Assisted reproductive technology in Japan: A summary report for 2021 by the Ethics Committee of the Japan Society of Obstetrics and Gynecology.

Purpose: The Japan Society of Obstetrics and Gynecology (JSOG) registry gathers comprehensive data from registered assisted reproductive technology (ART) facilities in Japan. Herein, we report 2021 ART cycle characteristics and outcomes. Methods: Descriptive statistics were used to summarize and analyze 2021 data. Results: In 2021, 625 ART facilities participated in the registry; 27 facilities did not conduct ART cycles and 598 registered treatment cycles. In total, 498 140 cycles were registered, and there were 69 797 neonates (increases of 10.7% and 15.5%, respectively, from the previous year). The number of freeze-all in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) cycles decreased in 2021; the number of neonates born was 2268 for IVF-embryo transfer (ET) cycles and 2850 for ICSI cycles. Frozen-thawed ET (FET) cycles increased markedly from 2020 (11.2% increase). In 2021, 239 428 FET cycles were conducted, resulting in 87 174 pregnancies and 64 679 neonates. For fresh transfers, the total single ET, singleton pregnancy rate, and singleton live birth rates were 82.7%, 97.0%, and 97.3%; for FET, these rates were 84.9%, 96.9%, and 97.1%. Conclusions: The 2021 Japanese ART registry analysis showed marked increases in both total treatment cycles and live births from the previous year.

Novel therapeutic strategies for injured endometrium: intrauterine transplantation of menstrual blood­derived cells from infertile patients.

BACKGROUND: Menstrual blood-derived cells show regenerative potential as a mesenchymal stem cell and may therefore be a novel stem cell source of treatment for refractory infertility with injured endometrium. However, there have been few pre-clinical studies using cells from infertile patients, which need to be addressed before establishing an autologous transplantation. Herein, we aimed to investigate the therapeutic capacity of menstrual blood-derived cells from infertile patients on endometrial infertility. METHODS: We collected menstrual blood-derived cells from volunteers and infertile patients and confirmed their mesenchymal stem cell phenotype by flow cytometry and induction of tri-lineage differentiation. We compared the proliferative and paracrine capacities of these cells. Furthermore, we also investigated the regenerative potential and safety concerns of the intrauterine transplantation of infertile patient-derived cells using a mouse model with mechanically injured endometrium. RESULTS: Menstrual blood-derived cells from both infertile patients and volunteers showed phenotypic characteristics of mesenchymal stem cells. In vitro proliferative and paracrine capacities for wound healing and angiogenesis were equal for both samples. Furthermore, the transplantation of infertile patient-derived cells into uterine horns of the mouse model ameliorated endometrial thickness, prevented fibrosis, and improved fertility outcomes without any apparent complications. CONCLUSIONS: In our pre-clinical study, intrauterine transplantation of menstrual blood-derived cells may be a novel and attractive stem cell source for the curative and prophylactic therapy for injured endometrium. Further studies will be warranted for future clinical application.

Ethanol potentiates follicle-stimulating hormone action in ovarian granulosa cells.

There are many previous reports on the effects of ethanol on physiological function, including reports of elevated blood estrogen levels in women who drank alcohol. However, the mechanism of ethanol's effects on ovarian functions, such as follicle development and hormone secretion, has not been fully clarified. Therefore, in this study, we investigated the impacts of ethanol on these phenomena and their mechanisms using a primary culture system of rat ovarian granulosa cells (GCs). In the present experiment, groups were created in which follicle-stimulating hormone (FSH) or ethanol was added alone or FSH and ethanol were co-added, and mRNA and protein expression in each group was measured for luteinizing hormone receptor (LHR) and sex steroid hormone synthase, as well as for estradiol (E2) production, cAMP production, and FSH receptor (FSHR) internalization rate. The addition of FSH induced mRNA expression of LHR and aromatase, which led to membrane LHR expression and E2 production. The coexistence of ethanol enhanced all these responses. The action of FSH is exerted via cAMP, and the co-addition of ethanol enhanced this cAMP production. Ethanol alone did not induce cAMP production. The enhancing effect of ethanol was also observed for cAMP induced by cholera toxin. Ethanol had no significant effect on the internalization rate of FSHR. In conclusion, ethanol increased FSH-stimulated cAMP production by increasing the activity of adenylyl cyclase, which enhanced FSH actions in rat GCs. Alcohol is an exacerbating factor in several female hormone-related diseases, and the mechanism of ethanol-induced increase in estrogen secretion revealed in this study may be involved in the pathogenesis of these diseases.

Assisted reproductive technology in Japan: A summary report for 2020 by the ethics Committee of the Japan Society of obstetrics and gynecology.

Purpose: Since 1986, the Japan Society of Obstetrics and Gynecology assisted reproductive technology (ART) registry system has collected data on national ART use and outcomes trends in Japan. Herein, we describe the characteristics and outcomes of ART cycles registered during 2020 and compare the results with those from 2019. Methods and Results: In 2020, 621 ART facilities participated in the registration. The total number of registered cycles was 449 900, and there were 60 381 live births, which decreased from the previous year (1.79% and 0.36% decrease, respectively). The number of freeze-all in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) cycles increased in 2020, and the number of neonates born was 2282 for IVF-embryo transfer (ET) cycles and 2596 for ICSI cycles, which had decreased from the previous year. Frozen-thawed ET (FET) cycles had slightly increased from 2019 (0.04%). In 2020, 215 285 FET cycles were conducted, resulting in 76 196 pregnancies and 55 503 neonates. Single ET was performed in 81.6% of fresh transfers and 85.1% of frozen-thawed cycles, respectively, resulting in over 97% singleton pregnancies/livebirths rates. Conclusion: Despite the COVID-19 pandemic during 2020, the overall number of ART cycles and neonates born demonstrated only a slight decrease in 2020 compared with 2019.

Pregnancy Outcome and Postnatal Chromosome Analysis of the Cord Blood and Chorionic Villi in Two Cases after Intrauterine Transfer of Mosaic Aneuploid Blastocysts.

The long-term prognosis and genetic mechanism of pregnancy after intrauterine mosaic aneuploid blastocyst transfer remain unknown. We report the case of two babies after the aforementioned procedure and chromosomal analysis of their cord blood and chorionic villi. Case Report 1. A 41-year-old primipara, with two previous spontaneous abortions, was pregnant after intrauterine transfer of a blastocyst carrying 40% mosaicism of long-arm monosomy of chromosome 5. The amniocentesis results were 46,XX. A cesarean section was performed at 39 weeks. The female infant was 3,315 g at birth. Case Report 2. A 44-year-old primipara, with two spontaneous abortions, was pregnant after intrauterine transfer of a blastocyst carrying 40% mosaicism of long-arm monosomy of chromosome 9 and monosomy of chromosome 14. After genetic counselling, she decided not to undergo amniocentesis. No abnormalities were found by ultrasound. A cesarean section was performed at 38 weeks. The male infant was 3,340 g at birth. Chromosome analyses of postnatal cord blood and chorionic villi were performed using SNP arrays. The cord blood and chorionic villi showed no chromosomal structural abnormalities or mosaicism. For both, no disorders were observed at 10 months of age. We experienced the birth of babies after intrauterine transfer of mosaic aneuploid blastocysts.

Assisted reproductive technology in Japan: A summary report for 2019 by the Ethics Committee of the Japan Society of Obstetrics and Gynecology.

Purpose: The Japan Society of Obstetrics and Gynecology records online annual cycle-based information for assisted reproductive technology (ART). This report presents the characteristics and treatment outcomes of ART cycles registered during 2019. Methods: The Japanese ART registry includes cycle-specific information from 619 participating facilities, including treatment and pregnancy outcomes. Descriptive analyses were conducted for cycles registered during 2019. Results: In 2019, 458 101 treatment cycles and 60 598 neonates were reported, both of which increased from 2018. The number of fresh cycles, including in vitro fertilization and intracytoplasmic sperm injection, decreased, while frozen-thawed embryo transfer (ET) cycles increased. The mean maternal age was 37.9 years (standard deviation ± 4.7). Of 239 348 oocyte retrievals, 123 690 (51.7%) involved freeze-all-embryos cycles; fresh ET was performed in 41 831 cycles (a decreasing trend since 2015). In 2019, there were 211 597 frozen-thawed ET cycles, resulting in 74 882 pregnancies and 54 168 neonates born. Single ET was performed in 82.6% of fresh transfers and 85.1% of frozen-thawed cycles, with singleton live birth rates of 97.3% for both. Conclusions: The number of fresh cycles decreased but frozen cycles increased in 2019. Single ET was performed in >80% of cases, and the proportion of babies born from frozen-thawed ET increased.

A new deep-learning model using YOLOv3 to support sperm selection during intracytoplasmic sperm injection procedure.

Purpose: To create and evaluate a machine-learning model for YOLOv3 that can simultaneously perform morphological evaluation and tracking in a short time, which can be adapted to video data under an inverted microscope. Methods: Japanese patients who underwent intracytoplasmic sperm injection at the Jikei University School of Medicine and Keiai Reproductive and Endosurgical Clinic from January 2019 to March 2020 were included. An AI model that simultaneously performs morphological assessment and tracking was created and its performance was evaluated. Results: For morphological assessment, the sensitivity and positive predictive value (PPV) of this model for abnormal sperm were 0.881 and 0.853, respectively. The sensitivity and PPV for normal sperm were 0.794 and 0.689, respectively. For tracking performance, among the 51 objects, 40 (78.4%) were mostly tracked, 11 (21.6%) were partially tracked, and 0 (0%) were mostly lost. Conclusions: This study showed that evaluating sperm morphology while tracking in a single model is possible by training YOLO v3. This model could acquire time-series data of one sperm, which will assist in acquiring and annotating sperm image data.

A case of laparoscopically treated broad ligament ectopic pregnancy followed by spontaneous gestation.

We present a case of spontaneous pregnancy after laparoscopic surgery for a broad ligament pregnancy. A 34-year-old nulliparous woman presented with 6 weeks of amenorrhea. Due to the presence of an empty uterus with a 10 mm right adnexal mass on transvaginal ultrasonography and elevated serum human chorionic gonadotropin (hCG), ectopic pregnancy was suspected. Upon diagnostic laparoscopy, the presence of a 2 cm broad ligament ectopic pregnancy was confirmed. Laparoscopic removal of the gestational tissues was performed. Six months after surgery, a spontaneous pregnancy was established. At the 40th week of gestation, a cesarean section was performed due to arrested labor, resulting in live birth. To the best of our knowledge, there have been no reports of a spontaneous pregnancy occurring after laparoscopic surgery for broad ligament pregnancy. Laparoscopic surgery as a treatment option for broad ligament pregnancy may be useful in early gestational age because it can be completed without complications.

Smartphone application improves fertility treatment-related literacy in a large-scale virtual randomized controlled trial in Japan.

People of reproductive age have unmet needs related to deficiencies in fertility literacy. Here, we aimed to investigate whether providing fertility-related information via a smartphone application could improve fertility treatment-related literacy in participants. We performed a randomized control-group pretest posttest study and recruited participants between June 18 and 25, 2020. Participants' fertility treatment-related literacy was assessed with a pretest that comprised of 28 questions and participants were allocated with stratified randomization to either intervention group or control group. The intervention comprised a one-week smartphone application-based provision of information on fertility-related information and the control group received general information about women's healthcare. Effectiveness of intervention was assessed using a posttest. A total of 4137 participants were administered the questionnaire and pretest, among which 3765 participants (91.0 %) responded and were randomly allocated into either the intervention group (N = 1883) or the control group (N = 1882). A significantly higher posttest mean score was observed for the intervention group compared to the control group (P = 0.0017). We also observed that posttest scores were significantly improved compared to pretest scores in both the intervention and control group (P < 0.001). When examining by specific test question, the proportion answering correctly increased at posttest compared to pretest for both intervention and control groups (P < 0.001). Furthermore, the intervention group showed a greater mean difference between posttest and pretest scores than the control group (P < 0.001). In conclusion, educational intervention using a smartphone application contributed to enhancing fertility treatment-related literacy.

A rare case of heterotopic pregnancy after a single embryo transfer: A case report and literature review.

Heterotopic pregnancy (HP) is a rare but life-threatening disease. We report a rare case of HP that occurred after single embryo transfer (SET) with spontaneous natural pregnancy possibly due to sexual intercourse (SI) during assisted reproductive technology treatment and reviewed previous reports. A 39-year-old woman at 7 weeks 5 days' gestation with anti-sperm antibody who underwent a single frozen-thawed embryo transfer in her natural cycle presented with lower abdominal pain and vaginal bleeding. She had several SIs before the day of SET. A viable intrauterine fetus and an extrauterine mass at the right adnexa were detected on transvaginal ultrasonography. An emergent laparoscopic surgery showed a swollen right fallopian tube, and right salpingectomy was performed. Unfortunately, intrauterine fetal death was diagnosed at 19 weeks' gestation. In conclusion, the possibility of HP should be considered in patients with SIs around the day of SET.

Analysis of Surgical Procedure of Four Cases of Ovarian Pregnancies Treated with Laparoscopic Surgery.

Ovarian pregnancy is a rare disease, accounting for 0.5%-3% of ectopic pregnancies. Ovarian pregnancy risk factors and preoperative diagnosis have been extensively reported. However, its histopathology and surgical findings have been poorly studied. To examine appropriate surgical procedures, we investigated the clinical features, surgical findings, and histopathological examinations of four ovarian pregnancy cases treated in our hospital. In histopathological examination, most specimens did not contain ovarian tissues; in some cases, villous tissues were buried in a clot. Therefore, evaluating the appropriateness of surgical resection range from histopathological images was difficult. However, the postoperative course was favorable; no cases manifested complications. Considering all these facts, we regarded the surgical procedures of the four cases in this study as appropriate. For the treatment of ovarian pregnancies, especially for the outward development type, a sufficient therapeutic effect may be achieved even without extensive excision of the ovarian tissues by laparoscopic surgery.

Follicular development during hormone replacement therapy in patients with premature ovarian insufficiency.

PURPOSE: To provide information about the relationship between follow-up period and follicular development in patients with infertility due to premature ovarian insufficiency (POI) who are undergoing hormone replacement therapy (HRT). It is necessary to detect follicle development for artificial insemination or in vitro fertilization. METHODS: This retrospective cohort study was conducted at a university hospital in Tokyo, Japan, from April 2014 to February 2019 in 20 patients [follicular development group, 11 women (55%); non-follicular development group, 9 women (45%)] with POI; their follicular development was followed up weekly. Background characteristics, including age, follicle-stimulating hormone (FSH) and anti-Mullerian hormone levels (AMH), the period from the last spontaneous menstruation to hormone replacement therapy initiation, and follow-up period during HRT were investigated. The period without follicular development was tabulated, and the subsequent cumulative follicular development detection rate was calculated. RESULTS: At least 1-year follow-up, the cumulative follicular development rate was 70%; follicular development was observed with a probability of 49.1% at 3 months, 33.4% at 6 months, and 8.3% at 12 months in the follow-up period. CONCLUSIONS: The results show that the longer the non-follicle development period, the lower the probability of subsequent follicular development in patients with POI during HRT.

Endometrial regeneration with endometrial epithelium: homologous orchestration with endometrial stroma as a feeder.

BACKGROUND: Thin endometrium adversely affects reproductive success rates with fertility treatment. Autologous transplantation of exogenously prepared endometrium can be a promising therapeutic option for thin endometrium; however, endometrial epithelial cells have limited expansion potential, which needs to be overcome in order to make regenerative medicine a therapeutic strategy for refractory thin endometrium. Here, we aimed to perform long-term culture of endometrial epithelial cells in vitro. METHODS: We prepared primary human endometrial epithelial cells and endometrial stromal cells and investigated whether endometrial stromal cells and human embryonic stem cell-derived feeder cells could support proliferation of endometrial epithelial cells. We also investigated whether three-dimensional culture can be achieved using thawed endometrial epithelial cells and endometrial stromal cells. RESULTS: Co-cultivation with the feeder cells dramatically increased the proliferation rate of the endometrial epithelial cells. We serially passaged the endometrial epithelial cells on mouse embryonic fibroblasts up to passage 6 for 4 months. Among the human-derived feeder cells, endometrial stromal cells exhibited the best feeder activity for proliferation of the endometrial epithelial cells. We continued to propagate the endometrial epithelial cells on endometrial stromal cells up to passage 5 for 81 days. Furthermore, endometrial epithelium and stroma, after the freeze-thaw procedure and sequential culture, were able to establish an endometrial three-dimensional model. CONCLUSIONS: We herein established a model of in vitro cultured endometrium as a potential therapeutic option for refractory thin endometrium. The three-dimensional culture model with endometrial epithelial and stromal cell orchestration via cytokines, membrane-bound molecules, extracellular matrices, and gap junction will provide a new framework for exploring the mechanisms underlying the phenomenon of implantation. Additionally, modified embryo culture, so-called "in vitro implantation", will be possible therapeutic approaches in fertility treatment.

Uterine Artery Embolization Combined with Dilation and Curettage for the Treatment of Cesarean Scar Pregnancy: Efficacy and Future Fertility.

PURPOSE: To investigate the efficacy and safety of uterine artery embolization (UAE) followed by dilation and curettage (D&C) as a treatment for cesarean scar pregnancy (CSP) and to assess pregnancy outcomes after the treatment. MATERIALS AND METHODS: We retrospectively analyzed 33 CSP patients treated with UAE followed by D&C. The serum level of beta human chorionic gonadotropin (ß-hCG) normalization, hospitalization, menstruation, and successful pregnancy after treatment was assessed as clinical and pregnancy outcomes. RESULTS: A total of 33 patients were initially treated without severe complications. However, four patients required additional systemic chemotherapy. ß-hCG normalization took 35.5 ± 14.9 days (range 13-79), and the hospitalization was 6.5 ± 2.5 days (2-15). All patients resumed normal menstruation after 36 ± 19.2 days (12-86). Of 16 of 33 patients who desired pregnancy after the treatment, seven patients (43.8%) had uneventful parturition. CONCLUSIONS: UAE combined with D&C was efficient and safe for CSP management. This minimally invasive procedure may be considered as one of the treatment options which enable preservation of fertility after treatment.

High Expression of Ubiquitin C-terminal Hydrolase L1 Is Associated With Poor Prognosis in Endometrial Cancer Patients.

OBJECTIVE: The ubiquitin C-terminal hydrolase L1 (UCHL1) plays a key role in tumor invasion and metastasis. Ubiquitin C-terminal hydrolase L1 is overexpressed in various cancers and reported to be correlated with a poor prognosis. The objective of this study was to determine the prognostic significance of UCHL1 in endometrial cancer. METHODS: The expression of UCHL1 in endometrial cancer was assessed using quantitative reverse transcription polymerase chain reaction and immunohistochemistry in 56 and 215 resected tumor specimens, respectively. RESULTS: The 4-year survival rates of the high UCHL1 messenger RNA expression group and high UCHL1 protein expression group were 78% and 71%, respectively, compared with 96% and 95% for the low UCHL1 messenger RNA expression group and low UCHL1 protein expression group, respectively. Kaplan-Meier and log-rank tests indicated a significant correlation between expression of UCHL1 and disease-free survival and overall survival. Moreover, multivariate stepwise Cox proportional hazard regression model analysis showed that UCHL1 was a significant independent marker for predicting a poor disease-free survival and overall survival. In 43 patients with metastatic lesions, immunohistochemical analysis of metastatic lesions revealed that the recurrence rate and mortality rate were 62% and 41%, respectively, in 29 UCHL1-positive patients and 36% and 29%, respectively, in 14 UCHL1-negative patients. CONCLUSIONS: The results of this study suggest that high UCHL1 expression is a strong marker of poor prognosis of endometrial cancer. Furthermore, we suggest that UCHL1 may be involved in the development of distant metastasis in endometrial cancer.

Expression of the gonadotropin receptors during follicular development.

Background: Gonadotropins induce follicular development that leads to ovulation and luteinization. In women, the level of gonadotropins, along with the expression of their receptors, changes dynamically throughout the menstrual cycle. This study aimed to clarify the mechanisms underlying these phenomena. Methods: The literature was reviewed, including that published by the authors. Main findings Results: Follicle-stimulating hormone receptor expression in the granulosa cells was induced by androgens that were derived from growth differentiation factor-9-stimulated theca cells. In the theca cells, luteinizing hormone receptor (LHR) expression was noted from their appearance. In the granulosa cells, follicle-stimulating hormone (FSH) stimulation was essential for LHR expression. However, FSH alone was not sufficient to respond to the luteinizing hormone (LH) surge for oocyte maturation, ovulation, and subsequent luteinization. To achieve these stages, various local factors that were derived from the granulosa and theca cells in response to FSH and LH stimulation had to work synergistically in an autocrine/paracrine manner to strongly induce LHR expression. Following the LH surge, the LHR expression decreased markedly; miRNAs were involved in this transient LHR downregulation. Following ovulation, LHR expression drastically increased again toward luteinization. Conclusion: The expression of gonadotropin receptors is controlled by sophisticated and complicated systems; a breakdown of this system could lead to ovulation disorders.

Clinical outcomes of infertility treatment for women with adenomyosis in Japan.

Aim: A multicenter, retrospective survey was conducted in order to investigate the current clinical status of adenomyosis in Japan. Methods: The questionnaires covered the management of infertile women with adenomyosis and the outcomes of infertility treatment in women with adenomyosis. The questionnaires were sent to 1149 facilities in Japan. Results: The data were obtained on 535 infertile women with adenomyosis from 190 facilities. Regarding management, infertility treatment was performed without pretreatment for adenomyosis in 37 facilities, after medication in eight facilities, and after an operation in four facilities. Management policies were not established in 106 facilities. Regarding outcomes, the pregnancy rate was 41.7% and the abortion rate was 29.8%. Eighty-five patients received medication and 89 patients underwent surgery as a pretreatment before infertility treatment, while 361 patients had no pretreatment. In relation to the type of adenomyosis, 162 patients had the focal type and 336 patients had the diffuse type. The pregnancy rate and abortion rate were not affected by pretreatment or the type of adenomyosis. Conclusion: The management policy for infertile women with adenomyosis has not been established. The pregnancy rate of infertility treatment is about 40%. There were no data to suggest that medication or surgery as a pretreatment for adenomyosis increased the pregnancy rate in infertile women.

Complications and outcomes of pregnant women with adenomyosis in Japan.

Purpose: To investigate the impact of adenomyosis on the complications and outcomes of pregnancy in Japan. Methods: We carried out a multicenter retrospective questionnaire survey. A questionnaire regarding pregnancy complications and the outcomes of pregnancy was sent to 725 facilities. Results: Data were obtained on the cases of 272 pregnant women with adenomyosis from 65 facilities. The complications of pregnancy included miscarriage before 12 weeks of pregnancy (14.8%), miscarriage after 12 weeks of pregnancy (9.9%), preterm delivery (24.4%), fetal growth restriction (11.8%), pregnancy-induced hypertension (9.9%), intrauterine infection (7.3%), and cervical incompetency (5.3%). The rates of pregnancy complications in the three groups classified according to pretreatment for adenomyosis (no pretreatment, medication, surgery) did not differ to a statistically significant extent. The rates of miscarriage (>12 weeks) and cervical incompetency increased according to the size of the adenomyosis. The rates of pregnancy-induced hypertension and uterine infection in patients with diffuse-type adenomyosis were higher than that in patients with focal-type adenomyosis. Conclusions: Our results show that the increased size and diffuse type of adenomyosis are associated with adverse pregnancy outcome. We should be aware of the higher incidence of pregnancy-induced hypertension and uterine infection in patients with diffuse-type adenomyosis.

Retinoic acid enhances progesterone production via the cAMP/PKA signaling pathway in immature rat granulosa cells.

Retinoic acid (RA) is a metabolite of vitamin A and has important roles in development, differentiation, and reproduction. Activin has been shown to regulate the RA pathway and affect granulosa cell (GC) proliferation, suggesting that RA is important for early follicle development. However, little is known about the effects of RA on GC functions, particularly steroidogenesis, during the early follicle stage. The aim of this study was to investigate the effects of all-trans-RA (atRA) on progesterone production in immature rat GCs cultured without gonadotropin. Our results demonstrated that atRA enhanced progesterone production by upregulating the levels of steroidogenic acute regulatory protein (StAR) and cytochrome P450scc (Cyp11a1) mRNAs, but not 3ß-hydroxysteroid dehydrogenase mRNA in immature rat GCs. Additionally, analysis of the mechanisms through which atRA upregulated StAR and Cyp11a1 mRNAs revealed that atRA enhanced intracellular cAMP accumulation and phosphorylation of cAMP response-element binding protein (CREB). In addition, H-89, an inhibitor of protein kinase A (PKA), abolished the stimulatory effects of atRA, indicating that atRA enhanced progesterone synthesis through cAMP/PKA signaling. In conclusion, our data demonstrated that atRA has a crucial role in progesterone synthesis in rat GCs during the early follicle stage.