Zeaxanthin inhibits endothelial pyroptosis and alleviates atherosclerotic plaque formation by suppressing Ang2-mediated activation of TLR4/NLRP3 signaling pathway.

Clinical studies showed that zeaxanthin reduces oxidized lipid products and inflammation, and alleviates the formation of atherosclerotic plaques. However, the molecular mechanism by which zeaxanthin alleviates atherosclerosis (AS) progression remains unclear. This study aimed to explore the molecular mechanism of zeaxanthin alleviating AS progression. Human umbilical vein endothelial cells (HUVECs) and human arterial endothelial cells (HAECs) were treated with 100 μg/mL oxidized low density lipoprotein (ox-LDL) to construct AS cell model in vitro, then treated with 50 or 150 μM zeaxanthin for 24 h. The results showed that zeaxanthin inhibited pyroptosis of ox-LDL-treated HUVECs and HAECs in a dose-dependent manner. Moreover, zeaxanthin reduced Angiopoietin-2 (Ang2) protein levels. Silencing Ang2 inhibited pyroptosis of ox-LDL-treated HUVECs and HAECs, while Ang2 overexpression counteracted the effect of zeaxanthin on ox-LDL-treated HUVECs and HAECs. Mechanism studies revealed that Ang2 inhibits Tie2 phosphorylation to activate the TLR4/NLRP3 pathway, thereby promoting pyroptosis of ox-LDL-treated HUVECs and HAECs. Both disulfiram, an inhibitor of pyroptosis, and TAK-242, an inhibitor of TLR4, counteracted the promoting effect of Ang2 overexpression on endothelial pyroptosis. Finally, ApoE-/- mice were fed with high-fat diet for 8 weeks to establish an in vivo mouse AS model, then administered with zeaxanthin (10 mg/kg) and injected with endothelial cell (EC) specific Ang2 overexpression vectors (AAV9-Ang2). The results showed that zeaxanthin inhibited endothelial pyroptosis and atherosclerotic plaque formation in AS mice, while Ang2 overexpression accelerated AS progression in mice. In conclusion, zeaxanthin might inhibit endothelial pyroptosis by blocking Ang2-mediated activation of TLR4/NLRP3 pathway, thereby inhibiting AS progression.