Normal Ovarian Function Research Articles

Comparative intrafollicular and plasma iron, ferritin, and transferrin concentrations in cycling mares

Background and Aim: In females of various species and experimental animals, iron (Fe) status in follicular fluid (FF) is associated with local physiological reproductive events related to follicle development, steroidogenesis, and oocyte maturation. However, these mechanisms remain unknown. This study aimed to determine and compare the intrafollicular and plasma concentrations of Fe, ferritin (Ferr), and transferrin (TRF) in cycling mares. Materials and Methods: Sixty ovaries were collected during the breeding season from 30 clinically normal mares raised for slaughterhouse meat production. Blood samples were collected before slaughter. Follicles were classified into three categories according to size: Small (20–30 mm; n = 20), medium (≥31–40 mm; n = 20), and large (≥41 mm; n = 20). The FF samples, after collection, were immediately taken to the laboratory for processing and were centrifuged, and the Fe and Ferr concentrations in the supernatant and plasma were determined by spectrophotometry. Results: Although intrafollicular Fe and Ferr were similar to plasma, TRF was significantly higher in FF than in systemic circulation (p < 0.05). Follicular development does not modify the status of Fe in the mare. Conclusion: Based on this evidence, it is possible that the acquisition of this molecule possibly originated from a local de novo source, whereas their diffusion through ultrafiltration does not play a relevant role. These results provide new scientific insights into the status of follicle Fe, suggesting its involvement in normal ovarian functions in mares. Keywords: ferritin, follicular fluid, iron, mare, transferrin.

Trends and Regional Differences for Fertility Preservation Procedures in Women With Breast Cancer

IntroductionBreast cancer is the most common malignancy in women of reproductive age and chemotherapy protocols impair fertility, frequently necessitating fertility preservation (FP) referral. Embryo, oocyte, or ovarian tissue cryopreservation are established FP modalities in women with breast cancer but there are few data on their uptake over time. In this study our aim was to determine the regional time trends and utility differences for fertility preservation methods of reproductive tissue cryopreservation. MethodsThis multicenter study included 1623 women diagnosed with breast cancer from 7 tertiary centers in 6 countries (Brazil, Italy, Scotland, South Korea, Turkey, USA). Participant centers provided the details of FP cryopreservation approaches broken down annually from 2012 to 2021. Women with newly diagnosed breast cancer, aged 18-45 years who were referred for FP at participating centers and had normal ovarian function at the time were included. ResultsWe found a mean increase of 7% per year (P = .002, adjusting for centers) in the number of women referred for FP. Of those who were referred (n = 1623), a mean 38.7% underwent FP (n = 629), with a range of 12% in South Korea) to 95% in Brazil. The number of women undergoing ovarian stimulation for FP continually increased until 2021, with oocyte cryopreservation being the most common procedure throughout the study period (P = .014 for time trend). The proportion of random start ovarian stimulation cycles increased each year from 58.3% in 2012 to 86.8% in 2021, (P = .005 for time trend, and P = .04 for 2012 vs. 2021). ConclusionsThe utility of FP has steadily increased for young women with breast cancer over the last decade, although regional differences significantly influence FP practices. The findings of our study could have value for policy making in FP care for young women with breast cancer at the local, regional, or global level.

An overview of different methods to establish a murine premature ovarian failure model.

Premature ovarian failure (POF)is defined as the loss of normal ovarian function before the age of 40 and is characterized by increased gonadotropin levels and decreased estradiol levels and ovarian reserve, often leading to infertility. The incomplete understanding of the pathogenesis of POF is a major impediment to the development of effective treatments for this disease, so the use of animal models is a promising option for investigating and identifying the molecular mechanisms involved in POF patients and developing therapeutic agents. As mice and rats are the most commonly used models in animal research, this review article considers studies that used murine POF models. In this review based on the most recent studies, first, we introduce 10 different methods for inducing murine POF models, then we demonstrate the advantages and disadvantages of each one, and finally, we suggest the most practical method for inducing a POF model in these animals. This may help researchers find the method of creating a POF model that is most appropriate for their type of study and suits the purpose of their research.

A Review of the Types of Chemotherapy Drugs Used to Induce Premature Ovarian Failure in Mice

Introduction: The most common complication of chemotherapy is infertility due to premature ovarian failure (POF). POF is defined as the loss of normal ovarian function before age 40, characterized by increased gonadotropin levels, decreased estradiol levels, and diminished ovarian reserve, often leading to infertility. Despite the high impact of POF on general health and quality of life, the pathophysiology of this disease is unclear. For this purpose, animal models provide us with the opportunity to hypothetically investigate the pathogenesis of the disease comprehensively. The most common method of creating an animal model of premature ovarian failure is the use of chemotherapy drugs. In this study, the types of chemotherapy drugs and the relevant molecular pathways that play a role in creating the premature ovarian failure model in mice will be investigated Conclusion: According to the studies, cyclophosphamide drug is introduced as the most common gonadotoxic drug in order to induce POF model in mice.

Endometriosis: A Comprehensive Exploration of Inflammatory Mechanisms and Fertility Implications.

Endometriosis is a prevalent gynecological disorder characterized by the ectopic growth of endometrial-like tissue outside the uterus. This condition poses significant challenges due to its chronic nature, debilitating symptoms such as pelvic pain and infertility, and substantial impact on quality of life. Central to the pathogenesis of endometriosis are inflammatory mechanisms that perpetuate tissue proliferation, adhesion formation, and immune dysregulation within the pelvic cavity. Inflammation plays a pivotal role in the development and progression of endometriosis, influencing the severity of symptoms and complications associated with the disease. Dysregulated immune responses contribute to the persistence of ectopic endometrial implants, exacerbating pelvic pain and other symptoms experienced by affected individuals. Moreover, the inflammatory milieu created by endometriotic lesions disrupts normal ovarian function, impairs follicular development, and compromises reproductive outcomes, thereby posing challenges to fertility. This review comprehensively explores the inflammatory mechanisms underlying endometriosis and their implications for fertility. Synthesizing current research and clinical insights elucidates the intricate interplay between inflammation, disease progression, and reproductive health outcomes. Understanding these complex interactions is essential for developing targeted diagnostic strategies and optimizing therapeutic approaches tailored to alleviate symptoms and improve fertility outcomes in individuals with endometriosis. Ultimately, this review aims to enhance the understanding of endometriosis pathophysiology, inform clinical practice, and stimulate further research to advance personalized care and management strategies for this challenging condition.

Granulosa cell metabolism at ovulation correlates with oocyte competence and is disrupted by obesity and aging.

Is oocyte developmental competence associated with changes in granulosa cell (GC) metabolism? GC metabolism is regulated by the LH surge, altered by obesity and reproductive aging, and, in women, specific metabolic profiles are associated with failed fertilization versus increased blastocyst development. The cellular environment in which an oocyte matures is critical to its future developmental competence. Metabolism is emerging as a potentially important factor; however, relative energy production profiles between GCs and cumulus cells and their use of differential substrates under normal in vivo ovulatory conditions are not well understood. This study identified metabolic and substrate utilization profiles within ovarian cells in response to the LH surge, using mouse models and GCs of women undergoing gonadotropin-induced oocyte aspiration followed by IVF/ICSI. To comprehensively assess follicular energy metabolism, we used real-time metabolic analysis (Seahorse XFe96) to map energy metabolism dynamics (mitochondrial respiration, glycolysis, and fatty acid oxidation) in mouse GCs and cumulus-oocyte complexes (COCs) across a detailed time course in the lead up to ovulation. In parallel, the metabolic profile of GCs was measured in a cohort of 85 women undergoing IVF/ICSI (n = 21 with normal ovarian function; n = 64 with ovarian infertility) and correlated with clinical parameters and cycle outcomes. Our study reveals dynamic changes in GC energy metabolism in response to ovulatory LH, with mitochondrial respiration and glycolysis differentially affected by obesity versus aging, in both mice and women. High respiration in GCs is associated with failed fertilization (P < 0.05) in a subset of women, while glycolytic reserve and mitochondrial ATP production are correlated with on-time development at Day 3 (P < 0.05) and blastocyst formation (P < 0.01) respectively. These data provide new insights into the cellular mechanisms of infertility, by uncovering significant associations between metabolism within the ovarian follicle and oocyte developmental competence. A larger prospective study is needed before the metabolic markers that were positively and negatively associated with oocyte quality can be used clinically to predict embryo outcomes. This study offers new insights into the importance of GC metabolism for subsequent embryonic development and highlights the potential for therapeutic strategies focused on optimizing mitochondrial metabolism to support embryonic development. National Health and Medical Research Council (Australia). The authors have no competing interests. N/A.

Zuogui Pills alleviate cyclophosphamide-induced ovarian aging by reducing oxidative stress and restoring the stemness of oogonial stem cells through the Nrf2/HO-1 signaling pathway

Ethnopharmacological relevanceZuogui Pill (ZGP) is a traditional herbal formula of Chinese Medicine with a long history of use in alleviating ovarian aging. Aim of the studyTo examine the impact of ZGP on oxidative stress and the stemness of oogonial stem cells (OSCs) in cyclophosphamide (CTX)-induced ovarian aging, as well as its molecular mechanisms involving the nuclear factor erythroid 2-related factor 2 (Nrf2, NFE2L2)/heme oxygenase-1 (HO-1, Hmox1) pathway. Materials and methodsFemale Sprague-Dawley (SD) rats were randomly divided into seven groups: control, model (CTX), estradiol valerate (EV, 0.103 mg/kg), ZGP-L (low dose Zuogui Pill, 1.851 g/kg), ZGP-H (high dose Zuogui Pill, 3.702 g/kg), ML385 (30 mg/kg), and ML385+ZGP-L. After CTX modeling, the EV, ZGP-L, ZGP-H, and ML385+ZGP-L groups were treated by gavage for 8 weeks, while the ML385 and ML385+ZGP-L groups were administered the Nrf2 antagonist ML385 twice a week. OSCs were isolated after modeling and then treated with drug serum containing 10% ZGP or 10 μM ML385. The general conditions of the rats, including body weight, ovarian weight/body weight ratio, and estrous cycle, were observed. Ovarian ultrastructure, follicle and corpus luteum counts were assessed via hematoxylin and eosin (H&E) staining. Serum hormone levels were measured using enzyme-linked immunosorbent assay (ELISA). Nrf2/HO-1 pathway, stem cell, germ cell, and cell cycle biomarkers were analyzed by qPCR and Western blot. Cell viability was assessed by cell counting kit-8 (CCK-8) assay. Oxidative stress biomarkers were evaluated using flow cytometry and assay kits. Immunofluorescence was employed to detect and locate OSCs in the ovary, quantify the average fluorescence intensity, and identify OSCs. ResultsAfter ZGP treatment, rats with CTX-induced ovarian aging exhibited improved general condition, increased body weight, higher total ovarian weight to body weight ratio, and a restoration of the estrous cycle similar to the control group. Serum levels of estradiol (E2) and follicle stimulating hormone (FSH), two sex hormones, were also improved. Ovarian ultrastructure and follicle count at all stages showed improvement. Moreover, the viability and proliferation capacity of OSCs were enhanced following ZGP intervention.The Nrf2/HO-1 pathway was found to be down-regulated in CTX-induced aging ovarian OSCs. However, ZGP reversed this effect by activating the expression of Nrf2, HO-1, and NAD(P)H oxidoreductase 1 (NQO1), increasing the activity of antioxidant enzymes superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX), and reducing the accumulation of malonaldehyde (MDA) and reactive oxygen species (ROS), thus restoring resistance to oxidative stress.Additionally, ZGP improved the cell cycle of OSCs, up-regulated the expression of Cyclin D1 and Cyclin E1, restored cell stemness, promoted proliferation, enhanced the expression of cell stemness markers octamer-binding transcription factor 4 (Oct4) and mouse VASA homolog (MVH), and down-regulated the expression of P21, thereby inhibiting apoptosis.The therapeutic effects of ZGP against oxidative stress and restoration of cell stemness were attenuated following inhibition of the Nrf2 signaling pathway using ML385. ConclusionsZGP protected against CTX-induced ovarian aging by restoring normal ovarian function, alleviating oxidative stress in aging OSCs, promoting OSCs proliferation, and restoring their stemness in rats, possibly through regulating the Nrf2/HO-1 pathway.

M6A demethylase FTO-stabilized exosomal circBRCA1 alleviates oxidative stress-induced granulosa cell damage via the miR-642a-5p/FOXO1 axis

BackgroundPremature ovarian insufficiency (POI) is an important cause of female infertility and seriously impacts the physical and psychological health of patients. Human umbilical cord mesenchymal stem cell-derived exosomes (HucMSCs-Exs, H-Exs) have exhibited protective effects on ovarian function with unclear mechanisms.MethodsA comprehensive analysis of the Gene Expression Omnibus (GEO) database were used to identify POI-associated circRNAs and miRNAs. The relationship between HucMSC-derived exosomal circBRCA1/miR-642a-5p/FOXO1 axis and POI was examined by RT-qPCR, Western blotting, reactive oxygen species (ROS) staining, senescence-associated β-gal (SA-β-gal) staining, JC-1 staining, TEM, oxygen consumption rate (OCR) measurements and ATP assay in vivo and in vitro. RT-qPCR detected the expression of circBRCA1 in GCs and serum of patients with normal ovarian reserve function (n = 50) and patients with POI (n = 50); then, the correlation of circBRCA1 with ovarian reserve function indexes was analyzed.ResultsHerein, we found that circBRCA1 was decreased in the serum and ovarian granulosa cells (GCs) of patients with POI and was associated with decreased ovarian reserve. H-Exs improved the disorder of the estrous cycles and reproductive hormone levels, reduced the number of atretic follicles, and alleviated the apoptosis and senescence of GCs in rats with POI. Moreover, H-Exs mitigated mitochondrial damage and reversed the reduced circBRCA1 expression induced by oxidative stress in GCs. Mechanistically, FTO served as an eraser to increase the stability and expression of circBRCA1 by mediating the m6A demethylation of circBRCA1, and exosomal circBRCA1 sponged miR-642a-5p to block its interaction with FOXO1. CircBRCA1 insufficiency aggravated mitochondrial dysfunction, mimicking FTO or FOXO1 depletion effects, which was counteracted by miR-642a-5p inhibition.ConclusionH-Exs secreted circBRCA1 regulated by m6A modification, directly sponged miR-642a-5p to upregulate FOXO1, resisted oxidative stress injuries in GCs and protected ovarian function in rats with POI. Exosomal circBRCA1 supplementation may be a general prospect for the prevention and treatment of POI.Graphical

Primary oocytes with cellular senescence features are involved in ovarian aging in mice

In mammalian females, quiescent primordial follicles serve as the ovarian reserve and sustain normal ovarian function and egg production via folliculogenesis. The loss of primordial follicles causes ovarian aging. Cellular senescence, characterized by cell cycle arrest and production of the senescence-associated secretory phenotype (SASP), is associated with tissue aging. In the present study, we report that some quiescent primary oocytes in primordial follicles become senescent in adult mouse ovaries. The senescent primary oocytes share senescence markers characterized in senescent somatic cells. The senescent primary oocytes were observed in young adult mouse ovaries, remained at approximately 15% of the total primary oocytes during ovarian aging from 6 to 12 months, and accumulated in aged ovaries. Administration of a senolytic drug ABT263 to 3-month-old mice reduced the percentage of senescent primary oocytes and the transcription of the SASP factors in the ovary, in addition, led to increased numbers of primordial and total follicles and a higher rate of oocyte maturation. Our study provides experimental evidence that primary oocytes, a germline cell type that is arrested in meiosis, become senescent in adult mouse ovaries and that senescent cell clearance reduced primordial follicle loss and mitigated ovarian aging phenotypes.

Normal Ovarian Function in Subfertile Mouse with Amhr2-Cre-Driven Ablation of Insr and Igf1r.

Insulin receptor signaling promotes cell differentiation, proliferation, and growth which are essential for oocyte maturation, embryo implantation, endometrial decidualization, and placentation. The dysregulation of insulin signaling in women with metabolic syndromes including diabetes exhibits poor pregnancy outcomes that are poorly understood. We utilized the Cre/LoxP system to target the tissue-specific conditional ablation of insulin receptor (Insr) and insulin-like growth factor-1 receptor (Igf1r) using an anti-Mullerian hormone receptor 2 (Amhr2) Cre-driver which is active in ovarian granulosa and uterine stromal cells. Our long-term goal is to examine insulin-dependent molecular mechanisms that underlie diabetic pregnancy complications, and our conditional knockout models allow for such investigation without confounding effects of ligand identity, source and cross-reactivity, or global metabolic status within dams. Puberty occurred with normal timing in all conditional knockout models. Estrous cycles progressed normally in Insrd/d females but were briefly stalled in diestrus in Igf1rd/d and double receptor (DKO) mice. The expression of vital ovulatory genes (Lhcgr, Pgr, Ptgs2) was not significantly different in 12 h post-hCG superovulated ovaries in knockout mice. Antral follicles exhibited an elevated apoptosis of granulosa cells in Igf1rd/d and DKO mice. However, the distribution of ovarian follicle subtypes and subsequent ovulations was normal in all insulin receptor mutants compared to littermate controls. While ovulation was normal, all knockout lines were subfertile suggesting that the loss of insulin receptor signaling in the uterine stroma elicits implantation and decidualization defects responsible for subfertility in Amhr2-Cre-derived insulin receptor mutants.

YKL-40 Knockdown Decreases Oxidative Stress Damage in Ovarian Granulosa Cells.

Background: Oxidative stress has been implicated in the pathogenesis of polycystic ovarian syndrome (PCOS). To develop novel antioxidant drugs, it is necessary to explore the key regulatory molecules involved in oxidative stress in PCOS. Plasma YKL-40 levels are elevated in patients with PCOS; however, its role remains unclear. Methods: The follicular fluids of 20 women with PCOS and 12 control subjects with normal ovarian function were collected, and YKL-40 in follicular fluids was measured by enzyme-linked immunosorbent assay. A letrozole-induced PCOS rat model was established and the expression level of YKL-40 in the ovaries was detected by immunohistochemistry. KGN cells were treated with H2O2 to generate an ovarian granulosa cell (OGC) model of oxidative stress. The siRNA was transfected into the cells for knockdown. The effect of YKL-40 knockdown on H2O2-treated KGN cells was evaluated by measuring proliferation, apoptosis, activities of T-SOD, GSH-Px, and CAT, levels of MDA, IL-1β, IL-6, IL-8, and TNF-α, and the PI3K/AKT/NF-κB signaling pathway. Results: YKL-40 levels were elevated in the follicular fluids of women with PCOS compared with control subjects with normal ovarian function. The expression level of YKL-40 in the ovaries of rats with PCOS is obviously higher than that in the ovaries of the control group rats. H2O2 treatment enhanced YKL-40 mRNA expression and protein secretion. YKL-40 knockdown enhanced cell proliferation and antioxidant capacity while decreasing apoptosis and inflammatory factor levels in KGN cells following H2O2 treatment. The knockdown activated the PI3K/AKT signaling pathway and suppressed NF-κB nuclear translocation from the cytoplasm. Conclusion: YKL-40 levels were elevated in the follicular fluids of women with PCOS and the ovaries of rats with PCOS. YKL-40 expression can be induced by oxidative stress, and YKL-40 knockdown can decrease oxidative stress damage in OGCs.

Mayer–Rokitansky–Kuster–Hauser Syndrome Type II with Mullerian Duct Aplasia–Renal Dysplasia–Cervical Somite Anomalies Association: A Case Report and Review of Literature

Mayer–Rokitansky–Kuster–Hauser (MRKH) syndrome, also known as Mullerian aplasia/Mullerian agenesis, is characterized by the absence of the uterus and upper two-thirds of the vagina in women who have normal ovarian function and secondary sexual characteristics. The syndrome occurs at a rate of approximately 1 in every 4500–5000 females. Its primary clinical presentation is primary amenorrhea. MRKH syndrome varies in severity and is classified into two types: type I, which is isolated, and type II, involving abnormalities in other organ systems, particularly the kidneys and skeleton. Due to the nature of this condition, individuals with MRKH syndrome may experience significant psychological challenges, highlighting the importance of counseling. This case report describes the diagnosis of type II MRKH syndrome in a 14-year-old girl with primary amenorrhea, also associated with Mullerian duct aplasia–renal dysplasia–cervical somite anomalies.

Tannic acid alleviates 3-nitropropionic acid-induced ovarian damage in Brandt's vole (Lasiopodomys brandtii).

Tannic acid (TA) is a polyphenol with antioxidant properties present in various plants. In this study, we explored the protective effect of TA against ovarian oxidative stress in Brandt's voles and its underlying mechanism. At various doses, 3-nitropropionic acid (3-NPA) was intraperitoneally injected into Brandt's voles to simulate ovarian oxidative stress. Thereafter, various doses of TA were intragastrically administered to examine the protective effect of TA against 3-NPA-induced ovarian damage. Changes in inflammation, autophagy, apoptosis, and oxidative stress-related factors were investigated through various biochemical and histological techniques. Ovarian oxidative stress was successfully induced by the intraperitoneal administration of 12.5 mg/kg 3-NPA for 18 days. As a result, the ovarian coefficient decreased and ovarian tissue fibrosis was induced. TA treatment effectively alleviated the increase in luteinizing hormone and follicle-stimulating hormone levels; the decrease in estradiol, progesterone, and anti-Müllerian hormone levels; and the decline in fertility induced by 3-NPA. Compared to that in the 3-NPA group, TA decreased the expression of autophagy-related proteins beclin-1 and LC3, as well as the level of apoptosis. It also activated the AKT/mTOR signaling pathway, downregulated PTEN and p-NF-κB expression, and upregulated Nrf2 expression. In conclusion, our findings indicate that TA could inhibit autophagy via the regulation of AKT/mTOR signaling, suppressing oxidative damage and inflammatory responses through Nrf2 to alleviate 3-NPA-induced ovarian damage. Collectively, the current findings highlight the protective effects of TA in Brandt's vole, where it promotes the maintenance of normal ovarian function.

Long-Term Ovarian Function Assessment After Haematopoietic Stem Cell Transplantation in Female Sickle Cell Anaemia Survivors.

Haematopoietic stem cell transplantation (HSCT) is a potentially curative treatment for sickle cell anaemia (SCA). While HSCT offers the possibility of disease remission, it can also lead to long-term complications, including gonadal dysfunction and premature menopause. We conducted a retrospective cohort study of female survivors who had hydroxyurea therapy and those who underwent post-HSCT follow-up for SCA at a teaching hospital in Lagos, Nigeria, between January 2019 and December 2022. Participants were eligible if they were at least five years post-HSCT or hydroxyurea treatment and had available serum samples for markers of ovarian function measurement. Demographic and clinical data were collected from the hospital register and patients' medical records. Serum levels of oestradiol, luteinizing hormone (LH), follicle-stimulating hormone (FSH), and anti-Müllerian hormone (AMH) were measured using the Abbott Architect i1000SR chemiluminescent immunoassay analyzer (Abbott Diagnostics, Abbott Park, IL). Descriptive statistics and inferential analyses were used to assess the relationship between markers of ovarian function (FSH and AMH) and clinical parameters. There were statistically significant differences in the median serum levels of all the assessed endocrine hormones between the HSCT and non-HSCT (hydroxyurea) groups of SCA survivors. Up to 82.6% of the SCA survivors experienced ovarian dysfunction after HSCT treatment. Impaired ovarian function in SCA survivors was associated with a longer median follow-up duration than in SCA survivors who had normal ovarian function (12.0 vs. 7.5 years, p = 0.048). There were higher odds of impaired ovarian function in the SCA survivors who had myeloablative regimens than in those who had reduced intensity conditioning regimens (94.1% vs. 50.0%, p = 0.040). Our study highlights the significant impact of HSCT on long-term ovarian function in female SCA survivors. However, further prospective studies with larger sample sizes and longer follow-up periods are required to confirm our findings and elucidate the factors influencing ovarian function in SCA survivors of HSCT. In addition, studies are also needed to further elucidate the optimal transplant protocols and fertility preservation strategies to minimize gonadal toxicity and preserve reproductive potential in female SCA patients undergoing HSCT.

The impact of chronic insomnia disorder on menstruation and ovarian reserve in childbearing-age women: A cross-sectional study.

Diminished ovarian reserve (DOR) is a disorder characterized by impaired ovarian function. Sleep disorders are disruptions of the circadian rhythm, which appears to be closely linked to reproductive systems. This study aimed to investigate the impact of poor sleep quality on the ovarian reserve of childbearing-age women. A cross-sectional study was conducted in China from June 2021 to March 2023. In total, 102 participants diagnosed with chronic insomnia disorder were included in the study. Questionnaires were administered to assess participants' menstrual patterns, insomnia severity, anxiety, and depression. The anti-Müllerian hormone level and the basal antral follicle count were measured for ovarian reserve evaluation. Correlation analysis and ordinal logistic regression analysis were conducted. The women with insomnia presented high percentages of hypomenorrhea, premenstrual syndrome, and dysmenorrhea (78.4%, 74.5%, and 46.1%, respectively). Severe sleep disorder in the past month was identified as an independent risk factor for hypomenorrhea and premenstrual syndrome (odds ratio [OR], 2.64 and OR, 2.688; p<0.05). The prevalence of DOR among women with insomnia (33.3%) was significantly higher than the average reported in previous studies for young women. Insomnia duration exceeding 1 year was determined to be an independent risk factor for DOR in women aged 36 to 40 years (OR, 4.5; p=0.033). This study highlights the association between sleep disorders and menstrual problems. Prolonged poor sleep quality in women aged 36 to 40 years was identified as a significant risk factor for DOR. We should pay more attention to improving sleep quality in order to maintain normal ovarian function.

Cellular senescence in the pathogenesis of ovarian dysfunction.

The follicular microenvironment is crucial for normal ovarian function, and intra-ovarian factors, in coordination with gonadotropins, contribute to its regulation. Recent research has revealed that the accumulation of senescent cells worsens the adverse environment of various tissues and plays critical roles in chronological aging and various pathological conditions. Cellular senescence involves cell-cycle arrest, a senescence-associated secretory phenotype (SASP), macromolecular damage, and dysmetabolism. In this review, I summarize the latest knowledge regarding the role of cellular senescence in pathological conditions in the ovary, in the context of reproduction. Specifically, cellular senescence is known to impair follicular and oocyte health in cisplatin- and cyclophosphamide-induced primary ovarian insufficiency and to contribute to the pathogenesis of polycystic ovary syndrome (PCOS). In addition, cellular senescence is induced during the decline in ovarian reserve that is associated with chronological aging, endometriosis, psychological stress, and obesity, but it remains unclear whether it plays a causative role in these conditions. Finally, I discuss the potential for use of cellular senescence as a novel therapeutic target. The modification of SASP using a senomorphic and/or the elimination of senescent cells using a senolytic represent promising therapeutic strategies. Further elucidation of the role of cellular senescence in the effects of various insults on ovarian reserve, including chronological aging, as well as in pathogenesis of ovarian pathologies, including PCOS, may facilitate a new era of reproductive medicine.

FOXL2 and NR5A1 induce human fibroblasts into steroidogenic ovarian granulosa-like cells.

Human granulosa cells in different stages are essential for maintaining normal ovarian function, and granulosa cell defect is the main cause of ovarian dysfunction. To address this problem, it is necessary to induce functional granulosa cells at different stages in vitro. In this study, we established a reprogramming method to induce early- and late-stage granulosa cells with different steroidogenic abilities. We used an AMH-fluorescence-reporter system to screen candidate factors for cellular reprogramming and generated human induced granulosa-like cells (hiGC) by overexpressing FOXL2 and NR5A1. AMH-EGFP+ hiGC resembled human cumulus cells in transcriptome profiling and secreted high levels of oestrogen and progesterone, similar to late-stage granulosa cells at antral or preovulatory stage. Moreover, we identified CD55 as a cell surface marker that can be used to isolate early-stage granulosa cells. CD55+ AMH-EGFP- hiGC secreted high levels of oestrogen but low levels of progesterone, and their transcriptome profiles were more similar to early-stage granulosa cells. More importantly, CD55+ hiGC transplantation alleviated polycystic ovary syndrome (PCOS) in a mouse model. Therefore, hiGC provides a cellular model to study the developmental program of human granulosa cells and has potential to treat PCOS.

L-carnitine fails to rescue chemotherapy injured ovaries by epigenetic changes of transcription factors

Cyclophosphamide (CP), as an anti-cancer drug, is frequently used to treat various types of cancer. A decreased number of ovarian follicles impaired normal ovarian function, and subsequent premature ovarian failure (POF) presented as a side effect of cyclophosphamide usage. These events may eventually affect the fertility rate of individuals. The present study showed the effect of cyclophosphamide on ovarian reserves and the protective effect of L-carnitine (LC) as an antioxidant to prevent POF.To design the study, six to eight-week-old NMRI female mice were divided into three groups: control, cyclophosphamide (CP), and cyclophosphamide + L-carnitine (CP + LC). Mice received drugs intraperitoneally (IP) for 21 days. In the following 24 h after the last injection, both ovaries were used to evaluate the expression of Sohlh1 and Lhx8 genes by Real-time PCR. Furthermore, the alteration of Lhx8 promoter methylation was examined by Methylation-sensitive high-resolution melting analysis (MS-HRM).The present data showed the negative effect of CP on regulator genes of oogenesis including Sohlh1 and Lhx8. In addition, an examination of the epigenetic status of the Lhx8 gene showed a change in promoter methylation of this gene following cyclophosphamide injection. Although, L-carnitine is an effective antioxidant in relieving oxidative stress caused by cyclophosphamide and its damage, in the present study, however, the use of L-carnitine failed to protect the ovaries from changes caused by CP injection. So, using cyclophosphamide can alter the expression of folliculogenesis genes through its effects on epigenetic changes and may cause POF. The results of the present study showed that L-carnitine consumption can’t protect the ovaries against the adverse effects of CP.

#140 : Combined Use of Letrozole, Cabergoline and GnRH Antagonist to Prevent Ovarian Hyperstimulation Syndrome Completely in Polycystic Ovarian Syndrome Treatment

Background and Aims: The main difficulty to treat polycystic ovarian syndrome (PCOS) is the possibility of ovarian hyperstimulation syndrome (OHSS). There are several clinical approaches to avoid dangerous complications of OHSS: laparoscopic ovarian drilling (LOD), in vitro maturation (IVM), and GnRH antagonist-GnRH agonist based controlled ovarian stimulation (COS) with low dose of gonadotrophin. However, clinical outcomes after using these methods are less satisfactory than COS in normal ovarian function. We started a new approach combining the use of three kinds of medicine, Letrozole, Cabergoline and GnRH antagonist from just after the oocyte pick up and continue for five consecutive days to prevent the occurrence of OHSS in the PCOS treatment. Method: We compared the severity of OHSS after using a new treatment (the novel COS) in 53 women (55 cycles) with the severity of OHSS in a group of 37 women (37 cycles) who received the most widely used conventional GnRH antagonist-GnRH agonist- based COS for PCOS in retrospective cohort study. Results: This table shows the comparison of clinical outcomes between two groups. Conclusion: PCOS patients have been treated with hypostimulation methods and IVM to prevent OHSS. However, our method is considered to be more effective, safer and more versatile than any other treatment method. It seems to be an important alternative treatment method for PCOS patients. We believe this new treatment is highly recommended for the complete prevention of OHSS.

#252 : Granulosa Cell Metabolism is Differentially Impacted by Obesity and Aging and Correlated With Oocyte Competence

Background and Aims: Aging and obesity are associated with female infertility, with mitochondrial dysfunction proposed as an underlying factor; but our understanding of energy metabolism within the ovary remains limited. Method: This study focused on the granulosa cells surrounding the oocyte and investigated mitochondrial function, glycolytic function, total ATP production, and fatty acid metabolism using real-time metabolic analysis (Seahorse XFe96) to understand the effects of obesity and aging on energy metabolism. Mouse models were used to precisely isolate the effects of aging and obesity, with parallel studies conducted on human granulosa cells. Results: In mice, aging impaired both mitochondrial function and glycolytic function, while obesity primarily reduced mitochondrial function. Mice with both aging and obesity showed massively impaired granulosa cell metabolism. Next, the metabolic profile of granulosa cells in the follicular fluid of a cohort of 130 women who underwent IVF/ICSI cycles was measured. Aging primarily impaired mitochondrial function, with ATP production biased towards glycolysis, and no correlation was found with glycolytic function. Obesity primarily affected ATP production efficiency in mitochondria. Correlations between granulosa cell energy metabolism and embryonic development were also found. Mitochondrial spare capacity was associated with more retrieved oocytes, glycolytic reserve capacity was associated with Day 3 embryo quality, and mitochondrial ATP production was associated with improved blastocyst formation rates. Analysis of patient diagnoses revealed that in patients with normal ovarian function, only ATP production efficiency was decreased by aging, whereas in patients diagnosed with conditions of impaired ovarian function, mitochondrial function was more important. Interestingly, patients with adequate mitochondrial function showed good embryonic development, whereas those with very high mitochondrial and glycolytic energy metabolism showed a negative correlation with embryonic development, which was also observed in patients with ovarian dysfunction. Conclusion: These results provide new insights into the underlying metabolic dysfunction that occurs with female infertility.