A nationwide survey of diagnostic procedures for abnormal uterine bleeding in Japan.

AIM: To clarify the diagnostic process of the causative disease of abnormal uterine bleeding (AUB) in Japan according to the International Federation of Gynecology and Obstetrics AUB diagnostic system. METHODS: Patients diagnosed with AUB were included in a nationwide survey of AUB conducted during any 2-week period between December 2019 and January 2020. The second survey included information on patient background, AUB symptoms, examinations for diagnosing AUB, the order in which they were performed, and the causative diseases of AUB. RESULTS: Correspondence analysis showed an association between hormonal testing, hysterosalpingography, and magnetic resonance imaging (MRI) in patients with amenorrhea, and heavy menstrual bleeding was strongly correlated with various examinations, such as coagulation tests, pelvic MRI, and endometrial cytology or biopsy. The results also indicated that each AUB causative disease can be diagnosed based on a specific examination profile. CONCLUSION: We clarified the process of diagnosing the causative disease of AUB in our country and determined that it was mainly diagnosed by imaging and pathological examination in cases of structural disease. The high rate of AUB-E and the low rate of AUB-C are possibly associated with specific examination trends in Japan. The results of this study will be useful for the development of a standard protocol for AUB diagnosis in our country.

Cut-off value for anti-Müllerian hormone in the diagnostic criteria for polycystic ovary syndrome in the Japanese population.

AIM: To establish cut-off values for anti-Müllerian hormone (AMH) and antral follicle count (AFC) in the diagnostic criteria for polycystic ovary syndrome (PCOS) applicable to the Japan Society of Obstetrics and Gynecology (JSOG) 2024 criteria and the Rotterdam/International Evidence-Based Guideline for the assessment and management of PCOS (IEBG) 2023 criteria based on a nationwide survey, respectively, taking into account age, assays, and structure of the diagnostic criteria. METHODS: Data were collected for 986 PCOS cases and 965 control cases using a national survey in Japan and used to establish cut-off values for AMH and AFC. RESULTS: Serum AMH levels were significantly higher in the PCOS group compared to the control group. Serum AMH showed a significant negative correlation with age and significant positive correlation with AFC in both groups. In multiple regression analysis, serum AMH level was independently affected by AFC and total testosterone. AMH cut-off values suitable for the JSOG 2024 criteria and the Rotterdam/IEBG 2023 criteria were separately established for the 20-29 and 30-39 years of age groups, respectively, and for Access, Lumipulse and Elecsys/ECLusys, respectively. AFC cut-off values suitable for the JSOG 2024 criteria and Rotterdam/IEBG 2023 criteria were also established separately. AFC exhibited statistically greater variability than AMH. CONCLUSION: The serum AMH level is the biochemical representation of ovarian findings in PCOS and considered objective and highly reliable. Therefore, it could serve as a surrogate for AFC as a marker of polycystic ovarian morphology in diagnostic criteria.

Diagnosis of abnormal uterine bleeding based on the FIGO classification: A systematic review and expert opinions.

AIM: To present evidence- and consensus-based recommendations for the diagnosis abnormal uterine bleeding. METHODS: A literature search for the diagnosis of abnormal uterine bleeding was systematically conducted in PubMed from its inception to May 2024 to identify meta-analyses, reviews, randomized controlled trials, and clinical trials, followed by an additional systematic search using keywords. Based on this evidence, an expert panel developed background, clinical, and future research questions. RESULTS: Based on a systematic search and the collected evidence, we developed five background questions, three clinical questions, and one future research question, with recommendations and/or statements. Evidence and recommendations are provided for clinical questions. Additionally, we developed a flowchart for diagnosis showing the steps of the examinations to be performed. CONCLUSION: The flowchart and nine recommendations/statements specify an efficient diagnostic procedure to differentiate abnormal causative diseases of uterine bleeding optimized for actual Japanese situations.

National survey of abnormal uterine bleeding according to the FIGO classification in Japan.

AIM: To investigate the status of abnormal uterine bleeding (AUB) in Japan using the International Federation of Gynecology and Obstetrics (FIGO) classification (AUB system 1 and 2; PALM-COEIN) and to clarify the relationship between AUB symptoms and the diseases causing AUB. METHODS: In a nationwide study, we enrolled first-time patients who visited target facilities during two consecutive weeks from December 1, 2019 to January 31, 2020. The FIGO classification was used to investigate patients with symptoms and causative diseases of AUB. Based on the proportion of patients in the nationwide study, 373 cases were included in the detailed survey. Survey items included symptoms of AUB according to AUB system 1, examination details, and causative diseases according to the PALM-COEIN classification. RESULTS: Within the study period, we encountered 61 740 first-time patients, of which 8081 (13.1%) were diagnosed with AUB. Among them, 39.9% had abnormal menstrual cycles and 56.9% had abnormal menstrual bleeding. In the survey, PALM had the highest percentage of AUB-L and COEIN had the highest percentage of AUB-O. Correspondence analysis showed that COEIN was strongly associated with abnormal menstrual cycles and PALM with abnormal menstrual bleeding. CONCLUSION: We conducted the first nationwide survey of AUB in Japan. The FIGO classification was a useful tool for the diagnosis of AUB, with a strong correlation between symptoms of AUB by AUB system 1 and the causative disease of AUB by PALM-COEIN. Conversely, a high percentage of AUB-N and AUB-E suggests that AUB system 1 and PALM-COEIN are ambiguous as diagnostic tools.

Manual vacuum aspiration (women's MVA) for endometrial biopsy for patients with suspected endometrial malignancies.

AIM: Endometrial biopsy is generally performed with a metal uterine curette sonde; however, recently, many types of vacuum aspirators are available, including the manual vacuum aspiration (MVA) system. We used the women's MVA system for endometrial sampling and evaluated its effectiveness in determining the presence of endometrial malignancy. METHODS: Forty-seven samples were examined using the following procedures after measuring endometrial thickness by transvaginal ultrasonography: fractional curettage biopsy (Bx; 20 samples), total curettage under general anesthesia (T/C; 13 samples), and MVA (14 samples). The quality of the endometrial samples was classified into four types: 1-4, where 1 denoted poor and 4, good quality. RESULTS: The mean score of the MVA group was significantly higher than that of the partial curettage biopsy group (p = 0.0065). No differences were observed between the MVA and total curettage groups (p = 1.00). When patients were divided into two groups according to endometrial thickness (<10 mm or ≥10 mm) and analyzed, both the MVA and T/C groups did not show a significant difference in their scores compared to the Bx group when the endometrial thickness was <10 mm. However, when the endometrial thickness was ≥10 mm, the MVA and T/C groups had significantly better scores than the Bx group (p = 0.0225 and p = 0.0244, respectively). Vagal reflex, as an adverse event, was observed only in two patients in the Bx group (2/20, 10%). CONCLUSION: Considering its quality and safety, Karman-type MVA for endometrial sampling could be an alternative to fractional curettage using a metallic uterine curette sonde.

National survey of primary amenorrhea and relevant conditions in Japan.

AIM: In Japan, most of the patients with primary amenorrhea or related conditions, such as delayed menarche, are diagnosed by pediatricians or gynecologists; accordingly, the number of the patients and the ratio of the causes were unclear. To clarify them, we conducted a nationwide survey in both the departments for the first time. METHODS: We sent a questionnaire about the patients with chief complaint of no menarche whose first visit was from January 2015 to December 2017, to 596 training institutions for specialist physicians of the Japan Society of Obstetrics and Gynecology and 152 facilities to which councilors of the Japanese Society for Pediatric Endocrinology belong. RESULTS: We received replies from 283 (37.8%) institutions. During the 3 years, 1043 patients first visited pediatrics or gynecology for no menarche. In 303 patients under 16 years old at the first visit, 177 (58.4%) patients had menarche by the age of 16. Of them, 41 (13.5%) patients had menarche spontaneously. Among 308 patients aged 16 to 17 at the first visit, 216 patients were 18 years or older at the survey. Of them, 124 (57.4%) patients had menarche by the age of 18, and 21 (9.7%) of them had menarche spontaneously. The causes of amenorrhea were detected in 462 patients. Abnormal karyotype including Turner syndrome was the most common at 122 (26.4%), followed by Mullerian agenesis at 73 (15.8%). CONCLUSIONS: The first national survey revealed the number and causes of primary amenorrhea and related conditions. This report will provide better information for clinicians.

Annual report of Reproductive Endocrinology Committee, Japan Society of Obstetrics and Gynecology, 2019.

The Japan Society of Obstetrics and Gynecology Reproductive Endocrinology Committee summarizes the activities of each subcommittee below from April 2017 to March 2019. Current important issues regarding reproductive medicine were examined and discussed from social, political, ethical and scientific viewpoints. A nation-wide survey targeted at OB/GYN facilities revealed the usual procedure in diagnosis and management of fertility-desiring POI patients and fertility outcomes of the patients. How to introduce and adapt FIGO AUB systems to obstetric and gynecologic practices in Japan was examined and discussed.

Assisted reproductive technology in Japan: A summary report for 2016 by the Ethics Committee of the Japan Society of Obstetrics and Gynecology.

PURPOSE: The Japan Society of Obstetrics and Gynecology started an online cycle-based assisted reproductive technology (ART) registry system in 2007. This report presents the characteristics and treatment outcomes of ART registered for the cycles practiced during 2016. METHODS: Cycle-specific information for all ART cycles implemented in participating ART facilities were collected. A descriptive analysis was conducted for the registry database of 2016. RESULTS: In total, 447 790 treatment cycles and 54 110 neonates (one in 18.1 neonates born in Japan) were reported in 2016. The mean patients' age was 38.1 years (SD = 4.5). Among the egg retrieval cycles, 104 575 of 251 399 (41.6%) were freeze-all cycles without fresh embryo transfers (ET), while fresh ET was performed in 64 497 cycles (58.4%). A total of 187 132 frozen-thawed ET cycles were reported, resulting in 62 432 pregnancies and 44 484 neonates born. Single ET was selected for 81.0% of fresh transfers and 82.7% of frozen cycles, resulting in singleton pregnancy/live birth rates of 97.0%/96.4% and 96.7%/96.4%, respectively. CONCLUSION: The total ART cycles and subsequent live births continued to increase in 2016. Single ET was performed more than 80%, and ET has shifted from using fresh embryos to frozen ones.

Correction: Nationwide survey for current clinical status of amniocentesis and maternal serum marker test in Japan.

Since the publication of this paper, the authors noticed that Yosuke Fujii was assigned to the incorrect affiliation. The affiliation information is provided correctly, above.

Assisted reproductive technology in Japan: a summary report for 2015 by The Ethics Committee of The Japan Society of Obstetrics and Gynecology.

Purpose: The Japan Society of Obstetrics and Gynecology (JSOG) implemented an assisted reproductive technology (ART) registry system in 1986. Here are reported the characteristics and treatment outcomes of ART cycles that were registered in 2015. Methods: JSOG has requested all participating ART facilities to register cycle-specific information for all ART cycles since 2007. A descriptive analysis was performed by using the registry database for 2015. Results: In total, 424 151 cycles and 51 001 neonates (1 in 19.7 neonates born in Japan) were registered in 2015. The patients' mean age was 38.2 years (standard deviation = 4.5). Among the fresh cycles, 94 158 of 244 718 (38.5%) egg retrieval cycles were cycles with freeze-all embryos or oocytes, while fresh embryo transfer (ET) was performed in 70 254 cycles, signaling a decrease from 2014. There were 169 898 frozen-thawed ET cycles, resulting in 56 355 pregnancies and 40 599 neonates. Single ET was performed at a rate of 79.7% for fresh and 81.8% for frozen cycles and the singleton pregnancy/live birth rates were 96.9%/96.5% and 96.8%/96.4% for the respective cycles. Conclusion: The total ART cycles and live births resulting from ART has been increasing in Japan. Single ET was performed at a rate of almost 80% and ET cycles have shifted from fresh to frozen cycles.

Nomenclature of primary amenorrhea: A proposal document of the Japan Society of Obstetrics and Gynecology committee for the redefinition of primary amenorrhea.

AIM: The aim of this study was to provide medical terms to describe the condition of a girl who should be evaluated for primary amenorrhea in order to facilitate intervention at an appropriate time. METHODS: We performed a literature and clinical guidelines search for recent practices with regard to menarche and discussed relevant cases that had been experienced by committee members. Additionally, we theoretically reviewed medical terms defined in the Glossary Book of Obstetrics and Gynecology in Japan (Japan Society of Obstetrics and Gynecology, 3rd edition). RESULTS: The committee for the redefinition of primary amenorrhea proposed the introduction of two terms and the deletion of one term that had been defined by the Japan Society of Obstetrics and Gynecology, instead of changing the age definition of primary amenorrhea. 'Delayed menarche' was introduced to describe a condition in which a girl has never experienced cyclic menstruation (menarche) by 15-17 years of age. 'Late menarche' was also introduced to describe a condition in which a girl has experienced menarche at 15 years of age or older. 'Delayed menstruation,' which was defined as a condition in which a girl experiences menarche at 15-18 years of age, was deleted. CONCLUSION: The new terms 'delayed menarche' and 'late menarche' were introduced, and the term 'delayed menstruation' was deleted. The new system might help in the early detection and appropriate treatment of primary amenorrhea.

Assisted reproductive technology in Japan: a summary report of 1992-2014 by the Ethics Committee, Japan Society of Obstetrics and Gynecology.

Aim: The Japan Society of Obstetrics and Gynecology implemented a registry report system for the clinical practice of assisted reproductive technology in 1986. The aggregated results from 1992 to 2014 are reported herein. Methods and Results: The total number of registered treatments was 393 745 cycles, of which 66 550 were pregnancy cycles and 46 008 were cycles with a live birth. Compared to the number of registered treatments in 2008, when the cycle-based registry was newly introduced, there was a 2.07-fold increase in the total number of treatments and a 2.25-fold increase in the number of cycles with a live birth. As the average age of patients who receive assisted reproductive technology has become markedly higher year by year, the most common age of those patients who received assisted reproductive technology in 2014 was 40 years. Conclusion: The total numbers of both assisted reproductive technology treatments and assisted reproductive technology live births are likely to be higher in the future. In addition, the trend toward aging patients seems to be continuing into the future.

Nationwide survey for current clinical status of amniocentesis and maternal serum marker test in Japan.

Prenatal testing has been provided in Japan over the past several decades. However, it is difficult to assess the clinical status of amniocentesis (AC) and maternal serum markers (MSM) because obstetricians can perform these tests without registration. This study aims to investigate the current clinical status of AC and MSM in Japan. We conducted a questionnaire study that was intended for a total of 5622 Japanese obstetrics/gynecology facilities during October 2013 to January 2014. The response rate was 40.8% (2295/5622). Of the 2295 facilities, 864 performed MSM (37.7%), 619 performed AC (27.0%) and 412 performed both (18.0%). The average number of MSM tests was 2.0 per month (range 0-52), and the average number of AC tests was 2.4 per month (range 0-30). Involvement of genetic professionals, such as clinical geneticists (CGs) and certified genetic counselors (CGCs), contribute to a content-rich explanation and management of difficult issues and lengthened the explanation time. Nevertheless, relatively few facilities employed these specialists (MSM: 96/864 and AC: 128/619). This is the first study to highlight the current clinical status of AC and MSM tests in Japan. Active involvement of CGs and CGCs can provide more appropriate genetic counseling for prenatal tests.

Identification of epigenetic memory candidates associated with gestational age at birth through analysis of methylome and transcriptional data.

Preterm birth is known to be associated with chronic disease risk in adulthood whereby epigenetic memory may play a mechanistic role in disease susceptibility. Gestational age (GA) is the most important prognostic factor for preterm infants, and numerous DNA methylation alterations associated with GA have been revealed by epigenome-wide association studies. However, in human preterm infants, whether the methylation changes relate to transcription in the fetal state and persist after birth remains to be elucidated. Here, we identified 461 transcripts associated with GA (range 23-41 weeks) and 2093 candidate CpG sites for GA-involved epigenetic memory through analysis of methylome (110 cord blood and 47 postnatal blood) and transcriptional data (55 cord blood). Moreover, we discovered the trends of chromatin state, such as polycomb-binding, among these candidate sites. Fifty-four memory candidate sites showed correlation between methylation and transcription, and the representative corresponding gene was UCN, which encodes urocortin.

Case of chronic ectopic pregnancy diagnosed in which the complete shape of the fetus was visible by ultrasonography.

Preoperative diagnosis of chronic ectopic pregnancy is often difficult because of the high incidence of negative results on pregnancy tests as a consequence of the very small amount of live villi, subtle symptoms, and the poor specificity of ultrasonographic patterns. A 45-year-old woman was referred to our department for evaluation of a mass 8 cm in diameter with solid parts in the right adnexal area. Transvaginal ultrasonography showed a mass consisting of a cystic part with an irregular thick capsule distinct from the right ovary. In the center of the cystic part, a fetus-like image, 20 mm in length was seen. Preoperative diagnosis was confirmed by the laparoscopy, which revealed a swollen right tube containing a fetus with highly necrotic changes. This case was unique because chronic ectopic pregnancy was detected at an early stage before absorption of the conceptus occurred, which coincidentally is an appropriate time for morphological diagnosis.

hScrib, a human homologue of Drosophila neoplastic tumor suppressor, is a novel death substrate targeted by caspase during the process of apoptosis.

hScrib, human homologue of Drosophila neoplastic tumor suppressor, was identified as a target of human papillomavirus E6 oncoprotein for the ubiquitin-mediated degradation. Here, we report that hScrib is a novel death substrate targeted by caspase. Full-length hScrib was cleaved by caspase during death ligands-induced apoptosis, which generates a p170 C-terminal fragments in Hela cells. In vitro cleavage assay using recombinant caspases showed that hScrib is cleaved by the executioner caspases. DNA damage-induced apoptosis caused loss of expression of full-length hScrib, which was recovered by addition of capase-3 inhibitor in HaCat cells. TUNEL positive apoptotic cells, which were identified 4 h after UV irradiation in HaCat cells, showed loss of hScrib expression at the adherens junction. Mutational analysis identified the caspase-dependent cleavage site of hScrib at the position of Asp-504. Although MDCK cells transfected with GFP-fused wild-type hScrib showed loss of E-cadherin expression and shrinkage of cytoplasm by UV irradiation, cells transfected with hScrib with Ala substitution of Asp-504 showed resistance to caspase-dependent cleavage of hScrib and intact expression of E-cadherin. These results indicate that caspase-dependent cleavage of hScrib is a critical step for detachment of cell contact during the process of apoptosis.

Cellular mechanisms of growth inhibition of human endometrial cancer cell line by an antagonist of growth hormone-releasing hormone.

The expression of growth hormone-releasing hormone (GHRH) and its receptors has been demonstrated in peripheral tissues as well as CNS. Recently, the functional splice variant SV1 of GHRH receptor was identified in various human cancers and cancer cell lines. Although antineoplastic activity of GHRH antagonists has been clearly demonstrated, the mechanism of action is incompletely understood. The objective of this study was the investigation of direct anti-proliferative effect of GHRH antagonist MZ-5-156 on HEC-1A human endometrial cancer cell line and the elucidation of underlying mechanisms. RT-PCR revealed the expression of mRNA for GHRH and SV1 of GHRH receptor in HEC-1A cells. MZ-5-156, at concentrations between 10(-7) and 10(-5) M, had a dose-dependent antiproliferative effect on HEC-1A cells, as determined by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, (MTS) assay. Hoechst 33342 staining and flow cytometric analysis indicated that MZ-5-156, at 10(-6) M, induced apoptosis in HEC-1A cells after 48 h of treatment. Western blot analysis of apoptosis-related proteins demonstrated that treatment with MZ-5-156 (10(-6) M) for 48 h significantly increased the protein levels of Fas, phospho-p53 (Ser46), p53AIP1 (p53-regulated Apoptosis-Inducing Protein 1), and caspase-8, -9, and -3, and decreased the protein level of Bcl-2. These results demonstrate that MZ-5-156 can directly inhibit the proliferation of human endometrial cancer cells, which express mRNA for GHRH and SV1 of GHRH receptor, presumably through the induction of p53-dependent apoptosis coupled with the up-regulation of Fas, phospho-p53 (Ser46), p53AIP1, and caspase-8, -9, and -3, and the down-regulation of Bcl-2.

Combined phospho-Akt and PTEN expressions associated with post-treatment hysterectomy after conservative progestin therapy in complex atypical hyperplasia and stage Ia, G1 adenocarcinoma of the endometrium.

Young patients with complex atypical hyperplasia (CAH) or stage Ia, G1 adenocarcinoma (IaG1) of the endometrium, who desire to preserve fertility, can select the conservative therapy by oral progestin, medroxyprogesterone acetate (MPA). However, conservative treatments involve potential risks of progression and recurrence. In an attempt to find out molecular markers for sensitivity to MPA, we performed immunohistochemical analysis of PTEN, phospho-Akt, p53, ER and PgR in MPA-treated 31 cases with CAH or IaG1. Eleven of 12 cases (92%) with CAH and 15 of 19 cases (79%) with IaG1 demonstrated an initial complete response, while five patients underwent hysterectomy due to no response. Four of 11 responders (36%) with CAH and five of 15 responders (33%) with IaG1 later developed relapse. Five of nine patients (56%) with CAH and three of 11 patients (27%) with IaG1 became pregnant after infertility treatment. Immunohistochemical analysis revealed that phospho-Akt expression was significantly decreased by MPA administration (p=0.002). Furthermore, combination of two factors, weak phosho-Akt or PTEN-null expression, was found to be significantly associated with receiving hysterectomy (p=0.04), while each factor showed a trend without statistical significance (p=0.07 and 0.2, respectively). Strong expression of both ER and PgR significantly correlated with successful pregnancy after infertility treatment following complete response to MPA (p=0.02). Our observations in vivo suggest that anti-tumor action of MPA may be mediated by dephosphorylation of Akt, and that immunohistochemical evaluation of phospho-Akt and PTEN may be able to predict the outcome of MPA therapy.

High incidence of tubal dysfunction is determined by laparoscopy in cases with positive Chlamydia trachomatis antibody despite negative finding in prior hysterosalpingography.

Aim: Hysterosalpingography (HSG) is one of the most commonly used methods in order to evaluate the condition of fallopian tubes in infertility clinics. In the present paper, we retrospectively compared the findings of HSG and laparoscopy to elucidate the relationship between tubal dysfunction and background factors, such as Chlamydia trachomatis infection, endometriosis and previous surgery. Methods: We retrospectively reviewed clinical records of 314 patients who were examined by both HSG and laparoscopy between 1996 and 2001 in the Department of Obstetrics and Gynecology, University of Tokyo. Results: When HSG findings were evaluated in reference to those of laparoscopy, sensitivity and specificity for tubal patency were 0.63 and 0.79, respectively, whereas those for peri-tubal adhesion were 0.65 and 0.61, respectively. We compared the percentage of existence of background factors between the patients who were diagnosed as normal by both HSG and laparoscopy (Group L[+]) and those whose fallopian tubes were observed as patent by HSG, but were not patent by chromopertubation under laparoscopy (Group L[-]). The percentage of patients with positive chlamydial antibodies in Group L(-) (42.9%, 15/35) was significantly higher than that of patients with positive chlamydial antibodies in Group L(+) (22.8%, 44/193, P < 0.05). Conclusions: These finding suggested that even if HSG showed normally patent tubes in a patient with positive Chlamydia trachomatis antibodies, the possibility of tubal occulusion still remains high and further examination by laparoscopy is recommended. (Reprod Med Biol 2007; 6: 39-43).

Cross-Talk between Fas/Fas ligand system and nitric oxide in the pathway subserving granulosa cell apoptosis: a possible regulatory mechanism for ovarian follicle atresia.

Recent studies have shown the involvement of Fas/Fas ligand (FasL) system and nitric oxide (NO) in ovarian follicle atresia. Here we asked whether Fas/Fas ligand system interacts with NO using rat granulosa cell culture. Soluble recombinant Fas ligand (rFasL), at 100 ng/ml, significantly decreased cell viability, as measured by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay, in the presence of 200 U/ml interferon-gamma, whereas the concurrent addition of a caspase inhibitor, Z-VAD-FMK, at 20 microm, significantly inhibited rFasL-induced cytotoxicity. Hoechst 33342 staining and flow cytometric analysis confirmed the induction of apoptosis in granulosa cells by 100 ng/ml rFasL in the presence of interferon-gamma, which was blocked by the concomitant addition of an NO donor, S-nitroso-N-acetylpenicillamine. Western blot analysis demonstrated that rFasL significantly up-regulated caspase-3, -8, and -9 activities in granulosa cells, which were attenuated by concurrent treatment with S-nitroso-N-acetylpenicillamine. Real-time quantitative RT-PCR revealed a significant decrease in inducible NO synthase mRNA levels in rFasL-induced apoptotic granulosa cells. In conclusion, we demonstrated the involvement of Fas/FasL system in inducing apoptosis through activation of a caspase-mediated cascade in rat granulosa cells, which is coupled with a decrease in inducible NO synthase expression. We further showed that NO inhibited Fas/FasL system-induced apoptosis by suppressing activation of the caspases, pointing to a cross-talk between Fas/FasL system-induced apoptosis pathway and NO-mediated antiapoptotic pathway in ovarian follicle atresia.