Evidence for involvement of multiple iron species in DNA single-strand scission by H2O2 in HL-60 cells.
Some of the properties of cellular iron species which react with H2O2 to cause DNA single-strand breaks in HL-60 cells were characterized in control cells and in cells made deficient of iron using 4,7-phenylsulfonyl-1,10-phenanthroline (bathophenanthroline disulfonic acid or BPS) and ascorbate. Single-strand breaks were measured using alkaline elution of DNA of cells treated at 4 degrees to minimize repair during treatment. Strand breakage in the presence of 10% serum was only 40% of that in the absence of serum. This effect was traced to reaction of H2O2 with metals, most likely iron, in serum. Dimethyl sulfoxide (Me2SO) inhibited a maximum of 65% of breaks in control cells. The diffusion distance from the site of generation of hydroxyl radicals to the site of reaction with DNA for the Me2SO-inhibitable fraction was 6.9 nm. There was no significant alteration in the fraction of Me2SO-inhibitable strand breaks or in diffusion distance in iron-deficient cells, though total strand breaks decreased by 70%. When the effect of extracellular iron in serum was taken into account, 60 microM orthophenanthroline (OP) inhibited a maximum of 85% of strand breaks. In cells pretreated with 60 microM OP, the Me2SO-inhibitable fraction of the remaining strand breaks decreased to 32%, while the diffusion distance decreased to 4.1 nm. These data indicate the existence of a number of different iron species, as characterized by overlapping but not coincidental inhibition by OP and Me2SO, and by differing diffusion distances.
Byrnes
R W
RW
Department of Chemistry, University of Wisconsin, Milwaukee 53201, USA.
D001205:Ascorbic Acid; D002614:Chelating Agents; D004249:DNA Damage; D004273:DNA, Neoplasm / Q000187:drug effects*; D004277:DNA, Single-Stranded / Q000187:drug effects*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D018922:HL-60 Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D017665:Hydroxyl Radical / Q000378:metabolism; D007501:Iron / Q000378:metabolism*; D007502:Iron Chelating Agents / Q000494:pharmacology; D007700:Kinetics; D010618:Phenanthrolines / Q000494:pharmacology
D002614:Chelating Agents; D004273:DNA, Neoplasm; D004277:DNA, Single-Stranded; D007502:Iron Chelating Agents; D010618:Phenanthrolines; C017049:bathophenanthroline disulfonic acid; D017665:Hydroxyl Radical; D006861:Hydrogen Peroxide; D007501:Iron; D001205:Ascorbic Acid; C025205:1,10-phenanthroline; D004121:Dimethyl Sulfoxide