K_Ca3.1 upregulation preserves endothelium-dependent vasorelaxation during aging and oxidative stress

Endothelial oxidative stress develops with aging and reactive oxygen species impair endothelium-dependent relaxation (EDR) by decreasing nitric oxide (NO) availability. Endothelial K_Ca3.1, which contributes to EDR, is upregulated by H₂O₂. We investigated whether K_Ca3.1 upregulation compensates for diminished EDR to NO during aging-related oxidative stress. Previous studies identified that the levels of ceramide synthase 5 (CerS5), sphingosine, and sphingosine 1-phosphate were increased in aged wild-type and CerS2 mice. In primary mouse aortic endothelial cells (MAECs) from aged wild-type and CerS2 null mice, superoxide dismutase (SOD) was upregulated, and catalase and glutathione peroxidase 1 (GPX1) were downregulated, when compared to MAECs from young and age-matched wild-type mice. Increased H₂O₂ levels induced Fyn and extracellular signal-regulated kinases (ERKs) phosphorylation and K_Ca3.1 upregulation. Catalase/GPX1 double knockout (catalase^−/−/GPX1^−/−) upregulated K_Ca3.1 in MAECs. NO production was decreased in aged wild-type, CerS2 null, and catalase^−/−/GPX1^−/− MAECs. However, K_Ca3.1 activation-induced, N^G-nitro-l-arginine-, and indomethacin-resistant EDR was increased without a change in acetylcholine-induced EDR in aortic rings from aged wild-type, CerS2 null, and catalase^−/−/GPX1^−/− mice. CerS5 transfection or exogenous application of sphingosine or sphingosine 1-phosphate induced similar changes in levels of the antioxidant enzymes and upregulated K_Ca3.1. Our findings suggest that, during aging-related oxidative stress, SOD upregulation and downregulation of catalase and GPX1, which occur upon altering the sphingolipid composition or acyl chain length, generate H₂O₂ and thereby upregulate K_Ca3.1 expression and function via a H₂O₂/Fyn-mediated pathway. Altogether, enhanced K_Ca3.1 activity may compensate for decreased NO signaling during vascular aging.