent an important aspect to control the quality of information given to a couple previously to a TOP. Table 1 Anomalies found in 385 cases after TOP. Anomalies Number of cases Chromosome abnormalities 149 Numerical 122 Trisomy 13 16 Trisomy 18 21 Trisomy 21 67 Monosomy X 12 Other 6 Structural unbalanced 22 Euploidies 5 Nervous system anomalies 77 Neural tube defects 59 Other 18 Monogenic disorders 44 Familial syndromes 13 Hemophilia A 3 X- fragile syndrome 2 Muscular spinal atrophy 1 Beta-thalassemia 1 Sickle cell anemia 1 Mucopolysaccharidosis type I 1 Gangliosidosis type I 1 Metachromatic leucodystrophy 1 Walker-Warburg syndrome 1 Familial amyloidotic polyneuropathy 1 Diagnosis of index case 31 Frontonasal dysplasia 3 Fryns syndrome 3 Meckel-Gruber syndrome 2 Multiple pterygium syndrome 2 Roberts syndrome 2 Bardet-Biedl syndrome 1 Thanatophoric dysplasia 1 Diastrophic dysplasia 1 Apert syndrome 1 Osteogenesis imperfecta type IIA 1 Tetraamelia with pulmonary agenesis 1 Bartsocas-Papas syndrome 1 Mucopolysaccharidosis type VII 1 Pfeiffer syndrome 1 Proteus syndrome 1 Atelosteogenesis type I 1 X-linked hydrocephalus 1 Solitary median maxillary central incisor syndrome 1 Other 6 Sequences 37 Oligohydramnios 17 Amnion rupture 6 Early urethral obstruction 6 Laterality sequences 2 Other 6

Diagnosis of index case 31 Frontonasal dysplasia 3 Fryns syndrome 3 Meckel-Gruber syndrome 2 Multiple pterygium syndrome 2 Roberts syndrome 2 Bardet-Biedl syndrome 1 Thanatophoric dysplasia 1 Diastrophic dysplasia 1 Apert syndrome 1 Osteogenesis imperfecta type IIA 1 Tetraamelia with pulmonary agenesis 1 Bartsocas-Papas syndrome 1 Mucopolysaccharidosis type VII 1 Pfeiffer syndrome 1 Proteus syndrome 1 Atelosteogenesis type I 1 X-linked hydrocephalus 1 Solitary median maxillary central incisor syndrome 1 Other 6 Sequences 37 Oligohydramnios 17 Amnion rupture 6 Early urethral obstruction 6 Laterality sequences 2 Other 6 Polymalformative syndrome 20 Isolated congenital heart disease 16 Nonimmune hydrops 5 Infections 5 CMV 4 Parvovirus B19 1 Teratogen 5 Misoprostol 4 Warfarin 1 Other diagnoses 24 Table 2 Comparison of previous studies regarding total agreement and disagreement. N° of cases Total agreement Total disagreement Phadke and Gupta [11] 91 72.5% 2,2% (2/91) Ramalho et al. [8] 76 61.1% 0 Kaasen et al. [4] 228 58.4% — Kaiser et al. [12] 121 49% 2.4% (3/121) Boyd et al. [7] 309 55% — Sankar and Phadke [2] 206 59% 1% (2/206) Our study 385 42% 0.2% (1/385)

1. Introduction Antenatal care is believed to have a positive impact on pregnancy outcomes, either through early diagnosis and intervention for complications, or by contributing to the elimination and reduction of modifiable maternal risk factors. The recommended antenatal care programme in less developed countries is often the same as the programmes used in developed countries [1]. However, across the world there is wide variation in the proportion of women who receive antenatal care. Women differ in the access they have to antenatal healthcare, the perceived need for it, and the use they make of it [2]. Most women in the United Kingdom (UK) are aware that there are many health risks to themselves and the unborn child, and that there are many tests that may detect these abnormalities. For these reasons less than 1% of British pregnant women have no antenatal tests [3]. Most women make first contact with dedicated antenatal healthcare services between the 11th and 13th weeks, and NICE guidelines recommend seven–ten appointments [4]. The guidelines for South Africa are for a minimum of one visit in each trimester [5].

less than 1% of British pregnant women have no antenatal tests [3]. Most women make first contact with dedicated antenatal healthcare services between the 11th and 13th weeks, and NICE guidelines recommend seven–ten appointments [4]. The guidelines for South Africa are for a minimum of one visit in each trimester [5]. In contrast to the UK where comprehensive, free antenatal care began in 1948, South Africa only began offering universal antenatal care in 1994 [6]. Despite the widespread availability of antenatal healthcare most women in rural South Africa attend their first antenatal clinic in late pregnancy [7] at an average of up to 25 weeks [8]. Inadequate antenatal care, due to late booking and failure to follow up, contributes to large numbers of avoidable perinatal deaths and maternal complications, and is one of the avoidable factors underlying maternal mortality [8, 9]. Lack of easy physical access to healthcare and difficulty in recognizing pregnancy as well as educational factors have been identified as reasons for late use of antenatal healthcare by Hamilton et al. [10]. As the reasons for these differences in healthcare attendance are poorly understood, we developed an interview based on a structured questionnaire to measure antenatal healthcare use and identify important differences in knowledge and perceptions of antenatal healthcare that may account for differences in use. (See Questionnaire in Supplementary Material available online at doi: 10.5402/2011/36243.)

d, we developed an interview based on a structured questionnaire to measure antenatal healthcare use and identify important differences in knowledge and perceptions of antenatal healthcare that may account for differences in use. (See Questionnaire in Supplementary Material available online at doi: 10.5402/2011/36243.) 2. Material and Methods We designed a comparative cross-sectional pilot study of postnatal women in two populations; one at Birmingham women's hospital in the UK and the other in Pretoria academic hospital (now renamed Steve Biko academic hospital) in South Africa. Birmingham women's hospital is a state-run hospital for women in the West Midlands region of the UK primarily providing antenatal care for south Birmingham. Pretoria academic hospital is a state-run teaching hospital in the Tshwane district of Pretoria. The Pretoria data was collected 01–11/05/2007 and the Birmingham data was collected 25–29/06/2007. Thirty-one patients were interviewed at each location, this being the maximum number that could be interviewed in the 10-day period in Pretoria. Three women from Birmingham and five from Pretoria declined interview.

oria. The Pretoria data was collected 01–11/05/2007 and the Birmingham data was collected 25–29/06/2007. Thirty-one patients were interviewed at each location, this being the maximum number that could be interviewed in the 10-day period in Pretoria. Three women from Birmingham and five from Pretoria declined interview. Interviews were conducted on women whom had given birth to healthy babies within the last 24 hours (both normal vaginal delivery and caesarean section). Sampling was systematic where all patients who fell within the target population in the maternity wards on consecutive days were interviewed. Following approval by the south Birmingham ethics committee and the ethics committee of Pretoria academic hospital and trust approval and indemnity from Birmingham women's hospital, structured questionnaires were completed as bedside interviews, by the main author, next to mother and baby. Information was collected on sociodemographic details; timing and use of antenatal healthcare; reasons for and barriers to the use of antenatal healthcare. Data was a mixture of categorical sociodemographic data, quantitative variables on antenatal healthcare, use and qualitative comments. Nurses acted as interpreters on the ward for women who could not speak English. All variables were analysed using Mann-Whitney tests in Minitab 15.

s to the use of antenatal healthcare. Data was a mixture of categorical sociodemographic data, quantitative variables on antenatal healthcare, use and qualitative comments. Nurses acted as interpreters on the ward for women who could not speak English. All variables were analysed using Mann-Whitney tests in Minitab 15. 3. Results 3.1. Response and Sociodemographics Thirty-six patients from Pretoria and thirty-four patients from Birmingham were approached for interview of which and five and three declined, respectively. A total of thirty-one patients were interviewed at each location. In Birmingham, the interview population was 64% Caucasian, 23% Asian, 10% Afro-Caribbean, and 3% “other” with 2 Asian and 1 Caucasian woman declining interview. Of those interviewed in Pretoria 100% were black South Africans. Two white South Africans, two black South Africans, and one Asian woman declined interview. In both populations all patients gave an urban address. The Birmingham population had a median monthly income of £1500 (range £200–£15,000) whilst the Pretoria median monthly income was 1200 Rand (~£920) (range 140–2600 Rand) (P = .0001). The employment rate was 58% versus 39% for Birmingham and Pretorian women, respectively (P = .1325).

all patients gave an urban address. The Birmingham population had a median monthly income of £1500 (range £200–£15,000) whilst the Pretoria median monthly income was 1200 Rand (~£920) (range 140–2600 Rand) (P = .0001). The employment rate was 58% versus 39% for Birmingham and Pretorian women, respectively (P = .1325). Women from Birmingham had their first pregnancy at a median of 24 years (range 17–40) and women from Pretoria at a median of 21 years (range 15–28). Birmingham women were significantly older at first pregnancy (P = .0027), however there was no significant difference in terms of age at interview or in number of pregnancies or live births. Women from Birmingham received a higher level of education than those in Pretoria (Figure 1). 3.2. Healthcare Use and Awareness Women in Birmingham attended antenatal clinics earlier (P < .0001) in pregnancy, median 6 weeks after conception (range 0–17), than those from Pretoria, median 16 weeks (range 4–27). Women in Birmingham also had more frequent visits (P < .0001) than those from Pretoria: median 11 visits (range 5–30) during pregnancy in Birmingham; median 5 visits (range 2–18) in Pretoria. (Figures 2(a) and 2(b)). The two populations differ markedly in their knowledge of the problems that may occur during pregnancy. Women from Birmingham could name more conditions that may affect pregnancy than those from Pretoria with a median of 6 reasons (range 1–10) versus 3 reasons (range 0–8), respectively (P ≤ .0001). Women from Pretoria were more aware of HIV as a problem (Figure 3).

owledge of the problems that may occur during pregnancy. Women from Birmingham could name more conditions that may affect pregnancy than those from Pretoria with a median of 6 reasons (range 1–10) versus 3 reasons (range 0–8), respectively (P ≤ .0001). Women from Pretoria were more aware of HIV as a problem (Figure 3). 3.3. Reason for Seeking and Barriers to Healthcare Women from both regions stated that checking the health of the baby and their own health were the two most important reasons for seeking antenatal healthcare (Table 1). Some women from Pretoria felt that discovering their HIV status was important (9.7%), whereas none from Birmingham felt likewise. Indicating that South African women are more enlightened about the HIV infection and may seek antenatal care to check their HIV status. Furthermore, women from Birmingham placed much more emphasis than women from Pretoria on “advice” (35.5% versus 6.5%) and “reassurance” (16.1% versus 3.2%) as a function of antenatal healthcare. In both populations the majority of women had no problem with accessing antenatal healthcare (Figure 4). In the Birmingham population the main problem was bringing other siblings to the clinic 4/31 (13%). Women from Pretoria complained of problems with distance or cost of travel 5/31 (16%), and that there were too many people at a clinic 4/31 (13%). None of the Birmingham women stated these latter issues as problems.

4). In the Birmingham population the main problem was bringing other siblings to the clinic 4/31 (13%). Women from Pretoria complained of problems with distance or cost of travel 5/31 (16%), and that there were too many people at a clinic 4/31 (13%). None of the Birmingham women stated these latter issues as problems. 4. Discussion Overall women from Birmingham attended earlier and went more often to antenatal healthcare than did women from Pretoria (Figure 2). These findings are unlikely to be a type 1 error as the sample size of this study is sufficient to provide 80% power at the 5% significance level for a comparison of the two groups using a Mann-Whitney test when the true difference in means is equal to 0.74 times the within-group standard deviation. Birmingham women had a significantly higher income, nonsignificantly higher rate of employment and had a higher level of education (Figure 1). Late attendance has been associated with educational factors by previous research [9]. Lower general education levels in women from Pretoria may explain their later use of healthcare, because they are less aware of the complications of pregnancy. This is supported by the finding that women from Pretoria could name fewer medical problems that affect pregnancy than Birmingham women. Poorer education leading to decreased knowledge of medical problems affecting pregnancy (Figure 3) may be a significant reason for poorer use of healthcare services in pregnancy in women from Pretoria.

by the finding that women from Pretoria could name fewer medical problems that affect pregnancy than Birmingham women. Poorer education leading to decreased knowledge of medical problems affecting pregnancy (Figure 3) may be a significant reason for poorer use of healthcare services in pregnancy in women from Pretoria. Similar numbers of women from Birmingham and Pretoria stated they had no problem with accessing antenatal healthcare (Figure 4). The main problem for Birmingham women was bringing another sibling, though this was more of an inconvenience than a problem blocking access to antenatal healthcare, for example, one patient said: “It is difficult to get to clinics when you have to bring a child that wants to be anywhere but there” Patient 23—Birmingham interviewee. The barriers of cost or distance to travel and clinic overcrowding (Figure 4) may help to explain why women from Pretoria access antenatal care later and make fewer visits but also demonstrates the very different problems that women from the two populations face. Provision of free transport or clinics closer to where the women live in Pretoria and child entertainment facilities in Birmingham may help tackle these problems.

n from Pretoria access antenatal care later and make fewer visits but also demonstrates the very different problems that women from the two populations face. Provision of free transport or clinics closer to where the women live in Pretoria and child entertainment facilities in Birmingham may help tackle these problems. Women from Birmingham stated “advice” and “reassurance” as reasons to attend antenatal healthcare appointments. If women from Birmingham stated these reasons because they gain more advice and reassurance and feel they derive benefit from appointments in this respect, they may be more willing to attend clinics than those from Pretoria. Therefore focussing antenatal healthcare in South Africa on provision of pregnancy advice and reassurance may increase antenatal healthcare use. However the ability to provide increased antenatal visits in Pretoria may be a limiting factor.

s respect, they may be more willing to attend clinics than those from Pretoria. Therefore focussing antenatal healthcare in South Africa on provision of pregnancy advice and reassurance may increase antenatal healthcare use. However the ability to provide increased antenatal visits in Pretoria may be a limiting factor. HIV awareness is higher in women from Pretoria (Figure 3). The HIV infection rate in 2005 in the UK was 0.2% in adults aged 15–49 and 18.8% in South Africa [11]. In addition, in Johannesburg Hospital, which is less than 50 km from Pretoria the HIV prevalence is 29.4% [12]. This difference in infection rate may explain the differing concern about HIV during pregnancy, as women in South Africa are more likely to have personal or family experience of the problems of HIV/AIDS. This difference in concern about HIV exists despite both centres offering routine HIV screening and education [12, 13]. Although 21/31 (67.8%) women from Pretoria stated that HIV was a condition that could affect pregnancy, only 3/31 (9.7%) said that finding out their HIV status was an important function of antenatal healthcare. Indeed this trend is also seen in women from Birmingham where 5/31 (16.1%) stated HIV was a condition that could affect pregnancy but 0/31 (0%) felt that finding out their HIV status was a role of antenatal healthcare. This suggests that although women may be aware of the effect of HIV on pregnancy they may not see it as a risk that they identify with. The differences in perception of HIV/AIDS risk between the two regions, due partly to the difference in infection rates, but potentially also due to differences in local education policies may explain the differences in HIV/AIDS awareness.

ffect of HIV on pregnancy they may not see it as a risk that they identify with. The differences in perception of HIV/AIDS risk between the two regions, due partly to the difference in infection rates, but potentially also due to differences in local education policies may explain the differences in HIV/AIDS awareness. Research has shown that 97% of deliveries occur in NHS hospitals in the UK [2] and 83% of births are in health facilities in South Africa [8]. Indeed the study by Abrahams et al. [9] stated that the majority of South Africans give birth in healthcare facilities. Since most women in both populations give birth in health facilities, interviews carried out in the respective hospitals should be approximately representative of all births in the two populations. The Birmingham population has a diverse ethnic composition. In the UK the minority ethnic populations are concentrated in the large urban centres [14], thus the results from the Birmingham population may be representative of the general population of other large UK cities. The Pretoria population in this study is 100% black. A study by Sőderlund [15] showed that 60% of South Africans with an average per family member income of greater than R11,000 per annum (approx £800) use private healthcare. This study had six women with this income or above, indicating that there were approximately seven to nine patients missing from this income bracket due to the use of private healthcare. Thus the Pretoria results are at best representative of the black population who use public healthcare in large South African cities.

This study had six women with this income or above, indicating that there were approximately seven to nine patients missing from this income bracket due to the use of private healthcare. Thus the Pretoria results are at best representative of the black population who use public healthcare in large South African cities. All women in the Pretoria population had at least one visit to an antenatal clinic in this study. As a tertiary referral unit in South Africa, women must usually have attended one antenatal clinic in order to be referred to the Pretoria Academic hospital. Therefore these results may be an overestimation of the use of antenatal healthcare in the general Black population. This study showed that women use healthcare services earlier in pregnancy before the main antenatal screening tests began in the 10th–12th week [4]. This discrepancy occurs because this study recorded antenatal healthcare use as “any contact with a healthcare professional”, rather than the “first use of dedicated antenatal screening”. This study is thus concerned with antenatal healthcare not dedicated antenatal screening.

ing tests began in the 10th–12th week [4]. This discrepancy occurs because this study recorded antenatal healthcare use as “any contact with a healthcare professional”, rather than the “first use of dedicated antenatal screening”. This study is thus concerned with antenatal healthcare not dedicated antenatal screening. Six Birmingham women stated that they sought appointments initially to book antenatal healthcare screening, thus they were attending their GP before the start of dedicated antenatal screening. The Pretoria population first used healthcare services at a median of 16 weeks into pregnancy for the first appointment. These results were similar to the Abrahams et al. [9] study carried out in Cape Town. This indicates that urban Black populations in South Africa attend antenatal healthcare services later than urban populations in the UK. When asked why women from Pretoria booked at this time, none stated that they did so to book further healthcare and only 8/31 did so to confirm their pregnancy. This indicates that women from Pretoria found it less important to book antenatal healthcare. This lower priority for antenatal healthcare may explain their later attendance to these services.

a booked at this time, none stated that they did so to book further healthcare and only 8/31 did so to confirm their pregnancy. This indicates that women from Pretoria found it less important to book antenatal healthcare. This lower priority for antenatal healthcare may explain their later attendance to these services. Previous research addressing outcomes investigated in this study have indicated that reduction in number of antenatal healthcare visits is not always associated with decreased healthcare effectiveness [16]. Therefore, although this study showed a reduced number of antenatal healthcare visits by women from Pretoria, this cannot be taken as evidence of a suboptimal antenatal healthcare system in Pretoria. 5. Conclusions Women from Birmingham use healthcare services earlier (P ≤ .0001) and more often during pregnancy (P ≤ .0001). Women from Birmingham identified more medical conditions that may affect pregnancy (median 6 versus 3 reasons). Women from Pretoria are more aware of HIV infection and may seek antenatal care to check their HIV status. Women from Birmingham perceived antenatal healthcare as relatively more important for advice and reassurance about pregnancy, whilst women from Pretoria had more problems with cost of transport and travel distance and clinic overcrowding.

ore aware of HIV infection and may seek antenatal care to check their HIV status. Women from Birmingham perceived antenatal healthcare as relatively more important for advice and reassurance about pregnancy, whilst women from Pretoria had more problems with cost of transport and travel distance and clinic overcrowding. This study has identified a number of important differences between perceptions of antenatal healthcare in Birmingham and Pretoria, and this may help explain decreased antenatal healthcare uptake in Pretoria. Addressing lack of knowledge about medical problems in pregnancy and emphasising the importance of antenatal healthcare may increase antenatal healthcare use. In addition emphasising the aspect of advice and reassurance at antenatal healthcare clinics may help increase attendance. Problems of cost and distance of travel to clinic could be tackled by improved transport links whilst overcrowding in clinic would inevitably require increased medical staffing in Pretoria. A future study including perinatal or maternal morbidity and mortality would be needed to assess the effectiveness of the antenatal healthcare programs in the two locations and elucidate if reduced antenatal attendance is linked with poorer outcomes. A larger more geographically diverse population would be needed to find significance in these parameters and to confirm the findings of this study.

e needed to assess the effectiveness of the antenatal healthcare programs in the two locations and elucidate if reduced antenatal attendance is linked with poorer outcomes. A larger more geographically diverse population would be needed to find significance in these parameters and to confirm the findings of this study. Supplementary Material The supplementary data includes the finalised structured questionnaire used in the patient interviews, the patient consent form, ethical approval from the ‘South Birmingham Research Ethics Committee' and ‘University of Pretoria' and the Birmingham Women's Hospital health care trust approval and indemnity letter. Click here for additional data file. Acknowledgments The authors would like to thank the cooperation of the ward staff and nurses at Pretoria Academic Hospital (now Steve Bilko Academic Hospital) and Birmingham Women's Hospital for access to the wards and interpretation services. They would like to thank the Children's Research Fund, UK for contributing to the cost of this research project. Conflict of Interests Dr. M. R. Openshaw received less than $1000 from the Children's Research fund. No other conflict of interest is reported. The authors alone are responsible for the content and writing of the paper. Figure 1 Highest level of education achieved by women from Pretoria and Birmingham. Figure 2 Scatter-plots showing the time of first visit to a healthcare professional (a) and number of visits made (b) by pregnant women in Pretoria and Birmingham. Figure 3 Conditions identified by women that may affect pregnancy.

Conflict of Interests Dr. M. R. Openshaw received less than $1000 from the Children's Research fund. No other conflict of interest is reported. The authors alone are responsible for the content and writing of the paper. Figure 1 Highest level of education achieved by women from Pretoria and Birmingham. Figure 2 Scatter-plots showing the time of first visit to a healthcare professional (a) and number of visits made (b) by pregnant women in Pretoria and Birmingham. Figure 3 Conditions identified by women that may affect pregnancy. Figure 4 Barriers to antenatal healthcare identified by women from Pretoria (a) and Birmingham (b). Table 1 Reasons for women seeking antenatal healthcare. Birmingham Pretoria n (%) n (%) Check baby health 26 83.9 26 83.9 Check own health 15 48.4 14 45.2 Advice 11 35.5 2 6.5 HIV status check 0 0 3 9.7 Felt compelled to go 3 9.7 1 3.2 Confirm pregnancy 1 3.2 2 6.5 Reassurance 5 16.1 1 3.2 Check baby orientation 0 0 3 9.7 Meet people & share experiences 2 6.5 0 0

1. Introduction Cervical cancer remains the second most common cancer in women worldwide and the first in most developing countries, where 80% of cases occur. Cervical cancer is developed through precancerous stages, termed cervical intraepithelial neoplasia CIN, and caused by infection with human papillomavirus (HPV) [1]. Epidemiological studies have clearly demonstrated that in countries that achieve frequent screening, the incidence and the mortality rates of cervical cancer have decreased considerably over the last four decades [2–4]. Screening with the use of PAP test prevents cancer by detecting precancerous lesions (CIN), and early treatment of precancerous lesions can prevent progression to cancer [5, 6]. Conventional cytology allows the detection of abnormal findings leading to a colposcopy with biopsy for a definitive diagnosis if there is persistence of atypical squamous cells with undetermined significance or low-grade or high-grade squamous intraepithelial lesion (LSIL, HSIL). Generally, ASCUS corresponds to ill-defined abnormalities of superficial cells and represents 2% to 3% of all cervical smears. Additionally, biopsy shows CIN grades II-III in 5% to 10% of ASCUS cases. LSIL corresponds to mildly abnormal squamous cells and is found in 1% to 2% of all cervical smears. In most cases it regresses spontaneously, especially in young patients. HSIL corresponds moderately to severely abnormal squamous cells and represents 0.5% of all cervical smears [7].

II-III in 5% to 10% of ASCUS cases. LSIL corresponds to mildly abnormal squamous cells and is found in 1% to 2% of all cervical smears. In most cases it regresses spontaneously, especially in young patients. HSIL corresponds moderately to severely abnormal squamous cells and represents 0.5% of all cervical smears [7]. Management options for LSIL (CIN1 grade) on histology vary, ranging from simple observation to excisional therapies. Patients with persistent LSIL should be treated chiefly with the use of office-based ablative therapies. Management guidelines for HSIL (CIN2/3 grade) are well established and recommend colposcopy-directed biopsy. Cold-knife conization or electroconization should be performed in all patients with biopsy-confirmed HSIL in order to exclude invasive disease [1].

reated chiefly with the use of office-based ablative therapies. Management guidelines for HSIL (CIN2/3 grade) are well established and recommend colposcopy-directed biopsy. Cold-knife conization or electroconization should be performed in all patients with biopsy-confirmed HSIL in order to exclude invasive disease [1]. The annual incidence for CIN 1 and CIN 2, 3 in the US is 1.6 and 1.2 per 1,000 women, respectively. The incidence is highest among women aged from 21 to 30 years (3.3 and 3.6 per 1,000) and women aged from 31 to 40 years (2.9 and 2.7 per 1,000) [8]. In Spain, it is estimated that 7.6 million Pap smears are performed annually, with 3.5% of routine Pap smears to be abnormal and with 40,530 women to be annually diagnosed with CIN 1, 26,243 with CIN 2, and 28,423 with CIN 3 [9]. In France, Estimated 6,111,787 Pap smears were performed in 2004, including 222,350 abnormal (3.9%) and 63,616 followup smears. In total, 58,920 cervical biopsies and 52,525 HPV tests were performed after an abnormal Pap smear [10]. Estimated 6.4 million women aged between 25 and 69 years undergo screening annually in Italy (1.2 million and 5.2 million through organized and opportunistic screening programs, resp.). Approximately 2.4% of tests have positive findings, and there are approximately 21,000 cases of CIN1 and 7,000–17,000 cases of CIN2/3 [11]. In Greece, cervical scrapes from 841 women in 2006, which were obtained for cytological evaluation, showed that the normal Pap test results or benign cellular changes were 45,8% of the women, atypical squamous cells of undeterminated significance (ASCUS) were 23,2%, low-grade squamous intraepithelial lesion (LSIL) was 27,9%, and high-grade squamous intraepithelial lesion (HSIL) was 3,1% [12].

cytological evaluation, showed that the normal Pap test results or benign cellular changes were 45,8% of the women, atypical squamous cells of undeterminated significance (ASCUS) were 23,2%, low-grade squamous intraepithelial lesion (LSIL) was 27,9%, and high-grade squamous intraepithelial lesion (HSIL) was 3,1% [12]. This study aims to present the management of precancerous lesions of the uterine cervix, according to cytological and epidemiological aspects as well as the treatment methods adopted. 2. Methods The study population consisted of 238 women screened for a regular gynaecological control in the gynaecological outpatient clinic of two public primary health care centres in Greece (a general one where all patients are admitted and a hospital specialized on cancer cases). Registry was annual and lasted from January 2007 to January 2008. Possible factors associated with CIN were examined like women age, marital and educational status, and professional and insurance type. Other characteristics like smoking, HPV, the season of women's visit at the hospital were also considered. Statistical tests used for comparisons of factors were Fisher exact test and Pearson Chi-square in cases where the latter was not possible to be implemented. Twoindependent samples t-test and one-way ANOVA were applied for comparisons of continuous variables between groups. Quantitative variables are expressed as Mean  ±  SD, and P-values smaller than  .05 were considered statistically significant. Analysis was performed with the statistical software SPSS 17.

e implemented. Twoindependent samples t-test and one-way ANOVA were applied for comparisons of continuous variables between groups. Quantitative variables are expressed as Mean  ±  SD, and P-values smaller than  .05 were considered statistically significant. Analysis was performed with the statistical software SPSS 17. 3. Results Population age was 37.03  ±  10.47 years (Mean  ±  SD). Almost half of women (49.6%) had at least one child, and most of them had two children. As far as education is concerned, 55.9% completed high school, 42% higher education while the remaining 2.1% elementary. The marital status and occupation of women who participated in this study are presented in Figures 1 and 2, respectively.

f of women (49.6%) had at least one child, and most of them had two children. As far as education is concerned, 55.9% completed high school, 42% higher education while the remaining 2.1% elementary. The marital status and occupation of women who participated in this study are presented in Figures 1 and 2, respectively. Demographic features according to the stages of dysplasia and the type of medical treatment are described in Tables 1, 2, and 3. As shown, the majority of women were married, young (17–30 years old), and private employees, who had completed high school. The marital status of patients seems to affect the dysplasia grade (P = .02). Mostly unmarried women were diagnosed with CIN1 (49.0%), divorced women with CIN2 (12.2%), and married women and widows with the high-grade intraepithelial lesion CIN3, with rates 67.9% and 7.1%, respectively. A positive relation was observed when we compared the education status (P = .64), the occupation (P = .66), and the insurance (P = .99) with the grade of dysplasia of the studied woman, without reaching statistical significance. The majority of the women with higher education (57,7%) and public insurance (50.8%), occupied as private employees (44,2%), were diagnosed with higher grades of dysplasia, CIN3, and ASC/ASCUS.

.66), and the insurance (P = .99) with the grade of dysplasia of the studied woman, without reaching statistical significance. The majority of the women with higher education (57,7%) and public insurance (50.8%), occupied as private employees (44,2%), were diagnosed with higher grades of dysplasia, CIN3, and ASC/ASCUS. Moreover, there was a significant variation of the type of abnormalities according to the age group. Specifically, young women (17–30 years old) did not have hgsil, but atypical squamous cells (ASCs) or atypical squamous cells of undetermined significance (ASCUS) (35%). Older women (30–35 years) were detected with CIN1 and at older ages (36–50 years) with CIN3. Specifically there was a rate of 17.9% of 36–40 years old women with CIN detected, a rate of 25% at women of 41–45 years, and a rate of 28.6% at women of 46–50 years old. Middle aged ones (51–55 years old) had a reduced level of cervical intraepithelial neoplasia CIN2 (12.2%), and women over 55 years old did not have atypical squamous cells (ASCs) or atypical squamous cells ASC of undetermined significance (ASCUS) dysplasia (ASC/ASCUS) (4.2%). It should be mentioned that patients with ASC/ASC-US had HPV infection rate of 47 5%, patients with cervical intraepithelial neoplasia CIN1 had 49%, and finally patients with CIN2 and CIN3 had 48.8% and 42.9%, respectively, (data not shown).

s cells ASC of undetermined significance (ASCUS) dysplasia (ASC/ASCUS) (4.2%). It should be mentioned that patients with ASC/ASC-US had HPV infection rate of 47 5%, patients with cervical intraepithelial neoplasia CIN1 had 49%, and finally patients with CIN2 and CIN3 had 48.8% and 42.9%, respectively, (data not shown). Additionally, several other factors were considered in order to study possible correlations according to CIN. For that purpose, smoking status was examined, but with no statistically significant results. Twenty-two point one percent of women smokers developed cervical intraepithelial neoplasia CIN1, 17.1% CIN2, and CIN3 10.7%, compared to nonsmokers, while the number of daily consumption of cigarettes did not appear to be associated with dysplasia (data not shown). When the same analysis was performed for women with confirmed cancer, the majority of married women were treated with surgical procedures, such as LLETZ (83,3%), loop electrosurgical excisional procedure (54,8%), and diathermic conization electrosurgical excisional procedure (70.5%). A statistically significant correlation was established (P < .001) when we compared the management of the precancerous and cancerous lesions with marital status. A positive relation was observed when we compared the education status (P = .75), the occupation (P = .26), and the insurance (P = .07) with the management of cancerous lesions of the studied woman, without reaching statistical significance.

the management of the precancerous and cancerous lesions with marital status. A positive relation was observed when we compared the education status (P = .75), the occupation (P = .26), and the insurance (P = .07) with the management of cancerous lesions of the studied woman, without reaching statistical significance. 4. Discussion The development and implementation of population control can lead to prevention and early detection of precancerous lesions of the cervix. Cytological screening is the basic method in the early finding and screening of precancer and cancer of the cervix. The colposcopical method is widely used in the every-day practice and is mostly used after obtaining the abnormal Pap smear test or after positive result from HPV test. All these diagnostic procedures have to combine together with the purpose of better early diagnosis and adequate treatment of these disease.

he cervix. The colposcopical method is widely used in the every-day practice and is mostly used after obtaining the abnormal Pap smear test or after positive result from HPV test. All these diagnostic procedures have to combine together with the purpose of better early diagnosis and adequate treatment of these disease. Regarding the treatment method followed, we observed that the greater the degree of dysplasia of the cervix was, the higher the degree of treatment was, ranging from conservative management to surgical excision treatment. The largest proportion of patients treated conservatively with 6-month and annual retesting with Pap test and colposcopy was with simple informal ASC-ASCUS (89.17%). For CIN1, the percentages of patients treated conservatively with surgical resection of the cervix were at the same level, 46.94% and 53.07%, while for HGSIL (CIN2/3), the main treatment was surgical reconstruction with conical resection of the cervix. According to the guidelines referred to in other studies, the management of high-grade intraepithelial lesions (CIN2/3) is colposcopy, biopsy with or without intracervical curettage, and surgical treatment with cervical cone excision [13]. According to the marital status of our study population, dysplasia grade seems to be differentiated especially in married women, with the rate of 67.9% for the high-grade intraepithelial lesion. The risk of high-grade intraepithelial lesion is almost 2 times greater in married women than in single ones, who happen to be in higher median age, and the risk increases with time since the last normal smear or with lower frequency of screening. A study in China showed that, in Beijing, married women at the age of 30–34 are the high-risk group in CIN incidence compared to women in other marital status [14].

n in single ones, who happen to be in higher median age, and the risk increases with time since the last normal smear or with lower frequency of screening. A study in China showed that, in Beijing, married women at the age of 30–34 are the high-risk group in CIN incidence compared to women in other marital status [14]. The association between smoking status and the degree of dysplasia was not confirmed, in contrary to other studies, where smoking intensity is an independent risk factor for high-grade CIN in young women, after controlling the cervical HPV infection. Other papers also indicate that current smokers are as likely to be diagnosed twice with high-grade CIN as nonsmokers [15] and also that current cigarette smoking is a risk factor, closely interrelated in that the high-risk HPV types are significantly more frequent in current smokers than in nonsmokers [16]. Also, despite the fact that it cannot be concluded that smoking is a genuine cause of cervical neoplasia, results from other studies support the hypothesis that smoking is a true risk factor in cervical neoplasia [17–19].

he high-risk HPV types are significantly more frequent in current smokers than in nonsmokers [16]. Also, despite the fact that it cannot be concluded that smoking is a genuine cause of cervical neoplasia, results from other studies support the hypothesis that smoking is a true risk factor in cervical neoplasia [17–19]. Our study included women with positive test and cervical intraepithelial abnormalities, so results cannot be extrapolated to the total female population in Greece. However, since the outpatient clinics are referral ones, the results give us important information on the cytological and epidemiological aspects of precancerous lesions of the uterine cervix of Greek women and the therapy provided according to current medical practice. In addition, precancerous lesions of the cervix are treatable with minimally invasive surgical methods, so women in early stages of precancerous lesions or patients undergoing such treatment should be rechecked at regular intervals in order to prevent the recurrence or progression of the disease. Larger epidemiological studies in different regions of the country are needed in order to report on the epidemiology of precancerous lesions of the uterine cervix in Greece.

s or patients undergoing such treatment should be rechecked at regular intervals in order to prevent the recurrence or progression of the disease. Larger epidemiological studies in different regions of the country are needed in order to report on the epidemiology of precancerous lesions of the uterine cervix in Greece. 5. Conclusion In the absence of a national screening programme, the present clinical study is an effort of utmost importance for the early detection and prevention of cervical cancer. Additional effort is needed from the Greek Government to improve the information provided and the education of women on the prevention of cervical cancer and also to develop an organized national policy, by call and recall of all women for the regular diagnostic procedures. Acknowledgments The authors would like to thank Dr. Papadimitriou A., the director of Gynaecology Department in Saint Panteleimon General State Hospital Nikea, and Dr. Akrivos T., the director of Gynaecological Department in Piraeus Metaxa Memorial Cancer Hospital in Greece, for their valuable contribution in data collection of this study. Figure 1 The marital status of the study population. Figure 2 The occupation of the study population. Table 1 Patient demographics by hospital. Characteristics   Hospital General hospital (%) Cancer hospital (%) Marital status Single 49.5 50.3 Married 39.5 38.8 Widow-divorced 11.0 10.9 Education Illiterate 0.0 0.0 Elementary primary school 3.7 0.8 High school 63.3 49.6 University 33.0 49.6

Figure 2 The occupation of the study population. Table 1 Patient demographics by hospital. Characteristics   Hospital General hospital (%) Cancer hospital (%) Marital status Single 49.5 50.3 Married 39.5 38.8 Widow-divorced 11.0 10.9 Education Illiterate 0.0 0.0 Elementary primary school 3.7 0.8 High school 63.3 49.6 University 33.0 49.6 Occupation Civil servant 12.8 14.7 Private employee 46.8 39.5 Free professional 4.6 4.7 household 21.1 16.3 Other 14.7 24.8 Insurance fund Public sector 17.4 18.6 Social security institute 51.4 47.3 Insurance fund for craftsmen and small traders 13,8 15,6 Lawyers insurance fund 4.6 4.7 Mariners' pension fund 6.4 8.5 Agricultural insurance organization 6.4 5.4 Table 2 Patient demographics according to dysplasia grade. Characteristics CIN grade P-value* CIN1 CIN2 CIN3 ASC/ASCUS N % N % N % N % Marital status .02* Married 19 38.8 23 56.1 19 67.9 58 48.3 Single 24 49.0 11 26.8 5 17.9 53 44.2 Widow-divorced 6 12.2 7 17.1 4 14.2 9 7.5 Education .64* Illiterate 0 0.0 0 0.0 0 0.0 0 0.0 Elementary primary school 0 0.0 0 0.0 1 3.6 4 3.3 High school 25 51.0 22 53.7 17 60.7 69 57.5 University 24 49.0 19 46.3 10 35.7 47 39.2 Occupation .66** Civil servant 9 18.4 6 14.6 3 10.7 15 12.5 Private employee 20 40.8 18 43.9 11 39.3 53 44.2 Free professional 3 6.1 2 4.9 0 0.0 6 5.0 Household 4 8.2 9 22.0 6 21.4 25 20.8 Other 13 26.5 6 14.6 8 28.6 21 17.5

Education .64* Illiterate 0 0.0 0 0.0 0 0.0 0 0.0 Elementary primary school 0 0.0 0 0.0 1 3.6 4 3.3 High school 25 51.0 22 53.7 17 60.7 69 57.5 University 24 49.0 19 46.3 10 35.7 47 39.2 Occupation .66** Civil servant 9 18.4 6 14.6 3 10.7 15 12.5 Private employee 20 40.8 18 43.9 11 39.3 53 44.2 Free professional 3 6.1 2 4.9 0 0.0 6 5.0 Household 4 8.2 9 22.0 6 21.4 25 20.8 Other 13 26.5 6 14.6 8 28.6 21 17.5 Insurance Fund .99* Public sector 10 20.4 9 22.0 5 17.9 19 15.8 Social security institute 24 49.0 18 43.9 14 50.0 61 50.8 Insurance fund for craftsmen and  small traders 7 14,3 5 12,3 3 10.7 20 16.7 Lawyers insurance fund 3 6.1 2 4.9 1 3.6 5 4.2 Mariners' pension fund 3 6.1 4 9.8 2 7.1 9 7.5 (Hellenic) agricultural insurance  organization 2 4.1 3 7.3 3 10.7 6 5.0 Age (mean  ±  SD) 49 35  ±  11 41 39  ±  10 28 43  ±  8 120 35  ±  11 <.001*** *Fisher's exact test **Pearson Chi-square ***One-way ANOVA. Table 3 Demographics of patients by treatment path. Conservative treatment Cryotherapy Diathermic conization electrosurgical excisional procedure Loop electrosurgical excisional procedure LLETZ P-value N % N % N % N % N % Marital status <.001* Married 55 41.4 5 38.5 31 70.5 23 54.8 5 83.3 Single 64 48.1 7 53.8 7 15.9 14 33.3 1 16.7 Widow-divorced 14 10.5 1 7.7 6 13.6 5 11.9 0 0.0 Education .75* Illiterate 0 0.0 0 0.0 0 0.0 0 0.0 0 0.0 Elementary primary  school 3 2.3 0 0.0 1 2.3 1 2.4 0 0.0 High school 81 60.9 6 46.2 21 47.7 22 52.4 3 50.0 University 49 36.8 7 53.8 22 50.0 19 45.2 3 50.0

Conservative treatment Cryotherapy Diathermic conization electrosurgical excisional procedure Loop electrosurgical excisional procedure LLETZ P-value N % N % N % N % N % Marital status <.001* Married 55 41.4 5 38.5 31 70.5 23 54.8 5 83.3 Single 64 48.1 7 53.8 7 15.9 14 33.3 1 16.7 Widow-divorced 14 10.5 1 7.7 6 13.6 5 11.9 0 0.0 Education .75* Illiterate 0 0.0 0 0.0 0 0.0 0 0.0 0 0.0 Elementary primary  school 3 2.3 0 0.0 1 2.3 1 2.4 0 0.0 High school 81 60.9 6 46.2 21 47.7 22 52.4 3 50.0 University 49 36.8 7 53.8 22 50.0 19 45.2 3 50.0 Occupation .26** Civil servant 15 11.3 2 15.4 8 18.2 8 19.0 0 0.0 Private employee 54 40.6 9 69.2 18 40.9 20 47.6 1 16.7 Free professional 8 6.0 0 0.0 3 6.8 0 0.0 0 0.0 Household 24 18.0 2 15.4 9 20.5 6 14.3 3 50.0 Other 32 24.1 0 0.0 6 13.6 8 19.0 2 33.3 Insurance Fund .07** Public sector 19 14.3 2 15.4 11 25.0 11 26.2 0 0.0 Social security institute 63 47.4 10 76.9 19 43.2 23 54.8 2 33.3 Insurance fund for  craftsmen and small  traders 23 17.3 0 0.0 8 18,2 3 7.1 1 16.7 Lawyers insurance fund 8 6.0 1 7.7 1 2.3 0 0.0 1 16.7 Mariners' pension fund 12 9.0 0 0.0 4 9.1 2 4.8 0 0.0 (Hellenic) agricultural  insurance organization 8 6.0 0 0.0 1 2.3 3 7.1 2 33.3 Age (mean  ±  SD) 133 35  ±  11 13 34  ±  6 44 41  ±  10 42 39  ±  10 6 43  ±  10 <.001*** *Fisher's exact test **Pearson Chi-square ***One-way ANOVA.

1. Case History A 48-year-old lady, gravida 3, para 3, presented with right inguinal mass associated with menometrorrhagia for eight weeks. Previously, she had regular monthly periods and no dysmenorrhea. She had a history of period pain at the groin during menses. On examination, the right inguinal mass measured 4 × 5 cm and was tender and relatively fixed. The cervix was normal and the uterus was bulky, mobile, and nontender. Pelvic ultrasound was normal, apart from a slightly bulky uterus. Endometrial sampling revealed proliferative endometrium. There was no evidence of polyp, hyperplasia nor malignancy. Fine-needle aspiration (FNA) biopsy of the inguinal mass was performed and revealed endometriosis. In view of her vaginal bleeding, bulky uterus, and inguinal mass, the patient was advised to have a dilatation and curettage along with excision of the groin mass. A 6 × 5 cm solid, fibroid-like tumor was removed from the right groin (see Figures 1 and 2). Histopathology study of the tumor confirmed deposits of endometrial type gland and stroma. The uterine curettage revealed disordered proliferating endometrium.

e a dilatation and curettage along with excision of the groin mass. A 6 × 5 cm solid, fibroid-like tumor was removed from the right groin (see Figures 1 and 2). Histopathology study of the tumor confirmed deposits of endometrial type gland and stroma. The uterine curettage revealed disordered proliferating endometrium. 2. Comments Endometriosis is defined as the presence of endometrial glands and stroma outside the endometrial cavity and uterine musculature. Most common sites of endometriosis are within the pelvis, while unusual locations include bladder, intestine, surgical scars, diaphragm [1], umbilicus [2], and groin [3, 4]. Patients presenting with endometriosis of the inguinal region are few and far between. More specifically, the lesions can be found in the extraperitoneal portion of the round ligament [5], in the inguinal lymph nodes, in the subcutaneous adipose tissue, and even in the wall of sacs of inguinal or femoral hernias [6–9]. This paper aims to elucidate the incidence, pathogenesis, and management of this rare clinical problem.

, the lesions can be found in the extraperitoneal portion of the round ligament [5], in the inguinal lymph nodes, in the subcutaneous adipose tissue, and even in the wall of sacs of inguinal or femoral hernias [6–9]. This paper aims to elucidate the incidence, pathogenesis, and management of this rare clinical problem. Inguinal endometriosis was first reported by Allen in 1896, but since then only a little more than 30 cases have been described [3, 6, 10, 11]. Battista Candiani et al. [11] reported six cases of inguinal endometriosis and they noted the mean age at diagnosis was 31 years [11]. The size of the mass reported ranges from 1 to 6 cm in diameter [8–11]. Table 1 summarized the reported cases in the literature. About 90% of reported cases of extraperitoneal endometriosis occur in the right inguinal area [4, 6, 10, 11] as was noted on the patient in this case. Malignant transformation to carcinoma had been reported, with three cases of clear cell carcinoma documented [12]. Thus, it is of significance to remove the inguinal mass for pathology confirmation. The actual incidence of inguinal endometriosis is difficult to ascertain [11]. Due to the nonpelvic location of the lesion, patients would most often consult a general surgeon rather than a gynecologist, with a preoperative diagnosis of inguinal pathology. This could probably lead to underreporting of these cases.

Malignant transformation to carcinoma had been reported, with three cases of clear cell carcinoma documented [12]. Thus, it is of significance to remove the inguinal mass for pathology confirmation. The actual incidence of inguinal endometriosis is difficult to ascertain [11]. Due to the nonpelvic location of the lesion, patients would most often consult a general surgeon rather than a gynecologist, with a preoperative diagnosis of inguinal pathology. This could probably lead to underreporting of these cases. Battista Candiani et al. [11] describe six patients with inguinal endometriosis all whom have pelvic endometriosis on laparoscopy. They reported that 91% of inguinal endometriosis cases are associated with coexisting pelvic endometriosis [11]. Yet Inguinal endometriosis may be present without associated pelvic endometriosis [4]. Quagliarello et al. [8] reported a patient with inguinal endometriosis with no evidence of pelvic endometriosis on laparoscopy [8]. Laparoscopy was not performed on this patient because she was asymptomatic with no dysmenorrhoea or pelvic pain suggestive of the presence of pelvic pathology.

ed pelvic endometriosis [4]. Quagliarello et al. [8] reported a patient with inguinal endometriosis with no evidence of pelvic endometriosis on laparoscopy [8]. Laparoscopy was not performed on this patient because she was asymptomatic with no dysmenorrhoea or pelvic pain suggestive of the presence of pelvic pathology. Although the typical complaints of patients with pelvic endometriosis include dysmenorrhoea, menstrual irregularities, dyspareunia, and infertility, patients with extrapelvic endometriosis like inguinal endometriosis present with unusual symptoms and occasionally, a diagnostic dilemma [6]. Periodic menstrual pain at the mass is sometimes the primary complaint [3, 10, 11]. Patients often present with a painful lump in the groin and premenstrual tenderness and/or swelling [6, 9, 10]. A history of a previous gynecologic procedure or surgical trauma is also contributory [3, 6]. Some patients may have no history of dysmenorrhoea, pelvic pain, or dyspareunia [3] as in our patient. However, she experienced right inguinal pain during menses.

e groin and premenstrual tenderness and/or swelling [6, 9, 10]. A history of a previous gynecologic procedure or surgical trauma is also contributory [3, 6]. Some patients may have no history of dysmenorrhoea, pelvic pain, or dyspareunia [3] as in our patient. However, she experienced right inguinal pain during menses. On surgical exploration, a distinct firm mass was encountered [3]. Perez-Seoane et al. [7] described the gross findings of an inguinal endometrioma removed during surgery as multilobulated dark brown-colored tissue with hard consistency. Similar findings were earlier reported by Quagliarello et al. [8] in 1985. The histological diagnosis of endometriosis was based on the finding of characteristic endometrial glands with stromal cells within the fibrous connective tissue [3, 8, 9, 11] as was seen in this case. There was also chronic inflammatory cells [7] and hemosiderin-laden histiocytes [7, 11] found in some sections. Inguinal endometriosis may develop in the inguinal region as a mass from direct implantation, coelomic metaplasia, tubal regurgitation lymphatic spread [6]. However, Battista Candiani et al. [11] tend to dispute the plausibility of the latter two theories. They contest that, in view of the extraperitoneal location of the endometriotic lesions, tubal regurgitation seems unlikely. Furthermore, in terms of coelomic metaplasia, there does not appear to be an adequate embryologic explanation for the disproportionate predominance of endometriotic lesions on the right inguinal region, as compared to the left [11].

raperitoneal location of the endometriotic lesions, tubal regurgitation seems unlikely. Furthermore, in terms of coelomic metaplasia, there does not appear to be an adequate embryologic explanation for the disproportionate predominance of endometriotic lesions on the right inguinal region, as compared to the left [11]. These theories do not offer an adequate explanation for the pathogenesis of inguinal endometriosis, especially in terms of patients with neither history of pelvic endometriosis nor previous surgery. According to separate accounts by Quagliarello et al. [8] and Seydel et al. [3], inguinal endometriosis is difficult to explain if it is not found in the pelvic lymph nodes [3, 8]. In separate reports, several authors have documented cases of inguinal endometriosis without evidence of concomitant pelvic endometriosis [7–10]. The lesions were localized either in the round ligament [10] or the hernial sac [7–9]. The mode of spread of endometriosis in these cases are still in question. The predominance of right-sided inguinal endometriosis remains unknown. The asymmetrical lymphatic drainage in favor of the right inguinocrural region might explain this phenomenon but according to Battista Candiani et al. [11], anatomical descriptions and oncologic observations do not seem to support this hypothesis [11].

predominance of right-sided inguinal endometriosis remains unknown. The asymmetrical lymphatic drainage in favor of the right inguinocrural region might explain this phenomenon but according to Battista Candiani et al. [11], anatomical descriptions and oncologic observations do not seem to support this hypothesis [11]. Endometriosis of the inguinal canal can be difficult to detect, with the correct preoperative diagnosis made less than 50% of the time [3]. The presumptive diagnosis is most often confused with conditions such as incarcerated hernia, lymphadenopathy, suture granuloma, neuroma, abscess, hydrocoele of the inguinal canal, primary or metastatic cancer, lymphoma, lipoma, hematoma, sarcoma, and subcutaneous cyst [3, 10]. Very often, the final diagnosis is made only after the pathology, either from biopsy or at exploration, and demonstrates the histological presence of endometriosis [3]. A history of periodic menstrual pain and tenderness associated with an inguinal mass is important in distinguishing this condition from other inguinal pathology [10].

Endometriosis of the inguinal canal can be difficult to detect, with the correct preoperative diagnosis made less than 50% of the time [3]. The presumptive diagnosis is most often confused with conditions such as incarcerated hernia, lymphadenopathy, suture granuloma, neuroma, abscess, hydrocoele of the inguinal canal, primary or metastatic cancer, lymphoma, lipoma, hematoma, sarcoma, and subcutaneous cyst [3, 10]. Very often, the final diagnosis is made only after the pathology, either from biopsy or at exploration, and demonstrates the histological presence of endometriosis [3]. A history of periodic menstrual pain and tenderness associated with an inguinal mass is important in distinguishing this condition from other inguinal pathology [10]. Diagnostic modalities such as radiological studies, ultrasound, and computed tomography scan have not been specifically helpful in the diagnosis [3]. These modalities cannot differentiate an endometrioma from a mass of similar density such as hematoma, lymphoma, sarcoma, or desmoid tumor [6]. On the other hand, magnetic resonance imaging is more accurate since it can identify the presence of iron in the haemosiderin deposits contained in an endometrioma [6]. Battista Candiani et al. [11] advocate more intensive use of this technique to increase the diagnostic accuracy [11]. Fine needle aspiration biopsy has also been shown to aid in the diagnosis of a case of endometriosis associated with an inguinal hernia [6, 7]. Perez-Seoane et al. [7] believe that this procedure should probably be the first step in the diagnosis of inguinal endometriosis due to its ease in performing and accuracy in results [7]. The finding of small groups of nonatypical epithelial cells in an inflammatory and proteinaceous background in FNA biopsy was similar to those of endometriosis in other sites. The absence of atypia helps to differentiate inguinal endometriosis from metastatic carcinoma to the inguinal region [7].

for the residual endometriosis [11]. In terms of the usage of prophylactic hormonal therapy for endometriosis, Quagliarello et al. [8] (1985) cautioned that, due to its side effects, suppressive therapy with Danazol should be reserved only for patients with laparoscopic evidence of coexisting pelvic endometriosis [8]. 4. Need for Laparoscopy Majeski [6] recommends performing pelvic laparoscopy for all patients who have endometriosis in the extraperitoneal part of the round ligament or in a scar [6]. This view is shared by other authors (Goh and Flynn [10] 1994; Batistta Candiani et al. [11] 1991) because of the association with pelvic endometriosis and subfertility [10, 11]. On the other hand, Seydel et al. [3] did not see the need to perform laparoscopy in patients who do not present with signs of pelvic endometriosis [3]. For the patient in this case, simple excision was performed. It was not deemed necessary to perform laparoscopy since this patient did not present with signs of pelvic endometriosis. 5. Summary We present a case of a 48-year-old woman with right inguinal mass associated with inguinal pain during menses. Fine-needle aspiration biopsy of the mass revealed endometriosis. This was managed by simple excision. The patient has remained asymptomatic ever since. Simple excision is deemed as adequate management in this case.

sent a case of a 48-year-old woman with right inguinal mass associated with inguinal pain during menses. Fine-needle aspiration biopsy of the mass revealed endometriosis. This was managed by simple excision. The patient has remained asymptomatic ever since. Simple excision is deemed as adequate management in this case. Inguinal endometriosis is a rare clinical condition. It affects reproductive age women, with or without concomitant pelvic endometriosis. It usually presents as painful/tender inguinal mass and is most often mistaken for incarcerated hernia and other inguinal lesions. A history of catamenial pain often helps to rule out other inguinal pathology. Clinicians should not underscore the importance of a carefully taken history and physical examination, an increased awareness of this disease entity and a high index of suspicion in coming up with a correct diagnosis. Failure to recognize this syndrome and treatment with adequate excision may predispose to symptomatic recurrence. On the other hand, an accurate preoperative diagnosis may spare both the patient and physician the undue stress of performing an unnecessary emergency operation. Figure 1 Inguinal mass, right. Figure 2 Gross specimen, right inguinal mass. Table 1 Review of the literature. Author No. of patients History of surgery History of Pelvic endometriosis Laparoscopy Results of laparoscopy Site of lesion Majeski [6] 1 (+) previous CS (−) Done       (−) Scar tissue (+) previous hernia Seydel et al. [3] 2 (+) previous CS (−) Not done N.A. No mention

Figure 2 Gross specimen, right inguinal mass. Table 1 Review of the literature. Author No. of patients History of surgery History of Pelvic endometriosis Laparoscopy Results of laparoscopy Site of lesion Majeski [6] 1 (+) previous CS (−) Done       (−) Scar tissue (+) previous hernia Seydel et al. [3] 2 (+) previous CS (−) Not done N.A. No mention Singh et al. [4] 1 (−) No mention Not done N.A. Hernial sac 1 (+) previous CS No mention Not done N.A. Scar tissue Goh and Flynn [10] 1 No mention (−) Not done N.A. Round ligament Battista Candiani et al. [11] 6 No mention All (+) Done All (+) Round ligament Perez-Seoane et al. [7] 1 (−) (−) Not done N.A. Hernial sac Quagliarello et al. [8] 1 No mention (−) Done (−) Hernial sac Brzezinskiand Durst [9] 1 No mention (−) Not done N.A. Hernial sac

1. Introduction The human papillomaviruses (HPVs), especially the high-risk types (HPV types 16 and 18) have been implicated in the development of high-grade cervical intraepithelial neoplasia (CIN II/III) and invasive cervical cancer [1, 2]. Preventive strategies utilizing vaccines directed against structural components of this virus have been approved. While prevention of HPV infection is an important goal for future generations of women, such intervention will useless to the millions already infected with high risk genotypes. Low-grade lesions (CIN I) usually clear spontaneously, whereas high-grade CIN II and III often persist and may progress to carcinoma [3, 4]. Current treatment strategies such as a loop electrosurgical excision procedure (LEEP) or cone biopsy of the cervix are aimed to excision of visible lesions and may eliminate HPV-infected cells and associated disease. Such ablative procedures may activate cellular immune responses to the associated HPV antigens. However, when the size of the lesion is large treatment with LEEP may be incomplete which leads to the permanence of positive margins and the persistence of high-grade lesions. Moreover, patients with CIN who have undergone ablative surgery or even a hysterectomy may still have HPV infection [5], which may result in recurrent or persistent CIN. These patients require repetitive invasive procedures for treatment. This can be problematic for women who desire future fertility due to distortion of the cervix after invasive interventions and also for preterm birth [6].

hysterectomy may still have HPV infection [5], which may result in recurrent or persistent CIN. These patients require repetitive invasive procedures for treatment. This can be problematic for women who desire future fertility due to distortion of the cervix after invasive interventions and also for preterm birth [6]. Vaccination strategies that boost natural immunity to HPV might allow clearance of the virus leading to resolution of dysplastic lesions. This could avoid the need for invasive treatment and may lead to a lower risk of recurrence. Various vaccines aimed at inducing regression in women with biopsy proven CIN II/III have been studied. The most frequently targeted antigens are the E6 and E7 proteins, because they are oncogenic and sustained expression is required for the maintenance of the cancerous phenotype. Their immunogenicity has rendered these HPV proteins an attractive target for immunization strategies to prevent cervical carcinoma [1, 2]. However, these vaccines have generally not been effective enough to produce a real tumor rejection (for review, see [7, 8]). As with every biological system, tumors use opportunistic and redundant mechanisms to guarantee their survival and development. The robustness of these processes makes cancer immunotherapy a real challenge. The success of cancer immunotherapy will not depend only on optimal tumor-associated antigen but also of the use of appropriate adjuvants, while being nontoxic and safe (for review, see [9]).

isms to guarantee their survival and development. The robustness of these processes makes cancer immunotherapy a real challenge. The success of cancer immunotherapy will not depend only on optimal tumor-associated antigen but also of the use of appropriate adjuvants, while being nontoxic and safe (for review, see [9]). A new adjuvant approach has been previously developed in which gangliosides are incorporated into the N. meningitides outer membrane complex to form a nanoparticulate of very small size proteoliposomes (VSSP) [10]. Vaccination with the VSSP adjuvated HPV16 E7 (49–57) minimal CTL peptide protected mice against the HPV16 tumor model TC-1 challenge, induced regression of established tumors as well as E7-specific CD8+ T-cell responses [11]. On the basis of this preclinical rationale, “first-in-human” clinical trial with the CIGB-228 vaccine containing HLA-restricted HPV16 E7 epitope known to be recognized by CTLs (E7 peptide 86–93) [12] adjuvated with VSSP was done in patients with CIN II/III. In addition to the toxicity and tolerability of the CIGB-228 vaccine, immune, virological and clinical response end points were assessed.

with the CIGB-228 vaccine containing HLA-restricted HPV16 E7 epitope known to be recognized by CTLs (E7 peptide 86–93) [12] adjuvated with VSSP was done in patients with CIN II/III. In addition to the toxicity and tolerability of the CIGB-228 vaccine, immune, virological and clinical response end points were assessed. 2. Material and Methods 2.1. Patient Eligibility HLA-A2-positive patients, 18-to-65 years-old with histological confirmed HPV16-positive CIN II/III, with a larger diameter ≥3 mm by videocolposcopy were recruited at specialized services from seven gyneco-obstetric hospitals throughout Havana and other provinces after oral and written informed consent (Table 1). They were then evaluated at the National Reference Center for Cervical Cancer at the “Ramon Gonzalez Coro” Havana Hospital, where the eligibility criteria were verified. Eligibility also required the following criteria: leukocytes >3 × 109/L, lymphocytes >1 × 109/L, thrombocytes >100 × 109/L, and hematocrit >30%. Exclusion criteria were to have received any immunomodulator treatment up to 3 months before inclusion, psychiatric dysfunctions, pregnancy, and breastfeeding, decompensate chronic diseases such as asthma, epilepsy, autoimmune, or immunodeficiency diseases, hypertension, anemia, acute systemic or genital tract infections, and renal, hepatic, and cardiovascular disorders.

dulator treatment up to 3 months before inclusion, psychiatric dysfunctions, pregnancy, and breastfeeding, decompensate chronic diseases such as asthma, epilepsy, autoimmune, or immunodeficiency diseases, hypertension, anemia, acute systemic or genital tract infections, and renal, hepatic, and cardiovascular disorders. 2.2. Study Design The study followed the principles of the Declaration of Helsinki for investigations in humans. It was approved by the Ethics and Scientific Committees of the participant institutions and by the Cuban Regulatory Authority. This was a single-arm, open, and uncontrolled study. The main purpose of the trial was to evaluate the product's safety during local and systemic adverse events. Sample size was previewed as between 7 and 10 patients. This N range assured that if no severe adverse reaction appeared, the probability of its occurrence would be less than 20%, with an 80%–90% confidence interval. Patients received four vaccinations. The vaccine was administered subcutaneously at 1-week intervals (Figure 1). Subjects were subjected to colposcopy monthly and LEEP excision of lesions in the cervix 60 days after completion of the immunization protocol (Figure 2).

less than 20%, with an 80%–90% confidence interval. Patients received four vaccinations. The vaccine was administered subcutaneously at 1-week intervals (Figure 1). Subjects were subjected to colposcopy monthly and LEEP excision of lesions in the cervix 60 days after completion of the immunization protocol (Figure 2). 2.3. Composition of the Vaccine The CIGB-228 vaccine consists of two components. The HPV16 E7(86–93) peptide was synthesized and supplied by the Center for Genetic Engineering and Biotechnology (CIGB) in 0.12 mg vials, as a lyophilized powder. VSSP was produced and supplied by the Center of Molecular Immunology in 0.8 mg/0.5 mL vials. Prior to the vaccination, the HPV16 E7(86–93) synthetic peptide was reconstituted with water for injection and then adjuvated with VSSP. One dose of the CIGB-228 vaccine contained 0.1 mg of peptide and 0.16 mg of VSSP in a total volume of 0.5 mL.

nd supplied by the Center of Molecular Immunology in 0.8 mg/0.5 mL vials. Prior to the vaccination, the HPV16 E7(86–93) synthetic peptide was reconstituted with water for injection and then adjuvated with VSSP. One dose of the CIGB-228 vaccine contained 0.1 mg of peptide and 0.16 mg of VSSP in a total volume of 0.5 mL. 2.4. Clinical Assessment Subjects underwent a general physical, cytological, and a colposcopic examinations monthly and cervical biopsy prior to and at termination of the study by LEEP (90 days after the first vaccine administration). Blood samples were drawn for routine safety analysis prior to and 1 week after each vaccine administration and for immunological analysis prior to each vaccination and two weeks after the last dose (Figure 1). Patients were tested for pregnancy at each visit. Additionally, patients were followed clinically and colposcopically at 3, 6, 9, and 12 months after LEEP (Figure 2). Since the main purpose of the trial was to evaluate the product's safety, local and systemic adverse events were carefully assessed. Systemic toxicity was evaluated for 24 hours after each CIGB-228 vaccination, including continuous cardiovascular monitoring, temperature, respiratory frequency, and blood pressure measurements 30 min after each injection, then every hour during 4 hours. Patients completed diary cards of adverse events as auto report over the seven days following each vaccine administration. The medical terminology used for AE and their severity classification (in grades) were those of the Cancer Therapy Evaluation Program, Common Terminology Criteria [13].

n every hour during 4 hours. Patients completed diary cards of adverse events as auto report over the seven days following each vaccine administration. The medical terminology used for AE and their severity classification (in grades) were those of the Cancer Therapy Evaluation Program, Common Terminology Criteria [13]. Colposcopical evaluations were done using a video colposcopy device (Mediscope, Medison, Korea) with a calibrated rack to standardize imaging distance and illumination. Lesions were examined by two different specialists after applying 5% acetic acid for 3 min. They were described according to the Barcelona 2002 classification [14], and their morphometric analysis was done with validated software for quantitative digital image evaluation (MADIP V.4; Institute of Cybernetics, Mathematics, and Physics, Havana). Histological analyses were performed on colposcopically directed biopsies from abnormal areas taken before treatment and afterwards in the LEEP specimen. Permanent sections were subsequently prepared, stained with hematoxylin-eosin, and finally reviewed by two different pathologists who classified the lesions according to the FIGO classification. 2.5. Response Criteria and Clinical Activity The total lesion area (TLA) was the parameter evaluated for colposcopical response. Response was defined according to the RECIST criteria [15]. The percentage of decrease was measured as

Histological analyses were performed on colposcopically directed biopsies from abnormal areas taken before treatment and afterwards in the LEEP specimen. Permanent sections were subsequently prepared, stained with hematoxylin-eosin, and finally reviewed by two different pathologists who classified the lesions according to the FIGO classification. 2.5. Response Criteria and Clinical Activity The total lesion area (TLA) was the parameter evaluated for colposcopical response. Response was defined according to the RECIST criteria [15]. The percentage of decrease was measured as (1) %decrease=TLAinitial−TLAfinalTLAinitial×100. Complete response was considered as disappearance of all initial TLA; partial response if at least a 30% decrease of the initial TLA; progressive disease meant at least a 20% increase of the initial TLA and as stable disease if a reduction or an increase of the TLA compared to the initial value that is not enough for classification of the outcome as partial response or progressive disease. The histological response was based on the lesions phase changes. A complete response was defined as the absence of high-grade lesions after vaccination, partial response when a downstaging of the high-grade lesion occurred; stable disease when a lesion remains at the same initial stage, and progression if there was upstaging. 2.6. HLA Testing HLA-A2 allele was performed by PCR-SSP typing of HLA class I allele method [16].

The histological response was based on the lesions phase changes. A complete response was defined as the absence of high-grade lesions after vaccination, partial response when a downstaging of the high-grade lesion occurred; stable disease when a lesion remains at the same initial stage, and progression if there was upstaging. 2.6. HLA Testing HLA-A2 allele was performed by PCR-SSP typing of HLA class I allele method [16]. 2.7. Technique for HPV Detection The presence of HPV DNA in biopsies was examined prior to and at termination of the study (90 days after the first vaccine administration) by PCR using a pair of L1-consensus primers (GP5+/GP6+) [17]. The HPV DNA detection limit was about 310 viral copies. Additional PCR with specific HPV16 primers was performed [18] and HPV DNA levels were normalized with those corresponding to β-globin as housekeeping gene. 2.8. Peptides Peptides used for in vitro studies were synthesized at the CIGB. The peptide sequences were from HPV16 E7, encompassing amino acids 11–20, 82–90, and 86–93, and influenza M1 peptide (GILGFVFTL, aa58–66). 2.9. Lymphocytes Peripheral blood mononuclear cells (PBMC) were isolated from fresh K3-EDTA blood samples within 2 h after sample collection using Ficoll density gradient according to the manufacturer (Bio-Sciences AB) and seeded at 5–10×106 cells/mL in freezing medium consisting of nine parts fetal bovine serum (FCS; Hyclone,) and one part dimethylsulfoxide (Sigma). PBMC were stored overnight in 1°C freezing containers (Nalgene Nunc International) at −80°C and then transferred into liquid nitrogen until use.

cturer (Bio-Sciences AB) and seeded at 5–10×106 cells/mL in freezing medium consisting of nine parts fetal bovine serum (FCS; Hyclone,) and one part dimethylsulfoxide (Sigma). PBMC were stored overnight in 1°C freezing containers (Nalgene Nunc International) at −80°C and then transferred into liquid nitrogen until use. 2.10. HPV-Specific T-Cell Immunity Monitoring Briefly, cryopreserved pre- and postvaccine PBMC were thawed quickly in a 37°C water bath. Then, cells were washed twice with phosphate buffer saline (PBS) and seeded in three replicate wells at a density of 2 × 105 cell/well in 100 μL of RPMI medium (Sigma) enriched with 10% of human AB serum (Sigma) and 50 μL/well of 10 μg/mL of indicated HPV16 E7 peptides, 10 μg/mL of influenza M1 peptide and 0.1 μg/mL of anti-CD3 mAb CD3-2 (positive control) or medium (negative control) as corresponded in a multiscreen 96-well PVDF plate (Mabtech AB) coated with an IFN-γ-catching antibody (Mabtech AB). After two days of incubation at 37°C, the ELISPOT was performed according to the manufacturer's instructions (Mabtech AB). Spots were counted with a fully automated computer-assisted video-imaging analysis system (Carl Zeiss Vision). A positive response to antigen was defined as >20 spots (IFN-γ-producing cells)/106 PBMCs in response to antigen (after subtraction of background). A positive response after vaccination was recorded if the frequency of IFN-γ-producing cells postvaccination was greater than twice the prevaccination response to an antigen.

positive response to antigen was defined as >20 spots (IFN-γ-producing cells)/106 PBMCs in response to antigen (after subtraction of background). A positive response after vaccination was recorded if the frequency of IFN-γ-producing cells postvaccination was greater than twice the prevaccination response to an antigen. 3. Results 3.1. Study Population Between October 2007 and February 2008, 7 women were included. Clinical and demographic characteristics of the study patients are shown in Table 1. Age ranged from 24 to 43 years (median 29). All patients had high-grade CIN confirmed on their histological pre-study and lesion areas between 30 and 638 mm2 (Table 2). Moreover, all women were HPV16 and HLA-A2 positive. The treatment schedule was completed in all patients, which were also evaluated one year after their LEEP for recurrence. 3.2. Adverse Events Table 3 lists all adverse events assessed as possibly being associated with the vaccine. All patients reported local pain at the vaccination site and 6 patients reported burning sensation. Other events were reported with low frequencies as local redness and swelling. Systemic adverse events, including fever, tremors, and cramps were registered in few cases. No vaccine-related events exceeded grade 1 (mild) and recovered spontaneously. One patient reported lower abdominal pain as a result of urinary sepsis. This event was considered to be unrelated to the vaccination. There were no late adverse events either.

cluding fever, tremors, and cramps were registered in few cases. No vaccine-related events exceeded grade 1 (mild) and recovered spontaneously. One patient reported lower abdominal pain as a result of urinary sepsis. This event was considered to be unrelated to the vaccination. There were no late adverse events either. 3.3. Clinical Evaluations Individual results are shown in Tables 1 and 2. TLA decreased in all patients at colposcopy at 3 months of followup after CIGB-228 vaccination. Colposcopic response was evidenced in six of seven patients (85.7%), four of them (57.1%) complete and two (28.6%) partial response (Table 2). Histological analyses indicated that 57.1% of the patients (4/7) experienced full regression while 14.3% (1/7) had histological grade downstaging. Stable disease was observed in two patients (28.6%). Concomitant negativization of HPV16 from the original lesion sites was observed in three of the patients who had a complete response (Table 1). None of the patients showed confirmed lesion recurrence during the 12-months, followup.

had histological grade downstaging. Stable disease was observed in two patients (28.6%). Concomitant negativization of HPV16 from the original lesion sites was observed in three of the patients who had a complete response (Table 1). None of the patients showed confirmed lesion recurrence during the 12-months, followup. 3.4. Immunologic Responses PBMCs isolated from blood samples drawn weekly before each vaccination (first, second, third, fourth) and one week after the last vaccination were subjected to IFNγ ELISPOT analysis. Some preexisting HPV16 T cell immunity that was E7 peptides 11–20, 82–90 and 86–93-specific was detected in all patients (including fourth patients who had a complete response); this immunity was boosted by vaccination and coincided with enhanced IFNγ production for all patients. The immunologic analysis in which PBMCs were tested not only against the vaccination CTL peptide 86–93 but also two additionally E7 peptides (11–20, 82–90) not only measured the presence of specifically a vaccine-induced T-cell response but also allowed the calculation of the strength of the immune response, operationally defined as a combination of the magnitude and breadth of the T-cell response to all three peptide as intramolecular epitope spreading. After the vaccination, all seven patients responded to peptides 86–93 and 82–90. Six of seven patients were positive to peptide 11–20. The frequencies ranged between one HPV-specific T cell among 100,000 PBMCs up to 7 of 10,000 (see Table 4 for complete overview). The peak of response for each patient reached after two or three vaccinations, and the maximum response to HPV16 E7 86–93 was more vigorous than against HPV16 E7 82–90 and HPV16 E7 11–20 in the strength of response (Figure 3) and in general are correlated with the clinical outcome.

(see Table 4 for complete overview). The peak of response for each patient reached after two or three vaccinations, and the maximum response to HPV16 E7 86–93 was more vigorous than against HPV16 E7 82–90 and HPV16 E7 11–20 in the strength of response (Figure 3) and in general are correlated with the clinical outcome. These results suggest that the clinical efficacy of a therapeutic HPV16 vaccine may be determined by its capacity to induce strong and broad immune responses to the HPV16 oncoprotein E7. 4. Discussion The HPV is an attractive target for a vaccine strategy, since the HPV E7 transforming protein, which is critical for the maintenance of a transformed state in animal model systems and in humans, has been shown to encode epitopes that bind to class I alleles and are recognized by T cells. Virus-like particle vaccines have promise for primary prevention of cervical intraepithelial neoplasia and cancer; however, it is unlikely that they will impact on established disease or preexisting intraepithelial lesions, since the capsid proteins of HPV are not expressed by neoplastic cells in high-grade CIN. Women with HPV16 are more likely to develop CIN II/III than those with other HPV types, and these data provide a strong justification for devising immunization strategies against high-risk HPV type 16 to prevent progression of low-grade CIN to high-grade disease, to treat high-grade CIN, to prevent and treat the recurrence of high-grade CIN, and to prevent the occurrence of invasive cervical cancer.

ypes, and these data provide a strong justification for devising immunization strategies against high-risk HPV type 16 to prevent progression of low-grade CIN to high-grade disease, to treat high-grade CIN, to prevent and treat the recurrence of high-grade CIN, and to prevent the occurrence of invasive cervical cancer. HPV-specific T cytolytic immune responses have been demonstrated in patients with high-grade CIN and cervical cancer [19]. Three peptides were defined that were immunogenic both in transgenic mice and in CTL induction experiments using PBMCs from HLA-A2 healthy donors derived from the 11–20, 82–90, and 86–93 amino acid sequences [20]. E7-specific CTL cells have been generated from the peripheral blood and lymph nodal tissue of HPV16 positive women with cervical dysplasia and cervical cancer by in vitro restimulation with autologous antigen-presenting cells pulsed with HPV16 E7 peptides [20, 21].

the 11–20, 82–90, and 86–93 amino acid sequences [20]. E7-specific CTL cells have been generated from the peripheral blood and lymph nodal tissue of HPV16 positive women with cervical dysplasia and cervical cancer by in vitro restimulation with autologous antigen-presenting cells pulsed with HPV16 E7 peptides [20, 21]. Trials to generate T cell reactivity against HPV16 E7 have been carried out in women with cervical cancer or cervical intraepithelial neoplasia. In previous reports, women with stage IV cervical cancer were immunized with an E7 86–93 lipopeptide sequence, and only six patients mounted a weak immune response against the E7 86–93 peptide sequence, without evidence of clinical benefit [22–24]. On the basis of the clinical rationale of that, women with a lower disease burden and preinvasive disease are more logical candidates for an antigen-specific immunotherapy than women with bulky invasive disease, prior chemotherapy, poor performance status, and profound immunosuppression; a phase I trial with a similar CTL peptide vaccine has been conducted in women with high-grade CIN. In that study, peptides binding to HLA-A2 emulsified with Montanide ISA 51, an oil-based adjuvant, were used for the treatment; 9/17 has evidence of clinical response with modest rates of immunity. Although the results obtained in that study were more optimistic respect to the clinical benefit, only 3 of 17 patients evaluated had complete regression of their lesion pathologically [25]. Due to the low levels of immunity seen and lack of regression observed, there is a perception that the peptide approach has little promise for control of HPV-induced cancer. That led many researchers to test vaccines based on E7 with other formats such as viral vectors, DNA vaccines, and fusion proteins [26–36]. Although there have been some encouraging results with these vaccines, so far, there is no clinical efficacy study in patients with cervical neoplasia and is still far to the approval of a therapeutic vaccine against HPV.

s based on E7 with other formats such as viral vectors, DNA vaccines, and fusion proteins [26–36]. Although there have been some encouraging results with these vaccines, so far, there is no clinical efficacy study in patients with cervical neoplasia and is still far to the approval of a therapeutic vaccine against HPV. In this work, immunotherapy with CIGB-228 containing HPV16 E7 epitope adjuvated with VSSP is proven to be safe. The maximal toxicity seen was grade 2 and consisted of discomfort and swelling at the vaccination sites and low-grade fever in the first 24 h after injection. In the quoted studies, mostly, exact HLA class I-binding peptides were used for immunotherapy of metastatic melanoma, and the overall clinical results with such exact HLA-fitting peptides in patients with melanoma have been very disappointing worldwide [37]. However, our preclinical studies in mice indicated that immunization with exact MHC class-I-fitting peptide and VSSP is superior to that with peptide alone [11]. VSSP belong to the new generation of adjuvants based on pathogen-related molecules identified as “danger” signals that are recognized by the innate immune system [38]. VSSP have the ability to activate mouse and human DC, in vitro and in vivo, with the corresponding IL-12p40/p70, TNF-α, and IL-6 production [9, 10]. Therefore, VSSP is a potent adjuvant for DC activation and Th1 differentiation that provide the proper costimulatory context for productive immune responses.

system [38]. VSSP have the ability to activate mouse and human DC, in vitro and in vivo, with the corresponding IL-12p40/p70, TNF-α, and IL-6 production [9, 10]. Therefore, VSSP is a potent adjuvant for DC activation and Th1 differentiation that provide the proper costimulatory context for productive immune responses. The rate of clinical response observed in this trial was significant although in the absence of randomized data and control arm, it is possible that the regression of high-grade CIN may be due to spontaneous regression [39]. CIGB-228 vaccine treatment met the predetermined criteria of at least a 30% clinical response rate (CR + PR). The observed clinical colposcopic response rate (85.7%), and histological response rate (71.4%) in three months, was more than the estimated spontaneous regression rate of high-grade CIN alone which is estimated to be maximum 10% in 6 months. In the current study, immune responses were seen to class I-restricted epitopes of HPV16 E7 protein and regression of CIN as well as diminution of grade of CIN have been associated with the detection of CD8 T cell reactivity to HPV16 E7 protein. There was no clear correlation between HPV16 clearance and clinical response. The small number of patients studied and the known spontaneous regression rate of CIN prevents any definitive conclusion as to the utility of the vaccine we tested. Only in futures larger multi-institutional randomized study with a placebo control arm will permit definitive conclusions to be drawn about the effectiveness of CIGB-228 vaccine in the treatment of high-grade CIN.

eous regression rate of CIN prevents any definitive conclusion as to the utility of the vaccine we tested. Only in futures larger multi-institutional randomized study with a placebo control arm will permit definitive conclusions to be drawn about the effectiveness of CIGB-228 vaccine in the treatment of high-grade CIN. The potential of peptides as therapeutic HPV vaccines pass for the difficulties of using peptide antigens for treatment of naturally occurring HPV-induced tumors are the types of HPV causing the tumors (most frequently 16 and 18) and the genetic immunological makeup of the patient (HLA type). However, it is possible to screen patients for the types of HPV present as well as HLA type and it is possible to create a patient-specific vaccine with the appropriate peptide antigens, based on this information. Furthermore, these vaccines are easily produced, are chemically stable, and are devoid of oncogenic potential as well as free of bacterial/viral contaminating substances, hereby avoiding the antigenic competition often seen against viral vector-based vaccines expressing tumor-associated antigens or HIV antigens [40–43] or between simultaneously injected antigens such as in the case with fusion protein or gene products [27, 44, 45].

ll as free of bacterial/viral contaminating substances, hereby avoiding the antigenic competition often seen against viral vector-based vaccines expressing tumor-associated antigens or HIV antigens [40–43] or between simultaneously injected antigens such as in the case with fusion protein or gene products [27, 44, 45]. 5. Conclusion In conclusion, our study shows that CIGB-228 vaccination is safe and well tolerated. This vaccine generates promising results based in the levels of immunity and clinical response. It was found to induce robust IFNγ T-cell responses to the HPV16 oncoprotein E7 in women with high-grade CIN. CIGB-228 vaccination resulted in clinical activity, as evidenced by the colposcopy, histology and the HPV clearance. In patients with premalignant lesions, immunotherapy with CIGB-228 vaccine is attractive and a trial to confirm therapeutic benefit in such patients has been initiated. Funding The financial support was provided by HeberBiotec SA. Conflict of Interests None declared. Author's Contributions All the authors contributed to the design of the paper. Ana M. Solares, I. Baladron, T. Ramos, Z. Borbon, L. Gonzalez, S. Fanjull, D. Castillo and J. Esmir carried out the clinical work. C. Valenzuela led the statistical analysis. M. Granadillo, A. Batte, A. Cintado, M. Ale, M. E. F. de Cossio, Ania Ferrer and I. Torrens were responsible of the laboratory tests. P. Lopez-Saura contributed to the overall supervision of the clinical work and the paper. All authors contributed to the writing up of the paper that was led by I. Torrens.

cal analysis. M. Granadillo, A. Batte, A. Cintado, M. Ale, M. E. F. de Cossio, Ania Ferrer and I. Torrens were responsible of the laboratory tests. P. Lopez-Saura contributed to the overall supervision of the clinical work and the paper. All authors contributed to the writing up of the paper that was led by I. Torrens. Acknowledgments The authors are grateful to the participating women. They also acknowledge Ministry of Public Health of Cuba (MINSAP): Evelio Cabezas, Blanca Manzano and Agueda Santana. They are grateful to the technical support provided by the Center for Genetic Engineering and Biotechnology (CIGB): Gerardo Guillen, Osvaldo Reyes, Hilda Garay, Matilde Lopez, Dania Bacardi, Karelia Cosme, Marisol Cruz, Elizeth Garcia, Marcelo Nazabal, Daniel Palenzuela, and Lidia I. Novoa. Figure 1 Schematic overview of vaccination scheme. Figure 2 Schematic overview of the clinical trial procedure. Figure 3 Immune response before and after vaccination. Note: ELISPOT data as number of spots per 106 input CD8 cells is shown on the ordinate, with paired patient samples (prevaccine, white columns; postvaccine, black columns) indicated on the abscissa. Each panel is shown for the individual peptide indicated at the top. In each case, the maximum response postvaccination is shown. Table 1 Results of detection of HPV16 correlated with clinical and immune response.

Figure 3 Immune response before and after vaccination. Note: ELISPOT data as number of spots per 106 input CD8 cells is shown on the ordinate, with paired patient samples (prevaccine, white columns; postvaccine, black columns) indicated on the abscissa. Each panel is shown for the individual peptide indicated at the top. In each case, the maximum response postvaccination is shown. Table 1 Results of detection of HPV16 correlated with clinical and immune response. Patients Age HPV 16 before vaccine HPV 16 prior to LEEP Initial histology End histology after LEEP Clinical status Immune response 01 43 + + CIN II Koilocytosis CR + 02 38 + + CIN III CIN III SD + 03 24 + + CIN III CIN II PR + 04 25 + − CIN II Koilocytosis CR + 05 26 + − CIN III Koilocytosis CR + 06 25 + − CIN III Negative CR + 07 25 + + CIN III CIN III SD + Note: All patients were positive for HLA-A2. Initial histology was by cervical biopsy prior to the study. End histology was by LEEP at termination of the study (90 days after the first vaccine administration). Abbreviations CIN II-III: cervical intraepithelial neoplasia, grade II or III. CR: complete response; PR: partial response; SD: stable disease. Table 2 Colposcopic evaluation of patients. TLA (mm2) Patients 01 02 03 04 05 06 07 Initial 244.93 493.68 608.98 638.98 481.71 30.8 291.96 1 month 0 (100%) 462.46 (6.32%) 486.72 (20.07%) 0 (100%) 75.70 0 (100%) 182.10 (37.62%) 2 months 0 (100%) 441.12 (10.64%) 381.8 (37.3%) 0 (100%) 48.15 0 (100%) 149.78 (48.69%) 3 months 0 (100%) 403.2 (18.32%) 286.61 (52.93%) 0 (100%) 0 (100%) 0 (100%) 104.73 (64.12%) Response status CR SD PR CR CR

TLA (mm2) Patients 01 02 03 04 05 06 07 Initial 244.93 493.68 608.98 638.98 481.71 30.8 291.96 1 month 0 (100%) 462.46 (6.32%) 486.72 (20.07%) 0 (100%) 75.70 0 (100%) 182.10 (37.62%) 2 months 0 (100%) 441.12 (10.64%) 381.8 (37.3%) 0 (100%) 48.15 0 (100%) 149.78 (48.69%) 3 months 0 (100%) 403.2 (18.32%) 286.61 (52.93%) 0 (100%) 0 (100%) 0 (100%) 104.73 (64.12%) Response status CR SD PR CR CR CR PR Abbreviations TLA: total lesion area. TLA: total lesion area. Complete response (CR) was considered as disappearance of all initial TLA; (PR) partial response if at least a 30% decrease of the initial TLA; progressive disease (PD) meant at least a 20% increase of the initial TLA and as stable disease (SD) if a reduction or an increase of the TLA compared to the initial value that is not enough for classification of the outcome as partial response or progressive disease. %decrease = TLAinitial − TLAfinal/TLAinitial × 100. Table 3 Adverse events of patients after each vaccination. 1st vaccination 2nd vaccination 3rd vaccination 4th vaccination Patients p a r s sys p a r s sys p a r s sys p a r s sys 01 x x x x 02 x x x x x x x x x 03 x x x x x 1,2,3 04 x x x x 4 x x 05 x x x 06 x x x x x x x x 07 x x 2 x x x x x x Abbreviations p, pain at vaccination site. a, ardor at vaccination site. r, redness at vaccination site. s, swelling at vaccination site. sys, systemic responses. 1-fever; 2-tremors; 3-Lower abdomen pain; 4-cramps. Table 4 IFNγ ELISPOT analysis of PBMC before and after vaccinations of patients. Patients 01 02 03 04 05 06 07 Prevaccination 86–93 315 100 70 180 100 160 90 82–90 265 30 80 145 90 90 50

a, ardor at vaccination site. r, redness at vaccination site. s, swelling at vaccination site. sys, systemic responses. 1-fever; 2-tremors; 3-Lower abdomen pain; 4-cramps. Table 4 IFNγ ELISPOT analysis of PBMC before and after vaccinations of patients. Patients 01 02 03 04 05 06 07 Prevaccination 86–93 315 100 70 180 100 160 90 82–90 265 30 80 145 90 90 50 11–20 300 15 80 120 50 75 50 M1 85 85 60 75 50 85 60 After one vaccination 86–93 425 145 190 190 110 175 110 82–90 260 45 150 150 100 115 60 11–20 315 25 130 130 75 110 60 M1 90 90 75 75 45 90 55 After two vaccinations 86–93 540 305 125 530 320 460 220 82–90 435 185 85 360 220 420 100 11–20 335 115 115 315 230 425 105 M1 80 80 55 80 50 80 55 After three vaccinations 86–93 670 285 190 425 215 510 245 82–90 710 160 150 320 190 445 160 11–20 595 100 180 290 155 480 140 M1 80 80 55 60 55 80 55 After four vaccinations 86–93 655 195 275 450 190 450 200 82–90 725 140 160 335 180 375 110 11–20 550 105 165 300 130 410 85 M1 75 75 60 80 50 75 60 Note: The PBMCs were tested against three peptide of HPV16 E7 (86–93, 82–90 and 11–20). M1 was taken as positive control. In bold, the positive responses (definition is described in Section 2) are depicted as number of specific spots per 106 PBMCs.

1. Introduction An emergency can be defined as a situation of serious and often dangerous nature, developing suddenly and unexpectedly and demanding immediate attention in order to save life [1]. The maternal mortality ratio (MMR), expressed as maternal deaths per 100,000 live births over a given period, is a major measure of quality of obstetric care. According to World Health Organization (WHO) estimates, it varies up to 100-fold, from approximately 10 in developed countries to approximately 1,000 in least developed [2, 3]. Obstetric emergencies are the leading causes of maternal mortality worldwide and particularly in developing countries where literacy, poverty, lack of antenatal care, poor transport facilities and inadequate equipment/staffing combine to magnify the problem [4, 5]. Prevention where possible and prompt and effective treatment of obstetric emergencies will go a long way to reduce the magnitude of ever increasing maternal mortality which appears to have defied all proposed measures set to reduce it by WHO [6]. However, there are sparse data on the contribution of obstetric emergencies to maternal mortality in sub-Saharan Africa and developing countries like Nigeria where the maternal mortality ratios are very alarmingly high. This is one of the principal obstacles to appropriate distribution of resources targeted towards improving maternal healthcare. A study carried out by Nwobodo [7] in North-Western Nigeria showed that obstetric emergencies were responsible for 96.7% and 87% of the maternal and perinatal mortality, respectively. Although hospital-based studies have their limitations including referral bias, they are easy to perform in low-resource countries and can provide substantial and useful information [8, 9]. There have been several studies in South-Western Nigeria on maternal mortality [10–12] but none on obstetric emergencies in general and their influence on maternal and perinatal mortality. This study is therefore designed to explore this subject.

urce countries and can provide substantial and useful information [8, 9]. There have been several studies in South-Western Nigeria on maternal mortality [10–12] but none on obstetric emergencies in general and their influence on maternal and perinatal mortality. This study is therefore designed to explore this subject. 2. Subjects and Methods 2.1. Definition of Terms In this study, an obstetric emergency is defined as an obstetric complication or situation of serious and often dangerous nature, developing suddenly and unexpectedly and demanding immediate attention in order to save life [1]. Direct maternal deaths are those resulting from complications of the pregnant state (pregnancy, labour, and puerperium), from interventions, from omissions, from incorrect treatment, or from a chain of events arising from any of the above while indirect maternal deaths are those due to a previously existing diseases or disease that develop during pregnancy, and not due to direct obstetric causes but which were aggravated by the physiological effects of pregnancy. An “unbooked patient” refers to a woman who did not utilize the antenatal care services of OOUTH, Sagamu, Nigeria. Pregnancy-induced hypertension is when a pregnant woman, who was normotensive before getting pregnant, develops high blood pressure without proteinuria during pregnancy. Preeclampsia is when a pregnant woman develops high blood pressure and proteinuria, usually after 20 weeks of gestation. Eclampsia is occurrence of fits in a preeclamptic patient.

n is when a pregnant woman, who was normotensive before getting pregnant, develops high blood pressure without proteinuria during pregnancy. Preeclampsia is when a pregnant woman develops high blood pressure and proteinuria, usually after 20 weeks of gestation. Eclampsia is occurrence of fits in a preeclamptic patient. 2.2. Study Design, Data Collection, and Analysis This was a retrospective study in which the case records of all obstetric emergencies in OOUTH, Sagamu, Nigeria between January 2005 and December 2007 were obtained from the labour ward, antenatal ward, lying-in ward, and department of medical records of the hospital. Data extracted from the records include maternal age, parity, occupation, booking status, type of emergency and maternal/perinatal mortality. The data was subsequently analysed, and where applicable, subjected to statistical analysis using SPSS 15.0 for Windows Evaluation Version. Level of significance was set at P < .05 and determined by x-squared analysis. This study was approved by The Scientific and Ethical Committee of OOUTH, Sagamu, Nigeria. 2.3. Hospital Setting Ogun State is one of the 6 states in the southwestern region of Nigeria. The state has a land area of 2017 square kilometers and a population of 380,527. Ogun State has 20 local government areas of which Sagamu, with a population of 156,312 is one.

The data was subsequently analysed, and where applicable, subjected to statistical analysis using SPSS 15.0 for Windows Evaluation Version. Level of significance was set at P < .05 and determined by x-squared analysis. This study was approved by The Scientific and Ethical Committee of OOUTH, Sagamu, Nigeria. 2.3. Hospital Setting Ogun State is one of the 6 states in the southwestern region of Nigeria. The state has a land area of 2017 square kilometers and a population of 380,527. Ogun State has 20 local government areas of which Sagamu, with a population of 156,312 is one. Olabisi Onabanjo University Teaching Hospital, Sagamu, Nigeria is the only tertiary hospital for referral from all clinics, maternity homes and hospitals in all the Local Government Areas in Remo and Ijebu areas of Ogun State and adjoining areas of Lagos State. The hospital is funded by the government of Ogun State in South-Western Nigeria. It provides emergency obstetric services to women referred from other centres in addition to providing antenatal care and delivery services for low and high risk pregnant women from Sagamu community and neighboring towns. Patients are expected to pay directly for their services (except few that participate in National Health Insurance Scheme) though in emergency situations, they are managed within the means of existing resources before funds are made available. The hospital provides blood transfusion services from limited stock, and relatives of patients are requested to donate or provide donors when blood transfusion is indicated.

Health Insurance Scheme) though in emergency situations, they are managed within the means of existing resources before funds are made available. The hospital provides blood transfusion services from limited stock, and relatives of patients are requested to donate or provide donors when blood transfusion is indicated. 3. Results During the three-year period, there were 262 obstetric emergencies out of 1420 total deliveries giving an incidence of 18.5%. One hundred and four (39.7%) were booked while 158 patients (60.3%) were unbooked for antenatal care and delivery. The maternal age ranged from 15 to 45 years with a mean of 30 ± 2 years. The parity ranged from 0 to 8 with a mean of 3 ± 1. The leading emergencies as shown in Table 1 were prolonged/obstructed labour, postpartum haemorrhage, fetal distress, severe pregnancy-induced hypertension/eclampsia and antepartum haemorrhage (placental praevia/abruptio placenta). Other important types of emergencies include puerperial sepsis, ruptured uterus and retained second twin. There were 17 maternal deaths within the period, and obstetric emergencies accounted for 12 deaths (70.6%). Eleven of the maternal deaths occurred among 158 unbooked patients (6.9%) while only one out of 104 booked patients (0.9%) died. The leading cause of maternal death was obstructed labour. The maternal death rate was statistically significantly higher in unbooked than booked patients (11 deaths in 158 unbooked patients versus 1 death in 104 booked patients; P < .05). Out of 50 perinatal deaths during the study period, obstetric emergencies were responsible for 43 (86%). The perinatal death rate was also significantly higher in unbooked than booked patients (43 deaths in 115 unbooked patients versus 7 deaths in 97 booked patients; P < .05). At the time of presentation, only 105 (40%) could afford about thirty-five dollars (five thousand naira) deposit for admission. Out of 64 patients that required immediate blood transfusion, only 20 (31.2%) had it within 2 hours. The causes of delay in the other group were failure of their relatives to donate blood or organize donors and lack of funds to pay for necessary investigations.

-five dollars (five thousand naira) deposit for admission. Out of 64 patients that required immediate blood transfusion, only 20 (31.2%) had it within 2 hours. The causes of delay in the other group were failure of their relatives to donate blood or organize donors and lack of funds to pay for necessary investigations. 4. Discussion This study has shown that obstetric emergencies were relatively common in this centre and unbooked patients constituted a substantial bulk of cases. Obstetric emergencies were responsible for most of the mortality within the period of study, and maternal death rate was higher among unbooked patients. This picture is similar in many tertiary institutions in Nigeria and other developing countries [5–20]. Therefore, if health facilities were able to adequately and effectively respond to the needs of these women, many of the fatal cases would probably have survived. With reference to maternal mortality, similar findings were reported by other authors in Nigeria [5–17, 19, 20]. Interestingly, obstructed/prolonged labor, eclampsia, obstetric hemorrhage, puerperal sepsis and ruptured uterus (which were the most common emergencies in this study) are among the leading causes of maternal and perinatal mortality in the country [10–17, 19, 20]. Similar findings were obtained by Nwobodo in North-Western Nigeria in 2006 [7].

d/prolonged labor, eclampsia, obstetric hemorrhage, puerperal sepsis and ruptured uterus (which were the most common emergencies in this study) are among the leading causes of maternal and perinatal mortality in the country [10–17, 19, 20]. Similar findings were obtained by Nwobodo in North-Western Nigeria in 2006 [7]. This study reiterates the importance of proper antenatal care and delivery towards reducing maternal and perinatal mortality in this environment. Women unbooked for antenatal care and delivery were up to 22 times as likely to die in the hospital compared to booked patients [16, 20, 21]. The percentage of booked patients who died probably reflects the likely MMR if all women were to have adequate antenatal care and well-supervised delivery. The leading causes of maternal and perinatal deaths in this study are not significantly different from those identified in the developing countries for several decades [10, 12–17, 19]. This implies that our pregnant women are still dying from preventable causes of maternal and perinatal deaths and unlike suggested by some authors [22], no special technology or research is required to tackle the problem in this part of the world.

ntified in the developing countries for several decades [10, 12–17, 19]. This implies that our pregnant women are still dying from preventable causes of maternal and perinatal deaths and unlike suggested by some authors [22], no special technology or research is required to tackle the problem in this part of the world. Therefore it can be deduced that prevention (where possible) and effective management of obstetric emergencies will go a long way in reducing maternal and perinatal mortality in Nigeria. The strategies for achieving this objective will include the utilization of antenatal services, making budget for pregnancies and childbirth at family level (pending the time every family participates in National Health Insurance Policy), adequate funding of social welfare services to assist the indigent patients, the development of adequate blood banking system, liberal blood transfusion, and regular training of doctors and nurses on this subject.

ildbirth at family level (pending the time every family participates in National Health Insurance Policy), adequate funding of social welfare services to assist the indigent patients, the development of adequate blood banking system, liberal blood transfusion, and regular training of doctors and nurses on this subject. Limitations Inadequate documentation and the fact that some cases could have been missed because of the retrospective nature of this study made the compilation of statistics difficult in developing countries like ours. In addition, maternal deaths in the puerperium could have been underreported as postnatal clinic attendance in the hospital is generally poor and there is presently no measure to conduct home-based followup of parturients. This study only evaluates the effect of antenatal care (booked versus unbooked patients) on perinatal outcomes following an obstetric emergency among women admitted to hospital for labour and delivery. We do not know the outcome following an obstetric emergency among women excluded from hospital care for not being insured or able to pay for assistance. Conflict of Interests The authors declare that they have no conflict interests. Table 1 Frequency of obstetric emergencies.

Limitations Inadequate documentation and the fact that some cases could have been missed because of the retrospective nature of this study made the compilation of statistics difficult in developing countries like ours. In addition, maternal deaths in the puerperium could have been underreported as postnatal clinic attendance in the hospital is generally poor and there is presently no measure to conduct home-based followup of parturients. This study only evaluates the effect of antenatal care (booked versus unbooked patients) on perinatal outcomes following an obstetric emergency among women admitted to hospital for labour and delivery. We do not know the outcome following an obstetric emergency among women excluded from hospital care for not being insured or able to pay for assistance. Conflict of Interests The authors declare that they have no conflict interests. Table 1 Frequency of obstetric emergencies. Type of emergency n (%) Prolonged labour 46 (17.6) Obstructed labour 43 (16.4) Postpartum haemorrhage 37 (14.1) Fetal distress 28 (10.6) Severe pregnancy-induced hypertension 23 (8.8) Eclampsia 21 (8.0) Antepartum haemorrhage 20 (7.6) Puerperial sepsis 16 (6.1) Ruptured uterus 12 (4.6) Retained second twin 10 (3.8) Severe anaemia in pregnancy 2 (0.8) Cord prolapse 2 (0.8) Cord presentation 1 (0.4) Uterine inversion 1 (0.4) Total 262 (100)

1. Introduction Schistosomiasis was first described by Theodor Bilharz in 1851 [1]. It is estimated that more than 200 million people worldwide have schistosomiasis and that the infection is responsible for more than 200,000 deaths annually [2]. Environmental changes that result from the development of water resources and the growth and migration of populations can facilitate the spread of schistosomiasis [1]. Although most infections occur in residents of endemic areas, it has been clearly documented that brief freshwater exposure is sufficient to establish infection; thus, travellers may also be infected [3]. There is no accurate information about infection rates among returned travellers and immigrants because it is a rare disease in western countries [1]. Herein, we report a rare case of female genital schistosomiasis (FGS) in a European woman.

sufficient to establish infection; thus, travellers may also be infected [3]. There is no accurate information about infection rates among returned travellers and immigrants because it is a rare disease in western countries [1]. Herein, we report a rare case of female genital schistosomiasis (FGS) in a European woman. 2. Case Report A European 20-year-old woman was referred from our mucous clinic with a history of a vulvar mass evolving for three weeks. Her past medical and surgical history was unremarkable. This mass was painful and was increased in size. The patient went to Senegal and Mali three months ago, and she had a history of swimming in fresh water lakes in these countries. Clinically, we observed a swelling of the right labium minus (Figure 1). This mass was painful with a soft consistency. No other cutaneous or genital lesion was found. She had no fever, urticarial rash, angioedema, dry cough, or wheeze. No hematuria and no dysuria were reported. Hepatomegaly and splenomegaly were absent. Following laboratory parameters were within normal limits: complete blood cell count, serum electrolyte profile, and liver function tests. No antischistosomal antibodies were found in the serum, and there was no eosinophilia. Microscopy of urine and stools was negative for ova, cysts, and parasites and did not contain red blood cells or white blood cells. Biopsy of the vulvar lesion was made.

ell count, serum electrolyte profile, and liver function tests. No antischistosomal antibodies were found in the serum, and there was no eosinophilia. Microscopy of urine and stools was negative for ova, cysts, and parasites and did not contain red blood cells or white blood cells. Biopsy of the vulvar lesion was made. Histological examination at low-power view revealed a noncaseating granulomatous reaction to clusters of viable-appearing eggs in the dermis (Figure 2). The granuloma was composed of epitheloid histiocytes and a lot of eosinophils and plasma cells surrounded by a rim of lymphocytes (Figure 3). The epidermis was moderately acanthotic with hyperkeratosis. We observed foci of spongiosis and exocytosis in front of the infiltrate. At high-power view, histology revealed numerous schistosome ova with terminal spines, characteristic of Schistosoma haematobium. The ova contained multiple viable miracidia surrounded by a marked granulomatous inflammatory infiltrate (Figure 4). The diagnosis proposed was FGS. The patient was treated with oral praziquantel at a dose of 2400 mg for one day. She came back to our clinic six months later with disappearance of the vulvar lesion, and no others symptoms were reported.

miracidia surrounded by a marked granulomatous inflammatory infiltrate (Figure 4). The diagnosis proposed was FGS. The patient was treated with oral praziquantel at a dose of 2400 mg for one day. She came back to our clinic six months later with disappearance of the vulvar lesion, and no others symptoms were reported. 3. Discussion Schistosomiasis is the infection of humans by trematodes (a class of helminths). Three major species of schistosomes are described: S. haematobium, S. japonicum, and S. mansoni [4]. S. haematobium infection is acquired predominantly in North Africa, sub-Saharan Africa, the Middle East, Turkey, and India. The majority of S. mansoni infections are found in sub-Saharan Africa. S. japonicum infection still occurs in China, Indonesia, and the Philippines [1, 5]. Morphologically, S. haematobium ova have a spine in the apical position, whereas the spine of S. mansoni ova is on the lateral aspect, and S. japonicum ova have no spine [5]. FGS is the isolated chronic form of genital schistosomiasis. No other symptom is reported. S. haematobium is the organism most often identified in FGS [1, 4, 6]. In most cases, the lesions are found on the labia majora [6]. Isolated internal genital disease is less frequent [1]. In postmortem studies of the reproductive tract in endemic areas, ova are found in the vulva in 7 to 17% of cases [7].

is reported. S. haematobium is the organism most often identified in FGS [1, 4, 6]. In most cases, the lesions are found on the labia majora [6]. Isolated internal genital disease is less frequent [1]. In postmortem studies of the reproductive tract in endemic areas, ova are found in the vulva in 7 to 17% of cases [7]. The life cycle of schistosomiasis is complex and requires both intermediate and definitive hosts. When eggs excreted from infected humans contaminate bodies of fresh water, they hatch to release free-swimming miracidia, which in turn infect snails, the intermediate hosts. The miracidia mature inside the snails to become cercariae, which are released back into the water and can penetrate the skin of humans. Cercariae can survive up to 48 hours in water but are most infectious to humans in the first few hours after release from the snail. Cercariae pass via the lung and the liver into the portal venous and then mature into adult worms and unite. Pairs of worms then migrate to the superior mesenteric veins (in the case of S. mansoni), the inferior mesenteric and superior haemorrhoidal veins (in the case of S. japonicum), or the vesical plexus and veins draining the ureters and the urinary bladder (in the case of S. haematobium). Live schistosome eggs are excreted in the feces (in the case of S. mansoni and S. japonicum) or urine (in the case of S. haematobium). Direct retrograde spread of the adult worms from their usual sites into the venous system supplying vulvar skin leads to deposition of ova in the skin and subsequent formation of genital lesions, as in our patient. The life cycle is completed when the eggs hatch, releasing miracidia that, in turn, infect specific freshwater snails [1, 4, 5]. The adult worms remain in the blood vessels for life and survive for five to seven years and can even persist for up to 30 years [8].

bsequent formation of genital lesions, as in our patient. The life cycle is completed when the eggs hatch, releasing miracidia that, in turn, infect specific freshwater snails [1, 4, 5]. The adult worms remain in the blood vessels for life and survive for five to seven years and can even persist for up to 30 years [8]. Physiopathologically, 2 to 3 cm adult worms may cause venous obstruction where they reside, but the disease is more commonly caused by the daily deposition of numerous eggs by the female (hundreds to thousands eggs per day, depending on the species), which invades local tissues, where they release toxins and enzymes and provoke a TH-2-mediated immune response [4, 8, 9]. Inflammation and granuloma formation occur around deposited eggs, which can lead to extensive tissue damage (fibrosis and scarring) [4, 10]. Most patients infected with schistosomes of all species are asymptomatic. Acute symptoms tend to be more common in nonimmune individuals, such as travellers, due to a more intense immune response to exposure. By contrast, chronic complications require a higher burden of infection and, thus, are mainly seen in individuals from endemic areas [11]. The clinical picture of genital schistosomiasis and the pathological findings vary according to the organ affected. The clinical manifestations of FGS are nonspecific except the presence of sandy patches (areas of roughened mucosa surrounding egg deposits) on the cervical surface [7, 12]. It can include irregular bleeding, discharge, pelvic pain or tenderness, dyspareunia, and otherwise unexplained infertility. The lesions are polypoid or papillomatous tumorlike lesions on the vaginal wall and vulva. Macroscopically, lesions in FGS may mimic any neoplastic or infectious process (e.g., condyloma lata, syphilis, squamous cell carcinoma) in the female genital tract [1, 4, 13]. Lesions of the vulva and clitoris vary in size and presentation. Progressive and relapsing swelling, painful or painless ulceration, nodular surface, pruritus, and a hypertrophic clitoris with an eroded granular surface have been described. Genital schistosomiasis started with an irritation of the skin, followed by oedema and hyperaemia. Later small nodules developed under the skin and papillomatous lesions appeared forming masses resembling condylomata [14].

rface, pruritus, and a hypertrophic clitoris with an eroded granular surface have been described. Genital schistosomiasis started with an irritation of the skin, followed by oedema and hyperaemia. Later small nodules developed under the skin and papillomatous lesions appeared forming masses resembling condylomata [14]. In travellers returning from the tropics, due to the unspecific nature of gynaecological findings, delays of more than 24 months in diagnosis have been reported, the lesions being misdiagnosed as, for example, vulval warts [14]. The diagnosis of FGS is first suspected after taking a careful history and physical examination, making sure to consider travel history in endemic areas. It can be confirmed by microscopy with egg identification or by serology [4, 15]. Eggs can be directly visualized in urine or feces, but this test has limited use in the diagnosis of FGS, because FGS may exist independently and without signs of urinary schistosomiasis [4, 14, 16]. Studies in S. haematobium-endemic areas have also shown that up to 23% of women may have involvement of the lower reproductive tract even without schistosome ova in the urine [7].

test has limited use in the diagnosis of FGS, because FGS may exist independently and without signs of urinary schistosomiasis [4, 14, 16]. Studies in S. haematobium-endemic areas have also shown that up to 23% of women may have involvement of the lower reproductive tract even without schistosome ova in the urine [7]. The complications with S. haematobium are granulomatous inflammation, ulceration of the vesical and ureteral walls with subsequent fibrosis, functional bladder neck obstruction, hydroureter, hydronephrosis, and calcifications of the urinary tract and bladder. Genital involvement can cause infertility secondary to ovarian fibrosis or tubal occlusion. S. haematobium could be a cofactor for the development of cervical cancer and squamous cell carcinoma of the bladder, but a clear link has yet to be established. Finally, FGS may also facilitate the transmission of human immunodeficiency virus [1, 4, 7, 12, 16].

rtility secondary to ovarian fibrosis or tubal occlusion. S. haematobium could be a cofactor for the development of cervical cancer and squamous cell carcinoma of the bladder, but a clear link has yet to be established. Finally, FGS may also facilitate the transmission of human immunodeficiency virus [1, 4, 7, 12, 16]. The standard of care for treatment is a single dose of oral praziquantel, a pyrazinoisoquinoline derivative, at a dose of 40 mg/kg [1, 4, 13]. The drug's precise action on adult worms is unknown. Although praziquantel is not considered to be teratogenic or mutagenic, the drug is not recommended in pregnancy or in lactating women [14]. Reexamination of feces or urine one month after treatment is recommended in order to assess efficacy [1]. In our patient, this control is useless because the urinary and feces tests were already negative before the treatment. Meltzer et al. propose to screen and treat asymptomatic travellers with history of freshwater exposure in endemic countries [17]. In conclusion, female genital schistosomiasis is a rare disease in the western world. Our patient illustrates an unusual presentation of schistosomiasis via the localisation of the lesion and the patient's origin. It should be born in mind as a cause of granulomatous inflammation on the genitalia and must enter into the differential diagnosis of symptomatic genital lesions when the clinical history indicates travel in endemic areas. Eggs can be identified by cytologic or histologic methods and appropriate treatment can be initiated, avoiding substantial morbidity.

of granulomatous inflammation on the genitalia and must enter into the differential diagnosis of symptomatic genital lesions when the clinical history indicates travel in endemic areas. Eggs can be identified by cytologic or histologic methods and appropriate treatment can be initiated, avoiding substantial morbidity. Figure 1 Clinically, we observed a swelling of the right labium minus. Figure 2 A noncaseating granulomatous reaction to clusters of viable-appearing eggs in the dermis (Haematoxylin-eosin ×100). Figure 3 The granuloma is composed of epitheloid histiocytes and a lot of eosinophils and plasma cells (Haematoxylin-eosin ×200). Figure 4 Histology revealed numerous schistosome ova with terminal spines (arrow) characteristic of Schistosoma haematobium (Haematoxylin-eosin ×400).

1. Introduction Maternal syphilis and congenital syphilis are public health problems worldwide. An estimated 50% of pregnancies with maternal syphilis will end in fetal or perinatal death, low birth weight babies, or babies born with congenital syphilis [1]. Necrotizing funisitis, which is associated with increased rates of stillbirth, perinatal infection, and preterm delivery, is sometimes accompanied with congenital syphilis. Immunohistochemistry (IHC) has been used to detect Treponema pallidum in the placenta and umbilical cord. Guarner et al. [2] reported that 11% of neonates born from maternal syphilis had necrotizing funisitis with T. pallidum. Even though fetal heart rate (FHR) monitoring is widely used to monitor the well-being of the fetus, congenital syphilis is rarely associated with an abnormal FHR before labor [3]. In this case report, we describe a 31-week gestation neonate with congenital syphilis in which fetal bradycardia was shown as a clinical sign by FHR monitoring. The neonate was delivered by emergency cesarean section and demonstrated severe chorioamnionitis (CAM) and necrotizing funisitis with numerous T. pallidum by histopathologic examination.

cribe a 31-week gestation neonate with congenital syphilis in which fetal bradycardia was shown as a clinical sign by FHR monitoring. The neonate was delivered by emergency cesarean section and demonstrated severe chorioamnionitis (CAM) and necrotizing funisitis with numerous T. pallidum by histopathologic examination. 2. Case Report An unmarried 24-year-old woman (gravida 4, para 4), who had not attended any antenatal care (ANC), presented to our hospital with a complaint of abdominal discomfort. In the patient's four previous births, the first and second children were delivered by full-term spontaneous deliveries at another hospital without ANC, and the third and fourth children were delivered at the patient's home without medical assistance. The mother and her children had not been diagnosed with syphilis. During the examination, the patient appeared healthy with a blood pressure of 118/81 mmHg, a pulse rate of 78 bpm, and a body temperature of 37.2°C. The gestational age was estimated by her last menstrual period to be approximately 31 weeks. Abdominal examination revealed a soft, nontender uterus. Ultrasound evaluation confirmed a single fetus in cephalic presentation with an estimated body weight of 1,500 g and no evidence of placental abruption. Vaginal examination showed the cervix to be 30% effaced and 2 cm dilated with an intact membrane. Laboratory findings revealed elevated white blood cell counts (11,670/μL) and C-reactive protein levels (1.63 mg/dL).

n cephalic presentation with an estimated body weight of 1,500 g and no evidence of placental abruption. Vaginal examination showed the cervix to be 30% effaced and 2 cm dilated with an intact membrane. Laboratory findings revealed elevated white blood cell counts (11,670/μL) and C-reactive protein levels (1.63 mg/dL). FHR monitoring at the initial evaluation showed several early decelerations and bradycardia to 50 bpm for a one-minute period prior to recovery to normal baseline in association with a painless uterine contraction. The patient was immediately placed in the lateral position followed by the administration of oxygen via a face mask and a rapid intravenous infusion of fluid. The baseline variability was within a normal range. Twenty-five minutes later, another prolonged bradycardia to 70 bpm for a 7-minute period prior to recovery to normal baseline occurred without a uterine contraction. A decision was made to deliver immediately by cesarean section. Tocolysis was not performed during the course of treatment. A female neonate covered with a thick meconium weighing 1,423 g with Apgar scores of 2 and 8 at 1 and 5 minutes was delivered. Umbilical arterial pH was 7.28. After resuscitation, the neonate was transferred to the neonatal intensive care unit. The neonate was assessed to be 31 weeks at birth using the Dubowitz score and was diagnosed with an intrauterine infection. Routine investigations were performed to determine the nature of the infection.

mbilical arterial pH was 7.28. After resuscitation, the neonate was transferred to the neonatal intensive care unit. The neonate was assessed to be 31 weeks at birth using the Dubowitz score and was diagnosed with an intrauterine infection. Routine investigations were performed to determine the nature of the infection. Following cesarean section, the mother was diagnosed with syphilis by a serological test. Congenital syphilis was confirmed by serology of the neonate: rapid plasma region 32x, Treponema pallidum hemagglutination 1280x, and a positive immunoglobulin fluorescent treponemal antibody-absorption test. Histopathological examination of the placenta, umbilical cord, and membranes revealed severe CAM and necrotizing funisitis; inflammatory cells could be seen on the surface of the cord and within Wharton jelly. Numerous T. pallidum were detected by IHC, which uses an antibody against T. pallidum (Figure 1). Both mother and neonate were treated with penicillin with good results. The neonate did not have any skin lesions or radiological bone changes and was discharged after 56 days with no evidence of sequela.

in Wharton jelly. Numerous T. pallidum were detected by IHC, which uses an antibody against T. pallidum (Figure 1). Both mother and neonate were treated with penicillin with good results. The neonate did not have any skin lesions or radiological bone changes and was discharged after 56 days with no evidence of sequela. 3. Discussion Congenital syphilis can be prevented through antenatal screening and treatment. However, more than 1 million infants are born with congenital syphilis each year worldwide. The World Health Organization aims to reduce maternal morbidity, fetal loss, and neonatal morbidity and mortality due to syphilis and is conducting a global effort to eliminate congenital syphilis [1]. It is well known that syphilis in untreated or inadequately treated pregnant women is associated with neonatal morbidity and mortality and may lead to histological effects in the placenta such as necrotizing funisitis.

and mortality due to syphilis and is conducting a global effort to eliminate congenital syphilis [1]. It is well known that syphilis in untreated or inadequately treated pregnant women is associated with neonatal morbidity and mortality and may lead to histological effects in the placenta such as necrotizing funisitis. To our knowledge, there has been only one case report of congenital syphilis presenting with an abnormal FHR before labor. Savage and Reader [3] reported that antepartum FHR monitoring showed a loss of baseline variability and several decelerations in a 36-week gestation neonate with congenital syphilis and severe birth asphyxia. This case study is the first report of an intrauterine syphilis infection fully supported by antepartum abnormal FHR patterns and placental histopathological evidence. Marked CAM and necrotizing funisitis with numerous T. pallidum were confirmed even though the symptoms and signs of inflammation of the mother were not as serious according to the laboratory test. In this case, bradycardia detected during FHR monitoring was a critical sign of fetal health and was important in the decision to perform an emergency cesarean section. Therefore, without FHR monitoring, prompt intervention, and skilled neonatal intensive care, this neonate might not have survived. This case highlights the importance of raising the level of awareness that a fetus with early onset neonatal sepsis may be born when abnormal patterns are identified during antepartum FHR monitoring. Furthermore, it is important to process all investigations urgently for women who are not attending or attending insufficient ANC. The risk of performing cesarean section on preterm infants should be considered especially when adequate facilities to manage high-risk pregnancies (e.g., NICU) are not available.

nitoring. Furthermore, it is important to process all investigations urgently for women who are not attending or attending insufficient ANC. The risk of performing cesarean section on preterm infants should be considered especially when adequate facilities to manage high-risk pregnancies (e.g., NICU) are not available. Necrotizing funisitis, which is associated with infectious neonatal complications, occurred in 0.1% of deliveries greater than 20-week gestation [4]. It is difficult to diagnose necrotizing funisitis accurately during the antepartum period. In the current case, we could not identify suspicious findings in the cord before delivery. Thus, this case serves as a reminder of the challenges in diagnosing necrotizing funisitis and performing optimal assessment and management for a fetus with both a systemic inflammatory response and sepsis in utero. Furthermore, an abnormal FHR monitoring pattern in cases complicated by preterm birth is not a sensitive predictor of perinatal sepsis [5]. The current case emphasizes that histopathological examination of the placenta should be utilized for investigating the correlation among FHR monitoring patterns and a fetus whose mother was infected with syphilis according to the degree of intrauterine inflammation.

s not a sensitive predictor of perinatal sepsis [5]. The current case emphasizes that histopathological examination of the placenta should be utilized for investigating the correlation among FHR monitoring patterns and a fetus whose mother was infected with syphilis according to the degree of intrauterine inflammation. An association of CAM with cerebral palsy has been reported [6]. The current neonate was discharged with no evidence of sequelae despite severe CAM and necrotizing funisitis. The current case shows that congenital syphilis still occurs even though antenatal syphilis screening is performed for all pregnant women by public funds in Japan. We stress that it is essential to establish strategies for women with socioeconomic difficulties to encourage ANC, to enroll mothers in ANC earlier, and to improve the quality of programs to control sexually transmitted infections to reduce the frequency of congenital syphilis. These strategies could help prevent cerebral palsy-associated CAM, which may be caused by perinatal infections including syphilis. 4. Conclusions We emphasize that histopathological examination of the placenta should be utilized for investigating the correlation among FHR monitoring patterns and a fetus whose mother was infected with syphilis according to the degree of intrauterine inflammation. Conflict of Interests The authors have no conflict of interests of financial support for this work.

4. Conclusions We emphasize that histopathological examination of the placenta should be utilized for investigating the correlation among FHR monitoring patterns and a fetus whose mother was infected with syphilis according to the degree of intrauterine inflammation. Conflict of Interests The authors have no conflict of interests of financial support for this work. Figure 1 Histopathological examination of the placenta. Numerous spirochetes of Treponema pallidum in areas of the umbilical vein (a), chronic plate (b), deciduas (c) and villi (d) were noted by immunohistochemistry testing, which was performed using an antibody against T. pallidum.

1. Introduction Several hormonal changes are produced during pregnancy that can increasingly predispose to infections of the lower genital tract [1–4]. These infections are associated with a great number of gynecologic and obstetric complications, such as preterm birth [5–13], premature rupture of the membranes [14–16], chorioamnionitis [17, 18], postpartum endometritis [8], inflammatory pelvic disease [19, 20], intrauterine growth retardation [21], and low birth weight [13]. These maternal and perinatological complications could be partly triggered by the local immune response as part of the pathogenic mechanism generated by the infection [22]. However, it is essential to ascertain whether this immunological response of the vaginal mucosa is actually influenced by gestation. There is no literature available referring to the vaginal inflammatory status (VIS) during pregnancy expressed as leukocyte numerical range (LNR) by microscopic observation. The aim of this work was to assess VIS in pregnant women, whether symptomatic or asymptomatic, by leukocyte quantification in relation to the normal microbiota and the presence of candidiasis, trichomoniasis, and bacterial vaginosis (BV) during each trimester of pregnancy (T).

However, it is essential to ascertain whether this immunological response of the vaginal mucosa is actually influenced by gestation. There is no literature available referring to the vaginal inflammatory status (VIS) during pregnancy expressed as leukocyte numerical range (LNR) by microscopic observation. The aim of this work was to assess VIS in pregnant women, whether symptomatic or asymptomatic, by leukocyte quantification in relation to the normal microbiota and the presence of candidiasis, trichomoniasis, and bacterial vaginosis (BV) during each trimester of pregnancy (T). 2. Materials and Methods A thousand two hundred and forty-eight vaginal exudates from pregnant women (231, 359, and, 658 in the first, second and third T, resp.) who were consecutively and prospectively examined at the Obstetrics Clinic at Hospital de Clínicas of the University of Buenos Aires, Argentina, from July 1, 2005 to December 31, 2008, were analyzed. This study was approved by the Hospital Ethics Committee. All the patients underwent a clinical and colposcopic examination and a microbiological study of vaginal exudates. Symptomatic patients showed pruritus and/or an increase in vaginal discharge, which was thick, sticky, purulent, smelly, thin and/or homogeneous depending on the case. The microbiological study of vaginal exudates included the following examinations: smears for Gram and prolonged May-Grunwald Giemsa staining, microscopic wet smear examination with 1 mL of physiologic saline solution,

many countries in sub-Saharan Africa continue to have restricted access to skilled birth attendants [8]. While more than half of all births in sub-Saharan Africa occur without the presence of a skilled attendant, nearly all births in developed nations take place with the assistance of a skilled birth attendant [8, 9]. In many West African countries, maternal and neonatal mortality is highest in rural areas where access to EmOC service is inhibited by vast geographic distances to health facilities and scarce resources [10]. Despite the efforts by the Gambian Government in adopting the primary health care (PHC) strategy, the aim of which was to make health care more accessible and affordable to the majority of Gambians, physical access to health services has been hampered by a rapidly growing population, inadequate financial and logistic support, gross shortage of skilled human resource for health, high staff attrition, and an inefficient referral mechanism [11]. Poverty and ignorance have, in some instances, led to inappropriate health seeking delivery behaviour and contributed to ill health [11]. Although the percentage of population living within 5 km of a primary health care facility has improved over the years, the availability, accessibility, and quality of EmOC services in The Gambia are below required standards [12]. While the number of comprehensive EmOC facilities per 500,000 populations is within the recommended level, the number of basic EmOC facilities is far below the United Nations recommended level [13]. Hence, a large number of women in remote settings still continue to travel some distance to reach a basic EmOC facility or resort to home deliveries. It should therefore be noted that nearly two-thirds of all births in Gambia take place in home settings, mostly attended to by traditional birth attendants (TBAs) and/or family members [13]. Thus, most of the deaths and disabilities that women face in these settings could have been averted if the services of a skilled birth attendant were available and accessible to those women in need. However, The Gambia has yet to formulate a policy on skilled attendance at birth.

2. Materials and Methods A thousand two hundred and forty-eight vaginal exudates from pregnant women (231, 359, and, 658 in the first, second and third T, resp.) who were consecutively and prospectively examined at the Obstetrics Clinic at Hospital de Clínicas of the University of Buenos Aires, Argentina, from July 1, 2005 to December 31, 2008, were analyzed. This study was approved by the Hospital Ethics Committee. All the patients underwent a clinical and colposcopic examination and a microbiological study of vaginal exudates. Symptomatic patients showed pruritus and/or an increase in vaginal discharge, which was thick, sticky, purulent, smelly, thin and/or homogeneous depending on the case. The microbiological study of vaginal exudates included the following examinations: smears for Gram and prolonged May-Grunwald Giemsa staining, microscopic wet smear examination with 1 mL of physiologic saline solution, microscopic wet smear examination with sodium-acetate acetic-acid formalin (SAF)/methylene blue (0.5 mL methylene blue and 0.5 mL SAF) [23], pH determination of vaginal exudates, fishy-odor test with 1 mL of 10% KOH with posterior microscopic wet smear examination, liquid medium culture (modified tioglycolate medium) for Trichomonas vaginalis detection with seven-day incubation period at 37°C in an atmosphere of 5% CO2 [24], and solid medium culture (Modified Columbia Agar) in 5% human blood plates with 48 hour incubation period at 37°C in an atmosphere of 5% CO2. The sample was preserved in Stuart transport medium.

liquid medium culture (modified tioglycolate medium) for Trichomonas vaginalis detection with seven-day incubation period at 37°C in an atmosphere of 5% CO2 [24], and solid medium culture (Modified Columbia Agar) in 5% human blood plates with 48 hour incubation period at 37°C in an atmosphere of 5% CO2. The sample was preserved in Stuart transport medium. The presence of Gram-positive bacilli in the Gram staining and the development of small, point-like, and ά-hemolytic colonies in the solid medium culture (Modified Columbia Agar) in 5% human blood plates, was considered normal or lactobacillary microbiota. Candidiasis detection was performed by microscopic wet smear examination with 1 mL of 10% KOH and by Sabouraud and blood agar culture. The diagnosis of bacterial vaginosis (BV) was performed by means of the Nugent's method through the determination of a score ≥7 in the Gram stain [25], and by the Amsel's method, that is the presence of three or more of the following criteria [26]: “clue-cells” in the Gram stain, pH ≥ 4.5, positive fishy-odor test, and thin and homogeneous vaginal discharge. T. vaginalis detection was performed by direct microscopic examination with physiologic saline solution and SAF/methylene blue, prolonged May-Grunwald Giemsa staining, and modified thioglycolate medium, which was examined daily by wet smears for the detection of motile parasites. Leukocyte quantification (without clumping) was determined by May-Grunwald Giemsa staining and was expressed as LNR per field (400x), by observation of 10 nonadjacent microscopic fields.

T. vaginalis detection was performed by direct microscopic examination with physiologic saline solution and SAF/methylene blue, prolonged May-Grunwald Giemsa staining, and modified thioglycolate medium, which was examined daily by wet smears for the detection of motile parasites. Leukocyte quantification (without clumping) was determined by May-Grunwald Giemsa staining and was expressed as LNR per field (400x), by observation of 10 nonadjacent microscopic fields. 2.1. Exclusion Criteria Patients showing cervicitis in the colposcopic exam were not included in the study. 2.2. Statistical Analysis Fisher's and chi-square tests χ 2 were performed to assess the VIS in relation to microbiota, applying the calculation of adjusted residual (AR) in those cases with P ≤ .05 to identify the LNRs which yielded the significance levels obtained (Epi info 6.04). LNRs with AR of ≥±2 were considered those yielding the significance levels obtained. 3. Results Statistically significant differences resulting from the LNR were observed in the VIS of asymptomatic pregnant patients with normal microbiota compared with that of symptomatic ones: 10 to 15 for the first trimester, <10, 20, to 25 and >25 for the second trimester, and >25 for the third trimester (χ 2: 10.73, P:.03, AR: ±2.32; χ 2: 11.11, P:  .02, AR: ±2.32 ± 2.02 y ± 2.50; χ 2: 10.53, P:  .03, AR: ±3.04, resp.) (Table 1).

nant patients with normal microbiota compared with that of symptomatic ones: 10 to 15 for the first trimester, <10, 20, to 25 and >25 for the second trimester, and >25 for the third trimester (χ 2: 10.73, P:.03, AR: ±2.32; χ 2: 11.11, P:  .02, AR: ±2.32 ± 2.02 y ± 2.50; χ 2: 10.53, P:  .03, AR: ±3.04, resp.) (Table 1). In those patients with candidiasis, statistically significant differences in the LNR were observed in the VIS of asymptomatic patients compared with symptomatic ones: <10 for the first trimester, <10, 15 to 20, and >25 for the second trimester, and <10 and >25 for the third trimester (χ 2: 10.40, P:  .03, AR: ±2.63; χ 2: 17.60, P:  .00, AR: ±2.65 ± 2.58 y ± 2.14; χ 2: 14.24, P:  .00, AR: ±3.47 and ±2.09, resp.) (Table 2). With regard to patients with trichomoniasis (n: 38), statistically significant differences in the LNR were observed between symptomatic patients with LRN ≥ 10 and asymptomatic patients with LNR < 10 in the three pregnancy trimesters altogether (P Fisher:  .01) (Table 3). In women with BV, no statistically significant differences were observed in LNR of asymptomatic patients with respect to symptomatic ones in any of the three trimesters (χ 2: 7.05, P:  .13; χ 2: 3.86, P:  .43; χ 2: 5.17, P:  .27, resp.) (Table 4).

ants (TBAs) and/or family members [13]. Thus, most of the deaths and disabilities that women face in these settings could have been averted if the services of a skilled birth attendant were available and accessible to those women in need. However, The Gambia has yet to formulate a policy on skilled attendance at birth. The purpose of this study was to provide a better understanding of the barriers to timely access to emergency obstetric care in perinatal deaths in a rural setting in The Gambia.

With regard to patients with trichomoniasis (n: 38), statistically significant differences in the LNR were observed between symptomatic patients with LRN ≥ 10 and asymptomatic patients with LNR < 10 in the three pregnancy trimesters altogether (P Fisher:  .01) (Table 3). In women with BV, no statistically significant differences were observed in LNR of asymptomatic patients with respect to symptomatic ones in any of the three trimesters (χ 2: 7.05, P:  .13; χ 2: 3.86, P:  .43; χ 2: 5.17, P:  .27, resp.) (Table 4). 4. Discussion With advancing gestation, a gradual increase in LNR (first trimester: 10 to 15, second trimester: 20 to >25, and third trimester: >25) was observed in pregnant women with lactobacillary microbiota. Therefore, in the last pregnancy trimester, the >25 LNR was statistically significant in the absence of lower genital tract infection, mainly in asymptomatic patients. Other authors such as Yamada et al. [27] also described an increase in the number of leukocytes and interleukin 8 (IL-8) as gestation advanced. Furthermore, Gilbert et al. [28] reported that a significant increase of proinflammatory cytokines (IL-6 and IL-8) is produced in the third trimester. These cytokines act as chemoattractants of polymorphonuclear leukocytes stimulating the expression of prostaglandins in response to delivery preparation. Thus, these interleukins may be considered markers for subsequent normal or preterm labor [29].

ammatory cytokines (IL-6 and IL-8) is produced in the third trimester. These cytokines act as chemoattractants of polymorphonuclear leukocytes stimulating the expression of prostaglandins in response to delivery preparation. Thus, these interleukins may be considered markers for subsequent normal or preterm labor [29]. In addition, Gilbert et al. [28] observed that a decrease in the production of inflammatory cytokines may occur during the second trimester of pregnancy as part of the normal course of pregnancy. Therefore, an increase of those cytokines in this trimester would produce preterm labor probably related to an intrauterine or lower genital tract infection. Furthermore, Gilbert et al. [28] reported that even in the case of intrauterine bacterial infection, the increase of inflammatory cytokines is more pronounced than that expected in a normal pregnancy in response to labor preparation.

In addition, Gilbert et al. [28] observed that a decrease in the production of inflammatory cytokines may occur during the second trimester of pregnancy as part of the normal course of pregnancy. Therefore, an increase of those cytokines in this trimester would produce preterm labor probably related to an intrauterine or lower genital tract infection. Furthermore, Gilbert et al. [28] reported that even in the case of intrauterine bacterial infection, the increase of inflammatory cytokines is more pronounced than that expected in a normal pregnancy in response to labor preparation. According to the results of this study, the patients with symptomatic candidiasis developed colpitis and were associated with LNR greater than 25 in the second and third trimester with positive microscopic examinations, whereas the colposcopy was normal in asymptomatic candidiasis, showing LNR lower than 10 in the three pregnancy trimesters with generally negative microscopic examinations. Furthermore, the patients with symptomatic trichomoniasis developed colpitis and were associated with LNR greater than 10, with positive microscopic examinations. On the other hand, in patients with asymptomatic trichomoniasis, the colposcopy was normal and was only detected by culture, that is, with generally negative microscopic examinations and showing LNR under 10, considering the three trimesters altogether.

with LNR greater than 10, with positive microscopic examinations. On the other hand, in patients with asymptomatic trichomoniasis, the colposcopy was normal and was only detected by culture, that is, with generally negative microscopic examinations and showing LNR under 10, considering the three trimesters altogether. However, no statistically significant differences were observed in any of the LNR of either symptomatic or asymptomatic patients with BV, since this infection usually occurs in the absence of an inflammatory reaction. These results agree with those described in the literature referring to the association of proinflammatory interleukins (IL-6 and IL-8) with vaginitis, whereas in BV, there is an increase of IL-1β (though lower than in vaginitis) but not of the interleukins mentioned above [29, 30]. In addition, Nenadić et al. observed a correlation between leukocyte count and IL-8 concentration and proposed microscopic observation of the number of leukocytes as a useful tool for identifying of patients with vaginal inflammation [31]. It is worth noticing that this work is the first report in the literature referring to vaginal inflammatory status during pregnancy expressed as leukocyte numerical range by microscopic observation, a universally studied methodology used for the diagnosis of infections of the lower genital tract in primary health care.

It is worth noticing that this work is the first report in the literature referring to vaginal inflammatory status during pregnancy expressed as leukocyte numerical range by microscopic observation, a universally studied methodology used for the diagnosis of infections of the lower genital tract in primary health care. 5. Conclusion The vaginal inflammatory status is influenced by vaginal microbiota and does not only depend on the state of pregnancy but also on gestational age. Therefore, the pronounced leukocyte increase in asymptomatic patients in the absence of lower genital tract infection during the third trimester of pregnancy should be highlighted. Table 1 Distribution per trimester of leukocyte numerical ranges in pregnant patients, whether symptomatic or asymptomatic, with normal microbiota. 1st trimester l/f1 2nd trimester l/f1 3rd trimester l/f1 <10 10–15 15–20 20–25 >25 <10 10–15 15–20 20–25 >25 <10 10–15 15–20 20–25 >25 Asymptomatic 91 11 6 4 7 157 15 11 6 9 307 45 11 19 29 Symptomatic 7 4 0 2 0 8 1 1 2 3 11 2 1 0 5 AR2 1.77 −2.32 0.83 −1.98 0.90 2.32 0.13 −0.18 −2.02 −2.50 1.63 0.06 −0.67 0.96 −3.04 −1.77 2.32 −0.83 1.98 −0.90 −2.32 −0.13 0.18 2.02 2.50 −1.63 −0.06 0.67 −0.96 3.04 1Leukocytes/field. 2Adjusted residuals. Table 2 Distribution per trimester of leukocyte numerical ranges in pregnant patients, whether symptomatic or asymptomatic, with candidiasis. 1st Trimester l/f1 2nd Trimester l/f1 3rd Trimester l/f1

−2.50 1.63 0.06 −0.67 0.96 −3.04 −1.77 2.32 −0.83 1.98 −0.90 −2.32 −0.13 0.18 2.02 2.50 −1.63 −0.06 0.67 −0.96 3.04 1Leukocytes/field. 2Adjusted residuals. Table 2 Distribution per trimester of leukocyte numerical ranges in pregnant patients, whether symptomatic or asymptomatic, with candidiasis. 1st Trimester l/f1 2nd Trimester l/f1 3rd Trimester l/f1 <10 10–15 15–20 20–25 >25 <10 10–15 15–20 20–25 >25 <10 10–15 15–20 20–25 >25 Asymptomatic 25 3 0 0 1 27 7 1 1 2 52 8 2 2 7 Symptomatic 8 2 2 2 2 17 2 9 4 9 26 8 7 5 14 AR2 2.63 −0.22 −1.95 −1.95 −1.17 2.65 1.89 −2.58 −1.30 −2.14 3.47 −0.36 −2.00 −1.40 −2.09 −2.63 0.22 1.95 1.95 1.17 −2.65 −1.89 2.58 1.30 2.14 −3.47 0.36 2.00 1.40 2.09 1Leukocytes/field. 2Adjusted residuals. Table 3 Distribution in the three trimesters of leukocyte numerical ranges in pregnant patients, whether symptomatic or asymptomatic, with trichomoniasis. L/f <10 10–>15 Asymptomatic 12 4 Symptomatic 7 15 L/f: Leukocytes per field. Table 4 Distribution per trimester of leukocyte numerical ranges in pregnant patients, whether symptomatic or asymptomatic, with bacterial vaginosis. 1st trimester l/f1 2nd trimester l/f1 3rd trimester l/f1 <10 10–15 15–20 20–25 >25 <10 10–15 15–20 20–25 >25 <10 10–15 15–20 20–25 >25 Asymptomatic 19 1 0 0 0 13 3 2 0 1 29 8 3 0 5 Symptomatic 23 5 1 2 5 34 5 12 5 6 39 5 1 2 9 1Leukocytes/field.

1. Introduction For early detection of arteriosclerotic disease, several parameters for assessing vascular nature have been used. The pulse wave velocity (PWV) is one of these parameters and has been widely used mainly in the field of internal medicine, especially in the prediction of cardiovascular events [1]. The original PWV was measured as the velocity between the carotid and femoral artery. However, carotid-femoral PWV requires somewhat complicated technique to obtain an accurate waveform. Brachial-ankle PWV has been developed, which is reported to be correlated with carotid-femoral PWV and can be measured with a simpler technique. It was also reported that PWV is elevated in preeclamptic women [2]. However, it was pointed out that both carotid-femoral PWV and baPWV were strongly affected by the changes in blood pressure during measurements. For the purpose of using PWV as an indicator of vascular vessel wall stiffness, it is necessary to eliminate the effect of blood pressure change from PWV. Recently, a new index of vascular stiffness, the cardio ankle vascular index (CAVI), was developed. Considered to be an index independent of blood pressure, CAVI is a parameter of adjusted PWV for blood pressure, based on a stiffness parameter β [3] using Bramwell-Hill's formula [4]. CAVI is calculated as follows: (1) CAVI=a(2ρΔP)×ln (PsPd)×PWV2+b, where Ps indicates systolic pressure in brachial artery; Pd, diastolic pressure in brachial artery; ρ, density of blood; a, b, correction coefficient.

Recently, a new index of vascular stiffness, the cardio ankle vascular index (CAVI), was developed. Considered to be an index independent of blood pressure, CAVI is a parameter of adjusted PWV for blood pressure, based on a stiffness parameter β [3] using Bramwell-Hill's formula [4]. CAVI is calculated as follows: (1) CAVI=a(2ρΔP)×ln (PsPd)×PWV2+b, where Ps indicates systolic pressure in brachial artery; Pd, diastolic pressure in brachial artery; ρ, density of blood; a, b, correction coefficient. The usefulness of CAVI in the management of patients with hypertensive, diabetic or renal disorders was previously reported in [5–8], but we found no reports on CAVI in pregnancy. In our current study, we assessed CAVI in normotensive and hypertensive pregnant women. We believe that this is the world's first report on CAVI in pregnancy.

CAVI in the management of patients with hypertensive, diabetic or renal disorders was previously reported in [5–8], but we found no reports on CAVI in pregnancy. In our current study, we assessed CAVI in normotensive and hypertensive pregnant women. We believe that this is the world's first report on CAVI in pregnancy. 2. Subjects and Methods 2.1. Study Population The subjects of this study were 109 Japanese women consisting of 23 nonpregnant healthy women (group A), 45 normotensive pregnant women (group B), 28 pregnant women complicated with established preeclampsia (group C), and 13 pregnant women with chronic hypertension (group D). Eighty-six pregnant women (groups B, C, and D) were patients who visited Mie University Hospital. Twenty-three nonpregnant women (group A) were volunteers including nurses and midwives in Mie University Hospital. All the data in this study were obtained from January 2005 to March 2007. The diagnosis of preeclampsia was made according to the criteria of the International Society for the Study of Hypertension in Pregnancy [9], which includes a blood pressure >140/90 mmHg and >300 mg of protein in a 24-hour urine collection. Chronic hypertension was defined as hypertension diagnosed prior to conception or within the first 20 weeks of pregnancy. The women in group D did not have superimposed preeclampsia. In some cases, the blood pressure recorded during the CAVI examination was lower than the criteria value. The women in group C were normotensive in the first trimester of pregnancy and the blood pressure returned to normal by 12 weeks postpartum. No significant differences were noted in the gestational week and maternal age among the four groups. All the subjects in group A were uncomplicated and uneventful throughout the course of pregnancy and puerperium.

ive in the first trimester of pregnancy and the blood pressure returned to normal by 12 weeks postpartum. No significant differences were noted in the gestational week and maternal age among the four groups. All the subjects in group A were uncomplicated and uneventful throughout the course of pregnancy and puerperium. 2.2. Cardioankle Vascular Index The CAVI is based on the stiffness parameter (β), which expressed blood pressure independent vascular stiffness. Stiffness parameter (β) is calculated as follows: (A) β=[  ln (PsPd)  ]·(DΔD). ln indicates natural log; Ps, systolic pressure; Pd, diastolic pressure; D, vascular diameter; ΔD, change of vascular diameter. The relationship between volume elastic modulus and PWV is expressed by Bramwell-Hill's formula, and the relationship between vessel volume and diameter is given when a cylindrical model is used: (B) Z2=(ΔPρ)·(VΔV) (Bramwell-Hill′s  formula), (C) VΔV=D2ΔD. Z indicates cardioankle PWV (m/sec); ΔP, pulse puressure; ρ, blood density (1.03 × 103 kg/m3); V, volume of blood vessel; ΔV, change of V. The previous 3 formulas (A, B, and C) give the following equation: (2) β=(2ρΔP)·[In (PsPd)]·Z2. Thus, the stiffness parameter (β) can be easily calculated by measures of BP and cardioankle PWV. The new index CAVI is expressed as follows: (3) CAVI=aβ+b. a and b indicate the constants to convert β to a value that is approximate to conventional aortic PWV for each patient compatibility. Thus CVI is calculated in the following formula. (4) CAVI=a(2ρΔP)×ln (PsPd)×PWV2+b.

Thus, the stiffness parameter (β) can be easily calculated by measures of BP and cardioankle PWV. The new index CAVI is expressed as follows: (3) CAVI=aβ+b. a and b indicate the constants to convert β to a value that is approximate to conventional aortic PWV for each patient compatibility. Thus CVI is calculated in the following formula. (4) CAVI=a(2ρΔP)×ln (PsPd)×PWV2+b. CAVI has a compatibility with conventional PWV. CAVI is easily measured using Vasera VS-1000 (Fukuda Denshi, Tokyo, Japan), which is now becoming popular. The CAVI is automatically calculated by the instrument and reflects the stiffness parameter (β). The CAVI has the great merit that it is not influenced by BP, whereas conventional PWV is strongly influenced by BP. However, CAVI might have an error because of the variety of blood density in each case, where in the formula for calculation of CAVI, a standard value of blood density was used. 2.3. Measurement and Statistical Analysis The subject was placed in the supine position and blood pressure: baPWV and CAVI were recorded using a vascular screening system VaSera VS-1000 (Fukuda Denshi, Tokyo, Japan). In all cases, no symptomatic adverse effects were noted by supine position. Figure 1 shows an example of measurement result report in a preeclamptic woman. Mann-Whitney's U-test was used for statistical analysis of comparing values between the two groups. Probability (P) values less than  .05 were considered as significant.

2.3. Measurement and Statistical Analysis The subject was placed in the supine position and blood pressure: baPWV and CAVI were recorded using a vascular screening system VaSera VS-1000 (Fukuda Denshi, Tokyo, Japan). In all cases, no symptomatic adverse effects were noted by supine position. Figure 1 shows an example of measurement result report in a preeclamptic woman. Mann-Whitney's U-test was used for statistical analysis of comparing values between the two groups. Probability (P) values less than  .05 were considered as significant. 3. Results Successful recordings were obtained from all women. The demographic characteristics of the study subjects were shown in Table 1. The values of blood pressure shown in Table 1 were those recorded during the CAVI examination and in some cases the values were lower than the criteria values. Mean gestational age of group B was lower than those in groups C and D, but no significant differences were noted among groups A, B, C, and D in the maternal age, and no significant differences were noted among groups B, C, and D in the gestational age.

and in some cases the values were lower than the criteria values. Mean gestational age of group B was lower than those in groups C and D, but no significant differences were noted among groups A, B, C, and D in the maternal age, and no significant differences were noted among groups B, C, and D in the gestational age. Significant differences were noted between groups A and C, A and D, B and C, B and D in systolic, diastolic and mean blood pressures. As shown in Figure 2, baPWVs in groups A, B, C, and D were 1087 ± 132 cm/sec, 1119 ± 183 cm/sec, 1446 ± 224 cm/sec, and 1614 ± 225 cm/sec, respectively. Significant differences were noted between groups A and C, A and D, B and C, B and D in baPWV. No significant difference was noted between groups C and D in baPWV. CAVI in each group was shown in Figure 3. CAVI in groups A, B, C, and D were 7.1 ± 0.4, 7.1 ± 0.5, 8.4 ± 1.0, and 9.8 ± 1.8, respectively. Similar to baPWV, significant differences were noted between groups A and C, A and D, B and C, B and D in CAVI. Furthermore, CAVI in group D was significantly higher than that in group C.

n each group was shown in Figure 3. CAVI in groups A, B, C, and D were 7.1 ± 0.4, 7.1 ± 0.5, 8.4 ± 1.0, and 9.8 ± 1.8, respectively. Similar to baPWV, significant differences were noted between groups A and C, A and D, B and C, B and D in CAVI. Furthermore, CAVI in group D was significantly higher than that in group C. 4. Discussion To assess the vascular stiffness, several parameters have been used such as carotid-femoral PWV, baPWV, SPβ, and augmentation index (AI). Carotid-femoral PWV is affected by blood pressure and can be adjusted using a compensating nomogram for blood pressure. However, carotid-femoral PWV requires somewhat complicated technique to obtain an accurate waveform. It is also time-consuming, and it takes approximately 15 to 20 minutes for one measurement. baPWV has been developed, which can be assessed with a simpler technique in a short time using a measurement system but it is strongly affected by the changes in blood pressure during measurement. AI is strongly influenced by heart rate, and it is necessary to correct AI by heart rate for the comparison among subjects. SPβ is a parameter which is independent of blood pressure but requires a long time for the measurement, and the measuring equipment for SPβ (sonographic vessel tracking system) is expensive. Moreover, SPβ is a parameter of the local vessel (usually the abdominal aorta or common carotid artery is used for the measurement) and not a systemic vessel parameter.

of blood pressure but requires a long time for the measurement, and the measuring equipment for SPβ (sonographic vessel tracking system) is expensive. Moreover, SPβ is a parameter of the local vessel (usually the abdominal aorta or common carotid artery is used for the measurement) and not a systemic vessel parameter. CAVI is thought to be an index which eliminates these defects in the conventional vascular parameters. It was previously reported that CAVI is independent of blood pressure [10]. The measuring technique of CAVI is easy and CAVI system is not so expensive. It is a noninvasive method and the repeated measurement during pregnancy is possible. In our current study, we found no complications in the subjects related to CAVI measurements, including pregnant women themselves. In preeclamptic women, no atherosclerotic change of the vessel wall was produced, which is different from the vessel wall in chronic hypertensive women. In our current study, both in preeclamptic and chronic hypertensive groups, baPWV was elevated and no significant difference was noted between the two groups. On the other hand, CAVI was also elevated both in preeclamptic and chronic hypertensive groups, but the elevation was milder in the former group. Significant difference was noted between the two groups.

c and chronic hypertensive groups, baPWV was elevated and no significant difference was noted between the two groups. On the other hand, CAVI was also elevated both in preeclamptic and chronic hypertensive groups, but the elevation was milder in the former group. Significant difference was noted between the two groups. In chronic hypertensive women, blood pressure was elevated and (at least partially) structural change in the vessel wall may have been produced, resulting in both baPWV and CAVI elevations. In preeclamptic women, blood pressure was elevated and thus baPWV was elevated. In preeclamptic women, lesser vessel wall structural change was produced, and lesser elevation of CAVI was seen, which is lesser influenced by blood pressure elevation than baPWV. From our results, we believe that we can distinguish the vessel structural change between chronic hypertension and preeclampsia through baPWV and simultaneous CAVI measurements.

wall structural change was produced, and lesser elevation of CAVI was seen, which is lesser influenced by blood pressure elevation than baPWV. From our results, we believe that we can distinguish the vessel structural change between chronic hypertension and preeclampsia through baPWV and simultaneous CAVI measurements. However, if CAVI could be completely independent of blood pressure and could completely reflect the atherosclerotic change of the vessel wall, it might be difficult to explain the reason of significant elevation of CAVI in preeclampsia from normotensive pregnancy. Brodszki and colleagues reported that there were no differences in vessel wall stiffness between women with preeclampsia and healthy controls, through the assessment of common carotid artery, abdominal aorta, and popliteal artery by using ultrasonic echo-tracking system [11]. On the other hand, Ong and colleagues reported that structural change was observed in myometrial radial arteries in women with preeclampsia [12]. Thus, we hypothesize that CAVI might reflect the minimal change of the vessel wall, which caused the elevation of CAVI in preeclamptic women. Further work is in progress to evaluate the usefulness of CAVI in the management of preeclampsia.

ange was observed in myometrial radial arteries in women with preeclampsia [12]. Thus, we hypothesize that CAVI might reflect the minimal change of the vessel wall, which caused the elevation of CAVI in preeclamptic women. Further work is in progress to evaluate the usefulness of CAVI in the management of preeclampsia. 5. Conclusion Recently, CAVI, a new index of vascular stiffness, was developed, which is thought to be an index independent of blood pressure. We assessed CAVI in normotensive and hypertensive pregnant women as the world's first report on CAVI in pregnancy. We believe that we can distinguish the vessel structural change between chronic hypertension and preeclampsia through simultaneous baPWV and CAVI measurements. Acknowledgment One of the authors declares his/her heartfelt appreciation goes to Professor Mitsunao Kobayashi whose comments and suggestions were of inestimable value for his/her study. Figure 1 An example of measurement result report in preeclamptic women. In this case, remarkable elevation of baPWV was noted (baPWV = 1812) but CAVI was not markedly elevated (CAVI = 8.3). Actual report is written in Japanese. Figure 2 baPWV in each group. NS: not significant. Figure 3 CAVI in each group. NS: not significant. Table 1 Demographic characteristics of the study subjects. Group A Group B Group C Group D Normal healthy Normotensive pregnant Preeclampsia Chronic hypertension Number of cases 23 45 28 13 Age (years old) 28.3 ± 7.4 30.3 ± 4.9 31.7 ± 6.0 35.8 ± 3.8 Gestational age (weeks) — 30.4 ± 8.7 33.5 ± 4.8 33.6 ± 3.5 Arterial blood pressure

Table 1 Demographic characteristics of the study subjects. Group A Group B Group C Group D Normal healthy Normotensive pregnant Preeclampsia Chronic hypertension Number of cases 23 45 28 13 Age (years old) 28.3 ± 7.4 30.3 ± 4.9 31.7 ± 6.0 35.8 ± 3.8 Gestational age (weeks) — 30.4 ± 8.7 33.5 ± 4.8 33.6 ± 3.5 Arterial blood pressure Systolic (mmHg) 113 ± 8 111 ± 12 145 ± 17 151 ± 14 Diastolic (mmHg) 70 ± 7 68 ± 9 91 ± 18 96 ± 14

1. Introduction The Millennium Development Goal (MDG) 4 of the United Nations, to reduce under-five mortality by two thirds, and MDG 5, to reduce maternal mortality by three-quarters between 1990 and 2015, are closely related [1]. Every year, 4 million newborn babies die in the first month of life. Of these, 99% occur in low- and middle-income countries [2]. Maternal and perinatal mortality rates are still alarmingly high, especially in sub-Saharan Africa, where little progress has been made over recent decades [3]. More than 515,000 women die as a result of pregnancy-related complications each year, of which half are in Africa [4]. In addition, it is estimated that 880,000 babies are stillborn in sub-Saharan Africa, a fact which remains invisible in the policy agenda [5]. However, the majority of these newborn and maternal deaths are due to preventable causes [2]. Undoubtedly, maternal interventions do benefit children, particularly in relation to newborn survival [6], but preventing deaths in newborn babies has not been a major focus of child survival or safe motherhood programmes, particularly in Africa [2]. Deaths due to childbearing are not only personal tragedies, but can be ruinous to the family, especially to children and the community.

rly in relation to newborn survival [6], but preventing deaths in newborn babies has not been a major focus of child survival or safe motherhood programmes, particularly in Africa [2]. Deaths due to childbearing are not only personal tragedies, but can be ruinous to the family, especially to children and the community. Access to appropriate maternity care including prompt referrals to emergency obstetric care (EmOC) services and skilled birth attendance could significantly reduce both perinatal and maternal mortality and/or morbidity [4, 7]. However, women in many countries in sub-Saharan Africa continue to have restricted access to skilled birth attendants [8]. While more than half of all births in sub-Saharan Africa occur without the presence of a skilled attendant, nearly all births in developed nations take place with the assistance of a skilled birth attendant [8, 9].

ants (TBAs) and/or family members [13]. Thus, most of the deaths and disabilities that women face in these settings could have been averted if the services of a skilled birth attendant were available and accessible to those women in need. However, The Gambia has yet to formulate a policy on skilled attendance at birth. The purpose of this study was to provide a better understanding of the barriers to timely access to emergency obstetric care in perinatal deaths in a rural setting in The Gambia. 2. Model The fundamental constructs in understanding the causes of maternal deaths is the “three delays” model developed by Thaddeus and Maine [14]. These are (1) delay in recognising danger signs/decision to seek care, (2) delays in reaching a medical facility, and (3) delay in receiving appropriate care once a facility is reached. We adopted the “three delays” model as a framework in assessing the barriers to obstetric care in survivors of severe obstetric complications. Twenty survivors with stillbirth or perinatal death were question. The “three delays” model provides a valuable theoretical account in examining composite interactions between women's ability to recognise danger signs during pregnancy and child birth, the problems they encounter in reaching an EmOC facility and the quality of care received at the facility. The model identifies key time periods in pregnancy and childbirth during which delays can occur that have direct consequences on both maternal and neonatal survival. Because perinatal and maternal morbidity and mortality share many risk factors, the “three delays” model may assist in perinatal health outcome assessment. The model has been applied previously in analyses of newborn death in Tanzania and Uganda [15, 16].

have direct consequences on both maternal and neonatal survival. Because perinatal and maternal morbidity and mortality share many risk factors, the “three delays” model may assist in perinatal health outcome assessment. The model has been applied previously in analyses of newborn death in Tanzania and Uganda [15, 16]. 3. Material and Methods 3.1. Study Setting/Area The study was carried in Bansang hospital located on the south bank of The Gambia in the Central River Region (CRR). The CRR has an under-five mortality of rate of 165/1000, the highest in the country [11]. Bansang hospital is the oldest rural hospital and serves the eastern part of the country including the catchment areas of Upper River Region (URR), covering about one-third of the country's population, mainly subsistence farmers. The hospital, which was purposively selected for this study serves as a referral point for nearly sixteen peripheral health centres that provide first-level care to the inhabitants of CRR and URR. Basic and comprehensive EmOC service is not available at any of these health centres except one, which is not fully operational. Thus, Bansang hospital is the only facility that provides comprehensive EmOC services in this part of the country. Though the catchment area population of this comprehensive EmOC facility is below the 500,000 minimum outlined in the UN guidelines, reaching this hospital still remains a challenge for remote rural dwellers. Poor road conditions and/or transportation networks and river crossings are practical factors that compromise access and utilisation of emergency obstetric care services. Most of the maternity admissions and deliveries in this hospital are high risk and the bulk of the referrals are during labour. However, there are no Gambian medical doctors or gynaecologists in this hospital and as such they depend to a large extent on the expatriate doctors on technical missions. Since all the specialist doctors are non-Gambian, the withdrawal of such assistance could affect the quality of obstetric care services in this hospital. The institutional fresh stillbirth rate was calculated at 49.6 per 1000 births [17]. In addition to operating as a referral hospital, it also provides outpatient services as well as static and mobile reproductive and child health services. Antenatal care is provided by nurses and or midwives within here and on outreach clinic.

itutional fresh stillbirth rate was calculated at 49.6 per 1000 births [17]. In addition to operating as a referral hospital, it also provides outpatient services as well as static and mobile reproductive and child health services. Antenatal care is provided by nurses and or midwives within here and on outreach clinic. 3.2. Study Population/Design Individual in-depth interviews were carried out with women who sought obstetric care and delivered in Bansang hospital a month prior to data collection. The study population included surviving women with the following diagnostic criteria antepartum haemorrhage (placenta previa/abruption placenta n = 9), pregnancy-induced hypertension (pre-eclampsia/eclampsia n = 4), prolonged and/or obstructed labour (n = 5), and severe anaemia (n = 2). Key informants (family member(s) and traditional birth attendants, TBAs) were also interviewed. A total of 20 women who delivered between April 2010 and June 2010 were purposively selected based on the characteristics similar to women from a preceding study [17], resource availability, and geographic location. All of them had a stillbirth (n = 19) or neonatal death (n = 1). Prior to discharge from the hospital, individual consent for participation in the study was obtained from the women. In addition, telephone numbers and permanent addresses were also provided to facilitate the data collection process. The principal investigator (AJ), assisted by experienced community health workers, visited the selected participants in their homes and administered semistructured interview guides in the local language they understood in the presence of relative (s) and TBAs who also acted as key informants.

e data collection process. The principal investigator (AJ), assisted by experienced community health workers, visited the selected participants in their homes and administered semistructured interview guides in the local language they understood in the presence of relative (s) and TBAs who also acted as key informants. The interviews concentrated on the following respondents' ability in recognising obstetric complications, care seeking/decision making procedures, and the processes of getting care from home to the hospital. The interviews were translated into English and transcribed verbatim using content analysis [18]. The main topics identified were examined for similarities and phrases that represented similar topics or ideas were further clustered together in categories. Furthermore, typical statements were used for citation. Data analysis was done manually and continuously during the data collection period. Informed consent was gained from all respondents before commencement of interviews. This entailed an explanation of the purpose of the study, a guarantee of confidentiality relating to the information to be given and assurance that participation would not have any negative bearing on the availability and provision of health care to them or their families. The ethics committee of Norway and the Joint Gambia Government/Medical Research Council Review board approved the study.

onfidentiality relating to the information to be given and assurance that participation would not have any negative bearing on the availability and provision of health care to them or their families. The ethics committee of Norway and the Joint Gambia Government/Medical Research Council Review board approved the study. 4. Results 4.1. Characteristics of Women The mean age of the women was 29 years, with a range of 18–40 years. The mean parity was 4, and all except one attended antenatal care. None of the women ever attended formal school and all were married except one. All the twenty women delivered at the study site but only three sought initial care at another health centre. Of the twenty women, 19 had a stillbirth while one had an early neonatal death before discharge from the hospital. The first delays occurred in 30% of the cases, second delays in 50%, and the third delays occurred in 20% of the cases (Table 1).

the study site but only three sought initial care at another health centre. Of the twenty women, 19 had a stillbirth while one had an early neonatal death before discharge from the hospital. The first delays occurred in 30% of the cases, second delays in 50%, and the third delays occurred in 20% of the cases (Table 1). 4.2. Recognising Danger Signs during Pregnancy and Labour Awareness and/or recognition of an obstetric complication among pregnant women and within their communities is the first step to initiating appropriate and timely referrals to essential obstetric and newborn care, hence reducing the time needed to make a decision to seek care, which is called the “first delay.” However, most women and families in developing countries, particularly in rural settings seldom know in detail the danger signs of pregnancy and childbirth, or they misinterpret or “hope for the best.” As a result many do not seek health care because they do not know that they have a problem. A severely anaemic woman narrated her ordeal: “…When I was pregnant I usually feel tired especially during late evenings…I have had sleepless nights all the time…previous pregnancies I was always strong but this time I am weak and can't do anything…I fell down and hurt my legs but was taken to my mothers' house to rest.”

severely anaemic woman narrated her ordeal: “…When I was pregnant I usually feel tired especially during late evenings…I have had sleepless nights all the time…previous pregnancies I was always strong but this time I am weak and can't do anything…I fell down and hurt my legs but was taken to my mothers' house to rest.” Giving her version of the above scenario the mother-in-law commented: “In the village women are responsible for all household chores…I am sure the tiredness my daughter-in-law got when she was pregnant is due to tedious domestic work…she work [sic] from morning to evening without rest…And …this is not good for a pregnant woman.” An informed individual is better placed to make reasonable decisions in emergency situations. At times women reflect on experiences during their last pregnancy and try to apply this to circumstances surrounding them. A 29-year-old woman who had antepartum haemorrhage narrated: “Before labour started…I had a tensed abdomen but there was no pain and the baby is not moving as usual…I have no idea what the problem was…I have never given birth at a health centre or hospital so I thought everything was going to be okay.”

urrounding them. A 29-year-old woman who had antepartum haemorrhage narrated: “Before labour started…I had a tensed abdomen but there was no pain and the baby is not moving as usual…I have no idea what the problem was…I have never given birth at a health centre or hospital so I thought everything was going to be okay.” However, some participants were well informed and aware of an imminent danger threatening the life of the pregnant woman. Persuading and getting the consent of other family members to seek help outside of the home is sometimes not as difficult as one might think. Narratives of a sister-in-law to a 32-year-old woman with obstructed labour and history of previous caesarean section (C/S) supported this: “The problem started at home when she was in labour…I recognized that she was in pain for a very long time with no improvement…Her (referring to the woman) abdomen was very big and tensed…and I feared that she might not be able to deliver on her own again…she have [sic] had two previous operations (C/S) so I thought she would not be able to deliver on her own again…Her abdomen was too big…the pain was increasing yet there was no sign of delivery and everybody was worried…So I suggested that we go to the hospital so that they could assist to deliver her through operation (C/S) once more…The traditional birth attendant was present and everybody agreed.”

to deliver on her own again…Her abdomen was too big…the pain was increasing yet there was no sign of delivery and everybody was worried…So I suggested that we go to the hospital so that they could assist to deliver her through operation (C/S) once more…The traditional birth attendant was present and everybody agreed.” 4.3. Socio-Cultural Belief/Family Decision-Making Social factors tend to influence women's likelihood to seek health care during pregnancy and labour. In rural areas of The Gambia the decision to seek early delivery at a health centre and/or hospital is usually influenced by mother-in-laws. Visiting the hospital early in labour was viewed by most mother-in-laws as unnecessary; they would prefer to prepare traditional herbs while waiting for established labour. This they believed will make the pelvic bones flexible thus facilitating quick and safe delivery even without going to the hospital. A 29-year-old woman with obstructed labour narrated her encounter with the mother-in-law: “My mother-in-law argued that I should not report to the hospital when I started having abdominal pains…that…it was too early to go and as a result we may stay at the hospital for a long time before I deliver…she decided to cook traditional herbs for me while waiting for established labour…she believed the herbs would be useful in making the delivery quick and less painful.”

hospital when I started having abdominal pains…that…it was too early to go and as a result we may stay at the hospital for a long time before I deliver…she decided to cook traditional herbs for me while waiting for established labour…she believed the herbs would be useful in making the delivery quick and less painful.” Even in situations where awareness of danger signs during pregnancy and labour helps women and families to recognize a complication, this does not automatically translate into a prompt decision to seek help outside of the home. How severe they consider the problem is equally an important determinant of deciding when to act. Mother of an 18-year-old single eclamptic woman explained her ordeal: “She got pregnant before the traditional married ceremony was done…she hide [sic] the pregnancy and never attended antenatal care. She developed swollen legs latter on…and started complaining of severe headache...and heart burn…she was not feeling well the day she went into labour since the early hours of the morning…but I felt it was too early to go to the hospital so I prepared local medicine for her. We remained home until very late in the evening when she started fitting…and the baby died in the womb.”

of severe headache...and heart burn…she was not feeling well the day she went into labour since the early hours of the morning…but I felt it was too early to go to the hospital so I prepared local medicine for her. We remained home until very late in the evening when she started fitting…and the baby died in the womb.” This is another testimony from a husband of woman with antepartum haemorrhage: “…She (referring to his wife) started bleeding in the night…but we thought everything will be all right even though I know that bleeding in pregnancy is dangerous…while waiting for sunrise…We gave her holly [sic] water…but this does [sic] not help. After the local remedies failed…we thought the best thing to do was to take her to the hospital…”.

e) started bleeding in the night…but we thought everything will be all right even though I know that bleeding in pregnancy is dangerous…while waiting for sunrise…We gave her holly [sic] water…but this does [sic] not help. After the local remedies failed…we thought the best thing to do was to take her to the hospital…”. 4.4. Transportation, Distance, and Road Infrastructure The mode of transport to the hospital is a fundamental determinant in health seeking. This often presented a threat to the survival of women and newborns in developing nations. The common mode of transport in villages is by horse and donkey carts or at times bicycles. Most of these villages are in very remote rural areas with poor roads and far from regular routes for commuter taxis and/or private cars. The only available mode of transport is usually inconvenient for a woman in labour. Hence donkey carts were classified as too slow to address the immediate need of a woman in emergency. This is what a woman with haemorrhage had to say: “We left home late in the evening with a donkey cart in order to get to the main high way…this is the only means of transport we have. It was moving very slowly because the road condition was very bad…and…the place was getting dark. We reached the main high way after three hours…all this time I was bleeding with severe abdominal pains. Luckily…shortly after reaching the high way one private car gave us a lift to the hospital…but the baby was not born alive.”

moving very slowly because the road condition was very bad…and…the place was getting dark. We reached the main high way after three hours…all this time I was bleeding with severe abdominal pains. Luckily…shortly after reaching the high way one private car gave us a lift to the hospital…but the baby was not born alive.” Even where horse carts are the common mode of transportation which is seemingly faster than donkeys, the roads may by be submerged in water, especially in the rainy season. The roads may also be so bumpy and paddy that the horse has to move very slowly so that the woman seeking health care might not endure severe pains that may worsen her condition; “It was during the rainy season and the roads were bad…like…paddy and full of stagnant water. I have to move the horse very slowly and carefully to avoid being stuck in the mud…this caused a lot of delay…but have to continue because I was worried and needed help. After a long and tiring journey...almost three hours…we arrived at the main road…but have [sic] to wait for another one hour before getting a commercial vehicle to the hospital” (husband of 32-year-old woman with obstructed labour).

…this caused a lot of delay…but have to continue because I was worried and needed help. After a long and tiring journey...almost three hours…we arrived at the main road…but have [sic] to wait for another one hour before getting a commercial vehicle to the hospital” (husband of 32-year-old woman with obstructed labour). Even after surmounting a rough ride on a horse or donkey cart to get to the nearest health centre with a maternity, timely access to appropriate obstetric care is further hampered by inadequate logistics for evacuation of emergencies. Results from the interviews pointed out that there were instances when no ambulances were available for evacuation of patients because the one available had embarked on another referral. As a result, when a woman with an obstetric complication is referred to a second level, the family has to find alternative means of evacuation thus, costing more time and money. Others have to desperately resort to using the cart whatever the distance and time it may take which could lead to further complication. Narratives of husband of a 46-year-old woman with antepartum haemorrhage needing evacuation: “We used a horse cart to get to the nearest health centre…there I thought everything would be okay…but...that was not the case.…We were told to proceed to the next facility…but the ambulance was not there as it has taken another patient to the hospital…I have no option but to continue the journey with my horse…it was dark and has [sic] no money to pay for a taxi…after four hours we arrived at the second health facility…there she was examined and put in an ambulance to Bansang hospital…unfortunately, the baby did not survive.”

has taken another patient to the hospital…I have no option but to continue the journey with my horse…it was dark and has [sic] no money to pay for a taxi…after four hours we arrived at the second health facility…there she was examined and put in an ambulance to Bansang hospital…unfortunately, the baby did not survive.” Access problems still confronted women suffering from obstetric complications even if an ambulance was available at the first referral point en route to the main rural hospital. As many of the peripheral health centres in the study are on other side of the river, ambulances have to be ferried. However, this is usually hampered by high tides especially during the rainy season or mechanical problems with the ferry. It is not uncommon for rivers in the central and upper river regions in The Gambia to overflow their banks during mid rainy season hence rendering ferry service redundant. A 40-year-old woman explained her experience: “When we arrived at the river the ferry that was expected to take us to the other side was not there…some waiting passengers said it had a mechanical problem since morning…at that time I can't [sic] bear the pain any more…and the only means of crossing the river was to use a paddle canoe…and this caused a big delay.”

: “When we arrived at the river the ferry that was expected to take us to the other side was not there…some waiting passengers said it had a mechanical problem since morning…at that time I can't [sic] bear the pain any more…and the only means of crossing the river was to use a paddle canoe…and this caused a big delay.” Another woman who had haemorrhage explained her ordeal during evacuation when they reached a crossing point. “I started seeing blood in the afternoon after lunch…my husband took me to the main road with a donkey cart to look for transport to the health centre…we waited for a long time in vain as all the commercial vehicles in the area have [sic] already left for Basse…he then decided to continue the journey with the donkey to the nearest health centre. When we reached the health centre…I was examined by the nurse but she said they will refer me to Basse Health Centre [major health centre] 25km away. When we reached the crossing point the ferry was not working because the whole area was overflowed and…the water was moving very fast…we resorted to using a paddled canoe to get to the other bank where another ambulance was waiting.”

ut she said they will refer me to Basse Health Centre [major health centre] 25km away. When we reached the crossing point the ferry was not working because the whole area was overflowed and…the water was moving very fast…we resorted to using a paddled canoe to get to the other bank where another ambulance was waiting.” 5. Cost The respondents explained how lack of money influences pregnant and labouring women's health seeking behaviour. Women attend antenatal care and if problems are observed they are advised to deliver at the hospital. However, the husbands and/partners may not have enough money to be able to pay for transport and/or emergencies as well as extra money for food while at the hospital. The only alternative is to reach out to other people in the village for help, which does contribute to delays in seeking health care. An 18-year-old woman with obstructed labour narrated: “My husband took me to the village health centre…but the nurses said the problem cannot be managed there…so they have to take me to the main hospital. At that time we don't [sic] have any money with us…my husband went back home…not very far…to negotiate for money while the ambulance was waiting. He got only D100.00 (equivalent US$3.8) as it was during night and most people were sleeping.”

e problem cannot be managed there…so they have to take me to the main hospital. At that time we don't [sic] have any money with us…my husband went back home…not very far…to negotiate for money while the ambulance was waiting. He got only D100.00 (equivalent US$3.8) as it was during night and most people were sleeping.” Another woman with obstructed labour had this to say: “…when I was in labour what prevented me from seeking health care early was the lack of money to hire a vehicle…we live in a very remote village and only managed to reach the high way on a donkey cart…I have to rely on my husband to make arrangements (…borrowing money from the nearby village) to pay for a car to drive us from the highway to the hospital.” In a related case, a husband of a 40-year-old woman with severe pre-eclampsia living 85 km away from the main hospital explained his frustrations in raising money for emergency services: “When the decision was reached to take her (his wife) to the hospital…I did not have any money on me...I had to borrow some money from a neighbour that night which was not enough…it took me two hours to raise this amount…and to beg the taxi driver to assist and accept the money in order to safe [sic] wife's life.”

ces: “When the decision was reached to take her (his wife) to the hospital…I did not have any money on me...I had to borrow some money from a neighbour that night which was not enough…it took me two hours to raise this amount…and to beg the taxi driver to assist and accept the money in order to safe [sic] wife's life.” In some instances husbands were compelled to sell their assets to ensure that the woman gets to the hospital. A 20-year-old woman with severe pre-eclampsia/haemorrhage narrated: “He (her husband) gave us the little money he had on him…to raise more money he initially sold his goat and latter on his sheep…We are farmers so we barely have enough for the family upkeep not to mention about [sic] emergency funds”. A mother of a 26-year-old woman with antepartum haemorrhage also commented on the unavailability of money during emergencies: “…there used to be a community ambulance in this village that transport [sic] all women in labour to the hospital…but it had a breakdown at the time she (meaning her daughter) went into labour. The husband had no money at that time…and has [sic] to go out in the village to borrow money from shopkeepers in order to hire a bush taxi to get to the hospital…this was late in the night…”

ansport [sic] all women in labour to the hospital…but it had a breakdown at the time she (meaning her daughter) went into labour. The husband had no money at that time…and has [sic] to go out in the village to borrow money from shopkeepers in order to hire a bush taxi to get to the hospital…this was late in the night…” 5.1. Blood Transfusion Timely availability and access to life-saving interpositions such as blood transfusion are an essential component for the reduction of maternal and perinatal mortality. However, the hospital from which our patients were recruited did have a blood bank, but was not adequately stocked to meet the demands of the population. As a result, patients who require transfusions are compelled to obtain transfusion blood mostly from friends, relatives, and/voluntary donors, failure of which prolongs access to timely intervention. Scarcity of blood causes delays in receiving appropriate care even if family members are able to provide the required quantity, because of the time they need to go outside of the hospital in search of suitable donors. Thus, the timely delivery of appropriate emergency obstetric care services is further compromised. The following are testimonies from key informants during their stay at the hospital in their pursuit for potential blood donors: “…her blood was taken and sent to the machine (meaning laboratory)…latter on I was informed by the nurses that she lacks blood and will need blood transfusion…but…no blood was given. After waiting in vain…I went back to the nurses to enquire about the blood because my daughter-in-law was not improving…One nurse told me that the only thing to save her life is blood…so I should bring a blood donor as early as possible. I am old and can't donate blood…so I sent for her husband” (mother-in-law of a 31-year-old anaemic woman).

went back to the nurses to enquire about the blood because my daughter-in-law was not improving…One nurse told me that the only thing to save her life is blood…so I should bring a blood donor as early as possible. I am old and can't donate blood…so I sent for her husband” (mother-in-law of a 31-year-old anaemic woman). A husband of a 20-year-old woman who had eclampsia and antepartum haemorrhage also had this to say: “After she (meaning his wife) delivered I was informed that she lacks blood…at that moment there was no blood in the hospital…I donated one bottle of blood but that was not enough…it was very difficult for me to get another suitable blood donor because I don't know anybody in this area…I latter sent a message for my younger brother from the village 8 km away who also came and donated one bottle of blood.”

nt there was no blood in the hospital…I donated one bottle of blood but that was not enough…it was very difficult for me to get another suitable blood donor because I don't know anybody in this area…I latter sent a message for my younger brother from the village 8 km away who also came and donated one bottle of blood.” In rare circumstances women in critical conditions would be transfused a pint of blood on loan which the relatives have to replace. Getting a suitable donor is tiring and frustrating for the relatives as the people they usually bring are normally not suitable candidates, thus have to resort to the negotiating table with the nurses; “…the nurses told me that my wife lacked blood and needed to be transfused…I was then instructed to provide a blood donor…my son volunteered to donate but unfortunately he was not in the same blood group…after a long negotiation with the nurses…one bag of blood was provided and transfused on loan. … I have to go and look for donors to replace the blood…this is not an easy task for me” (husband of a 30-year-old woman with antepartum haemorrhage).

volunteered to donate but unfortunately he was not in the same blood group…after a long negotiation with the nurses…one bag of blood was provided and transfused on loan. … I have to go and look for donors to replace the blood…this is not an easy task for me” (husband of a 30-year-old woman with antepartum haemorrhage). A mother-in-law explained her encounter with the nurses at the hospital when the daughter-in-law with antepartum haemorrhage was found to be requiring blood; “…at the hospital we were told that her blood is very low so we should go and look for a blood donor…at that time her husband was not around and we are too old to donate…We begged them (nurses) to assist us safe [sic] her life by getting blood…latter on during the day one bottle of blood was provided and mounted on as a loan” (mother-in-law of 29-year-old woman with APH).

low so we should go and look for a blood donor…at that time her husband was not around and we are too old to donate…We begged them (nurses) to assist us safe [sic] her life by getting blood…latter on during the day one bottle of blood was provided and mounted on as a loan” (mother-in-law of 29-year-old woman with APH). 6. Discussions The current study applies the classic “three delay” model to explore delays of care in stillbirths and neonatal deaths. The majority of perinatal deaths that occurred in the current study were due to delay in reaching a health facility, transport/cost delays (delay 2). We also found household delays (delay 1) as the second important determinant of perinatal deaths. Most of the stillbirths were reported to be macerated, suggesting that the deaths probably occurred even before reaching the hospital. The first and second delays are significant because births still occur at home and access to appropriate care is still a major problem [15]. Thus, a call for strengthening community programmes seems warranted for improved newborn care in low-income countries [6], such as The Gambia. Our findings on the contribution of delays on perinatal deaths differ from studies conducted in Tanzania [15] and Uganda [16], where most newborn deaths were found to be associated with the first and third delays.

nity programmes seems warranted for improved newborn care in low-income countries [6], such as The Gambia. Our findings on the contribution of delays on perinatal deaths differ from studies conducted in Tanzania [15] and Uganda [16], where most newborn deaths were found to be associated with the first and third delays. Obstetric complications are similar around the world, but the risk of death is not. Ninety-nine of all maternal and perinatal deaths occur in developing countries [19]. Thus, women must have timely access to good quality obstetric care services at all times because complications are virtually inconceivable to anticipate and hard to forestall. In addition, families and/or communities must both realise and respect the available services. In our study, the inherent factors that contributed to stillbirths were similar to the classical list of the “three delays” in accessing EmOC [14]. However, in most cases these circumstances contribute first to the death of the child and later eventually also to the death of the mother. In stillbirths the women who survive and her relatives can tell their stories. The organisation of health care in poor countries is often fragmented based on subsided care packages. We emphasise that stillbirths share the same dilemmas as maternal deaths. Initiatives addressing mother and foetus as “one” and as such the pathways to solutions should be informed. The results of the current study illustrates that initiatives to make a change for maternal health would save many more lives than counted in a reduction of maternal mortality rate.

e dilemmas as maternal deaths. Initiatives addressing mother and foetus as “one” and as such the pathways to solutions should be informed. The results of the current study illustrates that initiatives to make a change for maternal health would save many more lives than counted in a reduction of maternal mortality rate. Medical care for women with obstetric complications begins with the recognition of danger signs [14]. Delays in recognizing danger signs, in seeking, reaching, and obtaining appropriate maternity care are key elements in maternal and perinatal deaths. However, recognition of the severity of a complication is an important determinant of the decision to seek help outside home. Pregnancy and childbirth are often considered as a natural process, thus signs and symptoms of complications are seldom recognized as reasons for concern. Thus, ineffective decision-making at the family level was identified as an important barrier in accessing timely EmOC in the present study. Similar findings have been reported elsewhere [20]. Birth preparedness and complication readiness (BP/CR), a strategy that promotes timely access and use of a skilled maternal and newborn care especially during childbirth, reduce delays in obtaining this care. Thus, birth preparedness motivates people to plan to have a skilled provider at every birth. If women and their families make the decision to seek health care before the onset of labour, and successfully follow through this plan, the woman will certainly reach and receive appropriate care before developing any potential complication during childbirth, thus averting the first two delays [21]. Even though women recognize danger signs, prompt decision to seek help outside of the home was not automatic as highlighted in the findings of the present study. Thus, it would be reasonable to speculate that failure in recognizing the severity of symptoms of obstetric complications could be one key reason for delay in seeking appropriate medical care in rural Gambia.

ecision to seek help outside of the home was not automatic as highlighted in the findings of the present study. Thus, it would be reasonable to speculate that failure in recognizing the severity of symptoms of obstetric complications could be one key reason for delay in seeking appropriate medical care in rural Gambia. Women's liberty and status are also reported to have an influence in the utilization of obstetric services [22]. Low levels of female education and lack of empowerment and autonomy to make decisions deter women from seeking obstetric care in time. In the current study, women's choice to deliver at the hospital was heavily influenced by their husbands. It is not the pregnant woman herself who often decides on the place of birth but rather her family. In rural areas of The Gambia the husband and/or mothers-in-law are the most influential decision-makers. They make the final decision even if others offer their opinion. However, traditional birth attendants also played a crucial role in facilitating swift decision to seek health care. They tended to refer to their experiences with previous women in relation to pregnancy and childbirth in order to convince the family to seek health care. In resource-poor countries, particularly in rural settings with limited access to healthcare, traditional birth attendants are an essential source of obstetric care [23]. Thus, a programme geared towards educating families, women, and traditional birth attendants to recognize danger signs during pregnancy and childbirth as well as dangers associated with delay in seeking health care in the event of an obstetric complication seems warranted.

re an essential source of obstetric care [23]. Thus, a programme geared towards educating families, women, and traditional birth attendants to recognize danger signs during pregnancy and childbirth as well as dangers associated with delay in seeking health care in the event of an obstetric complication seems warranted. Timely referral during an obstetric emergency is a critical element in maternal and child health survival strategies. Once a decision is made that a complication needs medical intervention, availability of transport and easy accessibility to a facility with EmOC capabilities become crucial. In rural areas of The Gambia antenatal care is provided by outreach teams from peripheral health centres many of which don't have a maternity with appropriate surgical capacity. Thus, health facilities with appropriate obstetric capacity are usually at considerable distance from women who develop obstetric complications. In our study, traditional birth attendants reiterated that they received women who needed referral to the hospital, but delay in seeking and getting care was compounded by the lack of reliable transport and poor road network. The significant contribution of transport and mobility to development and the livelihood of people is widely recognized. Thus, appropriate, affordable, and timely transport in accessing health care should be recognized as a major developmental goal. Proponents argued that improved transportation and mobility directly improve access to health services and indirectly enhance aspects of social and economic development that impact positively on health. However, transportation to health facilities is not often seen as an essential element of access to care, but an individual challenge to overcome.

ed transportation and mobility directly improve access to health services and indirectly enhance aspects of social and economic development that impact positively on health. However, transportation to health facilities is not often seen as an essential element of access to care, but an individual challenge to overcome. At the time of an obstetric emergency, every moment of delay in seeking and receiving skilled care increases the risk of maternal deaths/disability and stillbirths. However, unavailability and high cost of transportation, poor roads, and difficult river crossing and transportation from remote villages may increase the time to reach a hospital [24]. Long distance to the hospital and lack of appropriate means of transportation are major problems affecting the referral system in rural areas of The Gambia. During the rainy season most of the roads become difficult to navigate. For many remote rural dwellers in the Gambia, the uneven distribution of EmOC facilities means slow and long journeys that contribute to poor pregnancy outcomes and increased morbidity and mortality. Distance from health facilities and transport difficulties such as finding a motorised vehicle for patient transportation were found to be barriers in access of care [25]. Many preventable maternal and perinatal deaths continue to happen in remote rural settings in The Gambia.

mes and increased morbidity and mortality. Distance from health facilities and transport difficulties such as finding a motorised vehicle for patient transportation were found to be barriers in access of care [25]. Many preventable maternal and perinatal deaths continue to happen in remote rural settings in The Gambia. It has been documented that a functioning continuum of care between the home and the hospitals is needed to minimize pernicious delays and efficaciously connect pregnant women to skilled obstetric care [24]. Even though families in remote areas made a prompt decision to seek care poor, terrain and transportation difficulties posed unnecessary delays in reaching adequate care. Women trying to reach far-off facilities often fail, and those with life-threatening complications may suffer en route [14]. Evidence showed that a lack of mobility and appropriate transportation during an obstetric emergency contributes significantly to maternal and neonatal mortality in rural Gambia [26] and in Ghana, Nigeria, and Sierra Leon [14]. Earlier studies conducted in rural West African communities demonstrated transportation difficulties and distance from health facilities as major obstacles in accessing EmOC services [27]. Thus, it would be reasonable to presume that improved transportation during an obstetric emergency could directly ameliorate access to EmOC facilities that impact positively on maternal and neonatal health. We therefore believed that forging partnership between rural communities and local transport owners could be one key strategy in overcoming transport issues during emergency. This scheme was implemented in north west Nigeria and had led to significant diminution in transportation cost and maternal and perinatal mortality [28]. Though the availability of EmOC and motorised transport are vital for maternal and child survival during complications, women in remote settlements should be encouraged to move nearer to a health facility with a maternity as the delivery date approaches, and to stay there until the baby is born [29]; like in a maternal shelter or waiting home.

f EmOC and motorised transport are vital for maternal and child survival during complications, women in remote settlements should be encouraged to move nearer to a health facility with a maternity as the delivery date approaches, and to stay there until the baby is born [29]; like in a maternal shelter or waiting home. Cost is a key factor in determining the choice of place of delivery during an emergency, and it contributes to the first delay in seeking care outside home [20]. The cost of maternity care and the inability to meet the cost is a significant barrier to EmOC access, thus an important determinant of maternal and perinatal mortality [30]. Although maternity care is officially provided free of charge in public hospitals in The Gambia, indirect and/or opportunity costs still remain a substantial obstacle in timely access to obstetric care in rural settings. In their review article, Ensor and Cooper [25] considered distance and time as indirect and opportunity costs that influence uptake of health services in developing countries. Women's access to EmOC is further compromised if the care being sought is going to be very costly. The distance and time involved in seeking health care often determine how much cost a family may incur. Women seldom travel alone during an obstetric emergency; so accompanying family members increase the time lost in domestic work, cost of transportation and food during their stay in the hospital [22]. Thus, critical decisions are made to spend scarce resources in saving the life of women and babies. Poverty and high costs of health services has been reported by Essendi et al [20] as important barriers in the utilization of maternal health services among the poor. This is consistent with findings in the current study. Most families intended to access formal delivery services in time but, failed because they are poor and cannot afford the cost to pay for commercial vehicles at the time a decision was made during an emergency. In the Gambia, a majority of the rural poor lack stable and regular sources of income. As a result, they had to borrow money from shopkeepers and/or relatives to meet the cost of transportation and food while at the hospital. Others resorted to selling their farm products or animals. This is consistent with findings from studies conducted elsewhere in low-income countries [31].

regular sources of income. As a result, they had to borrow money from shopkeepers and/or relatives to meet the cost of transportation and food while at the hospital. Others resorted to selling their farm products or animals. This is consistent with findings from studies conducted elsewhere in low-income countries [31]. In many developing countries the direct costs of transport constitute a significant proportion of the total health care expenditure. In northeast Brazil [32] the costs of accessing health facilities have been estimated to represent 25% of the total expenditure on health. The current study highlighted lack of money as an impediment in making prompt decision to seek care and in reaching a medical facility. This is corroborated by findings from a previous study in rural Gambia [26] and Bangladesh [33, 34]. The abolition of maternity fees could be part of the strategies to improve outcome and utilization of services, but this cannot be successful without complementary supply-side and increase investment. Like any health intervention, EmOC requires resources; personnel, time, equipment, drugs, and supplies; and resources are always limited [35].

ity fees could be part of the strategies to improve outcome and utilization of services, but this cannot be successful without complementary supply-side and increase investment. Like any health intervention, EmOC requires resources; personnel, time, equipment, drugs, and supplies; and resources are always limited [35]. Blood transfusion is one of eight key life-saving interventions that should be available in first-level referral health care facilities providing EmOC [36]. Although the need for blood is universal, millions of patients requiring transfusion do not have timely access to safe blood [37]. Shortage of blood and difficulty obtaining blood for transfusion are still a major obstacle in the management of obstetric emergencies in rural hospitals in The Gambia. This was previously reported by another author on a study on maternal mortality in rural Gambia [26]. The lack of access to and shortage of safe blood for women in this rural hospital could be explained by the increased demand for blood among women of reproductive age in the Gambia, many of whom are anaemic [38]. Additionally, many of the women in this study were admitted with severe bleeding which required immediate blood transfusion. This put a strain on the limited stock of blood in the bank. For blood to be readily available in hospitals, people must be willing to donate voluntarily. However, cultural beliefs (taboos) coupled with the fear of being tested positive for HIV during the screening of blood deters people from donating blood in rural Gambia. As a result potential donors seldom make themselves available for blood donation, thus creating a big gap between supply and demand. Timely, appropriate, and safe blood transfusion during pregnancy and childbirth can make a real difference between life and death for many women and their newborns [36], especially in rural settings.

potential donors seldom make themselves available for blood donation, thus creating a big gap between supply and demand. Timely, appropriate, and safe blood transfusion during pregnancy and childbirth can make a real difference between life and death for many women and their newborns [36], especially in rural settings. Safe blood donors are the mainstay of safe and adequate supply of blood and blood products in hospitals. The safest blood donors are voluntary, nonremunerated blood donors from low-risk populations [39]. However, due to inadequate stock of blood supply, family/replacement or paid donors still provide a large quantity of blood collected in rural hospitals in The Gambia. As a result, timely access to transfusion service is jeopardised taking cognisant of the time the relatives have to spend in search for blood donors. It is important to note that the implication of enumerated blood donation increases the risk of transfusion transmitted infections, especially HIV and viral hepatitis [40]. Public health interventions to inform operating blood bank services seems warranted to reducing perinatal mortality rates in poor rural settings in the Gambia.

ant to note that the implication of enumerated blood donation increases the risk of transfusion transmitted infections, especially HIV and viral hepatitis [40]. Public health interventions to inform operating blood bank services seems warranted to reducing perinatal mortality rates in poor rural settings in the Gambia. 7. Methodological Considerations The broad approach taking into consideration the influence of various individual, socio-cultural, and community factors on accessibility to EmOC services in rural Gambia is a strength of the current study. In addition, conducting interviews in homes as opposed to facility setting allows the respondents to speak their minds freely, thus minimising information bias. The advantage of this study design, using stillbirths to address the shortcomings, is that the women themselves can give information. This is not the case with maternal mortality. Finally, the short window period for the follow-up interviews may have also eliminated recall bias.

reely, thus minimising information bias. The advantage of this study design, using stillbirths to address the shortcomings, is that the women themselves can give information. This is not the case with maternal mortality. Finally, the short window period for the follow-up interviews may have also eliminated recall bias. Selection bias could play a role in the current study. A little more than half (52%) of the births in The Gambia take place in health facilities suggesting that a substantial proportion of women still give birth in home settings. Since the study aimed at exploring barriers to EmOC services in the catchment area of Bansang hospital, the findings may not be representative of the of the entire country with diverse geographic and sociodemographic differences. Thus, the results should be interpreted with caution. Systemic analysis of perinatal deaths in developing countries is hampered by a scarcity of data, especially in countries with nonexistent or incomplete vital registration systems, and where some events take place in homes and villages, not hospitals. However, the findings illustrate useful information regarding barriers to EmOC services for women trying to reach the hospital which is vital for reproductive health managers in The Gambia.

with nonexistent or incomplete vital registration systems, and where some events take place in homes and villages, not hospitals. However, the findings illustrate useful information regarding barriers to EmOC services for women trying to reach the hospital which is vital for reproductive health managers in The Gambia. 8. Conclusion The barriers uncovered in this study are similar to the classical list of “three delays” in accessing life-saving emergency obstetric care interventions. Long distances to the hospital and lack of appropriate means of transportation were found to be major problems affecting timely evacuation of women with obstetric complications. Obtaining blood for transfusion was also identified as another impediment to the timely dispensation of obstetric care interventions. Maternal and perinatal deaths could be averted if all women delivered in a facility that has the capacity to provide EmOC and/or life-saving interventions, such as blood transfusion in the event of obstetric complications. Thus, improved and timely access to EmOC backed by emergency transportation seems warranted for improved maternal and neonatal survival in poor rural settings in The Gambia. 9. Key Issues/Observations The constraints for maternal and perinatal health are similar. Care for the infant starts with proper pregnancy care for the mother including access to good-quality maternity care.

8. Conclusion The barriers uncovered in this study are similar to the classical list of “three delays” in accessing life-saving emergency obstetric care interventions. Long distances to the hospital and lack of appropriate means of transportation were found to be major problems affecting timely evacuation of women with obstetric complications. Obtaining blood for transfusion was also identified as another impediment to the timely dispensation of obstetric care interventions. Maternal and perinatal deaths could be averted if all women delivered in a facility that has the capacity to provide EmOC and/or life-saving interventions, such as blood transfusion in the event of obstetric complications. Thus, improved and timely access to EmOC backed by emergency transportation seems warranted for improved maternal and neonatal survival in poor rural settings in The Gambia. 9. Key Issues/Observations The constraints for maternal and perinatal health are similar. Care for the infant starts with proper pregnancy care for the mother including access to good-quality maternity care. The “three delays” framework is a useful tool for addressing perinatal survival. These delays are addressed as individual problems, not part of an overall access to proper and good-quality public maternity care. Access issues are important in perinatal survival in resource-poor areas of the world such as rural Gambia. Conflict of Interests The authors report no competing interest. The authors are responsible for the writing and content of this paper.

The “three delays” framework is a useful tool for addressing perinatal survival. These delays are addressed as individual problems, not part of an overall access to proper and good-quality public maternity care. Access issues are important in perinatal survival in resource-poor areas of the world such as rural Gambia. Conflict of Interests The authors report no competing interest. The authors are responsible for the writing and content of this paper. Acknowledgments This project was funded by Section for International Health, Department of General Practice and Community Medicine, the Institute of Health and Society, University of Oslo, Norway. The authors' sincere thanks go to the women and families for their participation in this study. The authors would also like to thank the staff and management of Bansang hospital, especially the maternity staff. They cannot also forget the invaluable assistance they received from Mr. Alieu Jammeh and his staff at the National Aids Secretariat; the authors deeply appreciate their efforts. The authors would finally like to register their sincere gratitude to their research assistants and driver for their untiring efforts during the entire period. Table 1 Types of delay experienced by women (n = 20). Delays n (%) (1) First delays (i) Delay in recognising danger signs during pregnancy and labour 3 (15.0) (ii) Sociocultural belief/family decision-making delays 3 (15.0) (2) Second delays (i) Problems in transport, distance, and road infrastructure 5 (25.0) (ii) Cost (no money) 5 (25.0) (3) Third delays (i) Delays receiving blood transfusion 4 (20.0)

1. Introduction In developed countries, ovarian cancer is a commonly occurring neoplasm, ranking the 7th and 6th most frequent position for incidence and mortality, respectively [1]. High-incidence areas are Europe and North America, making it an important public health issue [2]. Epithelial ovarian tumors are thought to arise from the surface epithelium (mesothelium) of the ovary [3]. Most early-stage ovarian cancer produce no symptoms, and therefore the majority of the patients present with advanced disease, making prognosis poor. So far, pathologists have devoted very little attention to early ovarian cancer originating in a coexisting benign epithelial lesion [4]. The precise origin of this epithelium is controversial; one hypothesis argues that the mesothelium lining of the ovarian surface undergoes a Müllerian metaplasia; another one that the same epithelium is derived from the fallopian tube or uterus via passive transport [5].

ng in a coexisting benign epithelial lesion [4]. The precise origin of this epithelium is controversial; one hypothesis argues that the mesothelium lining of the ovarian surface undergoes a Müllerian metaplasia; another one that the same epithelium is derived from the fallopian tube or uterus via passive transport [5]. There are four main types of epithelial ovarian cancer: serous ovarian carcinoma (30% to 70%), mucinous ovarian carcinoma (5% to 20%), endometrioid ovarian carcinoma (10% to 20%), clear cell carcinoma (3% to 10%), and furthermore a minor group of transitional cell or Brenner tumor (1%), and mixed epithelial carcinoma (3%) [6–8]. As described by Mok et al. [9] the progression model of epithelial ovarian cancer is complicated, when the different histological subtypes of ovarian neoplasms are considered. One theory is that ovarian carcinoma arises from endometriosis, the ectopic occurrence of endometrial tissue [10]. Especially clear cell and endometrioid carcinomas are thought to arise from foci of endometriosis [11]. Associations between endometriosis and ovarian carcinoma have been found, but could also be explained by shared risk factors for the coexistence of the conditions [11]. The ovarian surface epithelium (OSE) is generally regarded as the precursor of epithelial ovarian tumors. Recent evidence, however, suggests that the fallopian tube could also be the source of some subtypes of epithelial ovarian cancers [12]. Since the ovaries and tubes are closely related, it is hypothesized that cells of the tubes can mimic ovarian cancer [13, 14].

regarded as the precursor of epithelial ovarian tumors. Recent evidence, however, suggests that the fallopian tube could also be the source of some subtypes of epithelial ovarian cancers [12]. Since the ovaries and tubes are closely related, it is hypothesized that cells of the tubes can mimic ovarian cancer [13, 14]. The aim of this study is to investigate the coexistence of different epithelial subtypes of ovarian carcinoma, with tumor-like conditions [15], benign tumors, and borderline malignancies, in relation to age under and above 55 years at diagnosis, as an approach to distinguish the development of pre- and postmenopausal pathways.

of this study is to investigate the coexistence of different epithelial subtypes of ovarian carcinoma, with tumor-like conditions [15], benign tumors, and borderline malignancies, in relation to age under and above 55 years at diagnosis, as an approach to distinguish the development of pre- and postmenopausal pathways. 2. Materials and Methods We used a random sample (1000 patients out of 9000 patients) out of the database of the nation-wide pathology database PALGA in the Netherlands [16] of the years 1996–2003, from which 824 patients appeared to be diagnosed with an epithelial ovarian cancer. Records in the PALGA database contain comprehensive summaries of the pathology reports and diagnostic codes similar to the systematized nomenclature of medicine (SNOMED) classification of the College of American Pathologists [16]. We scrutinized all reports of 20 lines on average, (>250 words) of the random sample of 1000 patients with an ovary-related diagnosis. Of these 824 patients, with an epithelial ovarian malignancy we collected all other records from PALGA linked with histopathological confirmed diagnoses of borderline malignancies, benign tumors, and tumor-like conditions (endometriosis, simple cysts, endometriotic cysts, and inclusion cysts (two ruptured cysts and one cyst wall)) in the ovary. In case of a diagnose of borderline malignancy as well as malignant carcinoma, in the same patient, the borderline tumor was seen as a coexisting condition.

nign tumors, and tumor-like conditions (endometriosis, simple cysts, endometriotic cysts, and inclusion cysts (two ruptured cysts and one cyst wall)) in the ovary. In case of a diagnose of borderline malignancy as well as malignant carcinoma, in the same patient, the borderline tumor was seen as a coexisting condition. Considering the impact of the findings to current medical practice, we also studied all PALGA receipts of the PALGA database of the pathology department at the Radboud University Nijmegen Medical Centre. It contained the records of 153 patients diagnosed with an ovary-related diagnosis. The Scientific Committee of PALGA approved the study protocol. 2.1. Data Analyses Descriptive analysis was applied to learn from the association between histopathologically subtypes of ovarian epithelial cancer and benign epithelial neoplasms and tumor-like conditions. Anticipating different developmental pre/postmenopausal pathways from benign to malignant tumors, age at diagnosis was dichotomized at 55 year. The Chi-square test was used to arrive at P-values, and 95% confidence intervals for proportions were calculated.

cer and benign epithelial neoplasms and tumor-like conditions. Anticipating different developmental pre/postmenopausal pathways from benign to malignant tumors, age at diagnosis was dichotomized at 55 year. The Chi-square test was used to arrive at P-values, and 95% confidence intervals for proportions were calculated. 3. Results We studied 824 patients with ovarian tumor; 806 had malignant carcinomas and 18 had borderline lesions without carcinoma lesions. In 135 women (16.4%) coexisting ovarian cancer lesions were observed (95% CI: 8.4%–24.4%). In 21 patients two different types of coexisting ovarian tumor lesions or tumor-like conditions were noticed. Of the 806 patients with malignant ovarian cancer, 129 patients (16.0%) had also a tumor-like condition (n = 38), a benign tumor (n = 47), or a borderline malignancy (n = 44). Of the 18 patients with borderline malignancies, 6 (33.0%) had coexisting lesions. The presence of possible precursors according to the different histological subtypes and for both age categories is shown in Table 1. Coexisting epithelial tumor conditions were more prevalent in the age category <55 years. Overall, the ratio of the prevalences of the under age 55 versus 55+ group was 1.71 (95% CI: 1.26–2.31). In particular the subtypes of endometrioid, mucinous, clear cell, and borderline malignancies showed excess prevalences for women under age 55 years.

mor conditions were more prevalent in the age category <55 years. Overall, the ratio of the prevalences of the under age 55 versus 55+ group was 1.71 (95% CI: 1.26–2.31). In particular the subtypes of endometrioid, mucinous, clear cell, and borderline malignancies showed excess prevalences for women under age 55 years. Of the 153 consecutive patients from the Department of Pathology of the UMCN in 2007, 54 out of 153 patients did not have coexisting ovarian tumor conditions or abnormalities. A number of 22 patients had nonepithelial lesions while for two lesions it could not be established whether the tumors were primarily linked to the ovaries or were metastases from elsewhere. The remaining 75 records concerned epithelial ovarian lesions ranging from tumor-like conditions (inclusion cysts and simple cysts n = 29), adenomas (n = 15), borderline malignancies (n = 6), to malignant tumors (n = 25); see Table 2. One-third were actually cancer, mainly serous carcinomas and adenocarcinomas not otherwise specified, of which 4 patients (16.0%) also showed coexisting epithelial tumor lesions or tumor-like conditions. This means that the hospital referral group revealed 35% (54 out of 153 patients) of coexisting epithelial ovarian tumor conditions.

ctually cancer, mainly serous carcinomas and adenocarcinomas not otherwise specified, of which 4 patients (16.0%) also showed coexisting epithelial tumor lesions or tumor-like conditions. This means that the hospital referral group revealed 35% (54 out of 153 patients) of coexisting epithelial ovarian tumor conditions. Of the lesions considered as coexisting epithelial tumor conditions, 58.0% (29/50) were tumor-like conditions, 40.0% (15/50) adenomas, and 12.0% (6/50) borderline malignancies. Of the carcinoma patients (25), 32.0% were younger than 55 years old while of the borderline malignancy patients 67.0% (6/10) were younger than 55. Of patients with adenomas or tumor-like conditions, 80.0% and 82.8% were under age 55 (Table 2). Coexisting conditions were almost similar to the principle ovary cancer in case of borderline malignancies (83%) and less similar in the other types of malignancies (46–64%).

Of the lesions considered as coexisting epithelial tumor conditions, 58.0% (29/50) were tumor-like conditions, 40.0% (15/50) adenomas, and 12.0% (6/50) borderline malignancies. Of the carcinoma patients (25), 32.0% were younger than 55 years old while of the borderline malignancy patients 67.0% (6/10) were younger than 55. Of patients with adenomas or tumor-like conditions, 80.0% and 82.8% were under age 55 (Table 2). Coexisting conditions were almost similar to the principle ovary cancer in case of borderline malignancies (83%) and less similar in the other types of malignancies (46–64%). 4. Discussion 4.1. The PALGA Database 1996–2003 One out of six patients with a malignant epithelial ovarian cancer did have a coexisting ovarian benign lesion. Tumor-like conditions and adenomas were the most commonly observed benign lesions whereas serous, endometrioid, and mucinous carcinomas were the most common malignant tumors. Tumor-like conditions, especially in endometrioid carcinomas, were very frequent (data not shown). One in three of these cases was an endometrioid cyst or endometriosis. The actual number of endometrioid cysts may have been even larger, indicating a relation between endometriosis, endometrioid cyst and endometrioid carcinoma [9, 11, 17]. Nevertheless, because of the cross-sectional nature of our study, causality cannot be deferred, which means that this relation may also be explained by shared risk factors [11].

trioid cysts may have been even larger, indicating a relation between endometriosis, endometrioid cyst and endometrioid carcinoma [9, 11, 17]. Nevertheless, because of the cross-sectional nature of our study, causality cannot be deferred, which means that this relation may also be explained by shared risk factors [11]. Ovarian cancer patients who had a malignant as well as a benign lesion were generally younger than women with only a malignant tumor. This may underpin the fact that benign tumors tend to occur at younger age than malignant tumors. As stated earlier, only part of the malignant epithelial ovarian tumors will arise from a coexisting benign epithelial lesion [4, 18]. As endometrioid, mucinous, clear cell, and borderline malignant epithelial tumors are more common in women with a benign and a malignant lesion, these subtypes are the most likely to develop through tumor transition. Recently it is hypothesized that most high-grade serous carcinomas (50% or more) arise from the distal fallopian tube (fimbria) [19]. Our results may not be in conflict with this hypothesis while only a small number (10.9%) of serous carcinomas had coexisting lesions. A subset will develop de novo from the ovarian surface epithelium or its inclusion cysts [20].

grade serous carcinomas (50% or more) arise from the distal fallopian tube (fimbria) [19]. Our results may not be in conflict with this hypothesis while only a small number (10.9%) of serous carcinomas had coexisting lesions. A subset will develop de novo from the ovarian surface epithelium or its inclusion cysts [20]. As described by Auersperg et al. [21] the surface epithelium of the ovary and tubal epithelium were considered as a unit, with an area of increased susceptibility to neoplastic progression encompassing the ovarian surface epithelium and the distal fimbriae. Progression along different histotypic pathways might then be influenced by microenvironmental factors and mutational events. 4.2. Findings of the PALGA Nijmegen 2007 Database Women with a tumor-like condition or adenoma appeared to be younger (82.8% at age <55 year and 80.0%, resp.) than women with a malignant tumor (32.0% at age <55 years). The majority of patients with a borderline malignancy also were younger than 55 years (66.6%).

As described by Auersperg et al. [21] the surface epithelium of the ovary and tubal epithelium were considered as a unit, with an area of increased susceptibility to neoplastic progression encompassing the ovarian surface epithelium and the distal fimbriae. Progression along different histotypic pathways might then be influenced by microenvironmental factors and mutational events. 4.2. Findings of the PALGA Nijmegen 2007 Database Women with a tumor-like condition or adenoma appeared to be younger (82.8% at age <55 year and 80.0%, resp.) than women with a malignant tumor (32.0% at age <55 years). The majority of patients with a borderline malignancy also were younger than 55 years (66.6%). Describing the occurrence of benign ovarian tumors in itself is also very difficult. As stated earlier, malignant ovarian tumors are often diagnosed in a late stage; benign tumors are often not detected at all, for example, when an obvious carcinoma is present within the ovary, most pathologists would not mention the presence of cortical inclusion cysts. Therefore, in daily gynecological practice most of the benign tumors may be detected coincidentally. This, however, only seems to be the tip of the iceberg: most benign tumors remain undetected. Consequently, it is hard to study at which age what kind of benign tumors occur.

on the presence of cortical inclusion cysts. Therefore, in daily gynecological practice most of the benign tumors may be detected coincidentally. This, however, only seems to be the tip of the iceberg: most benign tumors remain undetected. Consequently, it is hard to study at which age what kind of benign tumors occur. A limitation of the PALGA Nijmegen 2007 database was that it was unclear whether the patients without abnormality or benign lesion were representing the general population. Some of the patients had the ovaries removed because they were carriers of a mutation in BRCA1 or BRCA2, which sets them at high risk to develop ovarian cancer. Other patients were examined because they had symptoms. Due to this fact, the incidence of benign lesions is probably higher than in the general population. It is unknown whether the larger dataset also contains BRCA/high-risk women as well. 5. Conclusion The strength of this study is the analysis specified to the various histological subtypes in relation to possible precursors of ovarian cancer and malignancies. Overall, the prevalence of coexisting epithelial ovarian tumor conditions was 1.71-times (95% CI: 1.26–2.31) as high in patients <55 years versus patients ≥55 years. Unfortunately, sparse numbers of cases hamper to arrive at a meaningful conclusion on more aggressive malignancies in younger women. The histological subtypes endometrioid, mucinous, clear cell, and borderline malignancies showed the highest prevalences of coexisting lesions.

tients <55 years versus patients ≥55 years. Unfortunately, sparse numbers of cases hamper to arrive at a meaningful conclusion on more aggressive malignancies in younger women. The histological subtypes endometrioid, mucinous, clear cell, and borderline malignancies showed the highest prevalences of coexisting lesions. Weaknesses of the study lie in its retrospective design and the fact that coexisting pathologies are usually overlooked or better said underreported. Further research is needed to confirm the results in larger study populations, and possibly be combined with a study of (pre-)neoplasia arising in the Fallopian tube in relation to the hereditary serous ovarian carcinogenesis hypothesis from Piek [13]. The analysis of the PALGA Nijmegen 2007 database indicates that half of all patients referred to the University Medical Centre St. Radboud with tissue specimen sent to the pathology department, have epithelial ovarian tumor conditions. Two-thirds of the latter seem to have coexisting epithelial tumor conditions, adding to the progression to the malignant disease stage. These findings may prompt to not only reporting ovary cancer but also coexisting ovarian tumor conditions. The latter if detected in the absence of ovary cancer need follow-up surveillance. Funding No funding was received for this paper. Conflict of Interests None of the authors have any conflict of interests to declare. Table 1 The occurrence of coexisting ovarian tumor conditions in epithelial ovarian cancer according to subtypes and the age at diagnosis in 824 patients.

The analysis of the PALGA Nijmegen 2007 database indicates that half of all patients referred to the University Medical Centre St. Radboud with tissue specimen sent to the pathology department, have epithelial ovarian tumor conditions. Two-thirds of the latter seem to have coexisting epithelial tumor conditions, adding to the progression to the malignant disease stage. These findings may prompt to not only reporting ovary cancer but also coexisting ovarian tumor conditions. The latter if detected in the absence of ovary cancer need follow-up surveillance. Funding No funding was received for this paper. Conflict of Interests None of the authors have any conflict of interests to declare. Table 1 The occurrence of coexisting ovarian tumor conditions in epithelial ovarian cancer according to subtypes and the age at diagnosis in 824 patients. Ovarian cancer subtype and age category Total 1 or 2 lesions present* Ratio of prevalences (95% CI) n n %* All subtypes 824 135 16.4 <55 yr 274 62 22.6 1.71 (1.26–2.31) ≥55 yr 550 73 13.3 1 Serous carcinoma 385 42 10.9 <55 yr 113 15 13.3 1.33 (0.74–2.42) ≥55 yr 272 27 9.9 1 Endometrioid carcinoma 113 25 22.1 <55 yr 46 13 28.3 1.58 (0.79–3.14) ≥55 yr 67 12 17.9 1 Mucinous carcinoma 98 28 28.6 <55 yr 47 16 34.0 1.45 (0.76–2.73) ≥55 yr 51 12 23.5 1 Adenocarcinoma 73 12 16.4 <55 yr 17 3 17.6 1.10 (0.33–3.60) ≥55 yr 56 9 16.1 1 Clear cell carcinoma 56 11 19.6 <55 yr 23 7 30.4 2.51 (0.83–7.60) ≥55 yr 33 4 12.1 1 Mixed malignancy 34 5 14.7 <55 yr 11 2 18.2 1.39 (0.27–7.18) ≥55 yr 23 3 13.0 1 Borderline malignancy 18 6 33.3

<55 yr 47 16 34.0 1.45 (0.76–2.73) ≥55 yr 51 12 23.5 1 Adenocarcinoma 73 12 16.4 <55 yr 17 3 17.6 1.10 (0.33–3.60) ≥55 yr 56 9 16.1 1 Clear cell carcinoma 56 11 19.6 <55 yr 23 7 30.4 2.51 (0.83–7.60) ≥55 yr 33 4 12.1 1 Mixed malignancy 34 5 14.7 <55 yr 11 2 18.2 1.39 (0.27–7.18) ≥55 yr 23 3 13.0 1 Borderline malignancy 18 6 33.3 <55 yr 11 4 36.4 1.27 (0.31–5.20) ≥55 yr 7 2 28.6 1 Other 47 6 12.8 <55 yr 6 2 33.3 3.41 (0.79–14.80) ≥55 yr 41 4 9.8 1 *Lesions present: tumor-like condition (endometriosis, simple cysts, endometriotic cysts, inclusion cysts), benign tumor, and borderline malignancy. 95% CI, confidence interval. Table 2 Specific diagnoses of 75 consecutive tissue specimen from PALGA Nijmegen 2007 according to age. Diagnosis All ages <55 yr ≥55 yr n n % n % Total 75 48 64.0 27 36.0 Ovarian carcinoma 25* 8 32.0 17 68.0 Borderline malignancies 6 4 66.6 2 33.3 Adenoma 15 12 80.0 3 20.0 Tumor-like conditions** 29 24 82.8 5 17.2 *4 patients (16.0%) showed coexisting epithelial ovarian tumor conditions. **tumor-like conditions: simple cysts, endometriotic cysts, inclusion cysts. Coexisting epithelial ovarian carcinoma conditions: (4 + 6 + 15 + 29)/75 = 72%. Chi-square = 17.225, df = 3, P-value = 0.001.

1. Introduction Cervical cancer (CC) is the third most common cancer in women, and the seventh overall, with an estimated 530 000 new cases in 2008. More than 85% of the global burden occurs in developing countries, where it accounts for 13% of all female cancers. Cervical cancer is responsible for 275 000 deaths in 2008, about 88% of which occur in developing countries: 53 000 in Africa, 31 700 in Latin America and the Caribbean, and 159 800 in Asia [1]. However, CC is preventable disease, and much effort should be done to prevent it. It is now well-established that persistence of Human Papillomavirus (HPV) infection is the strongest epidemiologic factor associated with intraepithelial neoplasia and cancer of cervix [2–5], and it is considered a necessary cause for development of the disease but not sufficient. Exogenous or endogenous factors had been identified to, in conjunction with HPV infection, influence the risk for CC. As Castellsaguè and Muñoz [6] suggested, cofactors may be classified into three groups: (1) environmental or exogenous cofactors, including use of oral contraceptives (OCs), tobacco smoking, diet, cervical trauma, coinfection with human immunodeficiency virus (HIV) and other sexually transmitted agents; (2) viral cofactors, such as infection by specific types, coinfection with other types, HPV variants, viral load, and viral integration; (3) host cofactors, including endogenous hormones, genetic factors such as human leukocyte antigen and other host factors related to the host's immune response.

ly transmitted agents; (2) viral cofactors, such as infection by specific types, coinfection with other types, HPV variants, viral load, and viral integration; (3) host cofactors, including endogenous hormones, genetic factors such as human leukocyte antigen and other host factors related to the host's immune response. Winkelstein Jr. [7] in 1977 was the first to put the hypothesis that smoking is a risk factor for cervical cancer. Since then, the action of tobacco on cervical carcinogenesis has been a matter of scientific debate. The objective of this paper is to update the state of the science about smoking and cancer of the cervix. 2. Material and Methods It was checked up in PubMed/Medline and Embase databases, studies written in English with keywords: cervical cancer; cervix; CIN; HPV; tobacco; smoking and smoke. Only meta-analysis and or multi-institutional studies were considered for analysis. 3. Results Plummer et al. reported the first multicentric case-control study in 2003 [8]. In analyzing eight studies on invasive cancer and two on carcinoma in situ, conducted by International Agency for Research on Cancer (IARC), between 1985 and 1997, the authors conclude that ever-smokers have an excess risk of cervical cancer that persists after controlling for the strong effect of HPV and for other potential cofactors of progression from infection to cancer, and they suggest that squamous cell carcinoma of the cervix should be added to the list of tobacco associated cancers, while for adenocarcinoma, further data should be warranted.

ancer that persists after controlling for the strong effect of HPV and for other potential cofactors of progression from infection to cancer, and they suggest that squamous cell carcinoma of the cervix should be added to the list of tobacco associated cancers, while for adenocarcinoma, further data should be warranted. In 2004, IARC revisited its previous conclusions and listed cervical cancer among those causally related to smoking [9]. In 2006, a collaborative reanalysis of 12 studies on CC [10], enrolling 8,097 cases of squamous cell carcinoma, 1,374 women adenocarcinoma, and 26,445 women without carcinoma of the cervix, showed that there are no substantial differences between the two most common histological types of invasive cervical cancer with respect to the role of number of sexual partners, age at first intercourse, age at first birth, body mass index, and use of oral contraceptives. The exception was current tobacco smoking, which is associated with an increased risk of squamous cell but not for adenocarcinoma of the cervix, in agreement with other studies [11, 12].

the role of number of sexual partners, age at first intercourse, age at first birth, body mass index, and use of oral contraceptives. The exception was current tobacco smoking, which is associated with an increased risk of squamous cell but not for adenocarcinoma of the cervix, in agreement with other studies [11, 12]. Vaccarella et al. [13] in 2008 reported a pooled analysis of 13 IARC HPV prevalence survey in 11 countries worldwide, carried out between 1993 and 2005. They concluded that current tobacco smoking was associated with a significant, although moderate, increased risk of prevalent HPV infection. Among current smokers, the risk of being HPV-positive increased with increasing number of cigarettes smoked per day, and women who reported to smoke 15 or more cigarettes daily had a 2-fold risk of HPV positivity as compared with never-smokers. These results have shown that smoking habits interfere with prevalence of HPV infection in agreement with reported in previous studies [14–16].

increasing number of cigarettes smoked per day, and women who reported to smoke 15 or more cigarettes daily had a 2-fold risk of HPV positivity as compared with never-smokers. These results have shown that smoking habits interfere with prevalence of HPV infection in agreement with reported in previous studies [14–16]. Syrjänen et al. [17], in 2007, in analyzing a cohort of 3,187 women, conclude that cigarette smoking was not an independent risk factor of CIN2+, except for those patients who tested for HR-HPV (high-risk HPV), and current smoking remains an independent predictor for those patients in a multivariate model, in agreement with previous study of Harris et al. in 2004, [18] that found among women with oncogenic HPV infection, smoking was associated with risk for both CIN1 and CIN2-3. Of the three smoking measures (smoking status, pack years of exposure, and number of cigarettes per day), number of cigarettes per day (>10 cigarettes) was the most strongly associated with risk for CIN1 and CIN2-3. In this study, association between cigarettes smoked by day and CIN did not appear to be mediated by an immunologic response. Those studies suggest that smoking has a special adverse effect on HR-HPV driven cervical carcinogenesis.

per day (>10 cigarettes) was the most strongly associated with risk for CIN1 and CIN2-3. In this study, association between cigarettes smoked by day and CIN did not appear to be mediated by an immunologic response. Those studies suggest that smoking has a special adverse effect on HR-HPV driven cervical carcinogenesis. A recent study [19], published in 2010, conducted on 2,011 women, 15–19 years old, recruited from 1988 to 1992 then regularly followed until 1997, concluded that there is no evidence to suggest that the risk of acquiring a HPV infection of any type, or a HPV16 or HPV18 infection, increases with either pack years of exposure to smoking or duration of current smoking episode, suggesting that smoking is not a important risk factor for HR-HPV infection. Xi et al. in 2009 [20] reported an analysis of 1,050 women HPV16 and/or HPV18 positives for viral DNA load, enrolled into the ASCUS-LSIL Triage Study. The authors concluded that higher HPV16 and HPV18 DNA load was associated with status of current, but not former, smoker. Among current smokers, the viral load did not appear to vary appreciably by the intensity and duration of cigarette smoking, in accordance, with previous study of Gunnell et al. [21] in 2006, that in testing for HPV16 DNA presence in first archival cervical smears from 375 cases of in situ cervical squamous carcinoma (CIS) and in 363 controls, it was found that current smokers with a high HPV16 viral load at time of first smear were at a particularly increased risk (27-fold) compared with current smokers without HPV-infection.

HPV16 DNA presence in first archival cervical smears from 375 cases of in situ cervical squamous carcinoma (CIS) and in 363 controls, it was found that current smokers with a high HPV16 viral load at time of first smear were at a particularly increased risk (27-fold) compared with current smokers without HPV-infection. 4. Discussion Some molecular mechanisms have been suggested through why smoking may contribute towards cervical carcinogenesis: one involves direct exposure of the deoxyribonucleic acid (DNA) in cervical epithelial cells to nicotine and cotinine, and the other involves exposure to metabolic products resulting from the reactions of other components of cigarettes such as aromatic polycyclic hydrocarbons and aromatic amines. [22, 23]. Cervical mucus of smokers contains measurable amounts of cigarette constituents and their metabolites such as benzo[a]pyrene (BaP) [24], nicotine, and nicotine derived nitrosamines 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone [25]. BaP up regulation of HPV genome amplification may increase the probability of viral DNA integration into the host genome, a milestone in the development of cervical cancer [26]. The in vivo effects of long-term nicotine exposure could affect persistent cellular proliferation, inhibition of apoptosis, and stimulation of vascular endothelial growth factor, with increased microvessel density [27]. Other mechanisms that may explain smoking-related carcinogenesis are related with abnormalities in the systemic and peripheral immune systems of smokers, that include unbalanced systemic production of pro- and anti-inflammatory cytokines [28], elevated numbers of cytotoxic/suppressor T lymphocytes, suppression of T lymphocyte activity, diminished numbers of helper T lymphocytes, decreased numbers of natural killer lymphocytes and low levels of immunoglobulins other than immunoglobulin E (IgE) [29]. These effects may result from substantial decreased numbers of Langerhans cells in the cervix of smokers [30, 31]. Aberrant HPV-induced DNA methylation may be another mechanism to explain cervical smoking-related carcinogenesis. In vitro studies in untransformed and transformed cell lines show that short-term exposure to nicotine or cigarette smoke extract is followed by changes in the expression of the DNA methyltransferases DNMT1, DNMT3A,and DNMT3B.

DNA methylation may be another mechanism to explain cervical smoking-related carcinogenesis. In vitro studies in untransformed and transformed cell lines show that short-term exposure to nicotine or cigarette smoke extract is followed by changes in the expression of the DNA methyltransferases DNMT1, DNMT3A,and DNMT3B. Aberrant methylation of the tumour suppressor gene, p16 (CDNK2A), is strongly associated with current smoking in women with squamous cell cervical cancers and high-grade CIN [32]. Intriguing studies analyzing wood-smoke effect on cervical carcinogenesis have shown that, among HPV-positive women, a dose-response relationship is observed for exposure to wood smoke and cervical cancer that persisted in multivariate analysis [33], and chronic exposure to wood smoke significantly increased the risk of CIN III, suggesting that chronic inhalation of carcinogens derived from wood smoke could have an effect on the progression to cervical cancer, similar to that observed for cigarette smoking [34]. Probably the smoking effect on cervical carcinogenesis is not tobacco-specific. Smoking seems to affect negatively the early natural history of HPV infections. The regression of Low Squamous Intraepithelial Lesions (LSIL) within 2 years is significantly lower in smokers than in never-smokers [35]. Smoking affecting clearance HPV infection remains a conflicting issue. For some authors, smoking have no influence in duration of HPV infection [19], for others tobacco delays the clearance of HPV infection [36, 37].

Smoking seems to affect negatively the early natural history of HPV infections. The regression of Low Squamous Intraepithelial Lesions (LSIL) within 2 years is significantly lower in smokers than in never-smokers [35]. Smoking affecting clearance HPV infection remains a conflicting issue. For some authors, smoking have no influence in duration of HPV infection [19], for others tobacco delays the clearance of HPV infection [36, 37]. Covalent alteration of DNA to form DNA adducts is considered an early step in chemical carcinogenesis and, therefore, detection of DNA adducts provides evidence of exposure of the cervix to carcinogens. Prokopczyk et al. [38] had shown, in accordnce with previous studies, that significantly higher DNA adduct levels are present in the cervix of smokers as compared with nonsmokers, providing molecular evidence of smoking-related carcinogenic agents that affects the DNA of the cervical epithelium. However, the authors reported no significant differences in smoking-related DNA damage (DNA adduct levels) between HPV-positive and HPV-negative smokers, suggesting that smoking DNA damage is not related with HPV infectivity. Genetic susceptibility to smoking is an important issue. Cervical cancer risk in smokers may be modified by genetic variants, as that described to interleukin 2 [39] or to 8q24 chromosome polymorphisms [40]. In a recent study [41], the tumor suppressors p53, the fragile histidine triad and the interleukin-10 were under-expressed, and the cyclooxygenase-2 and the Ki-67 were over-expressed in smoking, compared with nonsmoking women with CIN.

s, as that described to interleukin 2 [39] or to 8q24 chromosome polymorphisms [40]. In a recent study [41], the tumor suppressors p53, the fragile histidine triad and the interleukin-10 were under-expressed, and the cyclooxygenase-2 and the Ki-67 were over-expressed in smoking, compared with nonsmoking women with CIN. Familial and social factors are determinant for smoking attitudes. Being older, being divorced, having friends/family who smoke, and having parents who smoke are all associated with ever smoking, and friends are the primary factor influencing ever smoking, especially among younger women [42]. Smokers seem to have lower compliance for cervical cancer screening, the most powerful weapon against cervical cancer mortality. Smokers held less positive attitudes towards cervical screening than did nonsmokers [43], and the level of nicotine dependence is also significantly related to compliance with screening recommendations; women with higher levels of dependence are less likely to be compliant [44]. Unhealthy life style, negatively rated life events, lack of social support, coping style, and distress, often associated with smoking, alcohol abuse, and illegal drugs addition, have been reported as risk factors for cervical cancer [45–47], especially among low-educated women [48, 49].

Familial and social factors are determinant for smoking attitudes. Being older, being divorced, having friends/family who smoke, and having parents who smoke are all associated with ever smoking, and friends are the primary factor influencing ever smoking, especially among younger women [42]. Smokers seem to have lower compliance for cervical cancer screening, the most powerful weapon against cervical cancer mortality. Smokers held less positive attitudes towards cervical screening than did nonsmokers [43], and the level of nicotine dependence is also significantly related to compliance with screening recommendations; women with higher levels of dependence are less likely to be compliant [44]. Unhealthy life style, negatively rated life events, lack of social support, coping style, and distress, often associated with smoking, alcohol abuse, and illegal drugs addition, have been reported as risk factors for cervical cancer [45–47], especially among low-educated women [48, 49]. Hormonal contraception is implicated on cervical cancer development. Appleby et al. [50], in analyzing 24 studies worldwide that included 16,573 women with cervical cancer and 35,509 without cervical cancer, concluded that the relative risk of cervical cancer is increased in current users of oral contraceptives, and that increased risk is higher in current smokers women.

er development. Appleby et al. [50], in analyzing 24 studies worldwide that included 16,573 women with cervical cancer and 35,509 without cervical cancer, concluded that the relative risk of cervical cancer is increased in current users of oral contraceptives, and that increased risk is higher in current smokers women. Nutrition is another important factor that seems to influence cervical carcinogenesis. Oxidative stress induced by deficiencies in antioxidant micronutrients is likely to change normal redox balance and transform the HPV-infected cells toward a carcinogenic process in the cervix [51]. Folate is a well-studied micronutrient with effect on cervical carcinogenesis. Reduced immunocompetence associated with deficiencies of folate and vitamin B12 could increase the risk of infection and persistence with multiple types or higher viral loads of HR-HPVs [52–54]. Higher prevalence rates of HPV have also been found in HIV-seropositive women, proportional to their level of immunosuppression [55–57]. HIV-seropositive women, and particularly the current smokers, are at significantly elevated risk for the development and recurrence of precancerous and cancerous cervical disease [58, 59], especially those who have lower levels of TH cells. It has been shown that TH cells are involved in the defense against HPV-transformed cells [60]. Hence, immunosuppression in general, and depletion of TH cells in particular, may interact with HPV infection to increase the risk of cervical cancer. Marijuana usage is another described factor that potencies tobacco immunosuppression [61].

that TH cells are involved in the defense against HPV-transformed cells [60]. Hence, immunosuppression in general, and depletion of TH cells in particular, may interact with HPV infection to increase the risk of cervical cancer. Marijuana usage is another described factor that potencies tobacco immunosuppression [61]. Smoking also affects survival among women diagnosed with cervical cancer. Recently, Coker et al. [62], in analysing 2661 women diagnosed with invasive cervical cancer from 1995–2005, found that, after adjustment for age and stage at diagnosis, cell type, rural residence, race, insurance coverage, and treatment-received, current smoker, were 35% more likely to die of any cause and 21% more likely to die of cervical cancer compared with known nonsmoking cases, in accordance with previous studies. Unfortunately, few smokers with cervical cancer quit or decreased consumption during treatment [63]. 5. Conclusions All meta-analyses and multi-institutional studies point out that smoking is an important cofactor for cervical squamous cancer and probably also for cervical adenocarcinoma. Acquisition of HR-HPV infection seems to be a smoking independent event; however, progression of the acquired infection is negatively affected by current smoking. Former smoking seems to be no so important.

5. Conclusions All meta-analyses and multi-institutional studies point out that smoking is an important cofactor for cervical squamous cancer and probably also for cervical adenocarcinoma. Acquisition of HR-HPV infection seems to be a smoking independent event; however, progression of the acquired infection is negatively affected by current smoking. Former smoking seems to be no so important. The action mechanisms of smoking on HPV driven cervical carcinogenesis seem to be complex and multifactorial. In current smokers, genetic, immunologic, and dietary factors may be associated with cervical epithelium susceptibility for HPV carcinogenic effect, but womens unhealthy lifestyles seem to be major factors associated with cervical carcinogenesis. What is much determinant for cervical cancer susceptibility: tobacco action by its own or women health or behaviour associated with smoking habits? Until now, a scientific-based answer is lacking. Epidemiologic, clinical, and laboratorial research is needed to elucidate this unsolved topic. In order to prevent cervical cancer and to improve therapy results, the practitioner should understand negative effects of tobacco on cervical carcinogenesis, and in those current smoking women with diagnosis of cervical HPV infection or CIN, and he must have an active intervention to invite them to quit smoking habits.

In order to prevent cervical cancer and to improve therapy results, the practitioner should understand negative effects of tobacco on cervical carcinogenesis, and in those current smoking women with diagnosis of cervical HPV infection or CIN, and he must have an active intervention to invite them to quit smoking habits. The researchers on cervical cancer should be aware that smoking habits are a major cofactor on cervical HPV driven carcinogenesis, and an important confounder factor for research. Smoking effects should be taken in account by all researchers in their studies. Conflict of Interests The author has no conflict of interests that are directly relevant to the content of this paper.

1. Introduction Most obstetrical inpatient stays are short—often no more than one or two days—and the involvement of the inpatient consultation-liaison (C-L) psychiatry service, when required, is necessarily brief. Regardless of its compendious quality, however, its impact can have significance for the patient.

1. Introduction Most obstetrical inpatient stays are short—often no more than one or two days—and the involvement of the inpatient consultation-liaison (C-L) psychiatry service, when required, is necessarily brief. Regardless of its compendious quality, however, its impact can have significance for the patient. We have previously reported on the characteristics of obstetrical inpatients referred for assessment by the C-L psychiatry service at Mount Sinai Hospital in Toronto [1]. Typically, C-L services do not follow up with patients after discharge. Borus et al. [2] argue that studies of C-L psychiatry's impact ought to be longitudinal and include examination of readmissions, persistence of physical and emotional dysfunction after hospital, adherence to prescribed medical regimens and rehabilitation programs, disability, and lost work days. While there is an extensive international literature on liaison mental health services, Callaghan et al. [3], in their review of 17 evaluative studies of these services, point out that most investigations examine the structure and process of liaison work, with little attention paid to outcome variables such as compliance with treatment recommendations and patient satisfaction. Furthermore, even in those instances where outcome is considered, patients' views tend not to be solicited. Where patients' views are sought, they tend to be elicited through satisfaction surveys. For example, Phillips et al. [4] surveyed former patients and the professionals who had referred them to a C-L psychiatry service in an obstetric-gynecology hospital. Mail-in questionnaires examined a number of variables, such as referral source and patient satisfaction with the consultation, and while their data provide a valuable contribution, the complexities of viewpoints are obscured through this forum.

erred them to a C-L psychiatry service in an obstetric-gynecology hospital. Mail-in questionnaires examined a number of variables, such as referral source and patient satisfaction with the consultation, and while their data provide a valuable contribution, the complexities of viewpoints are obscured through this forum. In a small number of studies, qualitative methods have been employed to garner patient evaluations of C-L psychiatry services and compliance with treatment recommendations. For example, an investigation by Rigatelli et al. [5] explores the effectiveness of a general hospital's C-L service; telephone interviews with 95 former inpatients and their primary care physicians 3–5 months after discharge indicated that 66% of patients reported high satisfaction with the psychiatric consultation, 22% were indifferent, and 11% expressed negative feelings. As for postdischarge compliance to treatment suggestions, the most common of which was psychopharmacology alone (68%), more than half of patients reported having filled their prescriptions and taken their medication, and half of these noted a positive outcome, a finding generally confirmed by their primary care physicians. These physicians, who had received a copy of the C-L consult note, agreed with the C-L psychiatric diagnosis in 81% of the study cases, and in 60% of cases, these primary care physicians assessed their patients' postdischarge conditions as improved. Rigatelli et al. conclude that “follow-up studies on outcome of psychiatric consultations are few and further analysis is strongly recommended.”

with the C-L psychiatric diagnosis in 81% of the study cases, and in 60% of cases, these primary care physicians assessed their patients' postdischarge conditions as improved. Rigatelli et al. conclude that “follow-up studies on outcome of psychiatric consultations are few and further analysis is strongly recommended.” A comparable study by Eales et al. [6] evaluated a C-L psychiatry service in an accident and emergency department and a general hospital setting. Service users, who had all presented with a psychiatric complaint, emphasized the necessity of excellent communication skills on the part of service providers; from the patients' perspective, this translates as feeling believed/listened to/taken seriously, receiving useful information, being responded to in a calming manner, receiving a novel perspective on their problems, and having an opportunity to talk without being interrupted by questions, especially during the beginning of the assessment. With regard to consultation outcome, some patients had felt involved in formulating decisions about their future and recalled having been told to return to the hospital if needed, while others had felt excluded from decision-making. The majority of patients felt the service could be improved if postdischarge followup was offered. Eales et al. conclude that it is important to engage service users in action planning to implement follow-up measures.

ng been told to return to the hospital if needed, while others had felt excluded from decision-making. The majority of patients felt the service could be improved if postdischarge followup was offered. Eales et al. conclude that it is important to engage service users in action planning to implement follow-up measures. In the interests of program evaluation and the development of best practice principles, we sought to ascertain the ongoing impact of our inpatient C-L Psychiatry service on the mental health of obstetrical inpatients after discharge, for example, whether they recollected and adhered to treatment recommendations made during their time in hospital and whether they developed or had a recurrence of mental health problems. We also wanted to understand from a phenomenological perspective patients' experience of the inpatient psychiatric consultation and to determine how the service could be rendered more effective. 2. Method Consent for the study was obtained from the Mount Sinai Hospital Research Ethics Board. Psychiatric consult notes were reviewed for all obstetrical inpatients referred to the C-L Psychiatry service at Mount Sinai Hospital, Toronto, Canada, during a six-month period from September 2007 to February 2008. Each of these patients was contacted by letter six months after discharge to invite participation in a semistructured, 30–60 minute telephone interview. The letter was immediately followed up by a phone call to set a date for the interview [1].

to, Canada, during a six-month period from September 2007 to February 2008. Each of these patients was contacted by letter six months after discharge to invite participation in a semistructured, 30–60 minute telephone interview. The letter was immediately followed up by a phone call to set a date for the interview [1]. Characteristics of the hospital and the C-L service are detailed in [1]. To summarize, Mount Sinai Hospital is a tertiary-level university-affiliated general hospital that cares for a large number of low-risk and high-risk pregnant women; there are approximately 7,000 deliveries per year. It is estimated that 40% of the hospital's obstetrical patients are “high-risk”, and it is not uncommon for high-risk antenatal patients to be transferred from smaller and/or remote communities in Ontario or even from other provinces in Canada. Data collected during the telephone interview included demographic information (age, relationship status, employment, level of education, ethnic or racial identification, religious affiliation, number of years of residency in Canada); reproductive history; participants' recollections of reason for referral (RFR) to psychiatry, diagnosis, treatment recommendations, psychotropic medications taken at the time of, or initiated as a result of, the consultation, and recommendations for postdischarge care/followup; participants' reasons for noncompliance with treatment recommendations; participants' evaluation of the inpatient psychiatry consultation; participants' recommendations for improving the service.

ns taken at the time of, or initiated as a result of, the consultation, and recommendations for postdischarge care/followup; participants' reasons for noncompliance with treatment recommendations; participants' evaluation of the inpatient psychiatry consultation; participants' recommendations for improving the service. The areas explored here include (1) congruence between data derived from consultation notes and participants' recollections pertaining to RFR, diagnosis, and treatment recommendations; (2) participants' postdischarge compliance with recommendations; (3) participants' concerns about mood after discharge; (4) participants' perceptions of the benefits and their critical assessments of the inpatient psychiatric consultation process; (5) participants' recommendations for improvement to the process, both inhospital and followup. 3. Results Between September 2007 and February 2008, 62 obstetrical inpatients were referred to the C-L Psychiatry service at Mount Sinai Hospital. Of these, 28 women (45%) agreed to participate in the current study; eight women could not be reached because they had moved and there was no forwarding address (in four of these cases there was “no fixed address”); in nine cases the phone number was no longer in service; four women who were contacted chose not to participate; five patient files were not accessible; eight women consistently did not answer their phone, and in the interests of privacy and confidentiality, a message was not left.

cases there was “no fixed address”); in nine cases the phone number was no longer in service; four women who were contacted chose not to participate; five patient files were not accessible; eight women consistently did not answer their phone, and in the interests of privacy and confidentiality, a message was not left. Of the 28 study participants, 21% were antenatal inpatients at the time of the psychiatry consultation while 79% were postpartum. Seventy-one percent (71%) of patients reported a past psychiatric history. Thirty-six percent (36%) of the sample currently had a normal delivery but were psychiatrically symptomatic and endorsed a past psychiatric history. Sixty-one percent (61%) of the total sample were experiencing an adverse perinatal event (e.g., premature birth, termination for fetal anomaly, unexpected Down Syndrome, preeclampsia, traumatic delivery); 55% of the total sample had had a previous adverse reproductive event (e.g., spontaneous abortion, termination of pregnancy for fetal anomaly, extreme prematurity). Demographic data for participants are shown in Table 1. Most notable is educational level: 71% had a university or college degree, and another 25% had attended university or college for at least one year. At six-month postpartum, 11% of participants had returned to the workforce, while 32% intended to remain at home as full-time homemakers. Most women (86%) were raised in Canada, and the majority reported their ethnic and/or religious affiliation as “None” (36%) or Jewish (21%). Over half (54%) of participants were primigravidae.

th postpartum, 11% of participants had returned to the workforce, while 32% intended to remain at home as full-time homemakers. Most women (86%) were raised in Canada, and the majority reported their ethnic and/or religious affiliation as “None” (36%) or Jewish (21%). Over half (54%) of participants were primigravidae. 3.1. Congruence between Patients' Recollections and C-L Psychiatry Notes (a) Reason for Referral In 79% of cases, women's perception of the RFR was congruent with the RFR stated in the psychiatry consult note. While terminology used by patients differed from that of the psychiatrist or psychiatric resident, the essence of their reasons was usually concordant, for example, consult note states: “High risk for PPD; crying, overwhelmed” and the patient describes her RFR this way: “I have a history of major depression, and on day one postpartum the emotions came fast and furious.”

t of the psychiatrist or psychiatric resident, the essence of their reasons was usually concordant, for example, consult note states: “High risk for PPD; crying, overwhelmed” and the patient describes her RFR this way: “I have a history of major depression, and on day one postpartum the emotions came fast and furious.” In some cases, contributory factors most pressing for the patient may not have been deemed so by the obstetrical or psychiatric staff. For example, a woman (GA 27 weeks) suffering from placenta previa had been airlifted to the hospital from a town several hundred miles away. One month after admission, while on antenatal bedrest, she was referred to the C-L psychiatry service. The RFR reported in the consult note was: “History PPD; currently mildly anxious; twins; planned C-section and hysterectomy; patient worried about her health”. During the study interview at six months post-discharge, the patient recalled the reasons for her referral as a history of PPD, but more important were the extreme loneliness she had been experiencing in the total absence of supports in Toronto, her strong feelings of insecurity about her relationship with her husband and other children, and her intense fear of dying in childbirth. From her perspective, the RFR was multifactorial, the very least of which was her past history of PPD.

eliness she had been experiencing in the total absence of supports in Toronto, her strong feelings of insecurity about her relationship with her husband and other children, and her intense fear of dying in childbirth. From her perspective, the RFR was multifactorial, the very least of which was her past history of PPD. In five cases there was a marked discrepancy between the stated RFR and the woman's perceived reason for referral. For example, RFR stated in the consult note: “High anxiety, mood lability; acute stress response following high-risk (HELLP) premature C-section” versus the patient's perceived RFR: “The social worker suggested the consult because my baby was premature and I was very sick, but I wasn't at all anxious, just sad to be separated from my other child.” Two women reported being asymptomatic from a psychiatric perspective at the time of their consult. Participants were asked to recall who had made the referral to Psychiatry. Approximately 25% did not know—a psychiatry resident and/or staff psychiatrist simply arrived at the bedside to do a consultation. In 11% of cases, the woman recalled that family members or she herself had asked for a referral. A member of the nursing staff was thought to have been instrumental in 29% of cases, and similarly there was a recollection of 29% involvement of either the antenatal or postpartum social worker. Physicians were less likely to be recalled as having made the referral (21%).

mbers or she herself had asked for a referral. A member of the nursing staff was thought to have been instrumental in 29% of cases, and similarly there was a recollection of 29% involvement of either the antenatal or postpartum social worker. Physicians were less likely to be recalled as having made the referral (21%). (b) Diagnosis Study participants were asked to recall whether they had been given a psychiatric diagnosis by the inpatient C-L service and whether they could recall the specific diagnosis: 61% of women could not recall being given a diagnosis; whereas 64% were assigned a DSM-IV-R diagnosis according to the consult note. Hence, 25% of the sample was unaware that a diagnosis had been made. Where women did recall, there was a very high degree of congruence with the diagnosis noted in the chart. (c) Treatment The recommendations for treatment made by the C-L Psychiatry service fell into three, nonmutually exclusive categories (see Table 2) (i) Participation in the “5-day/5-night programme” whereby women remain in hospital for 5 days and 5 nights, while neonates are cared for in the nursery overnight to allow new mothers as much consolidated sleep as possible [7]; (ii) postdischarge treatment; (iii) initiation or continuation of psychotropic medication. Twelve women were offered the 5-day/5-night programme, and 10 women accepted.

(c) Treatment The recommendations for treatment made by the C-L Psychiatry service fell into three, nonmutually exclusive categories (see Table 2) (i) Participation in the “5-day/5-night programme” whereby women remain in hospital for 5 days and 5 nights, while neonates are cared for in the nursery overnight to allow new mothers as much consolidated sleep as possible [7]; (ii) postdischarge treatment; (iii) initiation or continuation of psychotropic medication. Twelve women were offered the 5-day/5-night programme, and 10 women accepted. With regard to postdischarge treatment, at six months there was complete congruence between recommendations articulated in consult notes and participants' recall of these. Women accurately remembered whether C-L psychiatry had suggested contacting the C-L psychiatrist on a per needed outpatient basis (46%), whether the hospital had organized outpatient psychiatric followup prior to discharge (7%), whether followup with the preadmission mental health professional was recommended (14%), or whether no follow-up recommendation was made (25%). Sixty percent (n = 17) of the sample left the hospital with a prescription for psychotropic medication which had either been started or continued in hospital (see Table 3). There was an almost perfect congruence between medications noted in patients' charts and their specific recollections six months after discharge.

With regard to postdischarge treatment, at six months there was complete congruence between recommendations articulated in consult notes and participants' recall of these. Women accurately remembered whether C-L psychiatry had suggested contacting the C-L psychiatrist on a per needed outpatient basis (46%), whether the hospital had organized outpatient psychiatric followup prior to discharge (7%), whether followup with the preadmission mental health professional was recommended (14%), or whether no follow-up recommendation was made (25%). Sixty percent (n = 17) of the sample left the hospital with a prescription for psychotropic medication which had either been started or continued in hospital (see Table 3). There was an almost perfect congruence between medications noted in patients' charts and their specific recollections six months after discharge. 3.2. Compliance with Recommendations for Postdischarge Management Where postdischarge psychiatric care had been suggested (n = 21), 81% of participants followed through by making an appointment; the majority (71%) of these women did so within two months after leaving the hospital. Hence, more than half (54%) of the total sample sought outpatient psychiatric or psychotherapeutic care within eight weeks of leaving the hospital. Among the 19% (n = 5) who did not comply with recommendations for treatment after discharge, women articulated a variety of reasons. I'm still really depressed about a lot of things, but I'm stubborn, and I like to help myself through soul-searching.

3.2. Compliance with Recommendations for Postdischarge Management Where postdischarge psychiatric care had been suggested (n = 21), 81% of participants followed through by making an appointment; the majority (71%) of these women did so within two months after leaving the hospital. Hence, more than half (54%) of the total sample sought outpatient psychiatric or psychotherapeutic care within eight weeks of leaving the hospital. Among the 19% (n = 5) who did not comply with recommendations for treatment after discharge, women articulated a variety of reasons. I'm still really depressed about a lot of things, but I'm stubborn, and I like to help myself through soul-searching. I figured I'd eventually get over it (termination of pregnancy for fetal anomaly). I can't indulge myself; if I go down that path, I'll never get out of it. I felt myself slipping into depression in the hospital, and I hit rock bottom, so I know I can only go up from there. I didn't want a crutch. I wanted to get better. I don't like to leave the baby for long periods and I live fairly far from the hospital. After I go back to work, I'll contact the Psychiatry department.

I figured I'd eventually get over it (termination of pregnancy for fetal anomaly). I can't indulge myself; if I go down that path, I'll never get out of it. I felt myself slipping into depression in the hospital, and I hit rock bottom, so I know I can only go up from there. I didn't want a crutch. I wanted to get better. I don't like to leave the baby for long periods and I live fairly far from the hospital. After I go back to work, I'll contact the Psychiatry department. Regarding compliance with recommendations for psychotropic medication (n = 17), 70% filled the prescription and took the medication as suggested; almost all (91%) of these women had previous experience with psychotropic medication. Twelve percent filled the prescription but stopped taking the medication without consulting their doctor; 18% chose not to fill their prescription, citing the following reasons: worried about side effects on baby; disliked taking medication; felt she did not need medication. 3.3. Concerns Regarding Mood after Discharge When participants were asked whether they had had any concerns about their mood after discharge, 64% (n = 18) reported at least some concern. These fell into four categories. (a) Depression and/or Generalized Anxiety I'm extremely resentful, angry, and depressed. My husband works from 4:30 pm until 2:00 am, and my marriage is disintegrating because of it. I want him home for the bewitching hour, at 5:30.

3.3. Concerns Regarding Mood after Discharge When participants were asked whether they had had any concerns about their mood after discharge, 64% (n = 18) reported at least some concern. These fell into four categories. (a) Depression and/or Generalized Anxiety I'm extremely resentful, angry, and depressed. My husband works from 4:30 pm until 2:00 am, and my marriage is disintegrating because of it. I want him home for the bewitching hour, at 5:30. I'm monitoring my mood and slowly decreasing my medication. My mood is stable now. I try to keep busy. I get out all the time; otherwise, I'd be very low. Depression can consume you if you don't have people around. I have money and resources, but what about all those women who don't? The first few months were the hardest time I've ever had to deal with. (b) Grief Following Fetal/Neonatal Demise or having a Child with an Unexpected Congenital Condition I still feel a lot of sadness, anger, grieving for the baby I thought I'd have. I'm not as depressed as at the beginning. I wouldn't have been ready to go back to work at four months. Maybe now, but I'm not sure. I'm not as motivated in general as I want to be. “I went through the “why me?” I went home with nothing. By now, I'm all worried out.” (c) Anxiety Expressed as Heightened Concern for Health of Neonate/Infant I was a goner when I came home, crying all day long. I was up every hour-and-a-half. I'm still worried that something will happen to the baby—SIDS—so there's a breathing monitor in her crib. But I'm still looking over there all night, making sure she's breathing.

(c) Anxiety Expressed as Heightened Concern for Health of Neonate/Infant I was a goner when I came home, crying all day long. I was up every hour-and-a-half. I'm still worried that something will happen to the baby—SIDS—so there's a breathing monitor in her crib. But I'm still looking over there all night, making sure she's breathing. I'm still anxious, but doing better. I feel I can continue with (psychiatrist). She said I had postpartum anxiety, but was heading toward postpartum depression. It felt like terror. For me it's harder than for most people to keep “normal” in perspective, so if the baby gets sick, I might find it harder to cope. (d) Insomnia Sleeplessness and a lack of bonding were freaking me out. He's a very good baby, but I was so sensitive to his sounds. I was upset with him, upset with me. … I don't have enough money to hire a regular babysitter for relief during the day. Among women reporting some mood concerns, three participants (11%) reported developing depression after discharge, and five (18%) reported a continuation of prepregnancy anxiety.

Sleeplessness and a lack of bonding were freaking me out. He's a very good baby, but I was so sensitive to his sounds. I was upset with him, upset with me. … I don't have enough money to hire a regular babysitter for relief during the day. Among women reporting some mood concerns, three participants (11%) reported developing depression after discharge, and five (18%) reported a continuation of prepregnancy anxiety. I'm feeling guilty about my decision (to terminate for fetal anomaly), although I know it was the right decision. I'm crying all the time. My family is very supportive, but they don't know our secret. … I'm going to call one of the psychiatrists at (the hospital). I'm concerned about my mood. Last week I saw my family doctor. I have no patience with the kids, I'm crying, anxious, and I can't get the image of the fetus out of my head. I'm always wondering what the fetus would have, or could have, been. My husband holds it together for my sake, he's very supportive, but he breaks down occasionally. My doctor thinks I might have postpartum depression. … I have to go on because of my other children, but I often feel like curling up in a ball.

my head. I'm always wondering what the fetus would have, or could have, been. My husband holds it together for my sake, he's very supportive, but he breaks down occasionally. My doctor thinks I might have postpartum depression. … I have to go on because of my other children, but I often feel like curling up in a ball. I thought I was just being a normal mother, having temper tantrums, but my mother said this wasn't normal and that I needed pills. My family doctor said I have psychomotor depression and that's worse than postpartum depression. I have to be a mommy AND a nurse (baby born with renal failure, on dialysis, awaiting transplant). I used to sweat and have panic attacks when the baby cried, but now I'm more confident as a mother, and my boyfriend and I don't fight as much. The Cipralex makes me calmer and tolerant and easy-going. It makes me want to get out and do something, not just wallow. A significant proportion (66%) of women with post-discharge mood concerns were seeing either a mental health professional or their family physician with regard to this issue. Of concern is that 34% had not yet sought help, 17% intending to do so in the near future, and 17% not intending to do so at all. On the positive side, 36% of participants reported a relatively smooth postpartum transition, with no mood concerns; for example,

A significant proportion (66%) of women with post-discharge mood concerns were seeing either a mental health professional or their family physician with regard to this issue. Of concern is that 34% had not yet sought help, 17% intending to do so in the near future, and 17% not intending to do so at all. On the positive side, 36% of participants reported a relatively smooth postpartum transition, with no mood concerns; for example, I have down days, but I'm doing fine. No panic attacks since discharge. I felt different with this birth, no sadness. I was more prepared. The first pregnancy came too quickly after marriage, I was later in age than most mothers with my first, and I had to adjust from being single and independent to being a mother. “My mood is really good, I'm coping well. We're thriving!” Summary of Findings Regarding Congruence and Compliance: reason for referral: 79% congruence between report in consult notes and participants' recollections, diagnosis: 61% recalled no diagnosis given by the C-L psychiatrist (36% of consult notes report no diagnosis); excellent congruence between diagnosis noted and recalled, recommendation for postdischarge therapy: 75% of sample recalled the suggestion to seek therapy; strong congruence between report in consult note and participants' recollections regarding specific mental health professional (e.g., hospital psychiatrist, family doctor, mental health professional seen prior to pregnancy) to contact after discharge,

arge therapy: 75% of sample recalled the suggestion to seek therapy; strong congruence between report in consult note and participants' recollections regarding specific mental health professional (e.g., hospital psychiatrist, family doctor, mental health professional seen prior to pregnancy) to contact after discharge, compliance regarding postdischarge therapy: of women recalling the suggestion for postdischarge therapy, 81% made an appointment and saw either a mental health professional or a family physician, and of these, 71% (i.e., 54% of total sample) saw such a professional within two months after discharge; 19% chose not to seek therapeutic help, recommendation for psychotropic medication: 60% of participants reported having received a prescription before leaving hospital; this is congruent with the percentage reported in consult notes, compliance regarding psychotropic medication: of participants receiving prescriptions, 70% filled their prescription and took their medications as directed by the C-L psychiatrist (91% of these women had previous experience with psychotropic medications); 12% filled their prescription and stopped taking their medication without consulting their physician; 18% did not fill their prescription, mood concerns after discharge: 64% of the sample reported being concerned about their mood within the six months after discharge; 36% reported no such concerns,

compliance regarding psychotropic medication: of participants receiving prescriptions, 70% filled their prescription and took their medications as directed by the C-L psychiatrist (91% of these women had previous experience with psychotropic medications); 12% filled their prescription and stopped taking their medication without consulting their physician; 18% did not fill their prescription, mood concerns after discharge: 64% of the sample reported being concerned about their mood within the six months after discharge; 36% reported no such concerns, decision to seek help regarding concerns: of those women who expressed specific concerns about their mood after discharge, 66% sought help (44% from mental health professional; 22% from family doctor); 17% intend to seek help in the near future; 17% did not intend to seek help even though they had concerns, referral source: 25% could not recall referral source; 29% recalled nurse, 29% recalled social worker on the obstetrical unit, 21% recalled physician as referral source. 3.4. Perceived Benefits and Criticisms of C-L Psychiatry Service (a) Benefits The majority (75%) of participants reported finding the psychiatry consultation helpful in at least one of three ways. (i) Emotional Support/Empathy: At first I hesitated to have a consultation. I'm not crazy! But when it was explained to me, I embraced it. I can't say enough about it, both nursing and psychiatry. They were so interested in seeing me healthy. … You don't feel abandoned and alone. In some places, they just stick you in a room and forget about you.

(i) Emotional Support/Empathy: At first I hesitated to have a consultation. I'm not crazy! But when it was explained to me, I embraced it. I can't say enough about it, both nursing and psychiatry. They were so interested in seeing me healthy. … You don't feel abandoned and alone. In some places, they just stick you in a room and forget about you. I didn't have to hide my feelings; I didn't have to be strong. Nobody tried to say I was wrong to feel what I felt. You feel like a failure if it (pregnancy) doesn't work ‘cause it's your God-given right as a woman to have a baby. It was nice to have someone listen to you, and give you resources in case you needed them. It's a resource for support and help. I was in physical pain (post C-section) and paranoid. I had the sense that somebody cares—the nurses and psychiatrist and resident came often. The psychiatrists were respectful and they asked the right amount of questions. (ii) Initiation/Resumption of Psychotropic Medications. I felt relieved to have (C-L psychiatrist) confirm that I was on the right track by returning to Effexor (venlafaxine). It was extremely helpful because it helped me identify that I needed help. (The C-L psychiatrists) didn't know my history and I didn't want to bring another person into the mix, but they were a conduit to (my regular psychiatrist). They agreed to contact him and he told them to give me a prescription for Paxil. So they were treating symptoms, not root causes. … Social work and psychiatry worked as an integrated team.

idn't know my history and I didn't want to bring another person into the mix, but they were a conduit to (my regular psychiatrist). They agreed to contact him and he told them to give me a prescription for Paxil. So they were treating symptoms, not root causes. … Social work and psychiatry worked as an integrated team. (iii) 5-Day/5-Night Programme: This programme was recommended for 12 women (43%) in our sample, most of whom had either a psychiatric history such as PPD, postpartum psychosis, or bipolar disorder, or were experiencing a particularly low mood and were without adequate social supports. Several women who participated in the programme stated unequivocally that they had not been in shape psychologically to leave the hospital in the usual timely fashion (i.e., after 1-2 days), and the 5-day/5-night programme afforded them time, emotional support, and practical help with their newborn: I was in tears, and (C-L psychiatrist) talked me through it. She recommended the 5-day/5-night programme. I wanted to go home, but was accepting of her advice…. The psychiatrist and social worker worked together, talking with my husband and mom to formulate a plan to help support me. The programme was extremely helpful. It worked for me because I spoke out and this brought a lot of people together, working as a team: the psychiatrist, the resident, social worker, nurses, my family doctor. They discussed what was in my best interest. I was very glad to have it.

o formulate a plan to help support me. The programme was extremely helpful. It worked for me because I spoke out and this brought a lot of people together, working as a team: the psychiatrist, the resident, social worker, nurses, my family doctor. They discussed what was in my best interest. I was very glad to have it. The programme was so helpful. I felt totally taken care of. Someone came everyday from psychiatry, and showed up even over the holiday. The psychiatry resident wasn't a doctor; he said, “Your baby is beautiful”. He was human. Everyone from psychiatry was kind, compassionate, listened well; they were very practical. The human part was so important to me—they were human. Just before we left, I saw (the C-L psychiatrist) again and her resident, and she discussed my Prozac and made certain I had supports in place back home. The programme was wonderful, knowing I could nap when I needed and was close enough to see the baby. I turned down the 5/5 programme because I was feeling anxious about leaving my two-year-old at home. But when I got home, I realized I'd made a mistake. I felt my husband and mom were mad at me for being sad, and so they weren't supportive. (b) Criticisms of the C-L Service Satisfaction with the service provided by C-L Psychiatry was generally high, but there were aspects that women felt needed to be reconsidered or improved upon. One participant reported feeling “judged” by psychiatry; other women found that the consultation exacerbated their sad or negative feelings:

of the C-L Service Satisfaction with the service provided by C-L Psychiatry was generally high, but there were aspects that women felt needed to be reconsidered or improved upon. One participant reported feeling “judged” by psychiatry; other women found that the consultation exacerbated their sad or negative feelings: Talking makes me think more about my problems. I didn't want to cry any more. I'm comfortable with my family doctor; she's not judging me, but I feel psychiatrists judge you. When the psychiatrist and resident asked for my history (severe PPD 7 years earlier during which patient vigorously shook baby), I experienced a lot of guilt. I felt they were digging. I know I was sick in the past, but I hate to think about what I did to my first child. Going over the details made me feel guilty. I felt vulnerable and embarrassed in front of the resident and my husband and the psychiatrist—a whole crowd. I got very upset by the psychiatrist's questions because he was asking me about my stillbirth. I told the nurse after, I explained why I was so upset. When the psychiatrist came back and I explained and after that I felt a bit more comfortable. Then he was understanding and sympathetic. It was hard to open up to people I didn't know. Then it felt weird to leave the hospital, and I was always afraid that someone might think badly of me, that I was a bad mother because I was anxious and afraid to go home. Some women reported feeling not completely listened to/understood:

I got very upset by the psychiatrist's questions because he was asking me about my stillbirth. I told the nurse after, I explained why I was so upset. When the psychiatrist came back and I explained and after that I felt a bit more comfortable. Then he was understanding and sympathetic. It was hard to open up to people I didn't know. Then it felt weird to leave the hospital, and I was always afraid that someone might think badly of me, that I was a bad mother because I was anxious and afraid to go home. Some women reported feeling not completely listened to/understood: The psychiatrist should be aware of the signs when a person needs a soft place, and not be clinical. I could have used a softer touch, someone who could've laughed with me. I wanted to put him at ease. If a person cracks a joke, the psychiatrist should try to go with it. Some women who remained in hospital as part of the 5-day/5-night programme reported that they received mixed messages from nurses and/or lactation consultants; this barrage of confusing information/opinion was perceived as insensitivity to the woman's needs: The lactation consultants pressured me to wake up at night; only one of them acknowledged that I could both breast and bottle feed. If a mother is having a nervous breakdown, then you need to change the rules. It didn't help my depression to be pressured and pushed.

Some women who remained in hospital as part of the 5-day/5-night programme reported that they received mixed messages from nurses and/or lactation consultants; this barrage of confusing information/opinion was perceived as insensitivity to the woman's needs: The lactation consultants pressured me to wake up at night; only one of them acknowledged that I could both breast and bottle feed. If a mother is having a nervous breakdown, then you need to change the rules. It didn't help my depression to be pressured and pushed. I felt relieved after my initial consultation with (C-L psychiatrist) … she arranged for me to get sleep, but the nurses seemed unaware of this option of having the baby sleep in the nursery—for 3 nights they took the baby for an hour or two, but they didn't seem to know the alternate arrangement was possible. 3.5. Patients' Recommendations for Improvements to the C-L Psychiatry Service When asked how the C-L Psychiatry service could be improved, participants offered recommendations that fall into six categories. (a) Time-Related Have the psychiatry resident schedule an appointment with the new mother rather than showing-up unannounced. Set appointment times with patients rather than dropping-by. Have the woman stay an extra day (after termination) and then have the psychiatric consult. I would have been more receptive to information on the third day. I felt so rushed after the fetus was delivered; I needed time to recover. They (psychiatrist and residents) stayed too long, the three of them, for well over an hour. (b) Consistency of Personnel

Have the woman stay an extra day (after termination) and then have the psychiatric consult. I would have been more receptive to information on the third day. I felt so rushed after the fetus was delivered; I needed time to recover. They (psychiatrist and residents) stayed too long, the three of them, for well over an hour. (b) Consistency of Personnel I would've liked more consistency in the people I saw (on bedrest one month, followed by termination). I had to repeat my story so many times, for each resident and psychiatrist. You want to feel like you're important, that it's not just their job, that you're not a lab rat. They need to build trust. I was guarded sometimes because of all this changeover. They should talk to the existing (external) psychiatrist to get the patient's history, and then come in more prepared. Otherwise they make you feel like a textbook case. (c) Sensitivity to What Patient Feels Her Needs are First, ask if a girl wants to talk about it, or whether she's just asking for a sleeping aid. Don't drill into a person. Speak in a conversational, not a doctor, way. Don't say, “How does that make you feel?” Wow, I didn't realize psychiatrists actually say that, and not just on TV! It made me feel so awkward. So I told the psychiatrist what she wanted to hear because I was exhausted and just wanted the damn sleeping pills. A psychiatrist ought to be part of the team, along with the geneticist, social worker, and nurse, that tells the patient about bad news. (d) Group Support for Antenatal Women in Hospital for Extended Periods

First, ask if a girl wants to talk about it, or whether she's just asking for a sleeping aid. Don't drill into a person. Speak in a conversational, not a doctor, way. Don't say, “How does that make you feel?” Wow, I didn't realize psychiatrists actually say that, and not just on TV! It made me feel so awkward. So I told the psychiatrist what she wanted to hear because I was exhausted and just wanted the damn sleeping pills. A psychiatrist ought to be part of the team, along with the geneticist, social worker, and nurse, that tells the patient about bad news. (d) Group Support for Antenatal Women in Hospital for Extended Periods A formal support group should be offered to inpatients to share whatever they're going through. I would have appreciated a group session while I was an inpatient. The lactation course draws tons of women. It would be helpful to have an opportunity to talk about feelings with other women. (e) Consistency of Message between Disciplines Lactation consultants should be educated about the 5/5 programme: “breast is best” might be true, but the rules need to bend for 5/5 patients. Patients should be made aware that they're included in the 5/5 programme so they understand why the nurses want to take their baby away.

(e) Consistency of Message between Disciplines Lactation consultants should be educated about the 5/5 programme: “breast is best” might be true, but the rules need to bend for 5/5 patients. Patients should be made aware that they're included in the 5/5 programme so they understand why the nurses want to take their baby away. (f) Postdischarge Follow-Up When study participants were asked for suggestions regarding postdischarge followup, 92% strongly recommended a telephone call to check up on mental health status postpartum and to provide an offer of help where needed/requested. Although women differed in opinion as to timing (anywhere from 1 to 6 weeks postpartum), each stressed the importance of such a contact. A follow-up call from the hospital would take the pressure off the woman to take the initiative. I would've come in for an appointment, to keep the relationship with (the C-L psychiatrist), but it takes a lot of effort to find the number and make the call, but I would've come in if someone had called me to make an appointment. Do a follow-up call ideally within the first week. If (the woman) doesn't need it, then fine. Do just what you're doing now—how are you doing? Getting enough support? Getting out?—so much emphasis is placed on the baby, but not much attention is paid to the mom postpartum. Follow-up is an excellent idea after one week. Check in with the patient, offer resource information, and mainly offer validation to women. You're the only one I've told how I felt about coming home, that I didn't think how I felt was normal.

Do a follow-up call ideally within the first week. If (the woman) doesn't need it, then fine. Do just what you're doing now—how are you doing? Getting enough support? Getting out?—so much emphasis is placed on the baby, but not much attention is paid to the mom postpartum. Follow-up is an excellent idea after one week. Check in with the patient, offer resource information, and mainly offer validation to women. You're the only one I've told how I felt about coming home, that I didn't think how I felt was normal. Do follow-up calls at one month and so on to check on women…. After a baby is born, the sense of community is gone. You feel alone, like an immigrant. Motherhood is like being a new immigrant; you walk down the street and you're all alone. There shouldn't be too long a gap between discharge and (follow-up); on the other hand, the timing shouldn't be too short because a woman needs time to digest because she's so darned overwhelmed at first. It would have been helpful to have a psychiatrist come to the house shortly after returning home. Also, continuity of care is important—it meant a lot to me to be able to see (the same C-L psychiatrist) both in the hospital and after discharge. I just felt dismissed once we left (the hospital). I recommend a follow-up call at one month, not before, because women are just coming out of baby blues and realizing what's what and whether they need some help. One woman offered a dissenting view: I would not have liked a call because it would have made me feel like I must have a problem if someone is checking on me.

I just felt dismissed once we left (the hospital). I recommend a follow-up call at one month, not before, because women are just coming out of baby blues and realizing what's what and whether they need some help. One woman offered a dissenting view: I would not have liked a call because it would have made me feel like I must have a problem if someone is checking on me. 4. Discussion The current study, set in the obstetrical unit of a tertiary-level, university-affiliated general hospital, provides understanding of the C-L psychiatry consultation process from the perspective of the patient herself. We achieved a participation rate of 45% of all former obstetrical inpatients who had received a consultation during the six-month study period. This response rate is comparable with that reported by others [4, 5, 8]. We acknowledge that our findings are likely biased by the fact that our participants are relatively high functioning, the majority having a university or college qualification. A significant proportion of women could not be included in the study because they had no fixed address or telephone contact number at the time of the consultation. It is possible that these women would be less likely to participate even if we had been able to contact them, and their recall of the consultation with C-L Psychiatry may have been dissimilar to that of the women interviewed. Furthermore, willingness to engage with recommendations for postdischarge care might have been lower.

ossible that these women would be less likely to participate even if we had been able to contact them, and their recall of the consultation with C-L Psychiatry may have been dissimilar to that of the women interviewed. Furthermore, willingness to engage with recommendations for postdischarge care might have been lower. Within the group that did participate, compliance to treatment recommendations, both psychotherapeutic and psychopharmacological, was high. The extent to which compliance is buttressed by family physicians is unclear. We have not been in the practice of sending a copy of the consult note or postdischarge recommendations to the family physician. Rigatelli et al. [5] stress the importance of this step, arguing that the consultation is essentially useless in the absence of followup, since its effects are then limited to the short hospital stay. Given what we know of the impact of postpartum mental illness, and the fact that the involvement of the C-L psychiatrist is usually very brief, contact with family physicians should be considered crucial. Of the women who had concerns regarding mood after discharge, 34% had not yet (at 6 months after discharge) sought help. Bearing in mind that our group was quite highly educated and that considerable effort is made to highlight the importance of postpartum mental health, this figure is concerning. Had their family physicians been made aware of the assessment and recommendations by C-L Psychiatry, the figure might not have been as high.

. Bearing in mind that our group was quite highly educated and that considerable effort is made to highlight the importance of postpartum mental health, this figure is concerning. Had their family physicians been made aware of the assessment and recommendations by C-L Psychiatry, the figure might not have been as high. In general, study participants perceived their interaction with our C-L Psychiatry service in a positive light. A significant proportion did, however, refer to at least one way in which the interaction could be improved. For example, some women had no idea who initiated the consultation—a psychiatrist simply appeared at the bedside without forewarning or the patient's approval. This echoes Phillips et al. [4] who noted that patients were often unaware of the reason for a referral to Psychiatry and/or what was to be achieved from the consultation.

, some women had no idea who initiated the consultation—a psychiatrist simply appeared at the bedside without forewarning or the patient's approval. This echoes Phillips et al. [4] who noted that patients were often unaware of the reason for a referral to Psychiatry and/or what was to be achieved from the consultation. The most pressing issue among our participants was the perceived absence of postdischarge followup (this would have been suggested/arranged where deemed necessary). Some women felt “abandoned” by a system that worked hard to offer a show of concern, but turned its back after discharge, a time when support was most needed. This reflects the findings of Eales et al. [6] who noted that the majority of service users believed they should have been offered followup (a letter, telephone call, or follow-up clinic), and that any followup ought to include a reminder of the treatment plan. Mistiaen and Poot [9] have carried out a comprehensive review of studies which examined the impact of follow-up telephone calls by C-L psychiatry services in the first month after discharge. They found that while no adverse effects were reported, studies show clinically equivalent results between telephone follow-up groups and control groups. Whether or not this holds true for the postpartum population remains to be determined. The feedback received in the current study suggests that postdischarge calls would be perceived as supportive and might encourage some women to seek care in a more timely fashion. The limiting factor is lack of personnel. However, if it can be determined that this is a cost-effective and preventative measure for the postpartum population, a case could be made for ensuring staff (e.g., psychiatric nursing) availability as an essential part of a comprehensive C-L service.

in a more timely fashion. The limiting factor is lack of personnel. However, if it can be determined that this is a cost-effective and preventative measure for the postpartum population, a case could be made for ensuring staff (e.g., psychiatric nursing) availability as an essential part of a comprehensive C-L service. As a result of this study we are now seeking to implement a number of changes on our C-L psychiatry service with obstetrical inpatients, namely, ensure patient understands why we were consulted, by whom, and what she can expect from the consultation, ensure that all disciplines involved in a patient's care are aware of the treatment plan—obstetrical staff, nursing, social work, and where appropriate, lactation consultants, send a discharge note to the patient's primary care physician and/or community psychiatrist, ensure postdischarge followup with mental health services where deemed appropriate, develop an inpatient group for supportive psychotherapy for antenatal women who are in hospital for prolonged periods, provide a follow-up call to all obstetrical C-L patients approximately two weeks after discharge. Table 1 Demographics of study participants (N = 28). Age Relationship Education Current employment status Ethnic/racial affiliation Religious affiliation Years in Canada Other living children Mean age = 34 Married 75% Completed college/university 71% Parental leave 46% “None” 36% “None” 36% Entire life 86% None 54% 1–32% 2–10% 5–4% Common law 18% Some college/university 25% Full-time homemaker 32% Jewish 21% Jewish 21% Philippines 1983; Zimbabwe 1993; Iran 2000; Pakistan 2005

Age Relationship Education Current employment status Ethnic/racial affiliation Religious affiliation Years in Canada Other living children Mean age = 34 Married 75% Completed college/university 71% Parental leave 46% “None” 36% “None” 36% Entire life 86% None 54% 1–32% 2–10% 5–4% Common law 18% Some college/university 25% Full-time homemaker 32% Jewish 21% Jewish 21% Philippines 1983; Zimbabwe 1993; Iran 2000; Pakistan 2005 Single 7% High school 4% Part-time job 7% “White” 11% Roman catholic 18% Seeking employment 7% 4% each: Black, “Black-and-White”, Greek orthodox Iranian, Italian, Pakistani, Philippine, Portuguese 7% each: Muslim, Greek orthodox, Christian Full-time job 4% Protestant 4% Social assistance 4% Table 2 Treatment recommendations and compliance as reported by participants. Categories of treatment suggested (N = 28) Recommendations for postdischarge therapy followup Patients' compliance with treatment recommendations (n = 21) Time between discharge and first appointment with therapist (n = 17) Inpatient 5-day/5-night programme: offered to 12, accepted by 10 (36%) Patient to contact C-L psychiatrist if needed: 46% (n = 13) Made appointment with psychiatrist/therapist: 81% of women for whom any followup was recommended 1 month or less: 47% Postdischarge therapy followup (refer to next column) Postdischarge appointment with psychiatrist arranged by hospital: 7% (n = 2) Did not make appointment: 19% of women for whom any followup was recommended 1-2 months: 24%

Categories of treatment suggested (N = 28) Recommendations for postdischarge therapy followup Patients' compliance with treatment recommendations (n = 21) Time between discharge and first appointment with therapist (n = 17) Inpatient 5-day/5-night programme: offered to 12, accepted by 10 (36%) Patient to contact C-L psychiatrist if needed: 46% (n = 13) Made appointment with psychiatrist/therapist: 81% of women for whom any followup was recommended 1 month or less: 47% Postdischarge therapy followup (refer to next column) Postdischarge appointment with psychiatrist arranged by hospital: 7% (n = 2) Did not make appointment: 19% of women for whom any followup was recommended 1-2 months: 24% Psychotropic medication (refer to Table 3) Patient instructed to follow up with preadmission mental health provider: 14% (n = 4) More than 4 months: 18% Postdischarge therapy strongly urged: 7% (n = 2) No treatment recommendations made: 25% (n = 7) No answer: 12% Table 3 Psychotropic medications: recommendations and compliance as reported by participants. Recommendations and compliance % (N = 28) Left hospital with psychotropic Rx 60% (n = 17) Filled Rx and took medication 70% (91% of these had experience with psych drugs prior to hospital stay) Filled Rx but stopped meds without medical consultation 12% (all had been on meds in recent past) Did not fill Rx 18% No psych medications administered in hospital or prescribed for postdischarge 40% (n = 11)

1. Background Of the approximately four million global neonatal deaths that occur annually, 98% occur in developing countries, where most newborns die at home while they are cared for by mothers, relatives, and traditional birth attendants [1]. During the past two decades, infant mortality rate has declined very slowly. This is as a result of a very slowly declining neonatal mortality rate. There has been relatively little change in neonatal mortality despite proven cost-effective solutions to reduce neonatal mortality, such as promoting tetanus toxoid immunisation, skilled attendance during delivery, immediate and exclusive breastfeeding, and clean cord care [2, 3]. In Nigeria, approximately 67% of births occur at home [4]. In 2005, the infant mortality rate in Nigeria was 64 per 1000 live births, and the neonatal mortality rate was 39 per 1000 per live births [5]. In rural areas of Nigeria, the proportion of institutional deliveries is as low as four percent [4, 6]. Even in urban areas like Lagos, a significant proportion of women (19%) still deliver at home [7]. This is in spite of a relatively easy access to institutional maternity services in urban areas. Previous studies about home deliveries in urban and periurban areas of Abeokuta have reported poor maternal education, multiparity, and low socioeconomic status as the predictors of home deliveries [6]. A study carried out in Sagamu reported “cost” and “convenience” as the reasons for delivering at home [8].

ban areas. Previous studies about home deliveries in urban and periurban areas of Abeokuta have reported poor maternal education, multiparity, and low socioeconomic status as the predictors of home deliveries [6]. A study carried out in Sagamu reported “cost” and “convenience” as the reasons for delivering at home [8]. The World Health Organization (WHO) guidelines for essential newborn care include the following: hygiene during delivery, keeping the newborn warm, early initiation of breastfeeding, care of the eyes, care during illness, immunization, and care of low-birth-weight newborns [9]. Therefore it is necessary for the mother and her family to understand these aspects of childbirth and newborn care and be prepared to react to the potential danger signs. A study from rural areas of Oyo State, Nigeria reported that a very large proportion (>90%) took place at home. The study also reported that only six percent of home deliveries were attended by skilled government health workers and newborn care practices were unhygienic and of high risk [10]. Such high-risk practices have also been reported from remote Osun State, Nigeria [4]. Newborn care practices may change over time and may be different in urban areas. Studies from low socioeconomic settlements of Karachi, Pakistan [11], Bangladesh [12] and Delhi, India [13] have reported that traditional newborn care practices were of high risk and emphasized the need for community-based interventions to promote proper newborn care practices in urban areas.

different in urban areas. Studies from low socioeconomic settlements of Karachi, Pakistan [11], Bangladesh [12] and Delhi, India [13] have reported that traditional newborn care practices were of high risk and emphasized the need for community-based interventions to promote proper newborn care practices in urban areas. Implementation of an effective program for promotion of childbirth and newborn care practices requires understanding of the community and household traditional newborn care practices. Such information will enable the development of programs to promote culturally sensitive and acceptable change in practices. Information about the reasons for delivering at home is also necessary for health care planners to design appropriate maternity services. Information about reasons for delivering at home, home delivery, and newborn care practices in suburban areas of Nigeria is lacking. Therefore, this study was undertaken in a suburban population of Western Nigeria with the following objectives: to describe the home delivery and newborn care practices, to assess the reasons for delivering at home.

Implementation of an effective program for promotion of childbirth and newborn care practices requires understanding of the community and household traditional newborn care practices. Such information will enable the development of programs to promote culturally sensitive and acceptable change in practices. Information about the reasons for delivering at home is also necessary for health care planners to design appropriate maternity services. Information about reasons for delivering at home, home delivery, and newborn care practices in suburban areas of Nigeria is lacking. Therefore, this study was undertaken in a suburban population of Western Nigeria with the following objectives: to describe the home delivery and newborn care practices, to assess the reasons for delivering at home. 2. Methods 2.1. Study Setting Nigeria has a population of about 135 million people, 70% of whom live in rural areas [5]. With an estimated per capital income of $1000 per year, Nigeria is a “poor developing country” in Sub-Saharan region of Africa because of uneven distribution of wealth—few are extremely stinking rich while the majority are languishing in abject poverty. Life expectancy at birth of 47.5 years is low, and it is still lower than neighboring West African countries. Infant mortality rate is among the highest in the region. Due to high maternal mortality rate in women and high mortality from road traffic accident in men, life expectancy for women is slightly higher than that of men (49 versus 47). Gender disparities are also common in terms of literacy. The literacy rate of Nigerian women as a percentage of men is 77% [5]. Curative and preventive health care is organised primarily by the Ministry of Health through hospitals located at federal, state, and local government levels, and primary health centres, health posts, and subhealth posts located at the community level. Private hospitals and clinics exist mostly in urban areas. Missionary and nonprofit hospitals operate in a few areas [14]. Nigeria spends about five percent of its gross domestic product on health of which only one fifth comes from the public sector, and the remainder is paid for by the individual households [5].

evel. Private hospitals and clinics exist mostly in urban areas. Missionary and nonprofit hospitals operate in a few areas [14]. Nigeria spends about five percent of its gross domestic product on health of which only one fifth comes from the public sector, and the remainder is paid for by the individual households [5]. Ogun State is one of the 6 states in the western region of Nigeria. The state has a land area of 2017 square kilometers and a population of 380,527. Ogun State has 20 local government areas of which Sagamu, with a population of 156,312, is one. Sagamu local government is administratively divided into 18 wards. In each of these wards immunisation clinics are conducted once a month in the child health clinics of public and private health institutions. The child health clinics are managed by the national, state, and local governments, and private hospitals, each providing manpower, vaccines, medicines, and technical input, respectively. Since primary immunisation in Nigeria is completed at one year of age, the majority of the children attending these clinics were infants. A few children who missed Measles vaccine between 9 and 12 months are older than a year when they attend the immunisation clinic.

cines, medicines, and technical input, respectively. Since primary immunisation in Nigeria is completed at one year of age, the majority of the children attending these clinics were infants. A few children who missed Measles vaccine between 9 and 12 months are older than a year when they attend the immunisation clinic. 2.2. Study Design and Participants The institutional ethics committee of Olabisi Onabanjo University Teaching Hospital, approved this study. A cross-sectional study was carried out in the immunisation clinics of Sagamu local government. We included the mothers of all the infants who were brought for immunisation during the months of June and July 2008. The framework used for the design and presentation of our study was based on a similar study carried out in rural area of Nepal [14]. A semistructured questionnaire was developed for the purpose of this study and was pretested among 25 mothers during the month of June 2008. After pretesting, the questionnaire was modified according to local traditions and cultural sensitivity. The questionnaire sought information about sociodemographic characteristics of the family, planned or unplanned home delivery, reasons for delivering at home, and the details of events that took place at home from the onset of labour pains till delivery and after birth till initiation of breastfeeding. The details included attendance at delivery, cleanliness and hygiene practices during delivery, thermal control, and infant feeding. The information about the reasons for delivering at home was sought by both open- and closed-ended questions. Those women who reported that they had decided to deliver in a hospital but could not reach the hospital after the onset of labour pains due to various reasons were categorized as unplanned home deliveries. Those women who reported that they had decided to deliver at home were categorized as planned home deliveries.

s. Those women who reported that they had decided to deliver in a hospital but could not reach the hospital after the onset of labour pains due to various reasons were categorized as unplanned home deliveries. Those women who reported that they had decided to deliver at home were categorized as planned home deliveries. Two health workers were trained to administer the questionnaire during a one-day training conducted by the investigator at OOUTH. The health workers were stationed at the registration counter and enquired from the mothers about place of delivery. The mothers of the infants who reported that they delivered at home were invited to participate. Verbal consent was sought and the respondents were assured that the interviewers were not a part of the health service team and services would not be denied if they declined to participate in the interview. After obtaining verbal consent, the health worker carried out the interview and recorded the necessary information on a semistructured questionnaire. The chief investigator and a staff nurse who trained the interviewers supervised all the interviews. The data were coded and analyzed using the SPSS 15.0 for Windows Evaluation Version package (Statistical Package for Social Sciences). Frequencies and percentages of different variables were calculated.

d questionnaire. The chief investigator and a staff nurse who trained the interviewers supervised all the interviews. The data were coded and analyzed using the SPSS 15.0 for Windows Evaluation Version package (Statistical Package for Social Sciences). Frequencies and percentages of different variables were calculated. 3. Results Three hundred and seven out of 1420 infants were brought to the mmunization clinics during the study period. Three hundred and seven were born at home. Seven infants were brought by a family member other than the mother or a relative, and they could not provide reliable information. Hence they were excluded from the analysis. Two hundred (66.7%) of these 300 home deliveries were planned whereas 100 (33.3%) were unplanned. 3.1. Sociodemographic Profile of the Respondents The median age of the infants was 4 months (interquartile range: 4 months). One hundred and forty seven (49%) infants were males and 153 (51%) were females. The median age of the mothers was 26 years (interquartile range: 8 years). More than half of the mothers were Christians (171, 57%), followed by Muslims (99, 33%) and traditional religion worshipers (30, 10%). Forty-three (14.3%) respondents were illiterate, and the majority of the mothers (202, 67.3%) had education of high school and above. The mean monthly family income was 18,360 naira (approximately 122.4 USD).

others were Christians (171, 57%), followed by Muslims (99, 33%) and traditional religion worshipers (30, 10%). Forty-three (14.3%) respondents were illiterate, and the majority of the mothers (202, 67.3%) had education of high school and above. The mean monthly family income was 18,360 naira (approximately 122.4 USD). 3.2. Antenatal Care and Past Obstetric Performance of the Respondents Out of the 300 mothers interviewed, 76 (25.3%) had not gone for antenatal visit and about 91 (30.3%) mothers had at least four antenatal visits as recommended by National Safe Motherhood Program of Nigeria. The majority of women received antenatal care from the publicly funded Sagamu local government health centers. Fifty (16.7%) mothers did not receive tetanus toxoid vaccine during their previous pregnancies; >48 hours and 210 (70%) received at least two doses of tetanus toxoid as recommended by National Safe Motherhood Program. Of the multiparous women, 181 (83.8%) had delivered at home at least once before. Only 100 (46.3%) mothers had at least one institutional delivery in the past. Nine mothers reported of having had a stillbirth (3.0%), 18 a neonatal death (6.0%), and eight a postnatal death (2.7%) after their previous home deliveries.

m. Of the multiparous women, 181 (83.8%) had delivered at home at least once before. Only 100 (46.3%) mothers had at least one institutional delivery in the past. Nine mothers reported of having had a stillbirth (3.0%), 18 a neonatal death (6.0%), and eight a postnatal death (2.7%) after their previous home deliveries. 3.3. Birth Place and Attendance at Delivery The majority (277, 92.3%) of the deliveries took place either in a separate room or inside the house, and the remaining 23 deliveries (7.7%) took place outside the house, either at the backyard or other places. One hundred (33.3%) deliveries were attended by neighbors, 70 (23.3%) were attended by traditional birth attendants and 47 (15.7%) were auxiliary nurse midwife or health assistant, and 47 (15.7%) were attended by auxiliary nurse midwife or health assistant, and 40 (13.3%) were attended by family members (Table 1).