NITRIC-OXIDE INHIBITED PEROXYL AND ALKOXYL RADICAL FORMATION WITH CONCOMITANT PROTECTION AGAINST OXIDANT INJURY IN INTESTINAL EPITHELIAL-CELLS

A model compound of lipid peroxidation, tert-butyl hydroperoxide (tBOO H), was used in vitro to investigate (i). the generation of tBOOH-deri ved peroxyl and alkoxyl radicals by rat intestinal epithelial cells or enterocytes and (ii) the role of nitric oxide (NO) on cell-generated free radical formation and cellular cytotoxicity. Peroxyl, alkoxyl, an d methyl radicals were detected and characterized by direct and spin-t rapping electron paramagnetic resonance spectroscopy in incubations co ntaining tBOOH and hematin, enterocytes, or intestinal epithelial cell line-6 cells. The direct interactions of tBOOH-derived radicals and N O from nitrosoglutathione (GSNO), nitrosoacetyl penicillamine (SNAP), or 1-{b3-aminopropy-4-(3-aminopropylammonio)} butylamino-diazeniumdiol ate (SpNONOate) were demonstrated as their levels were depleted in the se incubations. SNAP, not GSNO or SpNONOate, was capable of trapping m ethyl radical produced during hematin-catalyzed decomposition of tBOOH . Cellular cytotoxicity expressed by percentage of dead cells and lact ate dehydrogenase was increased with tBOOH treatment. Addition of GSNO , SNAP, or SpNONOate suppressed tBOOH-induced elevation of cell cytoto xicity. The NO donor precursor glutathione, acetylpenicillamine, or sp ermine did not have any effects on tBOOH-derived radical generation or cell cytotoxicity. These findings demonstrated free radical-free radi cal reactions between NO- and tBOOH derived alkoxyl and peroxyl radica ls generated by enterocytes. These reactions, at least in part, descri be the protective role of NO from hydroperoxide-induced injury in inte stinal epithelial cells. (C) 1998 Academic Press.