EFFECT OF NITRIC-OXIDE AND CELL REDOX STATUS ON THE REGULATION OF ENDOTHELIAL-CELL XANTHINE DEHYDROGENASE

We have previously reported that endothelial cell (EC) xanthine dehydr ogenase/xanthine oxidase (XD/XO) activity correlates inversely with th e O-2 tension to which the cells are exposed. Whether this effect is r elated to the production of reactive O-2 species is unclear. We expose d bovine pulmonary artery EC to various conditions that altered the re dox status of the cells: 1) hypoxia (3% O-2) add normoxia (20% O-2); 2 ) menadione (MEN), known to generate O-2 radicals; 3) catalase (CAT) a nd reduced glutathione (GSH), which detoxify H2O2; and 4) various NO-g enerating systems. Changes in intracellular XO and XO+XD activities we re correlated with rates of extracellular H2O2 release from the same c ells. Conditions that decreased extracellular H2O2 release (hypoxia, C AT, and GSH) produced significant and parallel increases in intracellu lar XO and XO+XD activities in a time-dependent fashion. MEN treatment increased extracellular release of H2O2 and subsequently reduced intr acellular XO and XO+XD activities. NO-generating agents did not change extracellular release of H2O2 but significantly reduced XO and XO+XD activities. The latter effect was prevented by reduced hemoglobin. Sca vengers of hydroxyl radicals reversed the inhibition of XO and XO+XD a ctivities produced by MEN but not that produced by NO. While NO signif icantly inhibited XD/XO activity from rat epididymal fat pad, it did n ot affect XD/XO mRNA expression in these cells. We conclude that intra cellular XD/XO activity is sensitive to changes in oxidant-generating and protective systems. Inhibition of XD/XO activity by NO may be medi ated through direct binding of NO to the enzyme iron-sulfur moiety or to its sulfhydryl groups. The finding of an inhibition of XD/XO by NO provides an important and novel regulatory function for this molecule.