Inhibition of iNOS protects endothelial-dependent vasodilation in aged rats

Abstract

Aim: To examine whether iNOS contributes to endothelial dysfunction in aged rats.

Methods: Male Sprague Dawley rats were divided into three groups: young rats, aged rats treated with vehicle and aged rats treated with N-[3-(Aminomethyl) benzyl] acetamidine (1400W, 1 mg/kg, ip). Vasorelaxation was measured in isolated thoracic aorta. iNOS expression of thoracic aortic arteries was detected using immunohistochemistry and Western blot. Nitrotyrosine (a marker for peroxynitrite formation) content and expression in thoracic aortic tissue were determined using enzyme linked immunosorbent assay and immunohistochemistry.

Results: Maximal relaxation induced by acetylcholine (10⁻⁹ to 10⁻⁵ mol/L) in the aged rats treated with vehicle was significantly decreased (70%±15%, P<0.01), as compared with the young rats (95%±8%). However, the maximal relaxation induced by acidified NaNO2 (an endothelium-independent vasodilator) had no significant difference between the two groups. Moreover, iNOS and nitrotyrosine expression increased in the vessels of the aged rats. In the aged rats treated with 1400W (a highly selective iNOS inhibitor) nitrotyrosine expression was reduced and acetylcholine-induced vasorelaxation was markedly improved (maximal relaxation was increased to 87%±8%, P<0.05), but the acidified NaNO₂-induced vasorelaxation had no significant change.

Conclusion: Our study demonstrated that inhibition of iNOS by 1400W increased endothelium-dependent vasodilation in aged rats. The mechanism was related with attenuation of peroxynitrite formation.

Figure 1

Vasorelaxation responses of isolated thoracic aortic rings to each concentration of acetylcholine (A, ACh, an endothelium-dependent vasodilator) or acidified NaNO₂ (B, an endothelium-independent vasodilator). ^cP<0.01 vs young group; ^eP<0.05, ^fP<0.01 vs aging group. n=6−8 rats per group.

Figure 2

Effects of aging and 1400W treatment on vascular inducible nitric oxide synthase (iNOS) immunostaining (A, DAB staining, ×40) and iNOS expression (B, Western blot). ^cP<0.01 vs young group; ^eP<0.05 vs aging group. n=7 rats per group.

Figure 3

Effect of 1400W, a selective iNOS inhibitor, on nitrotyrosine (NT) staining (A, DAB staining, ×40) and vascular nitrotyrosine content (B) in aged rats. ^cP<0.01 vs young group; ^eP<0.05 vs aging group. n=7 rats per group.