Superoxide Anion From the Adventitia of the Rat Thoracic Aorta Inactivates Nitric Oxide

Abstract

Abstract-The purpose of this study was to determine whether superoxide anion is produced endogenously in the rat aortic adventitia and whether sufficient superoxide anion is produced to interfere with the response of the rat aorta to nitric oxide. Relaxation was measured in rings of the rat thoracic aorta, which were oriented so that the adventitial or luminal surface could be preferentially exposed to nitric oxide or sodium nitroprusside. To accomplish this, the rings were mounted (1) with the adventitia facing outward, (2) with the adventitia facing inward after inverting, or (3) with the adventitia facing outward after inverting twice (to control for the inverting procedure). The relaxation to nitric oxide, but not to sodium nitroprusside, was less in rings with the adventitia facing outward compared with those in which it faced inward. In contrast, the response to nitric oxide via either surface was similar when extracellular superoxide anion was scavenged with superoxide dismutase. Incubation of rings with nitro blue tetrazolium (NBT) resulted in blue formazan staining of the adventitia, and lucigenin chemiluminescence was significantly greater when detected from the adventitial compared with the intimal aspect of the artery. The reduction of NBT in intact aortic rings was 30Ϯ2 pmol ⅐ min Ϫ1 ⅐ mg Ϫ1 and was significantly decreased by superoxide dismutase to 19Ϯ2 pmol ⅐ min Ϫ1 ⅐ mg Ϫ1 and by a synthetic superoxide dismutase mimic, Euk-8, to 11Ϯ2 pmol ⅐ min Ϫ1 ⅐ mg Ϫ1. The NADPH oxidase inhibitor, diphenyleneiodonium, decreased NBT reduction to 9Ϯ1 pmol ⅐ min Ϫ1 ⅐ mg Ϫ1, whereas inhibitors of xanthine oxidase, mitochondrial oxidases, and nitric oxide synthase were ineffective. Immunohistochemical staining indicated the localization of NADPH oxidase proteins gp91 phox , p22 phox , p47 phox , and p67 phox almost exclusively in the adventitia of the rat aorta with no substantial staining in the media. These results indicate that NADPH oxidase located in the adventitia of rat thoracic aorta generates sufficient extracellular superoxide anion to constitute a barrier capable of inactivating nitric oxide. This study suggests that adventitial superoxide anion can play a role in the pathophysiology of the arterial wall. (Circ Res. 1998;82:810-818.)

Effects of chronic administration of the novel endothelin antagonist J‐104132 on endothelial dysfunction in streptozotocin‐induced diabetic rat

“…The details of this assay have been published previously (Wang et al, 1998). Brie¯y, aortae were cut into transverse rings 10 mm in length.…”

NAD(P)H oxidase p22phox gene C242T polymorphism, nitric oxide production, salt sensitivity and cardiovascular risk factors in Hispanics

“…1 This enzyme complex plays a major role in superoxide ion production in blood vessels. [1][2][3][4] Superoxide ions are known to avidly interact with nitric oxide (NO), reducing its bioactivity by generating peroxynitrite. [2][3][4] Increased activity of the NAD(P)H oxidase system is expected to be associated with reduced NO levels, vascular dysfunction, hypertension and salt sensitivity.…”

“…[1][2][3][4] Superoxide ions are known to avidly interact with nitric oxide (NO), reducing its bioactivity by generating peroxynitrite. [2][3][4] Increased activity of the NAD(P)H oxidase system is expected to be associated with reduced NO levels, vascular dysfunction, hypertension and salt sensitivity. [4][5][6] The latter, is known to be associated with reduced NO bioactivity [7][8][9][10][11][12][13] and increased superoxide production.…”

Nitric Oxide–Induced Increase in p21 Sdi1/Cip1/Waf1 Expression During the Cell Cycle in Aortic Adventitial Fibroblasts

“…10,11 Recent in vivo and in vitro studies have implicated the adventitial fibroblast as a major site for superoxide production in response to angiotensin II administration, 12,13 and we have recently shown that adventitial fibroblasts are a major source of the inducible form of NO synthase in aortic tissue after the in vivo administration of endotoxin to produce an inflammatory response. 14 To begin to evaluate the possible role of NO in modulating vascular remodeling during hypertension, we have used procedures for obtaining adult rat adventitial fibroblasts in culture and have studied the effects of NO on cell proliferation under standardized conditions in which quiescent cells were stimulated with FCS.…”