Superoxide anion formation is vital to the microbicidal activity of phagocytes. Recently, however, there is accumulating evidence that it is also involved in cell growth in vascular smooth muscle cells (VSMCs). We have shown that the hypertrophic agent angiotensin II stimulates superoxide production by activating the membrane-bound NADH/NADPH oxidase and that inhibition of this oxidase attenuates vascular hypertrophy. However, the molecular identity of this oxidase in VSMCs is unknown. We have recently cloned the cytochrome b558 alpha-subunit, p22(phox) (one of the key electron transfer elements of the NADPH oxidase in phagocytes), from a rat VSMC cDNA library, but its role in VSMC oxidase activity remains unclarified. Here we report that the complete inhibition of p22(phox) mRNA expression by stable transfection of antisense p22(phox) cDNA into VSMCs results in a decrease in cytochrome b content, which is accompanied by a significant inhibition of angiotensin II-stimulated NADH/NADPH-dependent superoxide production, subsequent hydrogen peroxide production, and [3H]leucine incorporation. We provide the first evidence that p22(phox) is a critical component of superoxide-generating vascular NADH/NADPH oxidase and suggest a central role for this oxidase system in vascular hypertrophy.

Recent studies suggest that superoxide production by the NADPH/NADH oxidase may be involved in smooth muscle cell growth and the pathogenesis of hypertension. We previously showed that angiotensin II (Ang II) activates a p22phoxbased NADPH/NADH oxidase in cultured rat vascular smooth muscle cells and in animals made hypertensive by infusion of Ang II. To investigate the mechanism responsible for this increased oxidase activity, we examined p22phox mRNA expression in rats made hypertensive by implanting an osmotic minipump that delivered Ang II (0.7 mg/kg per day). Blood pressure began to increase 3 days after the start of Ang II infusion and remained elevated for up to 14 days. Expression of p22phox mRNA in aorta was also increased after 3 days and reached a maximum increase of 338 +/- 41% by 5 days after pump implantation compared with the value after sham operation. This increase in mRNA expression was accompanied by an increase in the content of the corresponding cytochrome (twofold) and NADPH oxidase activity (179 +/- 11% of that in sham-operated rats 5 days after pump implantation). Treatment with the antihypertensive agents losartan (25 mg/kg per day) or hydralazine (15 mg/kg per day) inhibited this upregulation of mRNA levels and activity. Furthermore, infusion of recombinant heparin-binding superoxide dismutase decreased both blood pressure and p22phox mRNA expression. In situ hybridization of aortic tissue showed that p22phox mRNA was expressed in medial smooth muscle as well as in the adventitia. These findings suggest that Ang II-induced hypertension activates the NADPH/NADH oxidase system by upregulating mRNA levels of one or several components of this oxidase system, including the p22phox, and that the NADPH/NADH oxidase system is associated with the pathology of hypertension in vivo.

In this large-scale Japan-wide general population study, an absolute VFA value of about 100 cm(2) equated with obesity-related cardiovascular risk factor accumulation, irrespective of gender, age, and BMI.

Abstract-Recent studies have indicated that angiotensin II (Ang II) can stimulate oxidative stress. The present study was conducted to assess the contribution of oxygen radicals to hypertension and regional circulation during Ang II-induced hypertension. With radioactive microspheres, the responses of systemic and regional hemodynamics to the membranepermeable, metal-independent superoxide dismutase mimetic 4-hydroxy-2,2,6,6-tetramethyl piperidinoxyl (tempol) were assessed in conscious Ang II-infused hypertensive rats. Ang II-infused rats (80 ng/min SC for 12 days: nϭ25) showed higher mean arterial pressure (MAP: 161Ϯ4 mm Hg) and total peripheral resistance (TPR: 1.59Ϯ0.08 mm Hg ⅐ min Ϫ1 ⅐ mL Ϫ1 ) than vehicle-infused normotensive rats (116Ϯ3 mm Hg and 0.95Ϯ0.04 mm Hg ⅐ min Ϫ1 ⅐ mL Ϫ1 , respectively; nϭ23). The blood flow rates in the brain, spleen, large intestine, and skin were significantly reduced in Ang II-infused rats compared with vehicle-infused rats, whereas rates in the lung, heart, liver, kidney, stomach, small intestine, mesenterium, skeletal muscle, and testis were similar. Vascular resistance was significantly increased in every organ studied except the lung, in which the resistance was similar. Tempol (216 mol/kg IV) significantly reduced MAP by 30Ϯ4% from 158Ϯ7 to 114Ϯ5 mm Hg and TPR by 35Ϯ6% from 1.57Ϯ0.17 to 0.95Ϯ0.04 mm Hg ⅐ min Ϫ1 ⅐ g Ϫ1 in Ang II-infused rats (nϭ9) but had no effect on these parameters in vehicle-infused rats (nϭ8). In Ang II-infused rats, tempol did not affect regional blood flow but significantly decreased vascular resistance in the brain (29Ϯ6%), heart (31Ϯ6%), liver (37Ϯ7%), kidney (30Ϯ7%), small intestine (38Ϯ6%), and large intestine (47Ϯ7%). Ang II-infused hypertensive rats showed doubled vascular superoxide production (assessed with lucigenin chemiluminescence), which was normalized by treatment with tempol (3 mmol/L, nϭ7). Further studies showed that the NO synthase inhibitor, N -nitro-L-arginine methyl ester (11 mol ⅐ kg Ϫ1 ⅐ min Ϫ1 IV, nϭ11) markedly attenuated the systemic and regional hemodynamic responses of tempol in Ang II-infused rats. These results suggest that in this model of hypertension, oxidative stress may have contributed to the alterations in systemic blood pressure and regional vascular resistance through inactivation of NO. (Hypertension. 2001;37:77-83.)

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