Overexpression of cytochrome P450CYP2J2 protects against hypoxia-reoxygenation injury in cultured bovine aortic endothelial cells

CYP2J2 is abundant in human heart and its arachidonic acid metabolites, the epoxyeicosatrienoic acids (EETs), have potent vasodilatory, antiinflammato ry and cardioprotective properties. This study was designed to examine the role of CYP2J2 in hypoxia-reoxygenation-induced injury in cultured bovine a ortic endothelial cells (BAECs). Early passage BAECs were exposed to 24-h h ypoxia followed by 4-h reoxygenation (HR). HR resulted in cell injury, as i ndicated by significant increases in lactate dehydrogenase (LDH) release an d trypan blue stained cells (p < 0.01) and was associated with a decrease i n CYP2J2 protein expression. Transfection of BAECs with the CYP2J2 cDNA res ulted in increased CYP2J2 expression and arachidonic acid epoxygenase activ ity, compared with cells transfected with an irrelevant green fluorescent p rotein (GFP) cDNA. HR induced significant injury in GFP-transfected BAECs, as indicated by increases in LDH release and trypan blue-stained cells (p < 0.01); however, the HR-induced injury was markedly attenuated in CYP2J2-tr ansfected cells (p < 0.01). HR increased cellular 8-iso-prostaglandin F-2<a lpha> (P < 0.05), and decreased eNOS expression, L-arginine uptake and conv ersion, and nitrite production (p < 0.01) in GFP-transfected BAECs. CYP2J2 transfection attenuated the HR-induced increase in 8-iso-prostaglandin F-2 alpha (P < 0.05) and decreased the amount of extracellular superoxide detec ted by cytochrome c reduction under normoxic conditions (p < 0.05) but did not significantly affect HR-induced decreases in eNOS expression, L-arginin e uptake and conversion, and nitrite production. Treatment of BAECs with sy nthetic EETs and/or epoxide hydrolase inhibitors also showed protective eff ects against FIR injury (p < 0.05). These observations suggest: (1) HR resu lts in endothelial injury and decreased CYP2J2 expression; (2) transfection with the CYP2J2 cDNA protects against HR injury; and (3) the cytoprotectiv e effects of CYP2J2 may be mediated, at least in part, by antioxidant effec ts.