Atherosclerosis 141 (1998) 265 – 271 Ex vivo gene transfer of endothelial nitric oxide synthase to atherosclerotic rabbit aortic rings improves relaxations to acetylcholine Geza Mozes a , Iftikhar J. Kullo b , Tibor G. Mohacsi a , David G. Cable e , David J. Spector g , Thomas B. Crotty c , Peter Gloviczki a , Zvonimir S. Katusic d , Timothy O’Brien f, * a Diision of Vascular Surgery, Mayo Clinic, 200 First Street SW, Rochester, MN 55905, USA b Diision of Cardioascular Disease, Mayo Clinic, 200 First Street SW, Rochester, MN 55905, USA c Diision of Anatomic Pathology, Mayo Clinic, 200 First Street SW, Rochester, MN 55905, USA d Diision of Anesthesiology and Pharmacology, Mayo Clinic, 200 First Street SW, Rochester, MN 55905, USA e Diision of General Surgery, Mayo Clinic, 200 First Street SW, Rochester, MN 55905, USA f Diision of Endocrinology and Metabolism, Mayo Clinic, 200 First Street SW, Rochester, MN 55905, USA g Department of Microbiology and Immunology, Pennsylania State Uniersity College of Medicine, Hershey, PA, USA Received 22 January 1998; received in revised form 11 June 1998; accepted 16 June 1998 Abstract Cholesterol feeding results in impaired endothelium dependent vasorelaxation. The role of nitric oxide in this process is unclear. The aim of this study was to evaluate the role of nitric oxide in cholesterol-induced vasomotor dysfunction by examining the effect of overexpression of eNOS in the hypercholesterolemic rabbit aorta on vascular reactivity. Vascular rings from the thoracic aorta of hypercholesterolemic rabbits were exposed ex vivo either to an adenoviral vector encoding endothelial nitric oxide synthase (AdeNOS) or Escherichia coli  Galactosidase (Ad Gal). Transgene expression was examined by histochemistry for  galactosidase, immunohistochemistry for eNOS and cyclic GMP measurements and vasomotor studies were performed. Transgene expression was found to localize to the endothelium and adventitia. cGMP levels were significantly greater in AdeNOS compared to Ad Gal transduced rings. Acetylcholine mediated relaxation was significantly impaired in cholesterol fed rabbits and was markedly improved by overexpression of eNOS. These results suggest that reduced NO bioavailability observed in cholesterol-in- duced vascular dysfunction can be partially overcome by eNOS gene transfer. © 1998 Elsevier Science Ireland Ltd. All rights reserved. Keywords: Gene transfer; Nitric oxide synthase; Nitric oxide; Adenoviral vector; Hypercholesterolemia 1. Introduction Hypercholesterolemia is associated with impaired en- dothelium-dependent vasomotor relaxation [1]. A num- ber of mechanisms have been proposed to explain this observation. Enhanced breakdown of NO by interac- tion with increased superoxide levels in the vessel wall is supported by the observation that decreased endothe- lium-dependent relaxations associated with increased NO levels are present in cholesterol-fed rabbits [2]. The latter finding suggests that NOS activity is not impaired by cholesterol feeding and there is no deficiency of substrate or cofactors necessary for NO production. Furthermore, this suggests that accelerated NO degra- dation may play a role in cholesterol-induced vasomo- tor dysfunction. Indeed, cholesterol-induced impaired acetylcholine-mediated vasorelaxation is associated with increased endothelial superoxide anion production [3] and NO is rapidly destroyed by superoxide radical generating systems such as xanthine/xanthine oxidase [4] and is protected by superoxide dismutase [5]. In * Corresponding author. Tel.: +1-507-255-2415; fax: +1-507-255- 4828; e-mail: obrien.timothy@mayo.edu. 0021-9150/98/$ - see front matter © 1998 Elsevier Science Ireland Ltd. All rights reserved. PII:S0021-9150(98)00180-4