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-l (ET-I), Experiments were performed ex vivo. The arteries were exposed (30 minutes at 37 degrees C) to adenoviral vectors encoding eNOS gene (AdCMVeNOS) or P-galactosidase reporter gene (AdCMV beta-Gal), Twenty-four hours after transduction, transgene expression was evident mainly in the vascular adventitia, Rings of control (nontransduced), AdCMV beta Gal- 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 AdCMV beta-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 ETB 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 ETA receptor antagonist, but were not affected by BQ-788 (3x10(-7) mol/L), an ETB receptor antagonist. These results suggest that genetically modified adventitia can produce nitric oxide and cause relaxations in response to ET-I via activation of ETA 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.