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Although obesity-related cardiovascular disease and hypoxia are associated with erectile dysfunction, little is known about the direct effects of hypoxia on penile arteries. In the present study, the effects of acute hypoxia (P[O.sub.2] = ~10 Torr, 20 min) were investigated in isolated penile arteries to determine the influence of endothelium removal, nitric oxide (NO) synthase (NOS), cyclooxygenase (COX), NADPH oxidase, changes in reactive oxygen species (ROS), and a high-fat diet. Hypoxia-relaxed penile arteries contracted with phenylephrine by ~50%. Relaxation to hypoxia and acetylcholine was reduced by endothelium removal and by inhibition of NOS ([N.sup.[omega]]-nitro-L-arginine) and COX (indomethacin) but was enhanced by Tempol and by NADPH oxidase inhibition with apocynin and gp91ds-tat. Basal superoxide levels detected by lucigenin chemiluminescence were reduced by Tempol and gp91ds-tat and were enhanced by NOS blockade. Hypoxic relaxant responses were enhanced by catalase and ebselen. Exogenous peroxide evoked relaxations of penile arteries, which were partially inhibited by endothelium removal and by the inhibition of COX and extracellular signal-regulated mitogen-activated protein kinase (MAPK) but enhanced by p38 MAPK blockade. The NO-dependent component of relaxation to hypoxia was impaired in penile arteries from high-fat diet-fed, obese rats associated with increased superoxide production. Thus hypoxic relaxation of penile arteries is partially mediated by endothelial NO in a manner that is normally attenuated by endogenous ROS production. Obesity further increases superoxide production and impairs the influence of NO. Therefore, cardiovascular disease involving decreased NO bioavailability and/or enhanced ROS generation may contribute to erectile dysfunction through impairing the relaxation of penile arteries to hypoxia. diet-induced obesity; hypoxia; penile vascular regulation; superoxide doi: 10.1152/ajpheart.00382.2010.

Protein kinase C (PKC) stimulation of NAD(P)H oxidases (Nox) is an important component of multiple vascular disease processes; however, the relationship between oxidase activation and the regulation of vascular smooth muscle contraction by PKC remains poorly understood. Therefore, we examined the signaling cascade of PKC-elicited Nox activation and the role of superoxide and hydrogen peroxide in mediating PKC-induced vascular contraction. Endothelium-denuded bovine coronary arteries showed a PKC-dependent basal production of lucigenin (5 [micro]M)-detected Nox oxidase-derived superoxide, which was stimulated fourfold by PKC activation with 10 [micro]M phorbol 12,13-dibutyrate (PDBu). PDBu appeared to increase superoxide generation by Nox2 through both [p47.sup.phox] and peroxide-dependent Src activation mechanisms based on the actions of inhibitors, properties of Src phosphorylation, and the loss of responses in aorta from mice deficient in Nox2 and [p47.sup.phox]. The actions of inhibitors of contractile regulating mechanisms, scavengers of superoxide and peroxide, and responses in knockout mouse aortas suggest that a major component of the contraction elicited by PDBu appeared to be mediated through peroxide derived from Nox2 activation stimulating force generation through Rho kinase and calmodulin kinase-II mechanisms. Superoxide generated by PDBu also attenuated relaxation to nitroglycerin. Peroxide-derived from Nox2 activation by PKC appeared to be a major contributor to the thromboxane [A.sub.2] receptor agonist U46619 (100 nM)-elicited contraction of coronary arteries. Thus a [p47.sup.phox] and Src kinase activation of peroxide production by Nox2 appears to be an important contributor to vascular contractile mechanisms mediated through activation of PKC. hydrogen peroxide; NADPH oxidase; protein kinase C; Src kinases; coronary artery

The dietary polyphenolic compound resveratrol, by activating the protein deacetylase enzyme silent information regulator 2/sirtuin 1 (SIRT1), prolongs life span in evolutionarily distant organisms and may mimic the cytoprotective effects of dietary restriction. The present study was designed to elucidate the effects of resveratrol on cigarette smoke-induced vascular oxidative stress and inflammation, which is a clinically highly relevant model of accelerated vascular aging. Cigarette smoke exposure of rats impaired the acetylcholine-induced relaxation of carotid arteries, which could be prevented by resveratrol treatment. Smoking and in vitro treatment with cigarette smoke extract (CSE) increased reactive oxygen species production in rat arteries and cultured coronary arterial endothelial cells (CAECs), respectively, which was attenuated by resveratrol treatment. The smoking-induced upregulation of inflammatory markers (ICAM-1, inducible nitric oxide synthase, IL-6, and TNF-[alpha])in rat arteries was also abrogated by resveratrol treatment. Resveratrol also inhibited CSE-induced NF-[kappa]B activation and inflammatory gene expression in CAECs. In CAECs, the aforementioned protective effects of resveratrol were abolished by knockdown of SIRT1, whereas the overexpression of SIRT1 mimicked the effects of resveratrol. Resveratrol treatment of rats protected aortic endothelial cells against cigarette smoking-induced apoptotic cell death. Resveratrol also exerted antiapoptotic effects in CSE-treated CAECs, which could be abrogated by knockdown of SIRT1. Resveratrol treatment also attenuated CSE-induced DNA damage in CAECs (comet assay). Thus resveratrol and SIRT1 exert antioxidant, anti-inflammatory, and antiapoptotic effects, which protect the endothelial cells against the adverse effects of cigarette smoking-induced oxidative stress. The vasoprotective effects of resveratrol will likely contribute to its anti-aging action in mammals and may be especially beneficial in pathophysiological conditions associated with accelerated vascular aging. tobacco; polyphenol; stroke; inflammation; apoptosis; vascular aging; sirtuin 1

Monocrotaline (MCT)-induced pulmonary artery hypertension (PAH) in rats is preceded by an inflammatory response, progressive endothelial cell membrane disruption, reduction in the expression of caveolin-1, and reciprocal activation of STAT3 (PY-STAT3). Superoxide and NF-KB have been implicated in PAH. To evaluate the role of caveolin-1, PY-STAT3 activation, and superoxide in PAH, MCT-injected rats were treated daily with pyrrolidine dithiocarbamate (PDTC; starting on days 1, 3, and 14 x 2 wk), an inhibitor of inflammation and NF-[kappa]B activation. Hemodynamic data, the expression of inhibitory (I)-[kappa]B[alpha], caveolin-1, and Tie2 (a membrane protein), activation of PY-STAT3 and NF-[kappa]B, and superoxide chemiluminescence were examined. Rats developed progressive PAH at 2 wk post-MCT. There was progressive reduction in the expression of caveolin-1, Tie2, and activation of PY-STAT3 in the lungs. Reduction in I-[kappa]B[alpha] expression was present at 2 and 4 wk post-MCT. Superoxide chemiluminescence and NF-[kappa]B activation were observed only at 2 wk post-MCT and both decreased by 4 wk post-MCT despite progressive PAH. PDTC (starting on days 1 and 3) rescued caveolin-1 and Tie2, reversed MCT-induced PYSTAT3 activation, and attenuated PAH. In addition, PDTC restored I-[kappa]B[alpha] expression and reduced superoxide chemiluminescence at 2 wk but did not inhibit NF-[kappa]B activation despite attenuation of PAH. PDTC had no effect on established PAH. Increased superoxide chemiluminescence and NF-[kappa]B activation appear to be a transient phenomenon in the MCT model. Thus the disruption of endothelial cell membrane integrity resulting in caveolin-I loss and reciprocal activation of PY-STAT3 plays a key role in the MCT-induced PAH. caveolin-l; inhibitory [kappa]B[alpha]; inflammation; PY-STAT3; Tie2

Hyperglycemia increases glucose metabolism via the polyol pathway, which results in elevations of intracellular sorbitol concentration. Thus we hypothesized that elevated level of sorbitol contributes to the development of hyperglycemia-induced dysfunction of microvessels. In isolated, pressurized (80 mmHg) rat gracilis muscle arterioles (~150 [micro]m), high glucose treatment (25 raM) induced reduction in flow-dependent dilation (from maximum of 39 [+ or -] 2% to 15 [+ or -] 1%), which was significantly mitigated by an aldose reductase inhibitor, zopolrestat (maximum 27 [+ or -] 2%). Increasing doses of sorbitol ([10.sup.10]-[10.sup.-4] M) elicited dose-dependent constrictions (maximum 22 [+ or -] 3%), which were abolished by endothelium removal, a prostaglandin [H.sub.2]/thromboxane [A.sub.2] ([PGH.sub.2]/[TXA.sub.2]) receptor (TP) antagonist SQ-29548, or superoxide dismutase (SOD) plus catalase (CAT). Incubation of arterioles with sorbitol ([10.sup.-7] M) reduced flow-dependent dilations (from maximum of 39 [+ or -] 2% to 20 [+ or -] 1.5%), which was not further affected by inhibition of nitric oxide synthase by [N.sup.[omega]]-nitro-L-arginine methyl ester but was prevented by SOD plus CAT and mitigated by SQ-29548. Nitric oxide donor sodium nitroprusside-induced ([10.sup.-9]-[10.sup.-6] M) dilations were also decreased in a SQ-29548 and SOD plus CAT-reversible manner, whereas adenosine dilations were not affected by sorbitol exposure. Sorbitol significantly increased arterial superoxide production detected by lucigenin-enhanced chemiluminescence, which was inhibited by SOD plus CAT. Sorbitol treatment also increased arterial formation of 3-nitrotyrosine. We suggest that hyperglycemia by elevating intracellular sorbitol induces oxidative stress, which interferes with nitric oxide bioavailability and promotes [PGH.sub.2]/[TXA.sub.2] release, both of which affect regulation of vasomotor responses of arterioles. Thus increased activity of the polyol pathway may contribute to the development of microvascular dysfunction in diabetes mellitus. aldose reductase; sorbitol; flow-dependent dilation; oxidative stress; prostaglandin [H.sub.2]; thromboxane [A.sub.2]; nitric oxide