Dietary Methionine Restriction Alleviates Choline-Induced Tri-Methylamine-N-Oxide (TMAO) Elevation by Manipulating Gut Microbiota in Mice

Dietary methionine restriction (MR) has been shown to decrease plasma trimethylamine-N-oxide (TMAO) levels in high-fat diet mice; however, the specific mechanism used is unknown. We speculated that the underlying mechanism is related with the gut microbiota, and this study aimed to confirm the hypothesis. In this study, we initially carried out an in vitro fermentation experiment and found that MR could reduce the ability of gut microbiota found in the contents of healthy mice and the feces of healthy humans to produce trimethylamine (TMA). Subsequently, mice were fed a normal diet (CON, 0.20% choline + 0.86% methionine), high-choline diet (H-CHO, 1.20% choline + 0.86% methionine), or high-choline + methionine-restricted diet (H-CHO+MR, 1.20% choline + 0.17% methionine) for 3 months. Our results revealed that MR decreased plasma TMA and TMAO levels in H-CHO-diet-fed mice without changing hepatic FMO3 gene expression and enzyme activity, significantly decreased TMA levels and expression of choline TMA-lyase (CutC) and its activator CutD, and decreased CutC activity in the intestine. Moreover, MR significantly decreased the abundance of TMA-producing bacteria, including Escherichia-Shigella (Proteobacteria phylum) and Anaerococcus (Firmicutes phylum), and significantly increased the abundance of short-chain fatty acid (SCFA)-producing bacteria and SCFA levels. Furthermore, both MR and sodium butyrate supplementation significantly inhibited bacterial growth, down-regulated CutC gene expression levels in TMA-producing bacteria, including Escherichia fergusonii ATCC 35469 and Anaerococcus hydrogenalis DSM 7454 and decreased TMA production from bacterial growth under in vitro anaerobic fermentation conditions. In conclusion, dietary MR alleviates choline-induced TMAO elevation by manipulating gut microbiota in mice and may be a promising approach to reducing circulating TMAO levels and TMAO-induced atherosclerosis.