Role of lipid droplet-mediated lipid peroxidation by GPX4 in docosahexaenoic acid-induced EA.hy926 endothelial cell ferroptosis.

Endothelial cells (EC) form the inner lining of the blood vessels and are essential for the vascular homeostasis. EC death has been implicated in the pathology of vascular diseases. Our previous studies indicated that docosahexaenoic acid (DHA) induces EA.hy926 EC death by upregulating lipid droplet (LD) biogenesis and activating p38 MAPK, however, how LDs contribute to this process remains unclear. DHA belongs to the polyunsaturated fatty acids that are susceptible to lipid peroxidation, which is a common mechanism in lipid-induced cell injury and ferroptosis. Therefore, the current study is to investigate the mechanism of LD in the regulation of lipid peroxidation in relation to p38 MAPK in DHA-induced EA.hy926 endothelial ferroptosis. Results showed that DHA induced a time-dependent, bidirectional modulation of GPX4 expression accompanied by a progressive GSH and NADPH depletion and increased lipid peroxidation in ECs. Suppression of LD formation through diacylglycerol acyltransferases inhibition, siRNA, or promotion of LD lipolysis reduced lipid peroxidation and restored the GPX4 expression. Inhibiting lipid peroxidation chemically, activating GPX4 or supplementing GSH all prevented the cell death by DHA. Our findings reveal a novel role of LD in regulating GPX4-mediated lipid peroxidation and highlight the key role of p38 MAPK in downregulating GPX4 in lipid peroxidation-induced EC ferroptosis by DHA. In conclusion, the present study suggested that DHA-induced LD formation in ECs increases the membrane area, which could facilitate the lipid peroxidation reaction, disrupt the endogenous GPX4 antioxidant system, and thereby exacerbate the detrimental effects of lipid peroxidation, ultimately leading to cell ferroptosis.