Endothelial NLRP3 Inflammasome Activation and Neointima Formation in Mice by Gut Microbial Metabolite Trimethylamine‐N‐Oxide

Recent studies have demonstrated that trimethylamine‐N‐oxide (TMAO), an intestinal microbial metabolite induces atherosclerosis and enhances the risk for cardiovascular disorders. However, the precise mechanisms of action of TMAO in cardiovascular pathophysiology are yet to be elucidated. In the current study, we examined whether TMAO induces inflammasome activation and thereby contributing to endothelial dysfunction and vascular injury. Confocal microscopic analysis demonstrated the formation of Nlrp3 inflammasomes in the intima of the carotid arteries of Nlrp3+/+ mice but not in Nlrp3−/− mice. Correspondingly, TMAO‐induced increases in IL‐1β production in the carotid intima were abolished in Nlrp3−/− mice. The endothelial tight junction protein, ZO‐1 was reduced by TMAO in Nlrp3+/+ mice, but not in Nlrp3−/− mice. It was also found that TMAO significantly increased neointimal formation in partially ligated carotid arteries of Nlrp3+/+ mice, but not in Nlrp3−/− mice. In in vitro studies, we also found that TMAO treatment significantly increased the colocalization of NLRP3 with Asc or NLRP3 with caspase‐1 in endothelial cells compared to control cells. Pretreatment with Nlrp3 siRNA transfection 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 but not in Nlrp3 siRNA transfected cells. Furthermore, western blot analysis showed that TMAO treatment significantly decreased tight junction protein ZO‐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). Prior treatment with Nlrp3 siRNA attenuated the TMAO‐induced cell permeability. Together, these results suggest that TMAO‐induced formation and activation of NLRP3 inflammasomes in vascular endothelium thereby leading to endothelial dysfunction and atherosclerosis in mice. Support or Funding Information DK104031