Therapies that raise levels of HDL, which is thought to exert atheroprotective effects via effects on endothelium, are being examined for the treatment or prevention of coronary artery disease (CAD). However, the endothelial effects of HDL are highly heterogeneous, and the impact of HDL of patients with CAD on the activation of endothelial eNOS and eNOS-dependent pathways is unknown. Here we have demonstrated that, in contrast to HDL from healthy subjects, HDL from patients with stable CAD or an acute coronary syndrome (HDL CAD ) does not have endothelial antiinflammatory effects and does not stimulate endothelial repair because it fails to induce endothelial NO production. Mechanistically, this was because HDL CAD activated endothelial lectin-like oxidized LDL receptor 1 (LOX-1), triggering endothelial PKCβII activation, which in turn inhibited eNOS-activating pathways and eNOS-dependent NO production. We then identified reduced HDL-associated paraoxonase 1 (PON1) activity as one molecular mechanism leading to the generation of HDL with endothelial PKCβII-activating properties, at least in part due to increased formation of malondialdehyde in HDL. Taken together, our data indicate that in patients with CAD, HDL gains endothelial LOX-1-and thereby PKCβII-activating properties due to reduced HDL-associated PON1 activity, and that this leads to inhibition of eNOS-activation and the subsequent loss of the endothelial antiinflammatory and endothelial repair-stimulating effects of HDL.

Background-High-density lipoprotein (HDL)-raising therapies are currently under intense evaluation, but the effects of HDL may be highly heterogeneous. We therefore compared the endothelial effects of HDL from healthy subjects and from patients with type 2 diabetes mellitus and low HDL (meeting the criteria for metabolic syndrome), who are frequently considered for HDL-raising therapies. Moreover, in diabetic patients, we examined the impact of extended-release (ER) niacin therapy on the endothelial effects of HDL. Methods and Results-HDL was isolated from healthy subjects (nϭ10) and patients with type 2 diabetes (nϭ33) by sequential ultracentrifugation. Effects of HDL on endothelial nitric oxide and superoxide production were characterized by electron spin resonance spectroscopy analysis. Effects of HDL on endothelium-dependent vasodilation and early endothelial progenitor cell-mediated endothelial repair were examined. Patients with diabetes were randomized to a 3-month therapy with ER niacin (1500 mg/d) or placebo, and endothelial effects of HDL were characterized. HDL from healthy subjects stimulated endothelial nitric oxide production, reduced endothelial oxidant stress, and improved endothelium-dependent vasodilation and early endothelial progenitor cell-mediated endothelial repair. In contrast, these beneficial endothelial effects of HDL were not observed in HDL from diabetic patients, which suggests markedly impaired endothelial-protective properties of HDL. ER niacin therapy improved the capacity of HDL to stimulate endothelial nitric oxide, to reduce superoxide production, and to promote endothelial progenitor cell-mediated endothelial repair. Further measurements suggested increased lipid oxidation of HDL in diabetic patients, and a reduction after ER niacin therapy. Conclusions-HDL from patients with type 2 diabetes mellitus and metabolic syndrome has substantially impaired endothelial-protective effects compared with HDL from healthy subjects. ER niacin therapy not only increases HDL plasma levels but markedly improves endothelial-protective functions of HDL in these patients, which is potentially more important.

Accumulating evidence suggests a critical role of increased reactive oxygen species production for left ventricular (LV) remodeling and dysfunction after myocardial infarction (MI). An increased myocardial activity of the NAD(P)H oxidase, a major oxidant enzyme system, has been observed in human heart failure; however, the role of the NAD(P)H oxidase for LV remodeling and dysfunction after MI remains to be determined. MI was induced in wild-type (WT) mice (n=46) and mice lacking the cytosolic NAD(P)H oxidase component p47(phox) (p47(phox)-/- mice) (n=32). Infarct size was similar among the groups. NAD(P)H oxidase activity was markedly increased in remote LV myocardium of WT mice after MI as compared with sham-operated mice (83+/-8 versus 16.7+/-3.5 nmol of O(2)(-) x microg(-1) x min(-1); P<0.01) but not in p47(phox)-/- mice after MI (13.5+/-3.6 versus 15.5+/-3.5 nmol of O(2)(-) x microg(-1) x min(-1)), as assessed by electron-spin resonance spectroscopy using the spin probe CP-H. Furthermore, increased myocardial xanthine oxidase activity was observed in WT, but not in p47(phox)-/- mice after MI, suggesting NAD(P)H oxidase-dependent xanthine oxidase activation. Myocardial reactive oxygen species production was increased in WT mice, but not in p47(phox)-/- mice, after MI. LV cavity dilatation and dysfunction 4 weeks after MI were markedly attenuated in p47(phox)-/- mice as compared with WT mice, as assessed by echocardiography (LV end-diastolic diameter: 4.5+/-0.2 versus 6.3+/-0.3 mm, P<0.01; LV ejection fraction, 35.8+/-2.5 versus 22.6+/-4.4%, P<0.05). Furthermore, cardiomyocyte hypertrophy, apoptosis, and interstitial fibrosis were substantially reduced in p47(phox)-/- mice as compared with WT mice. Importantly, the survival rate was markedly higher in p47(phox)-/- mice as compared with WT mice after MI (72% versus 48%; P<0.05). These results suggest a pivotal role of NAD(P)H oxidase activation and its subunit p47(phox) for LV remodeling/dysfunction and survival after MI. The NAD(P)H oxidase system represents therefore a potential novel therapeutic target to prevent cardiac failure after MI.

Abstract-Prehypertension is a highly frequent condition associated with an increased cardiovascular risk. Endothelial dysfunction is thought to promote the development of hypertension and vascular disease; however, underlying mechanisms remain to be further determined. The present study characterizes for the first time the in vivo endothelial repair capacity of early endothelial progenitor cells (EPCs) in patients with prehypertension/hypertension and examines its relation with endothelial function. Early EPCs were isolated from healthy subjects and newly diagnosed prehypertensive and hypertensive patients (nϭ52). In vivo endothelial repair capacity of EPCs was examined by transplantation into a nude mouse carotid injury model. EPC senescence was determined (RT-PCR of telomere length). NO and superoxide production of EPCs were measured using electron spin resonance spectroscopy analysis. CD34 ϩ /KDR ϩ mononuclear cells and circulating endothelial microparticles were examined by fluorescence-activated cell sorter analysis. Endothelium-dependent and -independent vasodilations were determined by high-resolution ultrasound. In vivo endothelial repair capacity of EPCs was substantially impaired in prehypertensive/hypertensive patients as compared with healthy subjects (re-endothelialized area: 15Ϯ3%/13Ϯ2% versus 28Ϯ3%; PϽ0.05 versus healthy subjects). Senescence of EPCs in prehypertension/hypertension was substantially increased, and NO production was markedly reduced. Moreover, reduced endothelial repair capacity of early EPCs was significantly related to an accelerated senescence of early EPCs and impaired endothelial function. The present study demonstrates for the first time that in vivo endothelial repair capacity of early EPCs is reduced in patients with prehypertension and hypertension, is related to EPC senescence and impaired endothelial function, and likely represents an early event in the development of hypertension. (Hypertension. 2010;55:1389-1397.)

Background-MicroRNAs are key regulators of angiogenic processes. Administration of angiogenic early outgrowth cells (EOCs) or CD34 ϩ cells has been suggested to improve cardiac function after ischemic injury, in particular by promoting neovascularization. The present study therefore examines regulation of angiomiRs, microRNAs involved in angiogenesis, in angiogenic EOCs and circulating CD34 ϩ cells from patients with chronic heart failure (CHF) and the role for their cardiac repair capacity. Methods and Results-Angiogenic EOCs and CD34ϩ cells were isolated from patients with CHF caused by ischemic cardiomyopathy (nϭ45) and healthy subjects (nϭ35). In flow cytometry analyses, angiogenic EOCs were largely myeloid and positive for alternatively activated M2 macrophage markers. In vivo cardiac neovascularization and functional repair capacity were examined after transplantation into nude mice with myocardial infarction. Cardiac transplantation of angiogenic EOCs from healthy subjects markedly increased neovascularization and improved cardiac function, whereas no such effect was observed after transplantation of angiogenic EOCs from patients with CHF. Real-time polymerase chain reaction analysis of 14 candidate angiomiRs, expressed in angiogenic EOCs, revealed a pronounced loss of angiomiR-126 and -130a in angiogenic EOCs from patients with CHF that was also observed in circulating CD34 ϩ cells. Anti-miR-126 transfection markedly impaired the capacity of angiogenic EOCs from healthy subjects to improve cardiac function. miR-126 mimic transfection increased the capacity of angiogenic EOCs from patients with CHF to improve cardiac neovascularization and function. Conclusions-The present study reveals a loss of angiomiR-126 and -130a in angiogenic EOCs and circulating CD34 ϩ cells from patients with CHF. Reduced miR-126 expression was identified as a novel mechanism limiting their capacity to improve cardiac neovascularization and function that can be targeted by miR-126 mimic transfection. (Circulation. 2012;126:2962-2975.)