Gene Transfer of Endothelial NO Synthase and Manganese Superoxide Dismutase on Arterial Vascular Cell Adhesion Molecule-1 Expression and Superoxide Production in Deoxycorticosterone Acetate-Salt Hypertension

Abstract

Abstract-Enhanced vascular cell adhesion molecule-1 (VCAM-1) expression directly contributes to vascular dysfunction in hypertension. Decreased NO and/or increased superoxide are causative factors for such an event in the vessel wall. The present study was undertaken to determine whether gene transfer of endothelial NO synthase (eNOS) or manganese superoxide dismutase (MnSOD) affects VCAM-1 levels in arteries from hypertensive rats. Isolated carotid and femoral arteries from deoxycorticosterone acetate (DOCA)-salt hypertensive rats were transduced for 4 hours with adenoviral vectors encoding eNOS, MnSOD, or ␤-galactosidase reporter genes. Recombinant eNOS or MnSOD expression was evident morphologically and quantitatively 24 hours after gene transfer. Immunohistochemistry, ELISA, and Western blot techniques were used to determine VCAM-1 expression and levels. In addition, endogenous eNOS and MnSOD and in situ superoxide levels were analyzed by immunoblotting and fluorescence confocal microscopy, respectively. Arterial VCAM-1 expression was significantly higher in DOCA-salt hypertensive rats than in sham-operated rats; this expression was accompanied by decreased MnSOD but unaltered endogenous eNOS levels. VCAM-1 expression was significantly lower in MnSOD-and eNOS-transduced hypertensive arteries, with a concomitant reduction of superoxide level. These results suggest that gene transfer of MnSOD or eNOS suppresses arterial VCAM-1 expression in DOCA-salt hypertension by reducing the superoxide level. A lthough hypertension is one of the key risk factors for atherosclerosis, the underlying molecular and cellular mechanisms remain to be delineated. 1 Enhanced adhesion molecule expression is known to contribute directly to vascular dysfunction in hypertension. 2 Vascular cell adhesion molecule-1 (VCAM-1) is an early marker of endothelial activation and dysfunction, leukocyte infiltration, and vascular remodeling in the development of early atherosclerotic lesions (fatty streaks and fibrous plaques). 2-5 Although VCAM-1 is structurally similar to intercellular adhesion molecule-1 and other adhesion molecules, its pattern of expression is unique. It exhibits low to negligible expression under baseline conditions but is profoundly upregulated by proatherosclerotic conditions in animal models and in humans. [2][3][4][5] In addition, compared with other adhesion molecules whose expression often extends into uninvolved and/or lesion-protected regions of the vessel wall, VCAM-1 expression is largely restricted to atherosclerotic lesions and lesionpredisposed regions. 2-5 Consistent with these phenomena, a recent study has demonstrated that VCAM-1, but not intercellular adhesion molecule-1 (ICAM-1), plays a critical role in early atherogenesis. 6 Enhanced adhesion molecule expression has been ascribed to an imbalance between oxidative stress and antioxidant activity. Experimental evidence suggests that NO and superoxide are 2 key regulating factors for the expression of adhesion molecules, including VCAM-1. 7,8 In angiotensi...

Gene Transfer of Human Guanosine 5′-Triphosphate Cyclohydrolase I Restores Vascular Tetrahydrobiopterin Level and Endothelial Function in Low Renin Hypertension

“…16,17 Our recent studies indicate that endothelin-1 (ET-1) plays an important role in NADPH oxidase-derived O 2 Ϫ production in this model. [13][14][15] However, the interaction between ET-1-induced O 2 Ϫ and BH 4 on endothelial dysfunction and the underlying mechanisms are unknown. In the present study, we tested the hypothesis that ET-1-induced O 2 Ϫ reduces BH 4 and endothelium-dependent relaxation in carotid arteries of DOCA-salt rats, and gene transfer of human GTPCH I reverses the BH 4 deficiency and endothelial dysfunction by reducing O 2 Ϫ levels.…”

“…[13][14][15] Briefly, rats (250 to 275 g, Charles River, Portage, Mich) underwent uninephrectomy (flank incision, left side) and a silicon rubber DOCA implant (Sigma) (200 mg/kg) was placed subcutaneously between the shoulder blades. Sham rats were also uninephrectomized but received no implant.…”

“…[13][14][15] Some arterial segments from sham and DOCA-salt rats were incubated at 37°C for 4 hours with or without the respective agents before O 2 Ϫ , BH 4 , and GTPCH I measurements.…”

“…[13][14][15]18 The rings were then transferred to fresh Eagle minimal essential medium (EMEM) with 0.1% bovine serum albumin and incubated for 24 hours at 37°C. Some vessels from sham and DOCA-salt rats were incubated for 24 hours without transduction and served as normal negative controls.…”

“…[13][14][15] The oxidative fluorescent dye dihydroethidine was used to evaluate in situ O 2 Ϫ production. Unfixed frozen rings of carotid arteries were cut into 30 m sections and placed on a glass slide.…”