Ovarian reserve and recurrence 1 year post-operatively after using haemostatic sealant and bipolar diathermy for haemostasis during laparoscopic ovarian cystectomy.

RESEARCH QUESTION: Is there a difference in the ovarian reserve 1 year post-operatively in those who used a haemostatic sealant or bipolar diathermy for haemostasis during laparoscopic ovarian cystectomy for ovarian endometriomas? DESIGN: This was an extended follow-up observational study of a previous randomized controlled trial where women aged 18 to 40 years with 3-8 cm unilateral or bilateral endometriomas were randomized to receive haemostasis by a haemostatic sealant or bipolar diathermy following ovarian cystectomy. The primary outcome was the ovarian reserve as assessed by antral follicle count (AFC) 1 year post-operatively. Secondary outcomes included the recurrence rate of ovarian endometrioma, the change in anti-Müllerian hormone (AMH) and FSH concentrations, and reproductive outcomes. RESULTS: The significant increase in AFC at 3 months after initial surgery (P = 0.025) in the haemostatic sealant group compared with the diathermy group was sustained at 1 year (P = 0.024) but there was no difference in AMH or FSH concentrations between the groups throughout the follow-up period. The recurrence rate in the FloSeal group was 7.7% (n = 3/39) compared with 22.2% (n = 8/36) in the diathermy group (P = 0.060). The recurrence rate in women who had bilateral lesions was significantly higher than those with unilateral lesions (risk ratio 5.33, interquartile range 1.55-18.38). No difference in reproductive outcomes was found between the two groups. CONCLUSIONS: Applying haemostatic sealant after laparoscopic cystectomy of ovarian endometriomas produces a significantly greater improvement in AFC, which was apparent at 3-month follow-up, and was sustained at 1-year follow-up without compromising the recurrence rate.

Exacerbated mini-puberty of infancy in an ex-extreme preterm girl.

Vaginal bleeding can occur shortly after delivery in 3%-5% of newborns as a consequence of placental hormone withdrawal . Although usually benign, its differential diagnosis includes central precocious puberty, tumours and other pathological conditions. A girl born at 26 weeks of gestation presented with five episodes of vaginal bleeding, each lasting less than a week, initiated at 4 months of age. Luteinising hormone and oestradiol levels were in the pubertal range. Later, she exhibited breast development, with no other pubertal signs. An ultrasonography test revealed an impregnated endometrium and a right ovarian cyst with 43 mm of diameter. A cranioencephalic MRI was unremarkable. Clinicians adopted expectant management and there was clinical, hormonal and radiological resolution in 3 months. The spontaneous resolution suggested mini-puberty of infancy. This is usually an asymptomatic condition, but to date, four cases of an exacerbated form in extremepremature infants have been reported. Long-term follow-up data are missing.A girl born at 26 weeks of gestation presented with five episodes of vaginal bleeding, each lasting less than a week, initiated at 4 months of age. Luteinising hormone and oestradiol levels were in the pubertal range. Later, she exhibited breast development, with no other pubertal signs. An ultrasonography test revealed an impregnated endometrium and a right ovarian cyst with 43 mm of diameter. A cranioencephalic MRI was unremarkable. Clinicians adopted expectant management and there was clinical, hormonal and radiological resolution in 3 months. The spontaneous resolution suggested mini-puberty of infancy. This is usually an asymptomatic condition, but to date, four cases of an exacerbated form in extremepremature infants have been reported. Long-term follow-up data are missing.

The Abnormal Ovary: Evolving Concepts in Diagnosis and Management.

Ovarian lesions are common and require a consistent approach to diagnosis and management for best patient outcomes. In the past 20 years, there has been an evolution in the approach to abnormal ovarian lesions, with increasing emphasis on reducing surgery for benign disease, standardizing terminology, assessing risk of malignancy through use of evidence-based scoring systems, and triaging suspicious abnormalities to dedicated oncology centers. This article provides an evidence-based review of how these changes in diagnosis and management of ultrasound-detected abnormal ovarian lesions have occurred. Current recommended practices are summarized. The current literature on transvaginal screening for ovarian cancer also is reviewed and summarized.

Hemodynamic changes detected by Doppler ultrasonography in the ovaries of cattle during early development of cystic ovarian disease.

A common reproductive disease in dairy cattle is Cystic Ovarian Disease. To study its development, there was use of an experimental model of follicular persistence to detect hemodynamic changes occurring in ovaries by using Doppler ultrasonography. After estrous synchronization, control cows received no additional treatment and were evaluated at proestrus (CG), whereas treated cows (PG) received sub-luteal doses of progesterone for 15 days and were evaluated at proestrus, and after 0, 5, 10 and 15 days of follicular persistence. Spectral Doppler was used to evaluate blood flow in the ovarian artery, and power Doppler for evaluation of blood flow in the ovarian parenchyma and follicular wall of persistent and dominant preovulatory follicles. Findings using power Doppler signals indicated there were no differences between groups in the parenchyma of both right (P =  0.455) and left (P =  0.762) ovaries. In contrast, power Doppler signals of blood flow were less in walls of persistent follicles from day 0 to 15 when there was follicular persistence than in dominant follicles of the CG (P <  0.001). Blood flow in ovarian arteries was less (P <  0.05) in diastolic velocity and time averaged maximum velocity in all PG groups than in the CG. Peak systolic velocity was less (P <  0.05) in all PG than in the CG, with the exception of P15 (P >  0.05). These findings indicate there are marked changes in blood irrigation area of walls of persistent follicles during the 15 days of follicular persistence.

Contribution of the VEGF system to the follicular persistence associated with bovine cystic ovaries.

Cystic ovaries (CO) characterize a disorder frequently found in dairy cattle. However, despite the contributions by several researchers, the mechanism that leads to ovulatory failure has not yet been completely elucidated. Thus, the aim of this study was to examine the mRNA expression of bovine vascular endothelial growth factor (VEGFA)-164, VEGFA-164b and VEGF receptors (VEGFR1 and VEGFR2) by real-time PCR and protein expression by immunohistochemistry, immunofluorescence and Western blot in follicular fluid from dairy cows with spontaneous CO and in an experimental model of follicular persistence induced by prolonged treatment with progesterone. Results showed that both VEGFA isoforms and receptors were coexpressed in granulosa and theca interna cells and in follicular fluid of ovaries from all the groups evaluated. VEGFA-164, VEGFA-164b and VEGFR2 protein expression was higher in theca cells of persistent follicles from group P0 (expected time of ovulation) than in those from dominant follicles (as reference structure) from the control group (p < 0.05). Also, VEGFA-164 expression was higher in theca cells of cysts than in those of dominant follicles of the control group (p < 0.05). In follicular fluid, VEGFA-164 expression was higher in persistent follicles from group P5 (5 days of follicular persistence) than in the control, P0 and P15 groups, and higher in cysts than in dominant follicles from the control group (p < 0.05). This study provides evidence of an altered expression of VEGFA-164, VEGFA-164b and VEGFR2 during the formation of persistent follicles and cysts in cows. Together, these results evidence that early development of CO in cows is concurrent with an altered expression of these growth factors and that these alterations may contribute to the follicular persistence, angiogenic dysregulation and ovulatory failure found in cows with follicular cysts.

Ovarian function after the use of various hemostatic techniques during treatment for endometrioma: protocol for a randomized clinical trial.

BACKGROUND: Laparoscopic cystectomy is currently considered the gold standard for the treatment of ovarian endometrioma, resulting in an improvement in symptoms, a lower recurrence rate, and a higher pregnancy rate among infertile patients. However, this treatment is not free from risk, since it is associated with a reduction in ovarian reserve. There is still controversy in the literature regarding whether the cause of the reduction in ovarian reserve is due to damage caused by the coagulation energy during hemostasis or whether the procedure itself is the cause of the damage irrespective of the hemostatic method used. The aim of this study is to compare the effects of different hemostatic methods on the ovarian function of women subjected to laparoscopic surgery for ovarian endometrioma. METHODS: An open-label randomized clinical trial to be conducted at the Lauro Wanderley University Hospital between December 2017 and August 2020. Eighty-four patients will be randomly allocated to three groups according to the hemostatic technique used during laparoscopic surgery for ovarian endometrioma: bipolar coagulation; laparoscopic suture; and hemostatic matrix. Ovarian function will be assessed by serum anti-Müllerian hormone measurement and by performing an antral follicle count using ultrasound before surgery and one, three, and six months after surgery. The internal review board of the Medical Sciences Center, Federal University of Paraíba approved the study protocol under reference CAAE 71621717.9.0000.8069. DISCUSSION: Bearing in mind the need for more randomized clinical trials to clarify this issue, we hope to contribute with data that will determine whether there is any difference between hemostatic methods despite the rational use of bipolar energy or whether the procedure itself explains the ovarian damage irrespective of the hemostatic technique used. TRIAL REGISTRATION: ClinicalTrials.gov, NTC03430609 . Registered on XX.10/31/2017. ISRCTN Registry, ISRCTN11469394 . Registered on XX.17/12/2017. Unique Protocol ID: U1111-1203-2508.

Levels of selected cytokines and acute-phase proteins in the serum of dairy cows with cystic ovarian disease and those in follicular and luteal phases of normal ovarian cycle.

The aim of the study was to evaluate the concentration of selected cytokines (tumor necrosis factor [TNFα], interleukin [IL]-6, IL-10) and acute-phase proteins (APPs; haptoglobin [Hp] and serum amyloid A [SAA]) in the serum of cows with follicular and luteal ovarian cysts and in those in the follicular and luteal phases of a physiological ovarian cycle. The study was conducted on 24 Holstein-Friesian (HF) cows: eight cows in the follicular phase of the cycle; the same cows after 10 days when they were in the luteal phase; eight cows with follicular cysts; and eight cows with luteal cysts. Levels of progesterone, 17ß-estradiol, TNF-α, IL-6, IL-10, SAA, and Hp were evaluated in the serum of all examined cows. The concentration of the assessed parameters in the serum was determined using commercially available bovine ELISA kits that were specific for each parameter. The results obtained then showed that values of TNF-α and IL-6 were highest in cows with luteal cysts (p < .001), compared to those with other ovarian structures. The highest level of IL-10 was recorded in cows with two types of ovarian cysts. Furthermore, high values of SAA and Hp were found in cows with two types of cysts; however, these values were higher in cows with follicular cysts. The present study shows that an assessment of the levels of cytokines (pro- and anti-inflammatory) and APPs in the serum may be important in the investigation of the processes underlying the formation and differentiation of ovarian cysts in dairy cows.

Giant Ovarian Mucinous Cystadenoma Complicating Term Pregnancy.

Huge ovarian cyst are found in less than 1% of all ovarian cyst in pregnancy and are associated with poor feto-maternal outcome. A 28 years old G2P1 with history of normal vaginal delivery 3 years back was referred from local health post with complains of intermittent pain abdomen at 29 weeks. Her scan showed huge ovarian cyst of 18.9×17.6 cm with multiple thick septation. Woman was conservatively managed till term and elective surgery was planned however she presented in labour with breech presentation at 39 weeks and 4days. Emergency lower segment caesarian section along with left sided salpingo-oophorectomy was done along with delivery of 2.5 kg healthy female baby. Histopathology was suggestive of mucinous cystadenoma of ovary. Although antepartum removal of ovarian cyst has been recommended to ensure good pregnancy outcome, expectant management and timed intervention can be adopted for pregnancy with huge ovarian cysts. Keywords: mucinous cystadenoma; ovarian cyst; pregnancy.

Vaginal bleeding and a giant ovarian cyst in an infant with 21-hydroxylase deficiency.

BACKGROUND: Increased adrenal androgen hormones in congenital adrenal hyperplasia (CAH) can rarely cause giant ovarian cysts in the neonatal period. Although the exact mechanism of the development of ovarian cysts is unknown, it is thought that increased androgen levels stimulate folicle development by increasing follicle stimulating hormone (FSH) levels. CASE PRESENTATION: A 16-day-old newborn with ambiguous genitalia was presented to our clinic. Laboratory test results were as follows: sodium: 126 mEq/L, potassium: 5.4 mEq/L, renin: 132 pg/mL, adrenocorticotropic hormone (ACTH): 207 pg/mL, cortisole: 7.8 µg/dL, basal 17OH progesterone: 21 ng/mL, androstenedione: 5.1 ng/mL, testosterone: 1188 ng/dL and dehydroepiandrosterone sulfate (DHEAS)>1500 µg/dL. Karyotype analysis resulted in 46,XX. A homozygous mutation of R356W was detected in the CYP21A2 gene. The classical severe form of salt wasting 21 hydroxylase deficiency was diagnosed and treatment was started with hydrocortisone and fludrocortisone. Good metabolic control was ensured by monthly visits but the baby presented with vaginal bleeding as soiling at 4 months. The cystic lesion which extended to the epigastric area from the pelvis in the midline abdomen, had a size of 90×80×60 mm and medially, thin ovarian parenchyma was detected in ultrasonography. CONCLUSIONS: The findings in our patient suggest that a decline in adrenal androgens after glucocorticoid treatment resulted in an increase in gonadotropin levels and the giant cyst is developed by activation of gonadotropin cascade and increased gonadotropin receptors, instead of androgens.

Effect of menstrual phase on the surgical treatment of ovarian cysts.

This study aimed to determine whether the menstrual cycle affects operative bleeding and postoperative ovarian reserve in patients undergoing laparoscopic ovarian cystectomy. A total of 155 patients who had undergone laparoscopic ovarian cystectomy were classified into the follicular phase (n = 84) and luteal phase (n = 71) of the menstrual cycle. The primary outcome measures were operative blood loss and the rate of decline in ovarian reserve, as calculated by measuring serial serum anti-Müllerian hormone (AMH) levels preoperatively and 3 months postoperatively. No significant difference in the baseline demographics, operative blood loss (p = .984), the rate of decline in ovarian reserve (p = .945), and other surgical outcomes were observed between both the groups. These results demonstrate that the menstrual cycle had no influence on the operative blood loss and ovarian reserve during laparoscopic ovarian cystectomy. Therefore, the menstrual cycle is not an important factor to determine the optimal timing of ovarian cystectomy. Impact statement What is already known on this subject?: The menstrual cycle results in periodic changes in haemostasis and blood flow in the reproductive organs. What do the results of this study add?: These results demonstrate that the menstrual cycle had no influence on the operative blood loss and ovarian reserve during laparoscopic ovarian cystectomy. What are the implications of these findings for clinical practice and/or further research?: The menstrual cycle is not an important factor to determine the optimal timing of ovarian cystectomy.

Management of Endometriomas.

Ovarian endometriomas affect 17 to 44% of women with endometriosis, and are often associated with pelvic pain and infertility. Treatment options include expectant management, medical and/or surgical treatment, and in vitro fertilization and embryo transfer (IVF-ET). The choice of treatment depends mostly on the associated symptoms. In most cases, surgery is the preferred choice, since endometriomas do not respond to medical treatment, which may only treat associated pain. In case of infertility, IVF-ET may be a suitable alternative to surgery, particularly when there is no associated pain. According to the best available scientific evidence, laparoscopic excision of the endometrioma wall should be considered the procedure of choice. Concerns have been raised as to the possibility that surgical excision may damage the ovarian reserve, but recent evidences demonstrate that part of the damage may be due to the presence of the endometrioma itself. Indication to surgical treatment should balance the possible risks of damaging the ovarian reserve with the advantages of surgery in terms of satisfactory pain relief rates and pregnancy rates, and of obtaining tissue specimen for ruling out the rare cases of unexpected ovarian malignancy. A score system to guide the clinician in the decision to perform or withhold surgery is presented.

Parity and risk of ovarian cysts: Cross-sectional evidence from the Dongfeng-Tongji cohort study.

Little is known about the association between parity and the risk of ovarian cysts. The aim of this study was to examine the association between parity and the risk of ovarian cysts among a population of Chinese women. A total of 20 502 women aged 45-86 years from the Dongfeng-Tongji Cohort study completed baseline questionnaires, medical examination and provided baseline blood samples. Participants were categorized into four groups according to parity (one, two, three, and four or more live births). Logistic regression models were used to investigate the association between parity and the risk of ovarian cysts. The prevalence of ovarian cysts in the study population was 4.0% (816/20 502). Increasing parity was associated with decreasing risk of ovarian cysts without adjustment for any covariates and after age-adjusted model (P<0.001). After adjusting for potential confounders, women who had had four or more live births had lower risk of ovarian cysts (OR: 0.51; 95% CI: 0.27-0.96) compared with women who had had one live birth. There was a consistent but non-significant decreased risk of ovarian cysts for women who had had two, and three live births (OR: 0.85; 95% CI: 0.68-1.05) and (OR: 0.84; 95% CI: 0.59-1.20) respectively compared with women who had had one live birth. It was concluded that higher parity was associated with decreasing risk of ovarian cysts in this population of Chinese women. These findings could be helpful in decision making in clinical practice for gynecologists when evaluating women suspected to have ovarian cysts.

Large abdominal cystic masses: Report of seven cases.

Cystic abdominal tumours are encountered quite often and are diagnosed more frequently due to the availability of better imaging possibilities. Presentation of huge cysts has become rare as most of them are diagnosed and treated early. But we still have patients with enlarged abdominal cysts; majority with cases of serous cystadenomas of the ovary. Absolute diagnosis is only possible with laparotomy and histopathological findings. In this report, seven patients with enlarged gynaecological or mesenteric cystic masses and gastroenterological symptoms are reported. Four of these cases were serous cystadenoma, two were mucinous cystadenoma and one was a paratubal cyst. Gynaecological tumours and mesenteric cysts should not be missed in female patients showing gastrointestinal symptoms.

Advancing Urinary Protein Biomarker Discovery by Data-Independent Acquisition on a Quadrupole-Orbitrap Mass Spectrometer.

The promises of data-independent acquisition (DIA) strategies are a comprehensive and reproducible digital qualitative and quantitative record of the proteins present in a sample. We developed a fast and robust DIA method for comprehensive mapping of the urinary proteome that enables large scale urine proteomics studies. Compared to a data-dependent acquisition (DDA) experiments, our DIA assay doubled the number of identified peptides and proteins per sample at half the coefficients of variation observed for DDA data (DIA = ∼8%; DDA = ∼16%). We also tested different spectral libraries and their effects on overall protein and peptide identifications and their reproducibilities, which provided clear evidence that sample type-specific spectral libraries are preferred for reliable data analysis. To show applicability for biomarker discovery experiments, we analyzed a sample set of 87 urine samples from children seen in the emergency department with abdominal pain. The whole set was analyzed with high proteome coverage (∼1300 proteins/sample) in less than 4 days. The data set revealed excellent biomarker candidates for ovarian cyst and urinary tract infection. The improved throughput and quantitative performance of our optimized DIA workflow allow for the efficient simultaneous discovery and verification of biomarker candidates without the requirement for an early bias toward selected proteins.

Involvement of PAPP-A and IGFR1 in Cystic Ovarian Disease in Cattle.

Cystic ovarian disease (COD) is one of the main causes of infertility in dairy cattle. It has been shown that intra-ovarian factors, such as members of the insulin-like growth factor (IGF) system, may contribute to follicular persistence. The bioavailability of IGF to initiate its response by binding to specific receptors (IGFRs) depends on interactions with related compounds, such as pregnancy-associated plasma protein A (PAPP-A). The aim of this study was to determine IGFR1 and PAPP-A expression both in follicles at different stages of development and in cysts, to evaluate the roles in the etiopathogenesis of COD in cattle. The mRNA expression of PAPP-A was higher in granulosa cells of large tertiary follicles than in cysts, whereas the protein PAPP-A present in the follicular fluid from these follicles showed no differences. Although no PAPP-A mRNA expression was detected in smaller tertiary follicles, in their follicular fluid, this protease was detected in lesser concentration than in cysts. The mRNA expression of IGFR1 was lower in granulosa cells from cystic follicles than in those from tertiary ones. However, the protein expression of this receptor presented the highest levels in cystic structures, probably to increase the possibility of IGF response. The data obtained would indicate that animals with COD have an altered regulation of the IGF system in the ovary, which could be involved in the pathogenesis of this disease in cattle.

Synchronization of ovulation with human chorionic gonadotropin in lactating dairy cows with ovarian cysts during heat stress.

A study was conducted during hot season to determine the effect of synchronization of ovulation with human chorionic gonadotropin (hCG) on fertility of lactating dairy cows with ovarian cysts. Non cyclic Holstein dairy cows (n = 80) were stratified by parity and diagnosed as having an ovarian cyst. The cows were further identified as follicular or luteal cysts according to the plasma progesterone (P4) concentration and the cystic image of ultrasonography. Cystic cows were randomly assigned to receive treatments (Ovsynch as the control or Ovsynch plus 3000 IU hCG). All cows were artificially inseminated at 16-18 h after the second gonadotropin releasing hormone injection. Cows supplemented with hCG had a greater number of corpus luteum (1.8 ± 0.2 and 0.8 ± 0.3; P < 0.05) and had greater P4 concentration on day 12 than those control cows (6.3 ± 0.3 and 3.9 ± 0.4 ng/ml; P < 0.05). Concentration of cortisol did not differ between groups of cystic cows. No significant differences were found in overall conception rates between the treatments; however, significantly greater conception rate (P = 0.03) was observed in cows with luteal cysts receiving Ovsynch plus hCG. This study highlights that administration of hCG following the Ovsynch-based timed artificial insemination (AI) is more effective than the control Ovsynch by which the hCG affects corpus luteum (CL) development, P4 concentration, and thus improves conception rate in dairy cows with luteal cysts.

Exploratory study of pre-surgical medications with dienogest or leuprorelin in laparoscopic cystectomy of endometrial cysts.

AIM: The aim of this study was to compare the effects of pre-surgical medication with dienogest or leuprorelin on post-surgical ovarian function. MATERIAL AND METHODS: We conducted an exploratory study in two centers in Japan that comprised 30 patients with ovarian endometrial cysts for whom surgical excision was planned. Patients were enrolled and divided into pre-surgical medication groups with dienogest or leuprorelin for 12 weeks. Thereafter, patients were treated by laparoscopic cystectomy. The primary outcome was ovarian function post-surgery, as assessed by serum anti-Müllerian hormone (AMH) level, antral follicle count (AFC) and resumption of menses. Secondary outcome was the effect of pre-surgical medication, as assessed by the size of endometrial cysts and visual analog scale (VAS) score. Serum AMH, AFC, size of endometrial cysts, and VAS scores were measured at baseline (before medication), after medication (1 day before surgery), and at 4 and 12 weeks post-surgery. RESULTS: Serum AMH levels did not change after pre-surgical medication with either dienogest or leuprorelin. Although AMH decreased after surgery, it recovered by 12 weeks post-surgery in both groups with no statistically significant difference. Mean AFC did not change after surgery in either group. Menses returned by 12 weeks post-surgery in all patients except for those who were pregnant. The rate of reduction of endometrial cyst volume did not differ between the groups. Both dienogest and leuprorelin were associated with substantial reductions in VAS scores. CONCLUSION: There were no statistically significant differences between pre-surgical medication with dienogest and leuprorelin in post-surgical ovarian function. Both medications were effective in reducing endometrial cyst volume and VAS score.

Two types of ovarian cortical inclusion cysts: proposed origin and possible role in ovarian serous carcinogenesis.

Ovarian cortical inclusion cysts (CICs) have been long regarded as a possible site of origin of epithelial ovarian carcinoma. It has been proposed that they develop from invagination of ovarian surface epithelium (OSE) which then undergoes metaplasia to form mullerian-type tissue and then undergoes neoplastic transformation. Recent studies have challenged this view, at least for high-grade serous carcinoma, proposing that the latter arise from occult carcinomas in the fallopian tube. Although there is compelling evidence supporting this view, it does not account for the origin of all high-grade serous carcinomas. We have postulated that a subset of high-grade serous carcinoma may develop from CICs, but that they are derived from implantation of tubal epithelium when the OSE is disrupted at ovulation. If true, it would be expected that the number of CICs would increase with age and that CICs would not be present before menarche. To test this hypothesis we examined ovaries removed at autopsy for the presence of CICs and correlated their presence with age. In addition, we used immunohistochemistry for PAX8 (mullerian marker) and calretinin (mesothelial marker). CICs were defined as either ciliated (tubal-type, PAX8-positive) or flat (OSE-type, calretinin-positive). As it has been argued that steroid hormones convert mesothelial-derived OSE to mullerian-type tissue, we performed immunohistochemistry for estrogen and progesterone receptors. CICs lined by tubal-type epithelium were found only in postmenarchial women and 20/21 (95%) were PAX8-positive; none of the 5 flat cysts expressed PAX8 but 4/5 (80%) expressed calretinin. Estrogen receptor was expressed in 1 of 21 (5%) ciliated CICs, whereas it was negative in all 5 flat CICs. Progesterone receptor was expressed in 14 of 21 (66%) ciliated CICs, and in none of the 5 flat cysts. The findings suggest that there are 2 types of CICs, 1 from OSE and 1 from tubal epithelium that probably develop at the time of ovulation.