3,3'-diindolylmethane ameliorates non-alcoholic fatty liver disease by inhibiting the FMO3-TMAO axis in mice

Non-alcoholic fatty liver disease (NAFLD) is the leading chronic liver disease globally, characterized by steatosis, inflammation, and hepatocyte injury. While the Flavin-containing monooxygenase 3 (FMO3)-trimethylamine N-oxide (TMAO) axis is a known driver of atherosclerosis, its role in NAFLD progression remains unclear. Here, we report that FMO3 is significantly upregulated in the livers of choline-deficient, high-fat diet (CDA-HFD)-induced NAFLD mice, as revealed by transcriptomic profiling and validation. Mechanistically, FMO3 overexpression exacerbated lipid accumulation and inflammatory cytokine release in vitro, while its metabolite TMAO directly aggravated hepatic steatosis and inflammation in vivo. Notably, the natural compound 3,3'-diindolylmethane (DIM) significantly attenuated NAFLD phenotypes, including serum ALT/AST levels and hepatic lipid content; however, these protective effects were reversed by exogenous TMAO supplementation. These findings identify the FMO3-TMAO axis as a critical target for regulating lipid homeostasis and inflammation, suggesting DIM as a promising therapeutic candidate for NAFLD intervention.