Endogenous and exogenous nitric oxide enhance the DNA strand scission induced by tert-butylhydroperoxide in PC12 cells via peroxynitrite-dependent and independent mechanisms, respectively

A short-term exposure to tert-butylhydroperoxide (tB-OOH) promoted a concen tration-dependent formation of DNA single-strand breaks in PC12 cells. Thes e events were paralleled by an increase in the cytosolic concentration of C a2+ that was in part cleared by the mitochondria. Unlike the extent of Ca2 mobilization and/or mitochondrial Ca2+ clearance, the DNA strand scission evoked by the hydroperoxide was markedly reduced by the nitric oxide (NO) s cavenger 2-phenyl-4,4,5,5-tetramethylimidazolin-1-oxyl-3-oxide (PTIO) or by the NO synthase inhibitor N-nitro-L-arginine methylester (L-NAME), Inhibit ors of electron transport (rotenone and myxothiazol), ruthenium red (RR, a polycation which inhibits the calcium uniporter of mitochondria), or peroxy nitrite scavengers (Trolox and L-methionine) were as effective as PTIO or L -NAME in inhibiting the DNA-damaging response mediated by tB-OOH. Rotenone, RR or peroxynitrite scavengers did not further reduce the residual DNA cle avage observed following treatment with tB-OOH in L-NAME-supplemented cells . Exogenous NO also increased the DNA damage caused by tB-OOH in L-NAME-sup plemented cells and this response was blunted by RR or by inhibitors of ele ctron transport but was insensitive to peroxynitrite scavengers. We conclud e that both endogenous and exogenous NO enhance the DNA cleavage generated by tB-OOH in PC12 cells. However, only endogenous NO set the bases for an i nvolvement of peroxynitrite in this DNA-damaging response.