The gut microbiota improves reproductive dysfunction in obese mice by suppressing the NLRP3/ASC/caspase-1 axis

<b>Aim:</b> To explore the complex relationship between gut microbiota, obesity-related male reproductive impairments, and the NLRP3 inflammasome. <b>Methods:</b> A high-fat diet was administered to induce obesity in a mouse model, fecal microbiota transplantation or a high-dietary fiber diet (HDFD) was administered for 5 weeks to evaluate changes in parameters related to reproductive capacity, NLRP3, gut microbiota composition and metabolites in mice. <b>Results:</b> A high-fat diet induces obesity and decreases reproductive capacity in male mice. Fecal microbiota transplantation and HDFD can improve reproductive capacity in obese mice by adjusting the gut microbiota population to suppress the NLRP3/ASC/caspase-1 axis, thereby reducing IL-1β levels. <b>Conclusion:</b> This study offers a potential treatment for obesity-induced reproductive dysfunction by targeting the gut microbiota and the NLRP3 inflammasome pathway. This study looks at how gut bacteria, obesity and our immune system affect male reproductive health. We made mice obese by feeding them a high-fat diet. Then, we treated them with either a transplant of gut bacteria or a high-fiber diet for 5 weeks. We found that the high-fat diet made it harder for male mice to have babies. Both the transplant and the high-fiber diet helped improve their ability to reproduce. Changing the bacteria in their gut reduced inflammation by affecting the immune system. Our findings suggest that changing gut bacteria and focusing on this part of the immune system could help with reproductive problems caused by obesity. Obesity poses serious health threats and negatively affects male reproductive function. The NLRP3 inflammasome is implicated in obesity-related reproductive impairments. Dysbiosis in gut microbiota are linked to obesity but their role in male reproductive impairments and impact on NLRP3 is unclear. Mice were fed a high-fat diet (HFD) for 15 weeks to induce obesity. Obese mice were treated with fecal microbiota transplantation (FMT) or high-dietary fiber diet (HDFD) for 5 weeks. Changes in body weight, histological parameters, organ indices, reproductive ability parameters, and NLRP3-related biochemical parameters were assessed. 16S rRNA gene sequencing and UPLC-MS/MS technology were used to analyze changes in gut microbiota composition and metabolites. An HFD induces male mice to become obese, exhibiting severe hepatic fat accumulation and a higher fat-to-body weight ratio. HFD-induced obesity leads to “vacuole-like” changes in the seminiferous tubules of the testes, increased apoptosis of germ cells, decreased serum testosterone levels, disrupted normal spermatogenesis and reduced fertility and offspring numbers. HFD-induced obesity alters the composition of the gut microbiota, with significant differences observed between the HFD group and the control diet (CD) group. Specific bacterial families, such as <i>Lachnospiraceae</i> and <i>Muribaculaceae</i>, showed significant changes in abundance due to HFD feeding. FMT and HDFD both inhibit weight gain and fat accumulation caused by HFD. Mice treated with FMT and HDFD showed seminiferous tubule structures more similar to those fed with CD, with more orderly arrangement of germ cells, reduced apoptosis of spermatogenic cells, increased serum testosterone levels and improved fertility and offspring numbers. The gut microbiota structure of the FMT and HDFD groups became similar to that of the CD group. The <i>Bacillaceae</i>, <i>Staphylococcaceae</i>, <i>Streptococcaceae</i>, <i>Lactobacillaceae</i> and <i>Lachnospiraceae_NK4A136_group</i> significantly increased in the FMT group. The <i>Proteobacteria</i>, <i>Gammaproteobacteria</i> and <i>Bacteroidetes</i> significantly increased in the HDFD group. In the testes of HFD group mice, the expression of NLRP3, caspase-1, cleaved-caspase-1 and ASC increased, and the levels of IL-1β in the testes and serum were high, indicating activation of the NLRP3/ASC/caspase-1 axis. After FMT and HDFD treatment, the NLRP3 inflammasome pathway was inhibited, reducing IL-1β levels and improving the reproductive capacity of obese mice. FMT upregulated 93 metabolites, mainly steroids such as cortisol 21-acetate, hydrocortisone succinate, nandrolone and estrone. HDFD upregulated 71 metabolites, primarily steroid compounds. FMT or HDFD can improve reproductive dysfunction in HFD-induced obese mice by regulating gut microbiota and inhibiting the NLRP3/ASC/caspase-1 axis. Altering gut microbiota can enhance male fertility in obese mice but does not fully restore it to the level of healthy males.