Nitroglycerin metabolism in vascular tissue: Role of glutathione S-transferases and relationship between NO(·) and NO₂^- formation

Nitroglycerin is a commonly employed pharmacological agent which produces vasodilatation by release of nitric oxide (NO(·)). The mechanism by which nitroglycerin releases NO(·) remains undefined. Recently, glutathione S-transferases have been implicated as important contributors to this process. They are known to release NO₂^- from nitroglycerin, but have not been shown to release NO(·). The present studies were designed to examine the role of endogenous glutathione S-transferases in this metabolic process. Homogenates of dog carotid artery were incubated anaerobically with nitroglycerin, and NO(·) and NO₂^- production was determined by chemiluminescence. The role of glutathione S-transferases was studied by incubating homogenates with nitroglycerin in the presence of 1 mM GSH or 1 mM S-hexylglutathione, a potent inhibitor of glutathione S-transferases. Homogenates released 163 pmol of NO(·)/h per mg of protein from nitroglycerin, and 2370 pmol of NO₂^-/h per mg. Adding GSH decreased NO(·) production by 82% and increased NO₂^- production by 98%. S-Hexylglutathione inhibited glutathione S-transferase activity by 96% and decreased NO₂^- production by 78%, but had no effect on NO(·) release. A linear relationship between glutathione S-transferase activity and NO₂^- production was observed, whereas glutathione S-transferase activity and NO(·) release were unrelated. Western-blot analysis demonstrated that dog carotid vascular smooth muscle contained Pi and Mu forms of glutathione S-transferases, with a predominance of the former. Purified preparations of human Pi and rat Mu isoforms metabolized nitroglycerin only to NO₂^- and not to NO(·). On the basis of these findings, we conclude that (1) glutathione S-transferases do not contribute to the bioconversion of nitroglycerin to NO(·), but instead act as a degradative pathway for nitroglycerin, and (2) the release of NO(·) from nitroglycerin is not dependent on the formation of NO₂^-.