Adventitial expression of recombinant endothelial nitric oxide synthase gene reverses vasoconstrictor effect of endothelin-1

The present study was designed to determine the effect of recombinant endothelial nitric oxide synthase (eNOS) gene expression on reactivity of canine basilar arteries to endothelin-1 (ET-1). Experiments were performed ex vivo. The arteries were exposed (30 minutes at 37°C) to adenoviral vectors encoding eNOS gene (AdCMVeNOS) or β-galactosidase reporter gene (AdCMVβ- Gal). Twenty-four hours after transduction, transgene expression was evident mainly in the vascular adventitia. Rings of control (nontransduced). AdCMVβGal- and AdCMVeNOS-transduced arteries with and without endothelium were suspended for isometric tension recording. Levels of guanosine 3',5'- cyclic monophosphate (cGMP) were measured by radioimmunoassay. During contractions to uridine 5'-triphosphate, ET-1 (10^-10) to 3x10^-9 mol/L) caused further increase in tension in control and AdCMVβ-Gal-transduced arteries. In contrast, ET-1 caused concentration-dependent relaxations of AdCMVeNOS-transduced arteries. The relaxations to ET-1 in AdCMVeNOS- transduced arteries were endothelium-independent. They were abolished by N(G)-nitro-L-arginine methyl ester or by chemical treatment of adventitia with paraformaldehyde before gene transfer. ET-1 (10^-9 mol/L) significantly increased intracellular cGMP levels in AdCMVeNOS-transduced arteries without endothelium. In arteries transduced with AdCMVeNOS, higher concentrations (10^-9 to 3x10^-8 mol/L) of ET-2 also caused relaxations, whereas ET-3 and sarafotoxin, a selective ET(B) receptor agonist, did not produce any relaxations. The relaxations to ET-1 in AdCMVeNOS-transduced arteries were strongly reduced by BQ-123 (10^-7 mol/L), an ET(A) receptor antagonist, but were not affected by BQ-788 (3x10^-7 mol/L), an ET₁₃ receptor antagonist. These results suggest that genetically modified adventitia can produce nitric oxide and cause relaxations in response to ET-1 via activation of ET(A) receptors. Our findings support a novel concept that successful transfer and expression of recombinant eNOS gene can lead to a qualitative change in responsiveness to vasoconstrictor substances.