ABSENCE OF ENDONUCLEASE ACTIVATION DURING ACUTE CELL-DEATH IN RENAL PROXIMAL TUBULES

The role of endonuclease and poly(ADP-ribose) polymerase activation in various types of cell injuries and death to rabbit renal proximal tub ule suspensions was examined. Proximal tubules were exposed to the mit ochondrial inhibitor antimycin A (0.1 muM), the protonophore carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone (FCCP, 1 muM), the calcium ionophore ionomycin (5 muM), or the oxidant t-butyl hydroperoxide (TB HP, 0.5 mM) in the absence or presence of the endonuclease inhibitor a urintricarboxylic acid or the poly(ADP-ribose) polymerase inhibitor 3- aminobenzamide. Lactate dehydrogenase (LDH) release was used as a mark er of cell death and analysis of genomic DNA for internucleosomal clea vage was used as a marker of endonuclease activation. Aurintricarboxyl ic acid and 3-aminobenzamide had no effect on the proximal tubule LDH release produced by 1 h exposure to antimycin A, FCCP, or ionomycin, o r 2 h exposure to TBHP. Furthermore, there was no evidence of DNA frag mentation with any compound prior to or after cell death began. As a p ositive control, proximal tubules exposed to digitonin in the absence of metabolic substrates resulted in the chelator-inhibitable fragmenta tion of DNA, indicating that the endonuclease is present in proximal t ubules. These results show that endonuclease activation did not occur prior to or after cell death began. Furthermore, these results suggest that endonuclease and poly(ADP-ribose) polymerase activation do not p lay a role in this model of acute renal proximal tubule cell injury an d death induced by agents that cause oxidative stress, mitochondrial d ysfunction, or increases in cytosolic free calcium.