OBJECTIVESirtuins (SIRTs) are NAD+-dependent deacetylases that regulate metabolism and life span. We used peripheral blood mononuclear cells (PBMCs) to determine ex vivo whether insulin resistance/metabolic syndrome influences SIRTs. We also assessed the potential mechanisms linking metabolic alterations to SIRTs in human monocytes (THP-1) in vitro.RESEARCH DESIGN AND METHODSSIRT1-SIRT7 gene and protein expression was determined in PBMCs of 54 subjects (41 with normal glucose tolerance and 13 with metabolic syndrome). Insulin sensitivity was assessed by the minimal model analysis. Subclinical atherosclerosis was assessed by carotid intima-media thickness (IMT). In THP-1 cells exposed to high glucose or fatty acids in vitro, we explored SIRT1 expression, p53 acetylation, Jun NH2-terminal kinase (JNK) activation, NAD+ levels, and nicotinamide phosphoribosyltransferase (NAMPT) expression. The effects of SIRT1 induction by resveratrol and of SIRT1 gene silencing were also assessed.RESULTSIn vivo, insulin resistance and metabolic syndrome were associated with low PBMC SIRT1 gene and protein expression. SIRT1 gene expression was negatively correlated with carotid IMT. In THP-1 cells, high glucose and palmitate reduced SIRT1 and NAMPT expression and reduced the levels of intracellular NAD+ through oxidative stress. No effect was observed in cells exposed to linoleate or insulin. High glucose and palmitate increased p53 acetylation and JNK phosphorylation; these effects were abolished in siRNA SIRT1–treated cells. Glucose- and palmitate-mediated effects on NAMPT and SIRT1 were prevented by resveratrol in vitro.CONCLUSIONSInsulin resistance and subclinical atherosclerosis are associated with SIRT1 downregulation in monocytes. Glucotoxicity and lypotoxicity play a relevant role in quenching SIRT1 expression.

Objective-Hyperglycemia is the main determinant of long-term diabetic complications, mainly through induction of oxidative stress. NAD(P)H oxidase is a major source of glucose-induced oxidative stress. In this study, we tested the hypothesis that rosiglitazone (RSG) is able to quench oxidative stress initiated by high glucose through prevention of NAD(P)H oxidase activation. Methods and Results-Intracellular ROS were measured using the fluoroprobe TEMPO-9-AC in HUVECs exposed to control (5 mmol/L) and moderately high (10 mmol/L) glucose concentrations. NAD(P)H oxidase and AMPK activities were determined by Western blot. We found that 10 mmol/L glucose increased significantly ROS production in comparison with 5 mmol/L glucose, and that this effect was completely abolished by RSG. Interestingly, inhibition of AMPK, but not PPAR␥, prevented this effect of RSG. AMPK phosphorylation by RSG was necessary for its ability to hamper NAD(P)H oxidase activation, which was indispensable for glucose-induced oxidative stress. Downstream of AMPK activation, RSG exerts antioxidative effects by inhibiting PKC. Conclusions-This study demonstrates that RSG activates AMPK which, in turn, prevents hyperactivity of NAD(P)H oxidase induced by high glucose, possibly through PKC inhibition. Therefore, RSG protects endothelial cells against glucose-induced oxidative stress with an AMPK-dependent and a PPAR␥-independent mechanism. (Arterioscler

These data show that the risk of hypertension associated with coffee intake varies according to CYP1A2 genotype. Carriers of slow *1F allele are at increased risk and should thus abstain from coffee, whereas individuals with *1A/*1A genotype can safely drink coffee.

Low expression of RGS2 contributes to increased G-protein-coupled signaling in hypertensive patients. The allele G is associated with low RGS2 expression and blood pressure increase in humans.

Prolonged ouabain administration (25 microg kg(-1) day(-1) for 5 wk) induces "ouabain hypertension" (OH) in rats, but the molecular mechanisms by which ouabain elevates blood pressure are unknown. Here, we compared Ca(2+) signaling in mesenteric artery smooth muscle cells (ASMCs) from normotensive (NT) and OH rats. Resting cytosolic free Ca(2+) concentration ([Ca(2+)](cyt); measured with fura-2) and phenylephrine-induced Ca(2+) transients were augmented in freshly dissociated OH ASMCs. Immunoblots revealed that the expression of the ouabain-sensitive alpha(2)-subunit of Na(+) pumps, but not the predominant, ouabain-resistant alpha(1)-subunit, was increased (2.5-fold vs. NT ASMCs) as was Na(+)/Ca(2+) exchanger-1 (NCX1; 6-fold vs. NT) in OH arteries. Ca(2+) entry, activated by sarcoplasmic reticulum (SR) Ca(2+) store depletion with cyclopiazonic acid (SR Ca(2+)-ATPase inhibitor) or caffeine, was augmented in OH ASMCs. This reflected an augmented expression of 2.5-fold in OH ASMCs of C-type transient receptor potential TRPC1, an essential component of store-operated channels (SOCs); two other components of some SOCs were not expressed (TRPC4) or were not upregulated (TRPC5). Ba(2+) entry activated by the diacylglycerol analog 1-oleoyl-2-acetyl-sn-glycerol [a measure of receptor-operated channel (ROC) activity] was much greater in OH than NT ASMCs. This correlated with a sixfold upregulation of TRPC6 protein, a ROC family member. Importantly, in primary cultured mesenteric ASMCs from normal rats, 72-h treatment with 100 nM ouabain significantly augmented NCX1 and TRPC6 protein expression and increased resting [Ca(2+)](cyt) and ROC activity. SOC activity was also increased. Silencer RNA knockdown of NCX1 markedly downregulated TRPC6 and eliminated the ouabain-induced augmentation; silencer RNA knockdown of TRPC6 did not affect NCX1 expression but greatly attenuated its upregulation by ouabain. Clearly, NCX1 and TRPC6 expression are interrelated. Thus, prolonged ouabain treatment upregulates the Na(+) pump alpha(2)-subunit-NCX1-TRPC6 (ROC) Ca(2+) signaling pathway in arterial myocytes in vitro as well as in vivo. This may explain the augmented myogenic responses and enhanced phenylephrine-induced vasoconstriction in OH arteries (83) as well as the high blood pressure in OH rats.