RESUMO
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.
Assuntos
Senescência Celular , Dano ao DNA , Reparo do DNA , Células da Granulosa , Insuficiência Ovariana Primária , Ubiquitina Tiolesterase , Feminino , Insuficiência Ovariana Primária/patologia , Insuficiência Ovariana Primária/metabolismo , Insuficiência Ovariana Primária/genética , Células da Granulosa/metabolismo , Células da Granulosa/efeitos dos fármacos , Células da Granulosa/patologia , Senescência Celular/efeitos dos fármacos , Animais , Humanos , Ubiquitina Tiolesterase/metabolismo , Ubiquitina Tiolesterase/genética , Reparo do DNA/efeitos dos fármacos , Camundongos , Adulto , Camundongos Endogâmicos C57BL , Linhagem CelularRESUMO
Premature ovarian failure (POF) is intricately linked to cellular fates such as senescence, apoptosis, and impaired granulosa cell (GC) differentiation, each of which contributes to ovarian dysfunction and follicular depletion. Autophagy is essential in preventing POF by maintaining cellular homeostasis through the degradation and recycling of damaged organelles and proteins, thereby preserving ovarian function and preventing follicular depletion. Recent studies have revealed that the targeted regulation and disruption of autophagy through various molecular mechanisms ultimately lead to the pathogenesis of POF. In this review, we provide a comprehensive analysis of the disruption in regulatory mechanisms of autophagy contributing to POF. Specifically, we elucidate the molecular mechanisms that can be targeted to restore autophagy homeostasis, offering therapeutic potential for the treatment of POF.
Assuntos
Autofagia , Insuficiência Ovariana Primária , Humanos , Insuficiência Ovariana Primária/metabolismo , Insuficiência Ovariana Primária/patologia , Feminino , Animais , Células da Granulosa/metabolismo , Células da Granulosa/patologiaRESUMO
Ovarian insufficiency is one of the common reproductive disorders affecting women with limited therapeutic aids. Mesenchymal stem cells have been investigated in such disorders before yet, the exact mechanism of MSCs in ovarian regeneration regarding their epigenetic regulation remains elusive. The current study is to investigate the role of the bone marrow-derived mesenchymal stem cells (BM-MSCs) lncRNA (Neat-1 and Hotair1) and miRNA (mir-21-5p, mir-144-5p, and mir-664-5p) in mitigating ovarian granulosa cell apoptosis as well as searching BM-MSCs in altering the expression of ovarian and hypothalamic IGF-1 - kisspeptin system in connection to HPG axis in a cyclophosphamide-induced ovarian failure rat model. Sixty mature female Sprague Dawley rats were divided into 3 equal groups; control group, premature ovarian insufficiency (POI) group, and POI + BM-MSCs. POI female rat model was established with cyclophosphamide. The result revealed that BM-MSCs and their conditioned media displayed a significant expression level of Neat-1, Hotair-1, mir-21-5p, mir-144-5p, and mir-664-5p. Moreover, BM-MSCs transplantation in POI rats improves; the ovarian and hypothalamic IGF-1 - kisspeptin, HPG axis, ovarian granulosa cell apoptosis, steroidogenesis, angiogenesis, energy balance, and oxidative stress. BM-MSCs expressed higher levels of antiapoptotic lncRNAs and microRNAs that mitigate ovarian insufficiency.
Assuntos
Apoptose , Ciclofosfamida , Fator de Crescimento Insulin-Like I , Células-Tronco Mesenquimais , MicroRNAs , Insuficiência Ovariana Primária , RNA Longo não Codificante , Ratos Sprague-Dawley , Animais , Feminino , MicroRNAs/genética , MicroRNAs/metabolismo , Células-Tronco Mesenquimais/metabolismo , Ciclofosfamida/efeitos adversos , Ratos , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Insuficiência Ovariana Primária/metabolismo , Insuficiência Ovariana Primária/genética , Insuficiência Ovariana Primária/induzido quimicamente , Fator de Crescimento Insulin-Like I/metabolismo , Fator de Crescimento Insulin-Like I/genética , Ovário/metabolismo , Células da Medula Óssea/metabolismo , AngiogêneseRESUMO
BACKGROUND: Oxidative damage to the ovaries is the primary cause of impaired reproductive functions in female animals. This study aimed to investigate the protective role of N-Acetyl-L-cysteine (NAC) in reducing oxidative damage in the ovaries of female rabbits. METHODS AND RESULTS: Female rabbit ovaries were treated in vitro with varying concentrations of D-galactose (D-gal): 0, 5, 10, and 15 mg/mL, and it was found that 10 mg/mL D-gal significantly disrupted follicular structures, causing disarray in granulosa cell arrangements and significantly reducing T-SOD and GSH levels (p < 0.01). Consequently, we selected 10 mg/mL D-gal to establish an ovarian failure model. These models were treated with multiple doses of NAC (0, 0.1, 0.3, 0.5 mg/mL). The results revealed that the disruption in granulosa cell arrangement caused by 10 mg/mL D-gal was effectively alleviated by 0.1 mg/mL NAC compared to the D-gal treatment group. Furthermore, 10 mg/mL D-gal significantly (p < 0.01) reduced GSH, T-SOD, and catalase (CAT) levels in the ovaries. However, 0.1 mg/mL NAC effectively (p < 0.01) suppressed these adverse effects. Moreover, the current results showed that 10 mg/mL D-gal alone significantly (p < 0.01) downregulated the expression of Nrf2, GPX, PRDX4, GSR, SOD1, and TAF4B, whereas 0.1 mg/mL NAC counteracted these suppressive effects (p < 0.01). CONCLUSIONS: It could be concluded that NAC may delay ovarian failure by reducing D-gal-induced ovarian oxidative damage in female rabbit, suggested NAC could be a promising therapeutic agent for protecting against ovarian failure and potentially delaying ovarian failure in female rabbits.
Assuntos
Acetilcisteína , Galactose , Ovário , Estresse Oxidativo , Animais , Coelhos , Feminino , Acetilcisteína/farmacologia , Galactose/efeitos adversos , Galactose/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Ovário/efeitos dos fármacos , Ovário/metabolismo , Ovário/patologia , Insuficiência Ovariana Primária/induzido quimicamente , Insuficiência Ovariana Primária/metabolismo , Insuficiência Ovariana Primária/patologia , Células da Granulosa/metabolismo , Células da Granulosa/efeitos dos fármacos , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Superóxido Dismutase/metabolismo , Glutationa/metabolismo , Catalase/metabolismo , Modelos Animais de DoençasRESUMO
<b>Background and Objective:</b> Platelet-Rich Plasma (PRP) is proposed to have a potential regenerative effect on the ovaries following premature ovarian insufficiency (POI), however, the main impact on the process of follicle development (folliculogenesis) remains unclear. Protein-protein interaction (PPI) or network interaction analysis enables us to predict the targets of PRP's biomolecules using biological data through public databases and platforms. Therefore, this method is more efficient in time and cost than traditional laboratory procedures. The purpose of this study was to predict the targets of PRP's biomolecules on folliculogenesis following POI using PPI analysis and to clarify if those predictive targets integrate into PI3K/Akt signaling pathway which is an important pathway in folliculogenesis. <b>Materials and Methods:</b> Mining data targets of POI, PRP and folliculogenesis was done by GeneCards. Only genes with "protein-coding" category were analyzed further. Network analysis was performed using Cytoscape and STRING. Finally, STRING, Enricher and ShinyGO platforms were conducted to analyze gene ontology, including biological processes, molecular function and cellular components, as well as pathways. <b>Results:</b> Network analysis with Cytoscape and STRING discovered 107 gene hubs for POI, PRP and folliculogenesis. Analysis of KEGG pathway using STRING, Enricher and ShinyGO identified 43 genes integrated into PI3K/Akt signaling pathway. From the KEGG pathway, PI3K and Akt were revealed as two main targets following PRP treatment for POI patients. <b>Conclusion:</b> Biomolecules in PRP may recover ovarian follicle development following POI through the PI3K/Akt signaling pathway.
Assuntos
Folículo Ovariano , Plasma Rico em Plaquetas , Insuficiência Ovariana Primária , Transdução de Sinais , Feminino , Insuficiência Ovariana Primária/terapia , Insuficiência Ovariana Primária/metabolismo , Humanos , Plasma Rico em Plaquetas/metabolismo , Folículo Ovariano/metabolismo , Folículo Ovariano/crescimento & desenvolvimento , Mapas de Interação de Proteínas , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fosfatidilinositol 3-Quinases/metabolismoRESUMO
Cis-dichlorodiammineplatinum(II) (CDDP), while widely utilized in tumor therapy, results in toxic side effects that patients find intolerable. The specific mechanism by which CDDP inflicts ovarian damage remains unclear. This study aimed to explore the involvement of ferrostatin-1 (FER-1) and ferroptosis in CDDP-induced ovarian toxicity. This study established models of CDDP-induced injury in granulosa cells (GCs) and rat model of premature ovarian failure (POF). CCK-8 assessed the effects of CDDP and FER-1 on GC viability. FerroOrange and Mito-FerroGreen, DCFH-DA and MitoSox-Red, Rhodamine 123 and Transmission electron microscopy (TEM) measured Fe2+, reactive oxygen species (ROS), mitochondrial membrane potential and the mitochondrial morphology in GC cells, respectively. Serum hormone levels; organ indices; malondialdehyde, superoxide dismutase, and glutathione analyses; and western blotting were performed to examine ferroptosis's role in vitro. Molecular docking simulation was evaluated the interaction between FER-1 and GPX4 or FER-1 and NRF2. Molecular docking simulations were conducted to evaluate the interactions between FER-1 and GPX4, as well as FER-1 and NRF2. The findings revealed that CDDP-induced ovarian toxicity involved iron accumulation, increased ROS accumulation, and mitochondrial dysfunction, leading to endocrine disruption and tissue damage in rats. These changes correlated with NRF2, HO-1, and GPX4 levels. However, FER-1 decreased the extent of ferroptosis. Thus, ferroptosis appears to be a crucial mechanism of CDDP-induced ovarian injury, with GPX4 as potential protective targets.
Assuntos
Cisplatino , Cicloexilaminas , Ferroptose , Simulação de Acoplamento Molecular , Fenilenodiaminas , Espécies Reativas de Oxigênio , Animais , Feminino , Ferroptose/efeitos dos fármacos , Cicloexilaminas/farmacologia , Ratos , Espécies Reativas de Oxigênio/metabolismo , Cisplatino/efeitos adversos , Fenilenodiaminas/farmacologia , Células da Granulosa/efeitos dos fármacos , Células da Granulosa/metabolismo , Ovário/efeitos dos fármacos , Ovário/metabolismo , Ovário/patologia , Insuficiência Ovariana Primária/induzido quimicamente , Insuficiência Ovariana Primária/metabolismo , Fosfolipídeo Hidroperóxido Glutationa Peroxidase/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Ratos Sprague-Dawley , Modelos Animais de Doenças , Humanos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismoRESUMO
Radiotherapy is one of the risk factors for radiation-induced premature ovarian failure and infertility in cancer patients. The development of methods for ovarian radioprotection remains relevant. Moreover, electrons are a little-studied and promising method of radiation with the least toxic effect on normal tissues. The assessment of intracellular mechanisms regulating the protective effects of leukocyte-poor platelet-rich plasma in a model of radiation-induced premature ovarian failure caused by electron irradiation. Wistar rats were divided into four groups, namely a control group, irradiation group (electron exposure), irradiation + leukocyte-poor platelet-rich plasma group, and only leukocyte-poor platelet-rich plasma group. Fragments of ovaries were removed and hormonal, oxidant, histological, and morphometric studies were carried out. The cell cycle of ovarian follicles and the inflammatory and vascular response were assessed using immunohistochemistry. The activity of MAPK, ERK, and PI3K pathways was also assessed using the RT-qPCR. We found that electron irradiation causes a decrease in the functional activity of the ovaries and the death of follicular cells through apoptosis. The administration of LP-PRP led to a partial restoration of the cytokine balance. In addition, minor ovarian damage and mild inflammation were observed in this group. Leukocyte-poor platelet-rich plasma components have anti-inflammatory, angiogenetic, and radioprotective effects, reducing the activation of the NOX4, caspase and cytokine cascades, and inflammatory response severity through the MAPK/p38/JNK signaling pathway. This leads to the induction of endogenous antioxidant protection, the repair of post-radiation follicular damage, and slowing down the development of radiation-induced premature ovarian failure after electron irradiation.
Assuntos
Elétrons , Plasma Rico em Plaquetas , Insuficiência Ovariana Primária , Ratos Wistar , Feminino , Insuficiência Ovariana Primária/etiologia , Insuficiência Ovariana Primária/metabolismo , Animais , Plasma Rico em Plaquetas/metabolismo , Ratos , Ovário/efeitos da radiação , Ovário/metabolismo , Ovário/patologia , Apoptose/efeitos da radiação , Protetores contra Radiação/farmacologia , Folículo Ovariano/metabolismo , Folículo Ovariano/efeitos da radiação , Citocinas/metabolismoRESUMO
BACKGROUND: Emerging evidence has highlighted the therapeutic potential of human umbilical cord mesenchymal stem cells (UC-MSCs) in chemotherapy-induced premature ovarian failure (POF). This study was designed to investigate the appropriate timing and molecular mechanism of UC-MSCs treatment for chemotherapy-induced POF. METHODS: Ovarian structure and function of mice were assessed every 3 days after injections with cyclophosphamide (CTX) and busulfan (BUS). UC-MSCs and UC-MSCs-derived extracellular vesicles (EVs) were infused into mice via the tail vein, respectively. Ovarian function was analyzed by follicle counts, the serum levels of hormones and ovarian morphology. The apoptosis and proliferation of ovarian granulosa cells were analyzed in vitro and in vivo. Label-free quantitative proteomics was used to detect the differentially expressed proteins in UC-MSC-derived EVs. RESULTS: After CTX/BUS injection, we observed that the ovarian function of POF mice was significantly deteriorated on day 9 after CTX/BUS infusion. TUNEL assay indicated that the number of apoptotic cells in the ovaries of POF mice was significantly higher than that in normal mice on day 3 after CTX/BUS injection. Transplantation of UC-MSCs on day 6 after CTX/BUS injection significantly improved ovarian function, enhanced proliferation and inhibited apoptosis of ovarian granulosa cells, whereas the therapeutic effect of UC-MSCs transplantation decreased on day 9, or day 12 after CTX/BUS injection. Moreover, EVs derived from UC-MSCs exerted similar therapeutic effects on POF. UC-MSCs-derived EVs could activate the PI3K/AKT signaling pathway and reduce ovarian granulosa cell apoptosis. Quantitative proteomics analysis revealed that clusterin (CLU) was highly expressed in the EVs of UC-MSCs. The supplementation of CLU proteins prevented ovarian granulosa cells from chemotherapy-induced apoptosis. Further mechanistic analysis showed that CLU-knockdown blocked the PI3K/AKT signaling and reversed the protective effects of UC-MSCs-derived EVs. CONCLUSIONS: Administration of UC-MSCs and UC-MSCs-derived EVs on day 6 of CTX/BUS injection could effectively improve the ovarian function of POF mice. UC-MSCs-derived EVs carrying CLU promoted proliferation and inhibited apoptosis of ovarian granulosa cells through activating the PI3K/AKT pathway. This study identifies a previously unrecognized molecular mechanism of UC-MSCs-mediated protective effects on POF, which pave the way for the use of cell-free therapeutic approach for POF.
Assuntos
Vesículas Extracelulares , Células-Tronco Mesenquimais , Fosfatidilinositol 3-Quinases , Insuficiência Ovariana Primária , Proteínas Proto-Oncogênicas c-akt , Transdução de Sinais , Cordão Umbilical , Feminino , Animais , Insuficiência Ovariana Primária/terapia , Insuficiência Ovariana Primária/metabolismo , Insuficiência Ovariana Primária/induzido quimicamente , Vesículas Extracelulares/metabolismo , Vesículas Extracelulares/transplante , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/citologia , Camundongos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Humanos , Fosfatidilinositol 3-Quinases/metabolismo , Cordão Umbilical/citologia , Clusterina/metabolismo , Apoptose , Transplante de Células-Tronco Mesenquimais/métodos , Ovário/metabolismo , Células da Granulosa/metabolismo , Proliferação de Células , Bussulfano/farmacologiaRESUMO
BACKGROUND: There has been a significant surge in animal studies of stem cell-derived extracellular vesicles (EVs) therapy for the treatment of premature ovarian failure (POF) but its efficacy remains unknown and a comprehensive and up-to-date meta-analysis is lacking. Before clinical translation, it is crucial to thoroughly understand the overall impact of stem cell-derived EVs on POF. METHODS: PubMed, EMBASE, Cochrane Library, Web of Science were searched up to February 18, 2024. The risk of bias was evaluated according to Cochrane Handbook criteria, while quality of evidence was assessed using the SYRCLE system. The PRISMA guidance was followed. Trial sequential analysis was conducted to assess outcomes, and sensitivity analysis and publication bias analysis were performed using Stata 14. RESULTS: Data from 25 studies involving 339 animals were extracted and analyzed. The analysis revealed significant findings: stem cell-derived EVs increase ovary weight (SMD = 3.88; 95% CI: 2.50 ~ 5.25; P < 0.00001; I2 = 70%), pregnancy rate (RR = 3.88; 95% CI: 1.94 ~ 7.79; P = 0.0001; I2 = 0%), count of births (SMD = 2.17; 95% CI: 1.31 ~ 3.04; P < 0.00001; I2 = 69%) and counts of different types of follicles. In addition, it elevates the level of AMH (SMD = 4.15; 95% CI: 2.75 ~ 5.54; P < 0.00001; I2 = 88%) and E2 (SMD = 2.88; 95% CI: 2.02 ~ 3.73; P < 0.00001; I2 = 80%) expression, while reducing FSH expression (SMD = -5.05; 95% CI: -6.60 ~ -3.50; P < 0.00001; I2 = 90%). Subgroup analysis indicates that the source of EVs, animal species, modeling method, administration route, and test timepoint affected efficacy. Trial sequential analysis showed that there was sufficient evidence to confirm the effects of stem cell-derived EVs on birth counts, ovarian weights, and follicle counts. However, the impact of stem cell-derived EVs on pregnancy rates needs to be further demonstrated through more animal experimental evidence. CONCLUSIONS: Stem cell-derived EVs demonstrate safety and efficacy in treating POF animal models, with potential improvements in fertility outcomes. TRIAL REGISTRATION: PROSPERO registration number: CRD42024509699.
Assuntos
Vesículas Extracelulares , Insuficiência Ovariana Primária , Feminino , Insuficiência Ovariana Primária/terapia , Insuficiência Ovariana Primária/metabolismo , Vesículas Extracelulares/metabolismo , Animais , Células-Tronco/metabolismo , Ovário/metabolismo , Humanos , Gravidez , Modelos Animais de DoençasRESUMO
BACKGROUND: Accumulating studies have highlighted the significant role of circulating metabolomics in the etiology of reproductive system disorders. However, the causal effects between genetically determined metabolites (GDMs) and reproductive diseases, including primary ovarian insufficiency (POI), polycystic ovary syndrome (PCOS), and abnormal spermatozoa (AS), still await thorough clarification. METHODS: With the currently most comprehensive genome-wide association studies (GWAS) data of metabolomics, systematic two-sample Mendelian randomization (MR) analyses were conducted to disclose causal associations between 1,091 blood metabolites and 309 metabolite ratios with reproductive disorders. The inverse-variance weighted (IVW) method served as the primary analysis approach, and multiple effective MR methods were employed as complementary analyses including MR-Egger, weighted median, constrained maximum likelihood (cML-MA), contamination mixture method, robust adjusted profile score (MR-RAPS), and debiased inverse-variance weighted method. Heterogeneity and pleiotropy were assessed via MR-Egger intercept and Cochran's Q statistical analysis. Outliers were detected by Radial MR and MR-PRESSO methods. External replication and metabolic pathway analysis were also conducted. RESULTS: Potential causal associations of 63 GDMs with POI were unearthed, and five metabolites with strong causal links to POI were emphasized. Two metabolic pathways related to the pathogenesis of POI were pinpointed. Suggestive causal effects of 70 GDMs on PCOS were detected, among which 7 metabolites stood out for strong causality with elevated PCOS risk. Four metabolic pathways associated with PCOS mechanisms were recognized. For AS, 64 GDMs as potential predictive biomarkers were identified, particularly highlighting two metabolites for their strong causal connections with AS. Three pathways underneath the AS mechanism were identified. Multiple assessments were conducted to further confirm the reliability and robustness of our causal inferences. CONCLUSION: By extensively assessing the causal implications of circulating GDMs on reproductive system disorders, our study underscores the intricate and pivotal role of metabolomics in reproductive ill-health, laying a theoretical foundation for clinical strategies from metabolic insights.
Assuntos
Estudo de Associação Genômica Ampla , Análise da Randomização Mendeliana , Metaboloma , Síndrome do Ovário Policístico , Insuficiência Ovariana Primária , Humanos , Síndrome do Ovário Policístico/genética , Síndrome do Ovário Policístico/sangue , Síndrome do Ovário Policístico/metabolismo , Feminino , Masculino , Insuficiência Ovariana Primária/genética , Insuficiência Ovariana Primária/sangue , Insuficiência Ovariana Primária/metabolismo , Metabolômica/métodos , Espermatozoides/metabolismoRESUMO
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.
Assuntos
Cisplatino , Fosfatidilinositol 3-Quinases , Insuficiência Ovariana Primária , Transdução de Sinais , Animais , Insuficiência Ovariana Primária/induzido quimicamente , Insuficiência Ovariana Primária/tratamento farmacológico , Insuficiência Ovariana Primária/metabolismo , Feminino , Humanos , Camundongos , Cisplatino/efeitos adversos , Transdução de Sinais/efeitos dos fármacos , Células HeLa , Fosfatidilinositol 3-Quinases/metabolismo , Camundongos Endogâmicos BALB C , Apoptose/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto , Hormônio Antimülleriano/sangue , Hormônio Antimülleriano/metabolismo , Antineoplásicos/farmacologia , Neoplasias do Colo do Útero/tratamento farmacológico , Neoplasias do Colo do Útero/metabolismo , Neoplasias do Colo do Útero/patologia , Receptores do FSH/metabolismo , Receptores do FSH/genéticaRESUMO
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.
Assuntos
Medicamentos de Ervas Chinesas , MicroRNAs , Estresse Oxidativo , Insuficiência Ovariana Primária , RNA Circular , Animais , Feminino , Ratos , Cicloexenos , Modelos Animais de Doenças , Medicamentos de Ervas Chinesas/farmacologia , MicroRNAs/metabolismo , MicroRNAs/genética , Ovário/efeitos dos fármacos , Ovário/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Insuficiência Ovariana Primária/tratamento farmacológico , Insuficiência Ovariana Primária/metabolismo , Insuficiência Ovariana Primária/genética , Ratos Sprague-Dawley , RNA Circular/genética , RNA Mensageiro/metabolismo , RNA Mensageiro/genética , Compostos de Vinila/farmacologiaRESUMO
Premature ovarian insufficiency (POI) critically affects female reproductive health, with obesity being a significant and recognized risk factor. Interleukin-27 (IL-27), known for its role in immune modulation and inflammation, has garnered attention in metabolic syndrome research. Nonetheless, the role of these immunometabolic factors on the initiation of POI remains to be unraveled. Our investigation delves into the influence of impaired IL-27 signaling on POI induction, particularly under the challenge of a high-fat diet (HFD). We analyzed patients' serum profiles and established a correlation of increased serum triglycerides with decreased IL-27 levels in POI cases. Experiments on C57BL/6 mice lacking the IL-27 receptor alpha (Il27ra-/-) revealed that when subjected to HFD, these mice developed hallmark POI symptoms. This includes escalated lipid deposition in both liver and ovarian tissues, increased ovarian macrophages cellular aging, and diminished follicle count, all pointing to compromised ovarian function. These findings unveil a novel pathway wherein impaired IL-27 signaling potentiates the onset of POI in the presence of HFD. Understanding the intricate interplay between IL-27, metabolic alterations, and immune dysregulation sheds light on potential therapeutic avenues for managing POI, offering hope for improved reproductive health outcomes.
Assuntos
Dieta Hiperlipídica , Macrófagos , Insuficiência Ovariana Primária , Receptores de Interleucina , Transdução de Sinais , Adulto , Animais , Feminino , Humanos , Camundongos , Senescência Celular , Dieta Hiperlipídica/efeitos adversos , Interleucinas/metabolismo , Macrófagos/metabolismo , Macrófagos/imunologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ovário/metabolismo , Ovário/patologia , Insuficiência Ovariana Primária/patologia , Insuficiência Ovariana Primária/metabolismo , Insuficiência Ovariana Primária/imunologia , Receptores de Interleucina/metabolismo , Receptores de Interleucina/genéticaRESUMO
In this study, a mouse model of premature ovarian failure(POF) was constructed by injecting D-galactose(200 mg·kg~(-1)) into the back of the neck for 6 weeks. The mice were randomly divided into a normal group(group N), a model group(group M), and a Qiwei Guibao Granules group(group A, 12.87 g·kg~(-1)). Starting from the 11th day of modeling, group A was treated with Qiwei Guibao Granules by gavage for 32 days, while group M and group N were given equal volume of saline. Metabolomics analysis was used to explore the mechanism of action of Qiwei Guibao Granules in the treatment of POF. The results showed that compared with group N, the group M exhibited decreased wet weight of bilateral ovaries, increased levels of LH and FSH in serum, and significantly decreased levels of E_2 and PROG. After treatment with Qiwei Guibao Granules, compared with the group M, the group A showed a significant increase in the wet weight of bilateral ovaries, a significant decrease in the levels of FSH and LH in serum, and a significant increase in the level of E_2. Metabolomics analysis revealed 55 differential metabolites identified between group N and group M(14 upregulated and 41 downregulated compared with group N) and 82 differential metabolites identified between group M and group A(56 upregulated and 26 downregulated compared with group M), with 5 metabolites showing consistent changes between the group N vs group M. After excluding these 5 metabolites, 77 metabolites that changed after intervention with Qiwei Guibao Granules were focused on. These mainly involved histidine metabolism, glycine, serine, and threonine metabolism, and glycerophospholipid metabolism. Among them, carnosine, 1-methyl-L-histidine, imidazoleacetic acid, choline, L-threonine, beta-hydroxypyruvic acid, phosphatidylcholine, and glycerol-3-phosphate were the major differential metabolites in these three metabolic pathways. Therefore, Qiwei Guibao Granules may exert therapeutic effects on POF mice by regulating amino acid metabolism and lipid metabolism in the mouse body.
Assuntos
Medicamentos de Ervas Chinesas , Metabolômica , Insuficiência Ovariana Primária , Animais , Feminino , Insuficiência Ovariana Primária/tratamento farmacológico , Insuficiência Ovariana Primária/metabolismo , Medicamentos de Ervas Chinesas/administração & dosagem , Medicamentos de Ervas Chinesas/farmacologia , Camundongos , Humanos , Ovário/efeitos dos fármacos , Ovário/metabolismo , Modelos Animais de DoençasRESUMO
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.
Assuntos
Proteínas de Ligação a DNA , Receptor beta de Estrogênio , Camundongos Endogâmicos C57BL , Mitocôndrias , Biogênese de Organelas , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo , Insuficiência Ovariana Primária , Fatores de Transcrição , Feminino , Animais , Receptor beta de Estrogênio/metabolismo , Receptor beta de Estrogênio/genética , Camundongos , Insuficiência Ovariana Primária/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Mitocôndrias/metabolismo , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Medicamentos de Ervas Chinesas/farmacologia , Proteínas Mitocondriais/metabolismo , Proteínas Mitocondriais/genética , Superóxido Dismutase/metabolismo , Proteínas de Grupo de Alta MobilidadeRESUMO
Premature ovarian insufficiency (POI), a major cause of female infertility, is defined as follicular atresia and a rapid loss of germ cells in women of reproductive age due to ovarian failure. Recently, findings from several studies have indicated that human umbilical cord mesenchymal stem cells (hUMSCs) can alleviate ovarian dysfunction resulting from POI. However, the mechanisms underlying this effect require further clarification. In this study, a mouse model of POI was established as achieved with an intraperitoneal injection of cyclophosphamide (CTX) into female C57BL/6J mice in vivo. These POI mice received a 1-week intervention of hUMACs. In addition, an in vitro POI model was also included. The cultured supernatants of hUMSCs and glycogen synthase kinase 3 beta (GSK3ß) inhibitor (SB216763) were used to treat theca cells (TCs) exposed to CTX. Hematoxylin and Eosin (H&E) staining and Enzyme-linked immunosorbent assay (ELISA) were used to assess ovarian structure and morphology, as well as endocrine function in these POI mice. Based on results from the ELISA and JC-1 labeling, CTX exerted significant detrimental effects on testosterone levels and the mitochondrial membrane potential in TCs. Subsequently, Western Blot, Immunofluorescence staining (IF), and Quantitative real-time polymerase chain reaction (qRT-PCR) were used to evaluate various indicators of testosterone synthesis function and mitochondrial dynamics in ovaries and TCs of POI mice. In vivo, dysfunctions in ovarian structure and function in the POI mouse model were effectively restored following hUMSCs treatment, and abnormalities in hormone synthesis were significantly reduced. Furthermore, when the stem cell supernatants of hUMSCs were applied to TCs in vitro we found that GSK3ß expression was reduced, the imbalance of mitochondrial dynamics was alleviated, and the ability of mitochondrial testosterone synthesis was increased. Taken together, our results indicate that hUMSCs treatment can restore the imbalance of mitochondrial dynamics and restart testosterone synthesis of TCs by suppressing GSK3ß expression, ultimately alleviating POI damage.
Assuntos
Glicogênio Sintase Quinase 3 beta , Células-Tronco Mesenquimais , Dinâmica Mitocondrial , Insuficiência Ovariana Primária , Células Tecais , Animais , Feminino , Glicogênio Sintase Quinase 3 beta/metabolismo , Camundongos , Células Tecais/metabolismo , Células Tecais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Humanos , Insuficiência Ovariana Primária/metabolismo , Insuficiência Ovariana Primária/terapia , Dinâmica Mitocondrial/efeitos dos fármacos , Modelos Animais de Doenças , Camundongos Endogâmicos C57BL , Ovário/metabolismo , Ovário/efeitos dos fármacos , Cordão Umbilical/citologia , Ciclofosfamida/farmacologia , Transplante de Células-Tronco Mesenquimais/métodos , Mitocôndrias/metabolismo , Mitocôndrias/efeitos dos fármacos , Testosterona , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Indóis , MaleimidasRESUMO
BACKGROUND: Premature ovarian failure (POF) is a clinical condition characterized by the cessation of ovarian function, leading to infertility. The underlying molecular mechanisms remain unclear, and no predictable biomarkers have been identified. This study aimed to investigate the protein and metabolite contents of serum extracellular vesicles to investigate underlying molecular mechanisms and explore potential biomarkers. METHODS: This study was conducted on a cohort consisting of 14 POF patients and 16 healthy controls. The extracellular vesicles extracted from the serum of each group were subjected to label-free proteomic and unbiased metabolomic analysis. Differentially expressed proteins and metabolites were annotated. Pathway network clustering was conducted with further correlation analysis. The biomarkers were confirmed by ROC analysis and random forest machine learning. RESULTS: The proteomic and metabolomic profiles of POF patients and healthy controls were compared. Two subgroups of POF patients, Pre-POF and Pro-POF, were identified based on the proteomic profile, while all patients displayed a distinguishable metabolomic profile. Proteomic analysis suggested that inflammation serves as an early factor contributing to the infertility of POF patients. For the metabolomic analysis, despite the dysfunction of metabolism, oxidative stress and hormone imbalance were other key factors appearing in POF patients. Signaling pathway clustering of proteomic and metabolomic profiles revealed the progression of dysfunctional energy metabolism during the development of POF. Moreover, correlation analysis identified that differentially expressed proteins and metabolites were highly associated, with six of them being selected as potential biomarkers. ROC curve analysis, together with random forest machine learning, suggested that AFM combined with 2-oxoarginine was the best diagnostic biomarker for POF. CONCLUSIONS: Omics analysis revealed that inflammation, oxidative stress, and hormone imbalance are factors that damage ovarian tissue, but the progressive dysfunction of energy metabolism might be the critical pathogenic pathway contributing to the development of POF. AFM combined with 2-oxoarginine serves as a precise biomarker for clinical POF diagnosis.
Assuntos
Biomarcadores , Metabolismo Energético , Vesículas Extracelulares , Metabolômica , Insuficiência Ovariana Primária , Proteômica , Humanos , Feminino , Vesículas Extracelulares/metabolismo , Biomarcadores/sangue , Metabolismo Energético/fisiologia , Insuficiência Ovariana Primária/sangue , Insuficiência Ovariana Primária/diagnóstico , Insuficiência Ovariana Primária/metabolismo , Proteômica/métodos , Adulto , Metabolômica/métodos , Progressão da Doença , Metaboloma/fisiologiaRESUMO
Granulosa cells (GCs), the largest cell population and primary source of steroid hormones in the ovary, are the important somatic ovarian components. They have critical roles in folliculogenesis by supporting oocyte, facilitating its growth, and providing a microenvironment suitable for follicular development and oocyte maturation, thus having essential functions in maintaining female fertility and in reproductive health in general. Pyroptotic death of GCs and associated inflammation have been implicated in the pathogenesis of several reproductive disorders in females including Premature Ovarian Insufficiency (POI) and Polycystic Ovary Syndrome (PCOS). Here, I reviewed factors, either intrinsic or extrinsic, that induce or inhibit pyroptosis in GCs in various models of these disorders, both in vitro and in vivo, and also covered associated molecular mechanisms. Most of these studied factors influence NLRP3 inflammasome- and GSDMD (Gasdermin D)-mediated pyroptosis in GCs, compared to other inflammasomes and gasdermins (GSDMs). I conclude that a more complete mechanistic understanding of these factors in terms of GC pyroptosis is required to be able to develop novel strategies targeting inflammatory cell death in the ovary.
Assuntos
Células da Granulosa , Síndrome do Ovário Policístico , Insuficiência Ovariana Primária , Piroptose , Feminino , Insuficiência Ovariana Primária/metabolismo , Insuficiência Ovariana Primária/patologia , Síndrome do Ovário Policístico/metabolismo , Síndrome do Ovário Policístico/patologia , Humanos , Piroptose/efeitos dos fármacos , Células da Granulosa/metabolismo , Células da Granulosa/efeitos dos fármacos , Animais , Inflamassomos/metabolismoRESUMO
Premature ovarian insufficiency (POI) is one of the important causes of female infertility. Yet the aetiology for POI is still elusive. FBXW7 (F-box with 7 tandem WD) is one of the important components of the Skp1-Cullin1-F-box (SCF) E3 ubiquitin ligase. FBXW7 can regulate cell growth, survival and pluripotency through mediating ubiquitylation and degradation of target proteins via triggering the ubiquitin-proteasome system, and is associated with tumorigenesis, haematopoiesis and testis development. However, evidence establishing the function of FBXW7 in ovary is still lacking. Here, we showed that FBXW7 protein level was significantly decreased in the ovaries of the cisplatin-induced POI mouse model. We further showed that mice with oocyte-specific deletion of Fbxw7 demonstrated POI, characterized with folliculogenic defects, early depletion of follicle reserve, disordered hormonal secretion, ovarian dysfunction and female infertility. Impaired oocyte-GCs communication, manifested as down-regulation of connexin 37, may contribute to follicular development failure in the Fbxw7-mutant mice. Furthermore, single-cell RNA sequencing and in situ hybridization results indicated an accumulation of Clu and Ccl2 transcripts, which may alter follicle microenvironment deleterious to oocyte development and accelerate POI. Our results establish the important role of Fbxw7 in folliculogenesis and ovarian function, and might provide valuable information for understanding POI and female infertility.
Assuntos
Proteína 7 com Repetições F-Box-WD , Oócitos , Folículo Ovariano , Insuficiência Ovariana Primária , Animais , Feminino , Insuficiência Ovariana Primária/genética , Insuficiência Ovariana Primária/metabolismo , Insuficiência Ovariana Primária/patologia , Proteína 7 com Repetições F-Box-WD/metabolismo , Proteína 7 com Repetições F-Box-WD/genética , Oócitos/metabolismo , Camundongos , Folículo Ovariano/metabolismo , Folículo Ovariano/crescimento & desenvolvimento , Folículo Ovariano/patologia , Modelos Animais de Doenças , Deleção de Genes , Camundongos Knockout , Infertilidade Feminina/genética , Infertilidade Feminina/metabolismo , Infertilidade Feminina/patologia , Cisplatino/efeitos adversosRESUMO
Cisplatin is a widely used chemotherapeutic drug that can induce ovarian damage. Icariin (ICA), a natural antioxidant derived from Epimedium brevicornum Maxim., has been found to protect against organ injury. The aim of the present study was to investigate whether ICA can exert an ovarian-protective effect on cisplatin induced premature ovarian failure (POF) and the underlying mechanism involved. The preventive effect of ICA was evaluated using body weight, the oestrous cycle, ovarian histological analysis, and follicle counting. ICA treatment increased body weight, ovarian weight, and the number of follicles and improved the oestrous cycle in POF mice. ICA reduced cisplatin-induced oxidative damage and upregulated the protein expression levels of Nrf2, GPX4 and HO-1. Moreover, ICA reduced the expression levels of Bax and γH2AX and inhibited ovarian apoptosis. In addition, ICA activated the Nrf2 pathway in vitro and reversed changes in the viability of cisplatin-induced KGN cells, reactive oxygen species (ROS) levels, lipid peroxidation, and apoptosis, and these effects were abrogated when Nrf2 was knocked down or inhibited. Molecular docking confirmed that ICA promotes the release of Nrf2 by competing with Nrf2 for binding to Keap1. The inhibitory effects of ICA on cisplatin-induced oxidative stress, ferroptosis, and apoptosis may be mediated by its modulatory effects on the Nrf2 pathway, providing a novel perspective on the potential mechanisms by which ICA prevents POF.