Effects of metformin on gut microbiota in mice with liver fibrosis revealed by 16S rRNA sequencing and resequencing

Background: Liver fibrosis is associated with gut dysbiosis. Metformin has emerged as a promising method for treating organ fibrosis. Here, we investigated whether metformin ameliorates liver fibrosis by gut microbiota in mice induced by carbon tetrachloride (CCl4) and explored the underlying mechanism. Materials and Methods: The liver fibrosis mouse model was established and therapeutic effects of metformin were observed. We performed the antibiotics treatment, fecal microbiota transplantation (FMT) and 16S rRNA-based microbiota analysis to evaluate the effects of gut microbiota on metformin-treated liver fibrosis. We cultured and isolated the bacterial strain preferably enriched by metformin, and assessed its anti-fibrotic effects in CCl4-treated mice. Results: Metformin treatment repaired gut integrity of CCl4-induced mice and restored the expressions of tight junction proteins, ZO-1 and Occludin. It reduced bacteria number in colon tissues and reduced portal vein lipopolysaccharide (LPS) levels. The FMT of feces from the metformin-treated CCl4 mice alleviated liver fibrosis and reduced portal vein LPS levels. The markedly changed gut microbiota was screened out from feces of the metformin-treated CCl4 mice, which was named Lactobacillus sp. MF-1 (L sp. MF-1). In CCl4-treated mice, the daily gavage of L sp. MF-1 maintained gut integrity, inhibited BT, and reduced liver fibrosis. Mechanistically, metformin or L sp. MF-1 inhibited the apoptosis of intraepithelial lymphocytes (IELs), restored CD3+ IELs in the ileum, and restored CD4+Foxp3+ lamina propria lymphocytes (LPLs) in the colon. Conclusions: Metformin and its enriched L sp. MF-1 can reinforce the intestinal barrier to alleviate liver fibrosis by restoring immune function. Keywords: Metformin, gut microbiota, bacterial translocation, liver fibrosis, Lactobacillus, fecal microbiota transplantation