The Highly Expressed and Inducible Endogenous NAD(P)H:quinone Oxidoreductase 1 in Cardiovascular Cells Acts as a Potential Superoxide Scavenger

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Abstract
It has recently been demonstrated that purified NAD(P)H:quinone oxidoreductase 1 (NQO1) is able to scavenge superoxide (O 2 •− ) though the rate of reaction of O 2 •− with NQO1 is much lower than the rate of enzymatic dismutation catalyzed by superoxide dismutase (SOD). This study was undertaken to determine if the endogenously expressed NQO1 in cardiovascular cells could scavenge O 2 •− . We observed that NQO1 was highly expressed in cardiovascular cells, including rat aortic smooth muscle A10 and cardiac H9c2 cells, as well as normal human aortic smooth muscle and endothelial cells. NQO1, but not SOD in the cardiovascular cells was highly inducible by 3H-1,2-dithiole-3-thione (D3T). Cytosols from H9c2 and human aortic smooth muscle cells (HASMCs) were isolated to determine the O 2 •− scavenging ability of the endogenously expressed NQO1 by using pyrogallol autooxidation assay. We showed that cytosols from the above cells inhibited pyrogallol autooxidation in an NADPH or NADH-dependent manner. The NADH/NADPH-dependent inhibition of pyrogallol autooxidation by the cytosols was completely abolished by the NQO1-specific inhibitor, ES936, suggesting that the endogenously expressed NQO1 could scavenge O 2 •− . In the presence of NADH/NADPH, cytosols from D3T-treated cells showed increased ability to scavenge O 2 •− as compared to cytosols from untreated cells. This increased ability to scavenge O 2 •− was also completely reversed by ES936. 5-(Diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide spin-trapping experiments using potassium superoxide as a O 2 •− generator further confirmed the ability of NQO1 from HASMCs to scavenge O 2 •− . The spin-trapping experiments also showed that induction of NQO1 by D3T in HASMCs augmented the O 2 •− scavenging ability. Taken together, these results demonstrate that the highly expressed and inducible endogenous NQO1 in cardiovascular cells may act as a potential O 2 •− scavenger.

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