Mechanism of action of Danlou tablets affecting MAFLD via KEAP1-mediated oxeiptosis

Metabolic fatty liver disease (MAFLD) is a significant risk factor for atherosclerotic cardiovascular disease. Several preliminary studies on MAFLD animal models have indicated the therapeutic potential of Danlou tablets (DLT), a primary Chinese medicine used for managing coronary artery disease. However, the underlying mechanism of DLT in the treatment of MAFLD remains elusive.

Clarify the potential effective components of DLT in the treatment of MAFLD, and preliminarily verify the molecular mechanism against MAFLD in vivo and in vitro experiments.

The composition of DLT and their content in DLT-treated rat serum were analyzed using UPLC/ESI- Q TRAP-MS/MS. Mice were given a high-fat diet to establish the MAFLD model. Then, the MAFLD mice were treated with DLT. Liver sections were taken for histopathological assessment. Furthermore, in vivo and in vitro alterations in the oxeiptosis pathway, de novo fatty acid synthesis, and Triglyceride catabolism were verified by qRT-PCR, Western Blot, and Immunofluorescence experiments. Moreover, how DLT modulated the oxeiptosis pathway was further investigated by rescue experimental strategies.

We isolated and detected a total of 1003 compounds from DLT, 109 of which were found in rat plasma, and hypothesized that 11 active ingredients represented by Tanshinone IIA might play a major role in anti-MAFLD. Furthermore, we found that DLT increased Triglyceride catabolism and suppressed de novo fatty acid synthesis in vivo and in vitro, thereby significantly attenuating hepatic lipid deposition. Mechanistically, DLT restored the phosphorylation of Protein Kinase B, promoted Triglyceride catabolism and inhibited the de novo fatty acid synthesis through the oxeiptosis pathway (KEAP1/PGAM5/AIFM1).

Our findings suggest that DLT promotes Triglyceride catabolism and inhibit de novo fatty acid synthesis by affecting the activation of the oxeiptosis pathway, suggesting a potential therapeutic strategy for ameliorating NAFLD.