Hedyotis diffusa Willd. extract alleviates CCl4-induced liver fibrosis via modulation of the gut microbiota and FXR/SHP/CYP7A1-mediated bile acid metabolism.

Liver fibrosis is a critical stage in the progression of chronic liver diseases, yet effective therapeutic agents are limited. Hedyotis diffusa Willd., a traditional Chinese medicine herb with heat-clearing and detoxifying properties, has long been used to treat inflammatory disorders, hepatic dysfunction and malignancies. Although accumulating studies suggest that Hedyotis diffusa Willd. Extract (HDW) possesses hepatoprotective and antifibrotic potential, the underlying mechanisms, particularly those involving gut microbiota and bile acid (BA) metabolism along the gut-liver axis, remain largely undefined. To investigate the protective effects of HDW against carbon tetrachloride (CCl4)-induced liver fibrosis in mice, and to determine whether its antifibrotic efficacy is mediated by modulation of the gut microbiota-bile acid-FXR/SHP/CYP7A1 axis. Liver fibrosis was induced by intraperitoneal injection of 10% CCl4 in olive oil for five weeks. Histopathological changes were evaluated using H&E, Sirius red and Masson staining. Liver function was assessed using serum levels of ALT, AST, ALP and γ-GT. Collagen deposition was evaluated by measuring hepatic hydroxyproline (HYP) and fibrosis-related markers (HAase, IV-C, LN and PC-III). Activation of hepatic stellate cells was determined by α-SMA and Col1a1 expression. The composition of the gut microbial was profiled using 16 S rRNA sequencing, and the necessity of gut microbiota for HDW efficacy was evaluated through an antibiotic (ABX) cocktail intervention. Targeted BA metabolomics quantified BA profiles, while RT-qPCR and Western blotting evaluated FXR/SHP/CYP7A1 pathway activity. Intestinal barrier integrity was assessed by villus morphology, tight junction protein levels (Claudin-1, ZO-1, Occludin), and serum lipopolysaccharide (LPS). HDW treatment markedly alleviated CCl4-induced liver fibrosis, demonstrated by improved hepatic architecture, reduced serum transaminases and ALP/γ-GT, decreased hydroxyproline levels, and downregulation of α-SMA and Col1a1. HDW reshaped the gut microbial composition by enriching beneficial taxa, whereas ABX treatment markedly attenuated its antifibrotic effects, indicating that the therapeutic action of HDW is largely microbiota-dependent. HDW restored BA homeostasis and significantly increased fecal odeoxycholic acid. Consequently, hepatic FXR/SHP/CYP7A1 signaling was upregulated at both the mRNA and protein levels. Furthermore, HDW strengthened the intestinal epithelial barrier by enhancing tight junction integrity and reducing serum LPS. HDW exerts significant antifibrotic effects in CCl4-induced liver fibrosis via a gut microbiota-dependent mechanism involving the restoration of BA metabolism and the activation of the hepatic FXR/SHP/CYP7A1 axis. Given its ability to modulate gut microbial ecology, BA homeostasis and intestinal barrier integrity simultaneously, HDW is a promising therapeutic candidate for targeting the gut microbiota in the treatment of liver fibrosis.