Abstract-The bioavailability of nitric oxide is decreased in animal models and humans with diabetes mellitus.Hyperglycemia, in particular, attenuates endothelium-dependent vasodilation in healthy subjects. In vitro and in vivo animal studies implicate activation of protein kinase C␤ as an important mechanism whereby hyperglycemia decreases endothelium-derived nitric oxide. Accordingly, this study tested the hypothesis that inhibition of protein kinase C␤ would prevent impairment of endothelium-dependent vasodilation in healthy humans exposed to hyperglycemia. This study was a randomized, double-blind, placebo-controlled, crossover trial. Healthy subjects were treated with an orally active, selective, protein kinase C␤ inhibitor, LY333531, or matching placebo once a day for 7 days before vascular function testing. Forearm blood flow was measured using venous-occlusion, strain-gauge plethysmography. Endothelium-dependent vasodilation was measured via incremental brachial artery administration of methacholine chloride (0.3 to 10 g/min) during euglycemia and after 6 hours of hyperglycemic clamp. The forearm blood flow dose-response curve to methacholine was significantly attenuated by hyperglycemia after placebo treatment (Pϭ0.009 by ANOVA, euglycemia versus hyperglycemia) but not after treatment with LY333531. Inhibition of protein kinase C␤ prevents the reduction in endothelium-dependent vasodilation induced by acute hyperglycemia in healthy humans in vivo. These findings suggest that hyperglycemia impairs endothelial function, in part, via protein kinase C␤ activation. (Circ Res. 2002;90:107-111.)Key Words: protein kinase C Ⅲ nitric oxide Ⅲ hyperglycemia Ⅲ endothelium Ⅲ diabetes V ascular disease is the principal cause of morbidity and mortality in patients with diabetes mellitus. 1 Diabetes mellitus is associated with changes in endothelial cell function that augur the development of atherosclerosis. An important early change, decreased bioavailability of endothelium-derived nitric oxide, is linked to many of the pathological features of atherosclerosis including upregulation of leukocyte adhesion molecules, platelet activation, and an increased propensity for vasoconstriction. 2,3 Previous studies have demonstrated decreased endothelium-dependent vasodilation, a physiological marker of decreased bioavailability of nitric oxide in both conduit arteries and resistance vessels in experimental models of diabetes and humans with type 1 and type 2 diabetes. 4 -6 The central importance of hyperglycemia to the development of cardiovascular disease in diabetes mellitus is becoming increasingly evident. Population studies have revealed that an incremental risk of cardiovascular disease is associated with higher levels of blood glucose, beginning in the upper normal range. 7 Hyperglycemia, per se, impairs vasodilator function in animals and healthy humans, similar to that which occurs in patients with diabetes. 8,9 This cannot be attributed to downregulation of endothelial nitric oxide synthase (eNOS), since glucose inc...

Oxidative stress decreases the bioavailability of endothelium-derived nitric oxide in diabetic patients. We investigated whether impaired endothelium-dependent vasodilation (EDV) in diabetes can be improved by long-term administration of oral antioxidants. Forty-nine diabetic subjects [26 Type 1 (T1) and 23 Type 2 (T2)] and 45 matched healthy control subjects were randomized to receive oral vitamin C (1,000 mg) and vitamin E (800 IU) daily or matching placebo for 6 mo. Vascular ultrasonography was used to determine brachial artery EDV and endothelium-independent vasodilation (EIV). EDV was decreased in both T1 (4.9 +/- 0.9%, P = 0.015) and T2 (4.1 +/- 1.0%, P < 0.01) subjects compared with control subjects (7.7 +/- 0.7%). EIV was decreased in T2 (15.0 +/- 1.2%, P < 0.01) but not T1 subjects (18.5 +/- 2.3%, P = 0.3) compared with controls (21.8 +/- 1.8%). Administration of antioxidant vitamins increased EDV in T1 (by 3.4 +/- 1.4%, P = 0.023) but not T2 subjects (by 0.5. +/- 0.4%, P = 0.3). Antioxidant therapy had no significant affect on EIV. Oral antioxidant therapy improves EDV in T1 but not T2 diabetes. These results are consistent with the lack of clinical benefit in studies that have included primarily T2 diabetic patients.

Abstract-Cigarette smoking impairs endothelial function. Hydroxymethylglutaryl (HMG) CoA reductase inhibitors (statins) may favorably affect endothelial function via nonlipid mechanisms. We tested the hypothesis that statins would improve endothelial function independent of changes in lipids in cigarette smokers. Twenty normocholesterolemic cigarette smokers and 20 matched healthy control subjects were randomized to atorvastatin 40 mg daily or placebo for 4 weeks, washed out for 4 weeks, and then crossed-over to the other treatment. Baseline low-density lipoprotein (LDL) levels were similar in smokers and healthy subjects, 103Ϯ22 versus 95Ϯ27 mg/dL, respectively (PϭNS) and were reduced similarly in smokers and control subjects by atorvastatin, to 55Ϯ30 and 58Ϯ20 mg/dL, respectively (PϭNS). Vascular ultrasonography was used to determine brachial artery, flow-mediated, endothelium-dependent, and nitroglycerin-mediated, endothelium-independent vasodilation. To elucidate potential molecular mechanisms that may account for changes in endothelial function, skin biopsy specimens were assayed for eNOS mRNA, eNOS activity, and nitrotyrosine. Endothelium-dependent vasodilation was less in smokers than nonsmoking control subjects during placebo treatment, 8.0Ϯ0.6% versus 12.1Ϯ1.1%, (Pϭ0.003). Atorvastatin increased endothelium-dependent vasodilation in smokers to 10.5Ϯ1.3% (Pϭ0.017 versus placebo) but did not change endothelium-dependent vasodilation in control subjects (to 11.0Ϯ0.8%, PϭNS). Endothelium-independent vasodilation did not differ between groups during placebo treatment and was not significantly affected by atorvastatin. Multivariate analysis did not demonstrate any association between baseline lipid levels or the change in lipid levels and endothelium-dependent vasodilation. Cutaneous nitrotyrosine levels and skin microvessel eNOS mRNA, but not ENOS activity, were increased in smokers compared with controls but unaffected by atorvastatin treatment. Atorvastatin restores endothelium-dependent vasodilation in normocholesterolemic cigarette smokers independent of changes in lipids. These results are consistent with a lipid-independent vascular benefit of statins but could not be explained by changes in eNOS message and tissue oxidative stress. These findings implicate a potential role for statin therapy to restore endothelial function and thereby investigate vascular disease in cigarette smokers. Key Words: cigarette smoking Ⅲ endothelial nitric oxide synthase Ⅲ hydroxymethylglutaryl CoA reductase inhibitor Ⅲ endothelial function Ⅲ oxidative stress C igarette smoking is a reversible risk factor for atherosclerosis. Smoking impairs endothelial function and creates a vascular environment that contributes to the development of atherosclerosis. 1,2 Putative mechanisms through which cigarette smoking alters endothelial function and particularly the bioavailability of nitric oxide (NO) in humans include increased oxidative stress, 3 alterations in endothelial nitric oxide synthase (eNOS), 4 and activation of rho kinase...