Increased superoxide production in hypertensive patients with diabetes mellitus - Role of nitric oxide synthase

Background: Hypertension and diabetes are important independent risk factors for increased oxidative stress and increased cardiovascular risk. The combination of hypertension and diabetes results in a dramatic increase in cardiovascular risk. Enhanced oxidative stress in hypertension and diabetes is linked to decreased nitric oxide (NO) bioavailability because of its interaction with vascular superoxide (O-2(center dot-)), derived predominantly from NAD(P)H-dependent oxidases. When uncoupled from essential cofactors, NO synthase III (NOS III) can also produce O-2(center dot-). We studied platelet superoxide production in patients with hypertension alone and in patients with coexistent diabetes mellitus, investigating the contribution of NOS III uncoupling to platelet superoxide production. Methods and Results: Gel-filtered platelets were obtained and were stimulated with Phorbol 12-myristate 13-acetate, and O-2(center dot-) production was detected using lucigenin-enhanced chemiluminescence. Superoxide production was significantly higher in patients with diabetes and hypertension (6.4 +/- 1.6 pmol/min/10(8) platelets) than in patients with hypertension (1.6 +/- 0.6 pmol/min/10(8) platelets) (P <.04). After incorporation of N-omega-nitro-L-arginine methyl ester (L-NAME, 1 mmol/L), O-2(center dot-) detection increased in 40% of patients with diabetes and hypertension and in 87% of patients with hypertension. This expected response results from L-NAME inhibition of NO production preventing NO scavenging of O-2(center dot-). A reduction in O-2(center dot-). production in response to L-NAME occurred in the remaining patients and indicates O-2(center dot-) production by the uncoupled NOS III enzyme. Conclusions: This study provides first published evidence that NOS III can reside in the uncoupled state in patients with hypertension and, to a greater extent, in patients with coexisting hypertension and diabetes, and that it contributes significantly to increased superoxide production in these disease states.