(Verbatim from the application): The goal of this project is to evaluate effects of inducible nitric oxide synthase (iNOS) on vascular function. The investigators have been studying effects of iNOS, using pharmacological inhibitors and iNOS-deficient mice, and have made a recombinant adenovirus, which will provide a novel approach to study vasomotor effects of iNOS. Preliminary data are the first studies of vasomotor effects of iNOS, using adenovirus-mediated gene transfer. Studies are proposed (based on preliminary data) to test the hypothesis that iNOS at high levels of expression, produces superoxide in blood vessels and thereby impairs endothelial function. Studies also are proposed to test the hypothesis that, at low levels of expression, gene transfer of iNOS may generate primarily nitric oxide (NO) that quenches superoxide and thereby improves endothelial function. These latter studies will be performed in vessels with high levels of superoxide, after exposure to lipopolysaccharide or from diabetic rabbits. Studies are proposed to study vessels in vitro and in vivo, and to examine mechanisms by which iNOS impairs vasomotor function. Approaches that will be used to address these aims are gene transfer of iNOS in vitro and in vivo, generally to the carotid or basilar artery of rabbits; measurement of vasomotor responses ex vivo in vascular rings in an organ bath, and responses in vivo using sonimicrometry; measurement of activity of nitric oxide synthase with labeled citrulline; detection of superoxide with hydroethidine in the vessel wall in situ; and quantitation of superoxide with lucigenin (5 uM). It is well recognized that iNOS is expressed in vessels in response to injury and in disease states, and that these disease states generally are associated with endothelial dysfunction. The use of gene transfer of iNOS to vessels to study vasomotor function is novel, and is likely to provide new insight into mechanisms by which iNOS alters vasomotor function.
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