Gut microbiota dysbiosis-derived macrophage pyroptosis causes polycystic ovary syndrome via steroidogenesis disturbance and apoptosis of granulosa cells.

Gut microbiota dysbiosis is critical in the etiology of polycystic ovary syndrome (PCOS). However, the mechanisms of gut microbiota in PCOS pathogenesis have not been fully elucidated. We aimed to explore the role of gut microbiota-derived macrophage pyroptosis in PCOS. This study conducted dehydroepiandrosterone (DHEA) induced PCOS mice model, 16S rDNA sequencing, western blot, genetic knocking out, transcriptome and translatome profiling, et al. to evaluate the underlying mechanisms. 16S rDNA sequencing showed reduced gut Akkermansia and elevated gram-negative bacteria (Desulfovibrio and Burkholderia) abundances in DHEA induced PCOS mice, which was accompanied by increased serum lipopolysaccharide (LPS). LPS could induce macrophage pyroptosis in mice ovaries, also activated in PCOS. Gasdermin D (GSDMD) is the final executor of macrophage pyroptosis. We demonstrated that Gsdmd knockout in mice could dramatically ameliorate PCOS. Mechanistically, transcriptome and translatome profiling revealed that macrophage pyroptosis disrupted estrogen production and promoted apoptosis of granulosa cells. Interferon (IFN)-γ, which was elevated in PCOS mice serum and ovaries, enhanced macrophage pyroptosis and exacerbated its effect on estrogen receptor in granulosa cells. Inspiringly, we identified that disulfiram and metformin could augment gut Akkermansia abundance, reduce serum IFN-γ level, inhibit macrophage pyroptosis in ovaries, therefore ameliorating PCOS. Collectively, this study emphasizes that macrophage pyroptosis, which was induced by gut microbiota dysbiosis and enhanced by IFN-γ, plays a key role in PCOS pathogenesis through estrogen synthesis dysfunction and apoptosis of granulosa cells. Disulfiram and metformin, which enhanced gut Akkermansia abundance and suppressed macrophage pyroptosis, may be considered as potential therapeutic strategies for PCOS.

Huang J ，Chen P ，Xiang Y ，Liang Q ，Wu T ，Liu J ，Zeng Y ，Zeng H ，Liang X ，Zhou C ... -  《-》

Effects of gut microbiota on omega-3-mediated ovary and metabolic benefits in polycystic ovary syndrome mice.

Polycystic ovary syndrome (PCOS) is a common reproductive endocrine disorder that frequently exhibits low-grade inflammation, pro-oxidant activity, and gut dysbiosis. PCOS has become one of the leading causes of female infertility worldwide. Recently, omega-3 polyunsaturated fatty acids (PUFAs) have been proven to benefit metabolic disorders in PCOS patients. However, its roles in the regulation of metabolic and endocrinal balances in PCOS pathophysiology are not clear. In the present study, we aimed to explore how omega-3 PUFAs alleviate ovarian dysfunction and insulin resistance in mice with dehydroepiandrosterone (DHEA)-induced PCOS by modulating the gut microbiota. We induced PCOS in female mice by injecting them with DHEA and then treated them with omega-3 PUFAs. 16S ribosomal DNA (rDNA) amplicon sequencing, fecal microbiota transplantation (FMT) and antibiotic treatment were used to evaluate the role of microbiota in the regulation of ovarian functions and insulin resistance (IR) by omega-3 PUFAs. To further investigate the mechanism of gut microbiota on omega-3-mediated ovarian and metabolic protective effects, inflammatory and oxidative stress markers in ovaries and thermogenic markers in subcutaneous and brown adipose tissues were investigated. We found that oral supplementation with omega-3 PUFAs ameliorates the PCOS phenotype. 16S rDNA analysis revealed that omega-3 PUFA treatment increased the abundance of beneficial bacteria in the gut, thereby alleviating DHEA-induced gut dysbiosis. Antibiotic treatment and FMT experiments further demonstrated that the mechanisms underlying omega-3 benefits likely involve direct effects on the ovary to inhibit inflammatory cytokines such as IL-1β, TNF-α and IL-18. In addition, the gut microbiota played a key role in the improvement of adipose tissue morphology and function by decreasing multilocular cells and thermogenic markers such as Ucp1, Pgc1a, Cited and Cox8b within the subcutaneous adipose tissues. These findings indicate that omega-3 PUFAs ameliorate androgen-induced gut microbiota dysbiosis. The gut microbiota plays a key role in the regulation of omega-3-mediated IR protective effects in polycystic ovary syndrome mice. Moreover, omega-3 PUFA-regulated improvements in the ovarian dysfunction associated with PCOS likely involve direct effects on the ovary to inhibit inflammation. Our findings suggest that omega-3 supplementation may be a promising therapeutic approach for the treatment of PCOS by modulating gut microbiota and alleviating ovarian dysfunction and insulin resistance.

Zhang H ，Zheng L ，Li C ，Jing J ，Li Z ，Sun S ，Xue T ，Zhang K ，Xue M ，Cao C ，Ouyang L ，Qian Z ，Xu R ，He Z ，Ma R ，Chen L ，Yao B ... -  《Journal of Ovarian Research》

Chinese medicine compound prescription HeQi San ameliorates chronic inflammatory states and modulates gut flora in dehydroepiandrosterone-induced polycystic ovary syndrome mouse model.

Polycystic ovary syndrome (PCOS) is a common and complex endocrine disease in women, with a prevalence of 5% to 18% worldwide. HeQi San (HQS) is a Chinese medicine compound prescription, which has been applied to treat various endocrine and metabolic diseases. The study was intended to investigate the effect of HQS on PCOS, and clarify the potential mechanism via in vivo and in vitro experiments. The PCOS mouse model was established by injecting the dehydroepiandrosterone (DHEA) subcutaneously and fading high-fat diet for 3 weeks. After making model, PCOS mice were treated with HQS (8.75 g/kg and 17.5 g/kg, ig.) for 4 weeks. Firstly, we assessed the histopathological changes in ovary tissues and detected the hormone level. Subsequently, the study evaluated the capability of anti-inflammatory and regulating macrophage polarization of HQS in vivo and in vitro. The secretion of inflammation indicators was measured with Elisa kits, and the expression level of phosphorylated nuclear factor kappa-B (P-NFκB) and B-lymphocyte activation antigen B7 (CD80) was measured by immunofluorescence and Western blot. Meanwhile, the apoptosis of ovarian granulosa cells was detected via tunel staining and Western blot. The co-culture model in vitro was utilized to assess the effect between macrophage polarization and human ovarian granulosa cells (KGN cells) apoptosis. Furthermore, 16S rDNA sequencing was utilized to elevate gut microbiota change in PCOS mice. HQS reversed the abnormal hormone increase, ameliorated insulin resistance, and improved histopathological changes of the ovary tissue to exert the therapeutic effect. HQS inhibited the expression of P-NF-κB and decreased the production of interleukin 6 (IL-6) and tumor necrosis factor-α (TNF-α) to further prohibit the macrophage M1 polarization in ovary tissues and macrophages. The apoptosis-positive cells, Bcl-2 Assaciated X protein (BAX), and cleaved-caspase 3 expression were also decreased in the treatment group. The B-cell lymphoma-2 (Bcl2) expression was enhanced after HQS treatment in vivo. The co-culture experiments also verified that HQS could prevent the apoptosis of KGN cells. Furthermore, HQS mediated the abundance of gut flora. The abundance of bifldobacterium and parasutterella was increased and the abundance of lachnoclostridium was decreased. The study verified that HQS has the effect of anti-inflammation and inhibits macrophage M1 polarization. Besides, HQS could mediate the abundance of gut microbiota in mice with PCOS. Thus, this study would provide more reasonable basis of HQS for clinical use. In conclusion, HQS might be a potential candidate for PCOS treatment.

Li J ，Liu D ，Zhao H ，Zhang P ，Cai F ，Li H ，Chu S ... -  《-》

E. coli Nissle 1917 ameliorates mitochondrial injury of granulosa cells in polycystic ovary syndrome through promoting gut immune factor IL-22 via gut microbiota and microbial metabolism.

Gut microbiota and its metabolites have regulatory effects on PCOS related ovarian dysfunction and insulin resistance. Escherichia coli Nissle 1917 (EcN) is a genetically controlled probiotic with an excellent human safety record for improving gut microbiome metabolic disorders and immune system disorders. Here we focused to explore the application and effect of probiotic EcN on the gut microbiota-metabolism-IL-22-mitochondrial damage axis in PCOS. PCOS mice were constructed with dehydroepiandrosterone (DHEA) and treated with EcN, FMT or IL-22 inhibitors. Clinically control and PCOS subjects were included for further analysis. Serum and follicular fluid supernatant levels of sex hormones, insulin, glucose, cholesterol, and inflammatory factors were detected by ELISA and biochemical reagents. The pathological changes of ovarian tissues were observed by HE staining. The JC-1 level and COX4 gene expression in granulosa cells was detected by ELISA and RT-qPCR. The expressions of progesterone receptor A (PR-A), LC3II/I, Beclin1, p62 and CytC were detected by western blot. The number of autophagosomes in granulosa cells was observed by electron microscopy. 16S rRNA and LC-MS/MS were used to analyze the changes of gut microbiota and metabolism. EcN promoted the recovery of sex hormone levels and ovarian tissue morphology, promoted the expression of IL-22, COX4 and PR-A in granulosa cells, and inhibited mitophagy in PCOS mice. EcN decreased the number of gut microbiota, and significantly increased the abundance of Adlercreutzia, Allobaculum, Escherichia-Shigella and Ileibacterium in PCOS mice. EcN improved metabolic disorders in PCOS mice by improving Amino sugar and nucleotide sugar metabolism pathways. IL-22 was positively associated with Ileibacterium, Adlercreutzia and Progesterone, negatively associated with RF39, Luteinizing hormone, Testosterone, N-Acetylglucosamin, L-Fucose and N-Acetylmannosamin. FMT reconfirmed that EcN ameliorated mitochondrial damage in granulosa cells of PCOS mice by gut microbiota, but this process was blocked by IL-22 inhibitor. Clinical trials have further demonstrated reduced IL-22 levels and mitochondrial damage in granulosa cells in PCOS patients. EcN improved IL-22 level and mitochondrial damage of granulosa cells in PCOS mice by promoting the recovery of sex hormone levels and ovarian tissue morphology, inhibiting the amount of gut microbiota, and promoting amino sugar and nucleotide sugar metabolism.

Luo M ，Chen Y ，Pan X ，Chen H ，Fan L ，Wen Y ... -  《Frontiers in Immunology》

Evaluating the therapeutic potential of moxibustion on polycystic ovary syndrome: a rat model study on gut microbiota and metabolite interaction.

Polycystic ovary syndrome (PCOS) is a common systemic disorder related to endocrine disorders, affecting the fertility of women of childbearing age. It is associated with glucose and lipid metabolism disorders, altered gut microbiota, and insulin resistance. Modern treatments like pioglitazone, metformin, and spironolactone target specific symptoms of PCOS, while in Chinese medicine, moxibustion is a common treatment. This study explores moxibustion's impact on PCOS by establishing a dehydroepiandrosterone (DHEA)-induced PCOS rat model. Thirty-six specific pathogen-free female Sprague-Dawley rats were divided into four groups: a normal control group (CTRL), a PCOS model group (PCOS), a moxibustion treatment group (MBT), and a metformin treatment group (MET). The MBT rats received moxibustion, and the MET rats underwent metformin gavage for two weeks. We evaluated ovarian tissue changes, serum testosterone, fasting blood glucose (FBG), and fasting insulin levels. Additionally, we calculated the insulin sensitivity index (ISI) and the homeostasis model assessment of insulin resistance index (HOMA-IR). We used 16S rDNA sequencing for assessing the gut microbiota, 1H NMR spectroscopy for evaluating metabolic changes, and Spearman correlation analysis for investigating the associations between metabolites and gut microbiota composition. The results indicate that moxibustion therapy significantly ameliorated ovarian dysfunction and insulin resistance in DHEA-induced PCOS rats. We observed marked differences in the composition of gut microbiota and the spectrum of fecal metabolic products between CTRL and PCOS rats. Intriguingly, following moxibustion intervention, these differences were largely diminished, demonstrating the regulatory effect of moxibustion on gut microbiota. Specifically, moxibustion altered the gut microbiota by increasing the abundance of UCG-005 and Turicibacter, as well as decreasing the abundance of Desulfovibrio. Concurrently, we also noted that moxibustion promoted an increase in levels of short-chain fatty acids (including acetate, propionate, and butyrate) associated with the gut microbiota of PCOS rats, further emphasizing its positive impact on gut microbes. Additionally, moxibustion also exhibited effects in lowering FBG, testosterone, and fasting insulin levels, which are key biochemical indicators associated with PCOS and insulin resistance. Therefore, these findings suggest that moxibustion could alleviate DHEA-induced PCOS by regulating metabolic levels, restoring balance in gut microbiota, and modulating interactions between gut microbiota and host metabolites.

Lin Y ，Zeng H ，Lin J ，Peng Y ，Que X ，Wang L ，Chen L ，Bai N ... -  《Frontiers in Cellular and Infection Microbiology》