Carpaine alleviates NASH-related fibrosis by targeting Nid1 to inhibit IL-6/JAK/STAT3 signaling and macrophage M1 polarization

Non-alcoholic steatohepatitis (NASH)-related liver fibrosis arises from interacting metabolic and inflammatory disturbances, yet effective therapies remain limited. Carpaine (CP), a natural alkaloid derived from Carica papaya, exhibits anti-inflammatory, anti-thrombocytopenic, and cytoprotective activities, but its role in NASH or fibrosis remains unclear. This study aimed to elucidate the protective effects and molecular mechanisms of CP against NASH-related fibrosis. Integrated metabolomic and proteomic analyses revealed a marked decrease of CP during NASH progression. To verify its biological relevance, the CDAHFD-induced NASH mouse model, AAV8-mediated nidogen-1 (Nid1) knockdown, and co-culture systems of hepatic stellate cells (HSCs) and macrophages were applied to comprehensively evaluate the therapeutic efficacy of CP. CP levels were inversely correlated with Nid1, which was significantly upregulated in the fibrotic livers of both mice and patients with NASH. CP administration downregulated Nid1 expression, attenuated liver fibrosis, and improved liver function. Nid1 knockdown together with exogenous recombinant Nid1 supplementation further confirmed Nid1 as a crucial mediator of CP action. Mechanistically, CP impeded Nid1-dependent activation of the JAK2/STAT3 pathway and IL-6 autocrine signaling in HSCs, thereby indirectly attenuating macrophage M1 polarization and inflammatory amplification. Moreover, CP treatment or Nid1 silencing significantly modulated gut microbial composition, enriching beneficial taxa associated with metabolic homeostasis. Collectively, CP alleviates NASH-related liver fibrosis by modulating the Nid1-JAK2/STAT3-IL-6 axis, highlighting CP as a promising multi-target therapeutic candidate for chronic metabolic liver diseases.