Celastrol ameliorates metabolic dysfunction associated steatohepatitis by regulating the CYP7B1-mediated alternative bile acid synthetic pathway.

BACKGROUND: Metabolic dysfunction-associated steatohepatitis (MASH) is a progressive liver disease driven by steatosis, inflammation, and fibrosis. Celastrol (CEL), a natural bioactive compound, has exhibited promise in preclinical models in MASH treatment. However, its precise mechanism, particularly regarding bile acid (BA) metabolism, remains unclear.

PURPOSE: To elucidate the effects of CEL in MASH treatment and uncover the novel regulatory mechanism, focusing on the alternative BA synthetic pathway.

METHODS: MASH mouse models were induced by Western diet (WD) and high-fat diet (HFD). The effect of CEL was assessed through analyses of lipid levels, liver function, and histopathology. Proteomics and targeted metabolomics were performed to evaluate alterations in BA metabolism. Network pharmacology, transcriptomics, and molecular docking were used to predict CEL targets, which were further validated by RT-qPCR, Western blotting, and siRNA-mediated knockdown experiments in HepG2 cells.

RESULTS: CEL significantly improved liver morphology, reduced liver index, normalized lipid metabolism, decreased total bile acid levels, and alleviated liver injury in MASH models. Proteomics and targeted metabolomics revealed a significant increase in the ratio of 12α-hydroxylated (12-OH) to non-12-OH BA ratio in MASH, indicating a disruption in BA homeostasis, which was markedly corrected by CEL treatment. Further analyses showed suppression of the alternative BA synthetic pathway and downregulation of CYP7B1 in MASH, both of which were reversed upon CEL administration. Network pharmacology and molecular docking identified FXR and LXR as potential upstream regulators. Functional validation demonstrated that CEL upregulated CYP7B1 expression by activating FXR/LXR signaling, thereby recovering alternative BA synthesis and metabolic homeostasis.

CONCLUSION: Our study demonstrated CEL as a novel MASH therapeutic that uniquely targets the FXR/LXR-CYP7B1 axis to reactivate the alternative BA synthetic pathway, providing a mechanistic basis for CEL in MASH intervention.