Evodiamine targets ZO-1 to ameliorate cholestatic liver disease: intestinal homeostasis as the core mediator of gut-liver axis repair and bile acid metabolism remodeling

BackgroundCholestatic liver disease (CLD) is a complex and multifactorial chronic disorder that requires a systematic and integrative approach for effective management. Evodiamine (EVO), a natural alkaloid derived from Evodiae Fructus, has demonstrated significant therapeutic potential in ameliorating digestive diseases. However, the beneficial effects of EVO on CLD and the underlying mechanisms remain poorly understood.ObjectiveThis study aims to elucidate the mechanisms through which EVO modulates the progression of CLD, with a particular focus on the regulation of gut-liver axis homeostasis.MethodsThe therapeutic efficacy of EVO in bile duct ligation (BDL)- and α-naphthyl isothiocyanate (ANIT)-induced CLD rat models was systematically evaluated. Furthermore, an integrative approach combining network pharmacology with multi-omics analyses-including transcriptomic, metagenomic sequencing, targeted bile acid (BAs) metabolomics -was employed to identify significantly altered molecular networks. Meanwhile, fecal microbiota transplantation (FMT) was conducted to validate the functional role of gut microbiota in the hepato-intestinal protective effects of EVO. Additionally, direct molecular targets as well as the function validation were confirmed through fecal microbiota transplantation (FMT), molecular docking, pull-down assays, surface plasmon resonance, cellular thermal shift assay.ResultsEVO treatment achieved significant synchronous protection of the liver and intestine in both BDL- and ANIT-induced CLD rats: it markedly ameliorated hepatic injury and reduced hepatic fibrosis, downregulated pro-inflammatory cytokine levels in the liver, while preserving intestinal barrier integrity and alleviating intestinal inflammation. Mechanistically, EVO exerted its hepato-intestinal protective effects by enhancing the expression and stability of the TJ protein ZO-1, which was identified as a direct functional target of EVO. Furthermore, EVO restored intestinal microbial homeostasis, corrected dysregulated BA metabolism-specifically normalizing deoxycholic acid (DCA) levels-and reversed gut microbial dysbiosis, as confirmed by FMT experiments demonstrating that the synchronous hepato-intestinal beneficial effects of EVO were partially mediated by gut microbiota.ConclusionEVO exerts a protective effect against CLD by directly targeting ZO-1 to strengthen intestinal barrier function, thereby restoring gut microbial balance and rebalancing BAs metabolism (especially DCA levels) in the gut-liver axis. This study uncovers a novel ZO-1-dependent mechanism of EVO in CLD, highlighting EVO as a promising candidate for the treatment of CLD and providing new insights into gut-liver axis-targeted therapies.