Oxidant Stress Enhances Adenylyl Cyclase Activation

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Abstract
The generation of oxygen-derived free radicals has been implicated in the disordered vascular regulation of inflammation and reperfusion. In the vasculature, oxygen-derived free radicals are vasodilatory. The mechanisms underlying this effect remain unclear. To examine the cellular processes involved, we studied the effects of hydrogen peroxide (H2O2) on adenylyl cyclase activity in A10 cells, a murine vascular smooth muscle cell line. Pretreatment with H2O2 caused a dose-dependent enhancement of forskolin-stimulated adenylyl cyclase activity (ED50, 44 μmol/L to a maximum of 166% of control activity; n=4). This enhancement was attenuated by iron chelation with deferoxamine and by the intracellular hydroxyl scavenger dimethylthiourea and mimicked by preincubation with purine/xanthine oxidase either alone or in the presence of superoxide dismutase. The effects of H2O2 were completely blocked by the tyrosine kinase inhibitors genistein and tyrphostin A9 but not by its inactive analogue tyrphostin A1 (H2O2 alone, 149±13%; H2O2+tyrphostin A9, 100±9%; H2O2+tyrphostin A1, 171±21%; n=4). H2O2 comparably enhanced adenylyl cyclase activity stimulated by isoproterenol (166±17% of control, n=5) and sodium fluoride (177±18% of control, n=5). Thus oxygen-derived free radicals enhance adenylyl cyclase activation, probably via tyrosine kinase–mediated effects on the catalytic subunit of adenylyl cyclase. Sensitization of adenylyl cyclase activation may be an important mechanism by which free radicals modulate hormone-mediated vasodilation.