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Intraovarian injection of platelet-rich plasma (PRP) has been recently proposed, with encouraging results to provide an alternative option to patients diagnosed with POR or POI. However, the broad spectrum of PRP effects on the reproductive function and the mechanisms of action in follicular activation, response to stimulation, and embryo quality have not yet been studied. In this study, we first induced poor ovarian reserve (POR) and premature ovarian insufficiency (POI) ovarian phenotypes in CD1 mice undergoing PRP or sham intraovarian injection. PRP administration reduced those alterations induced by chemotherapy in ovarian stroma and follicle morphology in both the POR and POI conditions. After ovarian stimulation, we found that PRP did not modify the MII-oocyte yield. Nevertheless, the amount of obtained 2-cell embryos and fertilization rate were increased, being especially relevant for the POI model. Further in vitro embryo culture led to improved blastocyst formation rates and higher numbers of good quality blastocysts in PRP vs. sham females in both the POR and POI conditions. These positive results of PRP injection were also validated in the C57Bl/6 stain. Altogether, our findings suggest a possible effect on oocyte and embryo quality. This effect is likely due to the increase of local paracrine signaling through the released growth factors in PRP-treated ovaries.

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Objective: Diminished ovarian reserve (DOR) and premature ovarian insufficiency (POI) represent conditions of different severity, characterized by an earlier-than-expected decrease in ovarian activity. The present study aims to compare metabolic disturbances between women with DOR and patients with POI from a different origin. Materials and methods: A total of 226 women (28 healthy women; 77 individuals with DOR, and 121 patients with POI/36 with Turner syndrome [TS] and 85 with non-TS POI/) have been studied retrospectively. Data concerning anthropometric indices, and metabolic parameters were collected. Results: Patients with DOR, non-TS POI, and TS had increased blood pressure and liver enzymes, pronounced insulin resistance, and worse lipid profiles than controls (p < 0.008 for all). TS patients had significantly higher ASAT, GGT, and TSH levels compared to non-TS POI and DOR individuals. The prevalence of type 2 diabetes tended to be higher in TS women compared to other groups. The prevalence of previously diagnosed polycystic ovarian syndrome was lower in the non-TS POI patients than in the DOR patients (p = 0.005). Conclusions: patients with decreased ovarian function suffer from insulin resistance, abnormal lipid profile, and subtle hepatic disturbances, irrespective of the severity of the condition and the presence of chromosomal aberrations.

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Premature ovarian insufficiency (POI) is the main cause of infertility in women. Some cases of POI are thought to be caused by genetic defects and the clinical outcomes of these patients are unknown. Here, we performed whole-exome sequencing of the peripheral blood of a cohort of 55 subjects with POI and identified one heterozygous missense variant in FOXL2 (c.1045G>C; p.Arg349Gly) and two heterozygous missense variants in ERCC6 (c.379G>A; p.Val127Ile and c.4223A>C; p.Glu1408 Ala) in four POI patients. All of these heterozygous mutations were predicted to be deleterious and were parentally inherited from their heterozygous fathers. The mRNA and protein expression of FOXL2 and ERCC6 were absent or decreased in the patients. The patients carrying the variants of FOXL2 (c.1045G>C; p.Arg349Gly) and ERCC6 (c.379G>A; p.Val127Ile) failed to conceive in two and four assisted reproductive cycles, respectively. Another patient and her sister carrying the ERCC6 (c.4223A>C; p.Glu1408 Ala) variant achieved good clinical outcomes after assisted reproductive therapy. Our findings support the possible roles of FOXL2 and ERCC6 in POI and might contribute to the genetic counseling of POI patients.

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Premature ovarian insufficiency (POI, defined as age at menopause < 40 years) affects 1%-3% of postmenopausal women. It is positively associated with an increased risk of diabetes mellitus, arterial hypertension, cardiovascular disease, osteoporosis, fractures, cognitive impairment, and depression. Early menopause (EM, defined as age at menopause < 45 years) is also associated with these adverse health consequences, in most cases to the same degree as in POI. Therefore, a unifying term for EM and POI, such as 'premature menopause', may be proposed, using the age threshold of < 45 years. This could provide broader coverage of these women, substantiating the need for prompt administration of menopausal hormone therapy (in this case, 'hormone replacement therapy'). However, the benefits of this approach, which precludes a higher oestrogen dose up to the normal age of menopause, need to be proven in well-designed randomized controlled trials.

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BACKGROUND: Premature ovarian insufficiency (POI) is a condition characterized by a substantial decline or loss of ovarian function in women before the age of 40. However, the pathogenesis of POI remains to be further elucidated, and specific targeted drugs which could delay or reverse ovarian reserve decline are urgently needed. Abnormal DNA damage repair (DDR) and cell senescence in granulosa cells are pathogenic mechanisms of POI. Ubiquitin-specific protease 14 (USP14) is a key enzyme that regulates the deubiquitylation of DDR-related proteins, but whether USP14 participates in the pathogenesis of POI remains unclear. METHODS: We measured USP14 mRNA expression in granulosa cells from biochemical POI (bPOI) patients. In KGN cells, we used IU1 and siRNA-USP14 to specifically inhibit USP14 and constructed a cell line stably overexpressing USP14 to examine its effects on DDR function and cellular senescence in granulosa cells. Next, we explored the therapeutic potential of IU1 in POI mouse models induced by D-galactose. RESULTS: USP14 expression in the granulosa cells of bPOI patients was significantly upregulated. In KGN cells, IU1 treatment and siUSP14 transfection decreased etoposide-induced DNA damage levels, promoted DDR function, and inhibited cell senescence. USP14 overexpression increased DNA damage, impaired DDR function, and promoted cell senescence. Moreover, IU1 treatment and siUSP14 transfection increased nonhomologous end joining (NHEJ), upregulated RNF168, Ku70, and DDB1, and increased ubiquitinated DDB1 levels in KGN cells. Conversely, USP14 overexpression had the opposite effects. Intraperitoneal IU1 injection alleviated etoposide-induced DNA damage in granulosa cells, ameliorated the D-galactose-induced POI phenotype, promoted DDR, and inhibited cell senescence in ovarian granulosa cells in vivo. CONCLUSIONS: Upregulated USP14 in ovarian granulosa cells may play a role in POI pathogenesis, and targeting USP14 may be a potential POI treatment strategy. Our study provides new insights into the pathogenesis of POI and a novel POI treatment strategy.

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BACKGROUND: Due to the lack of oestrogen, premature ovarian insufficiency (POI) is an independent risk factor for ischaemic heart disease and overall cardiovascular disease. This study aimed to apply layer-specific myocardial strain for early quantitative evaluation of subclinical left ventricular myocardial systolic function changes in patients with POI. METHODS: Forty-eight newly diagnosed, untreated patients with POI (POI group) and fifty healthy female subjects matched for age, height and weight (control group) were enrolled. Standard transthoracic echocardiography was used to measure conventional parameters and layer-specific strain parameters.The layer-specific strain parameters included subendomyocardial global longitudinal strain (GLSendo), mid-layer myocardial global longitudinal strain (GLSmid), subepimyocardial global longitudinal strain (GLSepi), subendomyocardial global circumferential strain (GCSendo), mid-layer myocardial global circumferential strain (GCSmid), and subepimyocardial global circumferential strain (GCSepi). RESULTS: There were no significant differences in age, body mass index (BMI), blood pressure, or left ventricular ejection fraction (LVEF) between the two groups. The end-diastolic interventricular septal thickness (IVST) was greater in the POI group (8.29 ± 1.32 vs. 7.66 ± 0.82, P = 0.008), and the POI group had lower E, E/A, and lateral e' (all P < 0.05). As for systolic functions,the POI group had lower GLSendo, GLSmid, GLSepi, GCSendo, GCSmid, and GCSepi (all P < 0.05).The intraobserver and interobserver coefficients of GLSendo, GLSmid, GLSepi, GCSendo, GCSmid, and GCSepi were greater than 0.900. CONCLUSIONS: POI patients with normal LVEF may suffer from subclinical left ventricular myocardial systolic dysfunction. Echocardiography of layer-specific myocardial strain could more sensitively detect subclinical impairment of left ventricular systolic function in POI patients.

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It is well known that oocytes are produced during fetal development and that the total number of primary follicles is determined at birth. In humans, there is a constant loss of follicles after birth until about two years of age. The number of follicles is preserved until the resumption of meiosis at puberty and there is no renewal of the oocytes; this dogma was maintained in the last century because there were no suitable techniques to detect and obtain stem cells. However, following stem cell markers, several scientists have detected them in developing and adult human ovarian tissues, especially in the ovarian surface epithelial cells. Furthermore, many authors using different methodological strategies have indicated this possibility. This evidence has led many scientists to explore this hypothesis; there is no definitive consensus to accept this idea. Interestingly, oocyte retrieval from mature ovaries and other tissue sources of stem cells has contributed to the development of strategies for the retrieval of mature oocytes, useful for assisted reproductive technology. Here, we review the evidence and controversies on oocyte neooogenesis in adult women; in addition, we agree with the idea that this process may occur in adulthood and that its alteration may be related to various pathologies in women, such as polycystic ovary syndrome, premature ovarian insufficiency, diminished ovarian reserve and several infertility and genetic disorders.

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Over the past decade, research on the human microbiota has become a hot topic. Among them, the female reproductive tract (FRT) also has a specific microbiota that maintains the body's health and dynamic balance, especially in the reproductive aspect. When the FRT ecosystem is dysregulated, changes in immune and metabolic signals can lead to pathological and physiological changes such as chronic inflammation, epithelial barrier disruption, changes in cell proliferation and apoptosis, and dysregulation of angiogenesis and metabolism, thereby causing disruption of the female endocrine system. Premature ovarian insufficiency (POI), a clinical syndrome of ovarian dysfunction, is primarily influenced by immune, genetic, and environmental factors. New evidence suggests that dysbiosis of the FRT microbiota and/or the presence of specific bacteria may contribute to the occurrence and progression of POI. This influence occurs through both direct and indirect mechanisms, including the regulation of estrogen metabolism. The use of probiotics or microbiota transplantation to regulate the microbiome has also been proven to be beneficial in improving ovarian function and the quality of life in women with premature aging. This article provides an overview of the interrelationships and roles between the FRT microbiome and POI in recent years, to fully understand the risk factors affecting female reproductive health, and to offer insights for the future diagnosis, treatment, and application of the FRT microbiome in POI patients.

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Premature ovarian insufficiency (POI) is defined as onset of menopause characterized by amenorrhea, hypergonadotropism, and hypoestrogenism, before the age of 40 years. The POI is increasing, which seriously affects the quality of patients' life. Due to its diversity of pathogenic factors, complex pathogenesis and limited treatment methods, the search for finding effective treatment of POI has become a hotspot. Stem cells are characterized by the ability of self-renewal and differentiation and play an important role in the regeneration of injured tissues, which is therapy is expected to be used in the treatment of POI. The aim of this review is to summarize the pathogenic mechanisms and the research progress of POI treatment with stem cells from different sources.

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OBJECTIVES: To investigate the effect of Chinese medicine He's Yangchao recipe on premature ovarian insufficiency (POI) and its relationship with mitochondrial function of ovarian granulose cells in an animal model. METHODS: Thirty-six female C57BL/6J mice were randomly divided into blank control group, model group, low-, medium- and high-dose He's Yangchao recipe treatment group and coenzyme Q10 (Q10) treatment group (positive control). The POI model was induced by a single intraperitoneal injection of cyclophosphamide (90 mg/kg). The animals were sacrificed after 21 days. Primary granulose cells were obtained from POI mice and treated with He's Yangchao recipe, ERß inhibitor PHTPP, and He's Yangchao recipe+PHTPP in vitro for 24 h, respectively. Ovarian histopathological changes were observed by hematoxylin-eosin (HE) staining, ATP levels were detected by luciferase assay, mtDNA copy numbers were detected by quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR), mitochondrial structure changes were observed by transmission electron microscopy, protein and mRNA expression levels of estrogen receptor ß (ERß), peroxisome proliferator-activated receptor γ coactivator 1α (PGC1α), mitochondrial transcription factor A (TFAM), and superoxide dismutase 2 (SOD2) were detected by Western blotting and qRT-PCR. RESULTS: The ovarian tissue in model group exhibited few secondary and tertiary follicles, whereas the He's Yangchao recipe groups and Q10 group had abundant secondary and tertiary follicles. Compared with the blank control group, ATP and mtDNA levels in model group decreased (P<0.01), mitochondrial crista disappeared or abnormal vacuolated structure increased; the protein and mRNA levels of ERß, PGC1α, TFAM, and SOD2 decreased (all P<0.01). ATP production increased in granulose cells of high-dose He's Yangchao recipe group and Q10 group; mtDNA copy numbers increased (P<0.05 or P<0.01); abnormal mitochondrial structure was reduced; the protein and mRNA expressions of ERß, PGC1α, TFAM, and SOD2 increased (P<0.05 or P<0.01). Compared with the PHTPP intervention group, the proportion of normal mitochondrial structure in the granulose cells of He's Yangchao recipe + PHTPP group was higher; ATP content increased (P<0.05 or P<0.01); mtDNA copy numbers increased (P<0.05 or P<0.01); the protein and mRNA expression of ERß, PGC1α, TFAM and SOD2 increased (P<0.05 or P<0.01). CONCLUSIONS: He's Yangchao recipe can regulate mitochondrial biogenesis through ERß/PGC1α/TFAM pathway to improve ovarian function in POI mice.

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INTRODUCTION: Premature ovarian insufficiency [POI] is a disease characterized by a premature decline in ovarian function before the age of 40. In China, Ligustrum lucidum [FLL] has long been used to improve ovarian function and treat POI. METHODS: This study aims to verify the effect of FLL on POI through network pharmacology, molecular docking, and in-vitro cell experiments. RESULTS: A total of 13 active substances were screened in FLL, including including quercetin, taxifolin, luteolin, kaempferol, and beta-sitosterol. Then, network analysis found that FLL may exert effects on POI through 10 targets, including AR, ESR1, ESR2, KDR, CYP19A1, CLPP, GC, MMP3, PPARG, and STS. According to GO and KEGG enrichment analysis, FLL is associated with mechanisms related to estrogen, including steroid hormone biosynthesis, ovarian steroidogenesis, and the estrogen signaling pathway. Molecular docking confirms the interaction between the active ingredients of FLL and CYP19A1, which encodes aromatase. CCK8 experiment confirmed that quercetin and taxifolin can enhance the proliferation of KGN granulosa cells, while quercetin, taxifolin, and kaempferol can inhibit the apoptosis of KGN granulosa cells. ELISA experiments have confirmed that quercetin, taxifolin, luteolin, and kaempferol can increase the synthesis of estradiol in KGN granulosa cells. WB confirms that quercetin can increase the expression level of CYP19A1 in KGN cells. CONCLUSION: FLL can improve the proliferation, apoptosis, and synthesis of estradiol in ovarian granulosa cells, and has the potential to treat POI.

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Organophosphate esters (OPEs), increasingly used as new flame retardants and plasticizers in various products, have been found to have reproductive toxicity with overt endocrine disruption potential, yet the relationship between OPEs and early menopause remains unexplored. In the present study, we included 2429 women who participated in the U.S. National Health and Nutrition Examination Survey data (2011-2020) and had data of five urinary OPE metabolite levels and information of menopause characteristics, to investigate the associations of OPEs exposure with premature ovarian insufficiency (POI) and age of menopause. Multivariable adjusted linear and logistic regression were used to assess the associations of urinary OPE metabolites with age of menopause and POI, respectively. Quantile g computation (QGC) models were used to assess the relative contribution of individual metabolites to associations of OPE metabolites mixture. After adjusting for covariates, urinary bis(2-chloroethyl) phosphate (BCEP) concentration was inversely associated with menopause age (ß = - 0.21; 95% confidence interval (CI): 0.41, - 0.002). Higher urinary BCEP level (>median) was associated with earlier age at menopause (ß = -1.14, 95% CI: 1.83, - 0.46), and elevated odds of having POI (OR = 1.93; 95% CI: 1.02, 3.66). These associations were robust to the further adjustment of cardiometabolic diseases and related traits (e.g., body mass index). Further QGC analyses confirmed that BCEP was the dominant metabolite contributing most to the associations of OPEs mixture with age of menopause (weight = 49.5%) and POI (weight = 75.1%). No significant associations were found for the other four OPE metabolites. In this cross-sectional study, urinary BCEP level was associated with earlier menopause and increased odds of POI, highlighting the potential negative impacts of this chemical and its parent compound tris(2-chloroethyl) phosphate on ovarian function. Further studies are required to validate our findings and reveal potential underlying mechanisms.

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BACKGROUND: It is challenging to improve the effects of chemotherapy and reduce its adverse impact on the ovaries. Our previous study suggested that the combination of galaxamide could enhance the antitumor effect of cisplatin (CIS) in HeLa cell xenograft mice. However, their potential effects on ovarian tissues remain unknown. METHODS: The Hela tumor-bearing female BALB/c mice model was established and randomly divided into three groups: control group (PBS group), CIS group (0.3 mg/kg CIS group) and galaxamide group (0.3 mg/kg CIS + 3 mg/kg galaxamide-treated group). The serum sex hormones levels, ovarian morphology, functional and molecular characterisation were determined and compared with those of the control group. RESULTS: The hormonal effects indicated premature ovarian insufficiency (POI) associated with CIS-induced tumor-bearing mice. CIS induces the apoptosis in primordial and developing follicles and subsequently increases follicular atresia, eventually leading to follicle loss. After cotreatment, galaxamide significantly increased anti-Mullerian hormone (AMH) and follicle-stimulating hormone receptor (FSHR) expression and prevented the CIS-induced PI3K pathway, which triggers follicle activation, apoptosis or atresia. CONCLUSION: These findings demonstrate that galaxamide could attenuate CIS-induced follicle loss by acting on the PI3K signaling pathway by stimulating AMH and/or FSHR and thus provides promising therapeutic options for patients with cervical cancer.

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Chemotherapy-induced premature ovarian insufficiency (CIPOI) triggers gonadotoxicity in women undergoing cancer treatment, leading to loss of ovarian reserves and subfertility, with no effective therapies available. In our study, fecal microbiota transplantation in a cisplatin-induced POI mouse model reveals that a dysbiotic gut microbiome negatively impacts ovarian health in CIPOI. Multi-omics analyses show a significant decrease in Limosilactobacillus reuteri and its catabolite, ß-resorcylic acid , in the CIPOI group in comparison to healthy controls. Supplementation with L. reuteri or ß-RA mitigates cisplatin-induced hormonal disruptions, morphological damages, and reductions in follicular reserve. Most importantly, ß-RA pre-treatment effectively preserves oocyte function, embryonic development, and fetus health, thereby protecting against chemotherapy-induced subfertility. Our results provide evidence that ß-RA suppresses the nuclear accumulation of sex-determining region Y-box 7, which in turn reduces Bcl-2-associated X activation and inhibits granulosa cell apoptosis. These findings highlight the therapeutic potential of targeting the gut-ovary axis for fertility preservation in CIPOI.

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Premature ovarian insufficiency (POI) means menopause before 40 years of age affecting about 1 % of women. Approaches based on cell therapy and the paracrine effects of stem cells or bioproducts such as platelet-rich plasma have been proposed, but concerns remain about undesired systemic effects, as well as the need to optimize delivery methods through bioengineering methods. This study explores the efficacy of decellularized bovine ovarian cortex extracellular matrix (OvaECM) hydrogels alone and as a growth factor (GF) carrier (OvaECM+GF) in a chemotherapy-induced POI murine model. In vitro assays showed a gradual release of GF from the OvaECM sustained for two weeks. Chemotherapy drastically reduced follicle numbers, but OvaECM+GF treatment restored pre-antral follicle development. Moreover, this treatment notably regenerated the ovarian microenvironment by increasing cell proliferation and microvessel density while reducing chemotherapy-induced apoptosis and fibrosis. Whole-ovary RNA sequencing and gene set enrichment analysis revealed an upregulation of regeneration-related genes and a downregulation of apoptotic pathways. The OvaECM+GF treatment also yielded significantly better outcomes following ovarian stimulation and in vitro fertilization. After two consecutive crossbreeding cycles, OvaECM+GF-treated mice showed normal reproductive function. This research showcases the biocompatibility and efficacy of OvaECM to reverse POI in mice, setting a foundation to explore innovative bioengineering-based POI therapies. STATEMENT OF SIGNIFICANCE: Premature ovarian insufficiency (POI) affects about 1 % of women worldwide, causing early menopause before 40 years old. Current treatments alleviate symptoms but do not restore ovarian function. This study explores an innovative approach using ovarian cortex extracellular matrix hydrogels to deliver growth factors into the murine ovarian niche and reverse POI. In vitro release kinetic assays demonstrated a gradual and sustained release of growth factors. In a POI-induced mouse model, intraovarian injections of the hydrogel encapsulating growth factors restored pre-antral follicle development, increased cell proliferation, reduced apoptosis and fibrosis, and improved ovarian response and in vitro fertilization outcomes. Long-term benefits included larger litter sizes. This innovative technique shows promise in regenerating the ovarian environment and improving reproductive outcomes.

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Gonadotoxicity resulting from systemic and locoregional cancer treatments significantly threatens women's reproductive health, often culminating in premature ovarian insufficiency. These therapies, particularly alkylating agents and ionizing radiation, induce DNA damage and apoptosis in ovarian follicles, leading to infertility, amenorrhea, and estrogen deficiency, which exacerbate risks of osteoporosis and cardiovascular diseases. Existing fertility preservation methods do not prevent immediate ovarian damage, underscoring the need for innovative protective strategies. Menstrual blood-derived stem cells (MenSC) and their extracellular vesicles (EV) present promising regenerative potential due to their therapeutic cargo delivery and pathway modulation capabilities. Preclinical studies demonstrate that MenSC-derived EV ameliorate premature ovarian insufficiency by inhibiting granulosa cell apoptosis, promoting angiogenesis, and activating pivotal pathways such as SMAD3/AKT/MDM2/P53. However, comprehensive research is imperative to ensure the safety, efficacy, and long-term effects of MenSC-derived EV in clinical practice. In this review, we update the current knowledge and research regarding the use of MenSC-derived EV as a novel therapeutic weapon for ovarian regeneration in the context of gonadotoxicity induced by systemic anticancer treatment.

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CONTEXT: Turner syndrome (TS) is characterized by short stature and premature ovarian insufficiency (POI). The main long-term complication of POI is osteoporosis, which can be prevented by hormone replacement therapy (HRT). OBJECTIVE: The objective of our study was to compare initial bone mineral density (BMD) and progression between TS and idiopathic POI patients under HRT. METHODS: A single-center retrospective study was conducted between 1998 and 2018. All women had undergone at least two bone densitometry assessments at least 2 years apart. RESULTS: Sixty-eight TS patients and 67 idiopathic POI patients were included. Mean age at initial assessment was 27 years (IQR, 21-35.5 years) in TS patients and 31.5 years (IQR, 23-37 years) in idiopathic POI patients (P=0.1). Lumbar and femoral neck BMD were lower in the TS group than in the idiopathic POI group (respectively 0.89g/cm2 versus 0.95g/cm2, P=0.03; 0.70g/cm2 versus 0.77g/cm2, P<0.0001). Mosaic karyotype was associated with better BMD in TS patients while history of growth hormone treatment had no impact on BMD. Over time, a significant gain in vertebral BMD was observed in TS patients versus a loss of BMD in idiopathic POI patients (P=0.0009). CONCLUSION: TS patients had a lower BMD at baseline than idiopathic POI patients, at both spinal and femoral levels. Over time, on HRT, a significant gain in vertebral BMD was observed in patients with TS, compared with a loss of BMD in patients with idiopathic POI. We hypothesized that earlier initiation and longer duration of HRT played an important role in this finding. Long-term prospective follow-up to assess the incidence of fractures in TS would be useful.

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Premature ovarian insufficiency (POI) is defined as the depletion of ovarian function before the age of 40 years. The global prevalence of POI is 3.5%. To date, genetic factors account for 23.5% of the etiology of POI. Herein, a previously uncharacterized pathogenic homozygous variant of the chromosome segregation-1-like gene (CSE1L) was identified in POI patients via targeted panel sequencing. It is reported that dysregulated iron metabolism is involved in many reproductive endocrine disorders; however, its precise role in POI remains obscure. In this study, we identified CSE1L as a potential candidate gene that plays an important role in maintaining iron homeostasis. Deficiency of CSE1L led to ferroptosis in human granulosa cells, which was confirmed by transmission electron microscopy. Mechanistically, coimmunoprecipitation identified the direct interaction between CSE1L and FoxO1. Inhibition of CSE1L led to the excessive accumulation of FoxO1 in the nucleus via nucleocytoplasmic transport. Then, FoxO1 bound to the promoter region of NCOA4 and promoted its transcription, which was verified by a chromatin immunoprecipitation assay. Moreover, inhibition of CSE1L in cumulus cell monolayer could impede oocyte maturation, which might be associated with oxidative stress. Consequently, our study first revealed that CSE1L participated in ferroptosis in human ovarian granulosa cells via nucleocytoplasmic transportation, which might be helpful in revealing the molecular mechanism of CSE1L in the development of POI. Importantly, these findings might provide new insights into the application of ferroptosis inhibitors in the treatment of POI.

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RESEARCH QUESTION: Can atorvastatin, with its antioxidant, anti-inflammatory and anti-apoptotic properties, improve ovarian function and follicular reserve in rats with cyclophosphamide-induced premature ovarian insufficiency (POI)? DESIGN: In this experimental study, 24 adult female Wistar rats were divided into four groups: control; POI; POI + atorvastatin; and atorvastatin. After treatment with atorvastatin, serum concentrations of total antioxidant capacity, glutathione, malondialdehyde, FSH, oestradiol, anti-Müllerian hormone, tumour necrosis factor-alpha and interleukin-6 were evaluated. Additionally, mRNA and protein expression of Bax, Bcl-2 and VEGF-A; number of follicles; and total volume of the ovary, and volumes of the cortex and medulla were examined. RESULTS: The results showed that serum concentrations of total antioxidant capacity (P < 0.001), glutathione, oestradiol and anti-Müllerian hormone (P < 0.05); mRNA and protein expression of Bcl-2 and VEGF-A (P < 0.05); number of primordial and primary follicles (P < 0.001), and preantral and antral follicles (P < 0.01); and total volume of the ovary, and volume of the cortex (P < 0.05) increased significantly in the POI + atorvastatin group compared with the POI group. Serum concentrations of malondialdehyde, FSH, tumour necrosis factor-alpha and interleukin-6; and mRNA and protein expression of Bax decreased significantly in the POI + atorvastatin group compared with the POI group (P < 0.05). CONCLUSIONS: Atorvastatin reduces the detrimental effects of cyclophosphamide in the POI model significantly by reducing oxidative stress and pro-inflammatory cytokines; regulating the expression of Bax, Bcl-2 and VEGF-A; and improving ovarian function and follicular reserve.

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BACKGROUND: POI (premature ovarian insufficiency) refers to premature and rapid decline of ovarian reserve function in women before the age of 40, which can be manifested as menstrual disorders, endocrine abnormalities and low fertility. Bu-Shen-Ning-Xin decoction (BSNXD) has been found to have therapeutic effects on POI. Nevertheless, how it exerts therapeutic effects remains elusive. PURPOSE: This research aims to clarify the pharmacological mechanisms of BSNXD. METHODS: We applied Ultra Performance Liquid Chromatography (UPLC) to identify the main components of BSNXD.4-vinylcyclohexene diepoxide(VCD)was used to induce POI models. ELISA detected the serum level of hormones. H&E staining evaluated the morphology of ovarian tissues.CircRNA and mRNA expression profiles in the ovaries of both POI rats and those treated with BSNXD were detected. Then, dysregulated circRNAs and mRNAs that were potentially altered by BSNXD were screened. Network pharmacology analysis was performed to identify drug targets of BSNXD active ingredients. A circRNA-miRNA-mRNA network and an oxidative stress(OS)-related subnetwork were constructed. Expression of rno_circRNA_012284, rno_miR-760-3p, and HBEGF(Heparin-binding epidermal growth factor-like growth factor) was measured by RT-PCR and their binding were verified by dual-luciferase reporter assays. ROS was measured through DCFH-DA fluorescence probes. The HBEGF target was selected for molecular docking with key active ingredients.Surface plasmon resonance(SPR) was applied to verify the binding ability and affinity between components and HBEGF. RESULTS: UPLC analysis indicated that 6 chemical compounds including berberine, paeoniflorin, morroniside,gallic acid, loganin, baicalin were identified.Elevated FSH and LH levels, suppressed E2 and AMH levels in the serum, and inhibited follicles and corpus luteums in the ovarian tissues of VCD-induced rats were notably reversed by BSNXD.In total, 992 up- and 1135 down-regulated circRNAs, and 205 up- and 243 down-regulated mRNAs were found in POI rat ovaries following BSNXD administration. Furthermore, 198 drug targets of BSNXD were identified. An OS-related and BSNXD-targeted ceRNA subnetwork composed of rno_circRNA_012284/rno_miR-760-3p/HBEGF was established. rno_circRNA_012284 and HBEGF were up-regulated and rno_miR-760-3p was down-regulated in POI ovarian granulosa cells (OGCs) after BSNXD administration. rno_circRNA_012284 was a sponge of rno_miR-760-3p to elevate HBEGF expression. Moreover, rno_circRNA_012284 overexpression alleviated POI-induced excessive ROS generation in ovarian granulosa cells, while rno_circRNA_012284 inhibition exerted the opposite effect. Finally,molecular docking speculated active ingredients of each herb acted on HBEGF to reduce the OS. SPR tests showed that Berberine,Baicalein,Quercetin,Pachymic acid,Paeoniflorin exhibited satisfying affinity with HBEGF protein. CONCLUSION: This study demonstrates that BSNXD ameliorates POI partly by attenuating OS in ovarian granulosa cells via rno_circRNA_012284/rno_miR-760-3p/HBEGF axis, uncovering the pharmacological mechanisms of BSNXD in alleviating POI.

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