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The role of biotransformation and oxidative stress in 3,5-dichloroaniline (3,5-DCA) induced nephrotoxicity in isolated renal cortical cells from male Fischer 344 rats
- Racine, Christopher R., Ferguson, Travis, Preston, Debbie, Ward, Dakota, Ball, John, Anestis, Dianne, Valentovic, Monica, Rankin, Gary O.
- Toxicology 2016 v.341-343 pp. 47-55
- acetylcysteine, alpha-tocopherol, antioxidants, biotransformation, cytochrome P-450, cytotoxicity, free radicals, glutathione, indomethacin, kidneys, lactate dehydrogenase, males, metabolites, metyrapone, nephrotoxicity, oxidative stress, peroxidase, piperonyl butoxide, pyruvic acid, rats, unspecific monooxygenase
- Among the mono- and dichloroanilines, 3,5-dichloroaniline (3,5-DCA) is the most potent nephrotoxicant in vivo and in vitro. However, the role of renal biotransformation in 3,5-DCA induced nephrotoxicity is unknown. The current study was designed to determine the in vitro nephrotoxic potential of 3,5-DCA in isolated renal cortical cells (IRCC) obtained from male Fischer 344 rats, and the role of renal bioactivation and oxidative stress in 3,5-DCA nephrotoxicity. IRCC (∼4millioncells/ml) from male rats were exposed to 3,5-DCA (0–1.0mM) for up to 120min. In IRCC, 3,5-DCA was cytotoxic at 1.0mM by 60min as evidenced by the increased release of lactate dehydrogenase (LDH), but 120min was required for 3,5-DCA 0.5mM to increase LDH release. In subsequent studies, IRCC were exposed to a pretreatment (antioxidant or enzyme inhibitor) prior to exposure to 3,5-DCA (1.0mM) for 90min. Cytotoxicity induced by 3,5-DCA was attenuated by pretreatment with inhibitors of flavin-containing monooxygenase (FMO; methimazole, N-octylamine), cytochrome P450 (CYP; piperonyl butoxide, metyrapone), or peroxidase (indomethacin, mercaptosuccinate) enzymes. Use of more selective CYP inhibitors suggested that the CYP 2C family contributed to 3,5-DCA bioactivation. Antioxidants (glutathione, N-acetyl-l-cysteine, α-tocopherol, ascorbate, pyruvate) also attenuated 3,5-DCA nephrotoxicity, but oxidized glutathione levels and the oxidized/reduced glutathione ratios were not increased. These results indicate that 3,5-DCA may be activated via several renal enzyme systems to toxic metabolites, and that free radicals, but not oxidative stress, contribute to 3,5-DCA induced nephrotoxicity in vitro.