Repair of H2O2-induced DNA damage in bovine lens epithelial cell cultures.
H2O2 concentrations only slightly higher than normal physiological levels found in the lens and aqueous fluid produce a significant number of DNA single-strand breaks in lens epithelial cell cultures. In this investigation, the repair of DNA damaged by short-term, H2O2-induced oxidation was examined in bovine lens epithelial cell cultures. Repair was rapidly initiated and was almost completed in 30 min. A drop in NAD concentration was associated with the DNA damage. 3-Aminobenzamide inhibition of poly(ADP-ribose) polymerase, an enzyme believed to be stimulated by DNA oxidation and involved in DNA repair, prevented the loss of NAD. In contrast, a similar drop in ATP concentration was only slightly lessened by the presence of this inhibitor. Inhibition of the polymerase by 3-aminobenzamide primarily affected only the early recovery period. Overall, recovery occurred almost as effectively in the presence of the inhibitor as in its absence. Preincubation of lens cultures with o-phenanthroline, an iron chelator, prevented the drop in NAD levels associated with DNA damage. Since a hydroxyl radical is produced from H2O2 by a Fenton type reaction, this result supports the concept that the H2O2-induced oxidation of DNA is caused by hydroxyl radical. In contrast, peroxide-induced loss of activity of a cytosolic enzyme, glyceraldehyde-3-phosphate dehydrogenase, was unaffected by the presence of o-phenanthroline, suggesting direct H2O2 oxidation of this enzyme. The results of these experiments suggest that lens epithelium contains enzymes that rapidly repair single-strand DNA breaks induced by H2O2 insult.
Spector
A
A
Department of Ophthalmology, College of Physicians and Surgeons, Columbia University, New York, NY 10032.
Kleiman
N J
NJ
Huang
R R
RR
Wang
R R
RR
D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D004249:DNA Damage; D004260:DNA Repair; D004848:Epithelium / Q000502:physiology; D006861:Hydrogen Peroxide; D007908:Lens, Crystalline / Q000502:physiology*; D009243:NAD / Q000378:metabolism; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism
D009243:NAD; D006861:Hydrogen Peroxide; D011065:Poly(ADP-ribose) Polymerases