Diosgenin alleviates hepatic fibrosis via PI3K/Akt/mTOR mediated fatty acid metabolic reprogramming and NF-κB activation.

Dioscorea oppositifolia L., also called Shan Yao in traditional Chinese medicine, is widely used to treat diabetes, hyperthyroidism, and liver damage. Diosgenin (DIO), a natural steroidal sapogenin, found abundantly in Dioscorea oppositifolia L., has historically been used to treat liver disorders. This study aimed to investigate the effect of DIO against hepatic fibrosis and the underlying molecular mechanisms. We utilized a CCl4-induced mouse model of hepatic fibrosis and TGF-β1-stimulated LX-2 human hepatic stellate cells (HSCs) to determine the anti-fibrotic effect of DIO. Network pharmacology, molecular docking, cellular thermal shift assay (CETSA), and Western blotting were employed to identify the targets and elucidate mechanisms. In vivo, DIO markedly improved CCl4-induced liver damage, collagen deposition and α-SMA expression, while reducing serum levels of ALT, AST, ALP, and hydroxyproline. In vitro, DIO suppressed TGF-β1-induced proliferation, activation, and fibrogenic gene expression in LX2 cells. Network pharmacology and molecular docking identified mTOR and PIK3CA as direct binding targets of DIO, which was further validated by cellular thermal shift assay (CETSA). Consistently, DIO blocked the PI3K/Akt/mTOR signaling cascade in activated HSCs, subsequently inhibiting NF-κB activation and attenuating IL6 and IL8 production. Notably, DIO counteracted TGF-β1-induced fatty acid uptake and de novo lipogenesis (DNL), as evidenced by BODIPY 500/510 C1, C12 and Nile red staining. This regulatory effect of DIO on metabolic reprogramming was attributed to the downregulation of CD36, SREBP1, FASN, SCD1, and CPT1A. Additionally, DIO reduced intracellular reactive oxygen species (ROS) accumulation in TGF-β1-activated HSCs. The anti-fibrotic and fatty acid metabolism-regulating effects of DIO were abrogated by the mTOR agonist MHY1485. DIO alleviates hepatic fibrosis by suppressing HSCs activation and modulating fatty acid metabolism through inhibiting the PI3K/Akt/mTOR pathway. Our results suggest that DIO is a promising natural candidate for the treatment of hepatic fibrosis.