EFFECTS OF QUINE ACETOXYMETHYL ESTER ON H2O2-INDUCED DNA SINGLE-STRAND BREAKAGE IN MAMMALIAN-CELLS - H2O2-CONCENTRATION-DEPENDENT INHIBITION OF DAMAGE AND ADDITIVE PROTECTIVE EFFECT WITH THE HYDROXYL-RADICAL SCAVENGER DIMETHYL-SULFOXIDE

The cell-membrane-permeable calcium probe quin2 acetoxymethyl ester (quin2 AM) was ineffective, in comparison with o-phenanthroline, in protecting cells against H2O2-induced DNA single-strand breakage at H2O2 concentrations of about, and higher than, 0.5 mM. The present study shows that quin2 actually potentiated intracellular DNA damage at high H2O2 concentrations. H2O2 induced DNA breakage appeared within 5 min after exposure, and quin2 affected the induction of DNA breaks at both 0 degrees C and 37 degrees C. Aurintricarboxylic acid, an endonuclease inhibitor, or a decrease in extracellular Ca2+, did not reduce DNA damage. These facts strongly suggest that the breaks were not produced by a Ca2+-dependent nuclease. We showed previously that, in the presence of Fe3+ and H2O2, quin2 strongly potentiated the formation of oxidizing species as well as plasmid DNA breakage, and, as could be expected for a transition-metal chelator, quin2 inhibited the Fenton reaction when Cu2+ was tested instead of Fe3+ [Sandstrom, Granstrom and Marklund (1994) Free Radicals Biol. Med. 16, 177-185]. In the present work with cultured cells, titration with quin2 AM showed that, despite the fact that Cu2+ has three-to-four-orders- of-magnitude higher affinity for quin2 than has Fe3+, both inhibition and potentiation of H2O2-induced DNA damage occurred at quin2 AM concentrations of-about 100 nM: Thus inhibition appeared not to involve Cu2+. The combination of quin2 AM and dimethyl sulphoxide (DMSO) gave an additive effect on H2O2-induced DNA damage compared with the effect of quin2 AM or DMSO alone, whereas the combination of o-phenanthroline and DMSO gave about the same effect as o-phenanthroline alone. In conclusion, our results do not support a role for Ca2+ in the inhibiting effect of quin2 on H2O2-induced DNA damage. Instead, it is likely that inhibition and potentiation by quin2 involves interaction with Fe ions.