Trimethylamine‐N‐Oxide, an Intestinal Microbial Metabolite Instigates NLRP3 Inflammasome Activation and Endothelial Dysfunction

Plasma trimethylamine‐N‐oxide (TMAO), a product of intestinal microbial metabolism of dietary phosphatidylcholine has been recently associated with atherosclerosis and increased risk of cardiovascular diseases (CVD) in rodents and humans. However, the molecular mechanisms of how TMAO induces atherosclerosis and CVD progression are still unclear. The present study tested whether TMAO induces NLRP3 inflammasome formation and activation and thereby contributes to endothelial injury initiating atherogenesis. Confocal microscopic analysis showed that TMAO treatment significantly increased the colocalization of NLRP3 with Asc or NLRP3 with caspase‐1 in endothelial cells (EOMA) compared to control group. Pretreatment with caspase‐1 inhibitor, YVAD abolished the TMAO‐induced inflammasome formation in these cells. Correspondingly, TMAO treatment significantly increased the caspase‐1 activity (2.2 folds) and IL‐1β production (1.8 folds) compared to control cells. The YVAD treatment significantly attenuated the TMAO‐induced caspase‐1 activity and IL‐1β production. Further, Western blot analysis showed that TMAO treatment significantly increased cleaved caspase‐1 level compared to control cells. In addition, TMAO treatment significantly increased the cell permeability of EOMA cell layers compared to control cells (Control: 1 ± 0.1 vs TMAO: 2 ± 0.07). However, pretreatment with YVAD attenuated TMAO‐induced cell permeability. In animal experiments, direct infusion of TMAO by osmotic pump implanted subcutaneously in mice with partially ligated left carotid artery were found to have significantly increased neo‐intimal formation, which correlated with increased IL‐1β production in the intima. In conclusion, the formation and activation of NLRP3 inflammasomes by TMAO may be an important initiating mechanism to turn on the endothelial inflammatory response leading to endothelial dysfunction and atherogenesis in mice. Support or Funding Information supported by NIH grants DK104031 and HL‐75316.