The use of a sensitive assay for troponin I improves early diagnosis of acute myocardial infarction and risk stratification, regardless of the time of chest-pain onset.

Incubation of endothelial cells in vitro with high concentrations of glucose activates protein kinase C (PKC) and increases nitric oxide synthase (NOS III) gene expression as well as superoxide production. The underlying mechanisms remain unknown. To address this issue in an in vivo model, diabetes was induced with streptozotocin in rats. Streptozotocin treatment led to endothelial dysfunction and increased vascular superoxide production, as assessed by lucigenin-and coelenterazine-derived chemiluminescence. The bioavailability of vascular nitric oxide (as measured by electron spin resonance) was reduced in diabetic aortas, although expression of endothelial NOS III (mRNA and protein) was markedly increased. NOS inhibition with N G-nitro-L-arginine increased superoxide levels in control vessels but reduced them in diabetic vessels, identifying NOS as a superoxide source. Similarly, we found an activation of the NADPH oxidase and a 7-fold increase in gp91 phox mRNA in diabetic vessels. In vitro PKC inhibition with chelerythrine reduced vascular superoxide in diabetic vessels, whereas it had no effect on superoxide levels in normal vessels. In vivo PKC inhibition with N-benzoyl-staurosporine did not affect glucose levels in diabetic rats but prevented NOS III gene upregulation and NOS-mediated superoxide production, thereby restoring vascular nitric oxide bioavailability and endothelial function. The reduction of superoxide in vitro by chelerythrine and the normalization of NOS III gene expression and reduction of superoxide in vivo by N-benzoyl-staurosporine point to a decisive role of PKC in mediating these phenomena and suggest a therapeutic potential of PKC inhibitors in the prevention or treatment of vascular complications of diabetes mellitus. The full text of this article is available at http://www.circresaha.org. (Circ Res. 2001; 88:e14-e22.) Key Words: diabetes nitric oxide synthase protein kinase C uncoupling NADPH oxidase M yocardial infarction and stroke constitute major causes of death in patients with diabetes mellitus. Long-term diabetes is associated with macroangiopathy and microangi-opathy. Several hypotheses have been put forth to explain the adverse effects of hyperglycemia on the vasculature. These include the activation of the polyol pathway, nonenzymatic glycation, redox potential alterations, and stimulation of the diacylglycerol (DAG)-protein kinase C (PKC) pathway (for review, see Ishii et al 1). Although studies on the polyol pathway and nonenzymatic glycation remain inconclusive so far, more recent studies strongly point to a decisive role of the DAG-PKC pathway for the vascular complications associated with diabetes.

Background-Angiotensin II activates NAD(P)H-dependent oxidases via AT 1 -receptor stimulation, the most important vascular source of superoxide (O 2 ⅐ Ϫ ). The AT 1 receptor is upregulated in vitro by low-density lipoprotein. The present study was designed to test whether hypercholesterolemia is associated with increased NAD(P)H-dependent vascular O 2 ⅐ Ϫ production and whether AT 1 -receptor blockade may inhibit this oxidase and in parallel improve endothelial dysfunction. Methods and Results-Vascular responses were determined by isometric tension studies, and relative rates of vascular O 2 ⅐

Among patients with suspected acute coronary syndrome, hsTnI or cTnI determination 3 hours after admission may facilitate early rule-out of AMI. A serial change in hsTnI or cTnI levels from admission (using the 99th percentile diagnostic cutoff value) to 3 hours after admission may facilitate an early diagnosis of AMI.

Reactive oxygen species may alter endothelium-dependent vascular relaxation not only by the interaction of O(2)(*-) with NO(*) but also through interactions between peroxynitrite and tetrahydrobiopterin. Peroxynitrite oxidation of tetrahydrobiopterin may represent a pathogenic cause of "uncoupling" of NO synthase.

Abstract-Conditions associated with impaired nitric oxide (NO) activity and accelerated atherosclerosis have been shown to be associated with a reduced bioavailability of tetrahydrobiopterin (BH4). We therefore hypothesized that BH4 supplementation may improve endothelial dysfunction of chronic smokers. Forearm blood flow (FBF) responses to the endothelium-dependent vasodilators acetylcholine (ACh; 0.75, 1.5, and 3.0 g/100 mL tissue/min) or serotonin (5-HT; 0.7, 2.1, and 6.3 ng/100 mL tissue/min), to the inhibitor of endothelial nitric oxide synthase (NOS) N G -monomethyl-L-arginine (L-NMMA; 2, 4, and 8 mol/min), and to the endothelium-independent vasodilator sodium nitroprusside (SNP; 0.1, 0.3, and 1.0 g/100 mL tissue/min) were measured by venous occlusion plethysmography in controls and chronic smokers. Drugs were infused into the brachial artery, and FBF was measured before and during concomitant intra-arterial infusion of BH4, tetrahydroneopterin (NH4; another reduced pteridine), or the antioxidant vitamin C (6 and 18 mg/min). In control subjects, BH4 had no effect on FBF in response to ACh, 5-HT, and SNP. In contrast, in chronic smokers, the attenuated FBF responses to ACh and 5-HT were markedly improved by concomitant administration of BH4, whereas the vasodilator responses to SNP were not affected. L-NMMA-induced vasoconstriction was significantly reduced in smokers compared with controls, suggesting impaired basal NO bioactivity. BH4 improved L-NMMA responses in smokers while having no effect on L-NMMA responses in controls. Pretreatment with vitamin C abolished BH4 effects on ACh-dependent vasodilation. In vitro, NH4 scavenged superoxide created by the xanthine/xanthine oxidase reaction equipotent like BH4 but failed to modify ACh-induced changes in FBF in chronic smokers in vivo. These data support the concept that in addition to the free radical burden of cigarette smoke, a dysfunctional NOS III due to BH4 depletion may contribute at least in part to endothelial dysfunction in chronic smokers. The full text of this article is available at http://www.circresaha.org. (Circ Res. 2000;86:e36-e41.)

Endothelial dysfunction (ED) in the setting of cardiovascular risk factors such as hypercholesterolemia, hypertension, diabetes mellitus, chronic smoking as well as in patients with heart failure has been shown to be at least in part dependent on the production of reactive oxygen species (ROS) such as superoxide and the subsequent decrease in vascular bioavailability of nitric oxide (NO). Methods to quantify endothelial dysfunction include forearm plethysmography, flow-dependent dilation of the brachial artery, finger-pulse plethysmography, pulse curve analysis, and quantitative coronary angiography after intracoronary administration of the endothelium-dependent vasodilator acetylcholine. Superoxide sources include the NADPH oxidase, xanthine oxidase, and mitochondria. Superoxide produced by the NADPH oxidase may react with NO released by the endothelial nitric oxide synthase (eNOS) thereby generating peroxynitrite (ONOO-), leading to eNOS uncoupling and therefore eNOS-mediated superoxide production. The present review will discuss current concepts of how to assess endothelial function, prognostic implications of ED, mechanisms underlying ED with focus on oxidative stress and circulating biomarkers, which have been proposed to indicate endothelial dysfunction and/or damage, respectively.

Abstract-Studies with cGMP-dependent protein kinase I (cGK-I)-deficient human cells and mice demonstrated that cGK-I ablation completely disrupts the NO/cGMP pathway in vascular tissue, which indicates a key role of this protein kinase as a mediator of the NO/cGMP action. Analysis of the vasodilator-stimulated phosphoprotein phosphorylated at serine 239 (P-VASP) is a useful tool to monitor cGK-I activation in platelets and cultured endothelial and smooth muscle cells. Therefore, we investigated whether endothelial dysfunction and/or vascular NO bioavailability is reflected by decreased vessel wall P-VASP and whether improvement of endothelial dysfunction restores this P-VASP. Incubation of aortic tissue from New Zealand White Rabbits with the NOS inhibitor N G -nitro-L-arginine and endothelial removal strikingly reduced P-VASP. Oxidative stress induced by inhibition of CuZn superoxide dismutase increased superoxide and decreased P-VASP. Endothelial dysfunction in hyperlipidemic Watanabe rabbits (WHHL) was associated with increased vascular superoxide and with decreased P-VASP. Treatment of WHHL with AT 1 receptor blockade improved endothelial dysfunction, reduced vascular superoxide, increased vascular NO bioavailability, and increased P-VASP. Therefore, the level of vessel P-VASP closely follows changes in endothelial function and vascular oxidative stress. P-VASP is suggested to represent a novel biochemical marker for monitoring the NO-stimulated sGC/cGK-I pathway and endothelial integrity in vascular tissue. Key Words: cGMP-dependent kinase Ⅲ VASP Ⅲ nitric oxide Ⅲ hyperlipidemia Ⅲ AT 1 receptor blockade T he endothelium-derived relaxing factor, identified as NO or a closely related compound, induces vasorelaxation by activating the target enzyme soluble guanylyl cyclase (sGC) and by increasing tissue levels of the second messenger cGMP. 1 Cyclic GMP in turn activates cGMP-dependent protein kinase types I and II (cGK-I and -II) of which cGK-I is highly expressed in vascular smooth muscle cell levels. 2-4 cGK-I has mediated NO/cGMP-caused vasorelaxation, which involves phosphorylation of proteins that affect myosin light chains and intracellular Ca 2ϩ levels. 2-5 Studies with cGK-Ideficient mice demonstrated a complete disruption of the NO/cGMP signaling pathway in the vascular smooth muscle. 6 Therefore, the activity and/or expression of cGK-I critically modulate NO-induced vasorelaxation. Recently, Smolenski and colleagues 7,8 provided evidence that analysis of the phosphorylation of vasodilator-stimulated phosphoprotein (VASP) at serine 239 (P-VASP) is a useful biomonitor of cGK activity and therefore NO effects in intact cells such as platelets and cultured endothelial and smooth muscle cells. Whether this is also applicable to intact vascular tissue must still be established.Depressed vasodilation to endothelium-dependent vasodilators such as acetylcholine and to authentic NO is a hallmark of early stages of atherosclerosis. 9 The mechanisms underlying endothelial dysfunction are likely to be multifactori...