Inhibition of inducible nitric oxide synthase restores endothelium-dependent relaxations in proinflammatory mediator-induced blood vessels.

Abstract

Endothelium-dependent relaxations mediated by nitric oxide (NO) are attenuated in arteries exposed to proinflammatory mediators. Because proinflammatory mediators stimulate the expression of the inducible NO synthase (iNOS) in vascular cells, the role of iNOS-derived NO in the impaired endothelium-dependent relaxation was examined in arterial ring preparations. Exposure of rabbit carotid arteries to interleukin-1 beta (IL-1 beta; 100 U/mL for 7 hours) and porcine coronary arteries to a combination of tumor necrosis factor-alpha (1000 U/mL), interferon-gamma (500 U/mL), and lipopolysaccharide (10 micrograms/mL) for 15 hours (conditions that are associated with iNOS expression) markedly attenuated relaxations to receptor-dependent agonists, whereas those to the calcium ionophore A23187 and sodium nitroprusside were virtually unchanged. The impaired relaxation was not associated with a reduced level of the constitutive endothelial NOS (cNOS) but was accompanied by a reduced formation of biologically active NO as assessed in a bioassay system. The attenuated relaxation of carotid arteries to acetylcholine was not affected by superoxide dismutase and was neither found in arteries exposed to IL-1 beta for only 15 minutes nor in IL-1 beta-treated arteries for 7 hours followed by a 17-hour incubation period without the cytokine. Furthermore, no impaired relaxation was found in rings exposed to IL-1 beta in combination with either cycloheximide or N-alpha-tosyl-L-lysine chloromethyl ketone or pyrrolidine dithiocarbamate, treatments that prevent iNOS expression. In addition, selective inhibition of iNOS with S-methylisothiourea (10 mumol/L) completely restored acetylcholine-induced relaxations. These findings indicate that the continuous generation of NO induced by proinflammatory mediators plays a major role in the inhibition of endothelium-dependent relaxation, most likely by impairing a step in the signal transduction cascade that links activation of endothelial receptors to the calcium-calmodulin-dependent activation of NOS.