| Glutathione (GSH) conjugates of hydroquinone (HQ) and 2-bromohydroquinone (2-BrHQ) produce severe renal proximal tubular necrosis when administered to rats. These GSH conjugates are the putative nephrotoxic metabolites of HQ and 2-BrHQ. Rats and guinea pigs are sensitive to quinone-thioethers, whereas mice and hamsters are resistant. Our hypothesis is that quinone-thioethers produce nephrotoxicity by metabolism via the mercapturic acid pathway followed by covalent binding and oxidative stress. The first aim, which was to characterize the enzymatic activation of HQ to form GSH conjugates, showed that a portion of the liver microsomal oxidation of HQ was cytochrome(s) P-450 dependent. The second aim was to determine whether species differences in quinone-thioether metabolism of correlated with differences in susceptibility to nephrotoxicity. Species differences in {dollar}gamma{dollar}-glutamyl transpeptidase activity and rates of acetylation/deacetylation only partially explained differences in susceptibility to nephrotoxicity. In the third and fourth aims, covalent binding was assessed immunochemically using antibodies directed against quinone-thioethers. Antibodies cross-reacted with in vivo covalent protein adducts in kidney cytosol from Fischer F344 rats treated with 2-BrHQ, HQ, or their corresponding GSH conjugates. Common immunostaining proteins were detected by Western blot analysis at approximately 42 and 79 kilodaltons. Immunohistochemical staining revealed covalent binding by 2,3,5-tris-(GSyl)HQ in the S{dollar}sb3{dollar} segment of the renal proximal tubules in F344 rats, guinea pigs, and hamsters which corresponded to the area of necrosis in rats and guinea pigs. Western blot analysis revealed numerous immunostaining proteins in renal subcellular fractions in rats, mice, guinea pigs, and hamsters. Predominant immunostaining proteins in rats migrated at approximately 34 kilodaltons (mitochondria), 35 kilodaltons (nuclei), and 73 kilodaltons (urine). Renal protein carbonyls, which are an index of oxidative protein damage, reached 127% control 8 hr after treatment of F344 rats with 2-Br-bis-(GSyl)HQ. By 1 hr, renal 8-hydroxydeoxyguanosine, which is an index of oxidative DNA damage, was elevated to nearly 200% control, and returned to baseline levels by 8 hr. The relative significance of quinone-thioether induced covalent and oxidative modifications remains to be determined. |
