Attenuation of Angiotensin Ⅱ Signaling Recouples eNOS and Inhibits Nonendothelial NOX Activity in Diabetic Mice

摘要

Angiotensin Ⅱ (Ang Ⅱ) levels are increased in patients with diabetes, but mechanisms underlying its contribution to diabetic vascular diseases are incompletely understood. We recently reported that in aortic endothelial cells, Ang Ⅱ induces endothelial nitric oxide synthase (eNOS) uncoupling to produce superoxide (O_(2~·)~-) rather than nitric oxide (NO·), upon loss of the tetrahydrobiopterin (H_4B) salvage enzyme dihydrofolate reductase (DHFR). Here, we found that streptozotocin-induced diabetic mice had a marked increase in aortic O_(2~·)~- production, which was inhibited by N-nitro-L-arginine methyl ester hydrochloride, indicating uncoupling of eNOS. Ang Ⅱ receptor type 1 blocker cande-sartan or ACE inhibitor captopril markedly attenuated eNOS-derived O_(2~·) ~-and hydrogen peroxide production while augmenting NO· bioavailability in diabetic aortas, implicating recoupling of eNOS. O_(2~·)~- and NO· production were characteristically and quantitatively measured by electron spin resonance. DHFR expression was decreased in diabetic aortas but significantly restored by candesartan or captopril. Either also improved vascular H_4B content and endothelium-dependent vasorelaxation in diabetes. Racl-dependent NAD(P)H oxidase (NOX) activity was more than doubled in the endothelium-denuded diabetic aortas but was attenuated by candesartan or captopril, indicating that NOX remains active in nonendothelial vascular tissues, although uncoupled eNOS is responsible for endothelial production of O_(2~·)~-. These data demonstrate a novel role of Ang Ⅱ in diabetic uncoupling of eNOS and that Ang Ⅱ-targeted therapy improves endothelial function via the novel mechanism of recoupling eNOS. Dual effectiveness on uncoupled eNOS and NOX may explain the high efficacy of Ang Ⅱ antagonists in restoring endothelial function.