Smith, Leslie

Our previous ex vivo and in vivo studies reported that expression of the recombinant endothelial nitric oxide (NO) synthase (eNOS) gene in adventitial fibroblasts recovers NO production in arteries without endothelium in response to bradykinin. The present study was designed to characterize subtypes of bradykinin receptors on adventitial fibroblasts coupled to the activation of recombinant eNOS. Endothelium-denuded segments of canine basilar arteries were transduced with β-galactosidase (β-Gal) gene or eNOS gene ex vivo, using a replication-defective adenoviral vector (1010plaque-forming units/ml) for 30 min at 37°C. Twenty-four hours later, isometric force recording or cGMP measurement was carried out. B1bradykinin receptor agonist (des-Arg9-bradykinin, 10−10-10−8mol/l) did not significantly affect vascular tone in control or β-Gal gene-transduced canine basilar arteries without endothelium. In contrast, this agonist caused concentration-dependent relaxations in recombinant eNOS gene-transduced arteries without endothelium. Relaxations to B1receptor agonist in the eNOS arteries were abolished by B1receptor antagonist (des-Arg9-[Leu8]bradykinin, 6 × 10−9mol/l) but not by B2receptor antagonist (Hoe-140, 5 × 10−8mol/l). Bradykinin did not significantly alter vascular tone in control or β-gal arteries without endothelium, whereas this peptide (10−11-10−8mol/l) induced concentration-dependent relaxations, as well as an increase in cGMP formation in endothelium-denuded eNOS-transduced arteries. Stimulatory effects of bradykinin were prevented in the presence of a B2receptor antagonist but not in the presence of a B1receptor antagonist. B1and B2receptor antagonists had no effect on relaxations to substance P, confirming the selectivity of the compounds. Our results suggest that B1and B2bradykinin receptors are coupled to activation of recombinant eNOS expressed in adventitial fibroblasts.