FXR overexpression alleviates cholestasis via NLRC4 inflammasome suppression and bile acid homeostasis regulation.

Cholestasis, a major driver of liver disease progression, is characterized by toxic bile acid accumulation due to impaired bile flow, potentially leading to hepatic fibrosis, cirrhosis, and hepatocellular carcinoma. To date, the pathogenesis of cholestasis has remained incompletely understood. In the present study, we investigated the role of farnesoid X receptor (FXR) in modulating the NLR family CARD domain-containing protein 4 (NLRC4) inflammasome activity using in vitro (AML-12 hepatocytes) and in vivo (C57BL/6 mice) models of lithocholic acid (LCA)-induced cholestasis. LCA suppressed FXR expression, downregulated bile acid transporters (Bsep, Mrp2, Ntcp), and elevated serum biomarkers of liver injury. Through restored hepatic function by FXR lentiviral vectors, FXR overexpression reduced bile acid accumulation and mitigated inflammation and oxidative stress. In addition, FXR overexpression suppressed bile acid synthesis enzymes (Cyp7a1, Cyp8b1) by upregulating Shp and Fgf15, while enhancing detoxification through Ugt1a1 and Sult2a1. Interestingly, molecular docking analysis and Co-IP experiments demonstrated a direct interaction between FXR and NLRC4. Furthermore, FXR overexpression significantly inhibited NLRC4 inflammasome activation and decreased the expression of NLRC4, caspase-1, IL-1β, and IL-18, thereby attenuating inflammation and oxidative stress. Conversely, FXR knockdown reversed these effects. In addition, to delineate the contribution of NLRC4 inflammasome activation to IL-18 and IL-1β elevation, NLRC4-targeting siRNA-mediated knockdown and NLRC4-encoding plasmid-driven overexpression strategies were systematically employed. Furthermore, DCFH-DA staining was adopted to visualize reactive oxygen species (ROS). In conclusion, for the first time, we found that FXR overexpression alleviates LCA-induced cholestasis by regulating bile acid metabolism and inhibiting NLRC4 inflammasome activation, providing a novel therapeutic strategy for drug development targeting the FXR-NLRC4 axis.