DNA repair induction by UV irradiation and various mutagens requiring metabolic activation in rat hepatocytes maintained in culture for several days.
Freshly isolated rat hepatocytes cultivated in Williams Medium E (WME) quickly lose their cytochrome P-450-dependent metabolic activities. A significantly better maintenance of cytochrome P-450 was achieved by supplementing WME with hormones and nutrients and 2% DMSO and 10% fetal calf serum. After 72-h cultivation in this medium the cytochrome P-450 content was still 72% of the amount measured in freshly isolated cells. Over the same period of time the activity of 7-ethoxycoumarin-O-deethylase dropped to approximately 55% of its initial value. DNA repair induction by UV irradiation, hydrogen peroxide, benzo[a]pyrene and 2-acetylaminofluorene was measured. A clear dose-dependent increase of the incorporation of [3H]thymidine was found with all agents tested, regardless of whether the treatment of the hepatocytes began 4 or 52 h after cell isolation. In the case of UV light and hydrogen peroxide the maximum effects were slightly lower after prolonged cultivation, whereas little change or even a slight increase was found with benzo[a]pyrene and 2-acetylaminofluorene. These results indicated that cell cultivation led to a slight loss of DNA repair activity but did not, despite the loss of cytochrome P-450, cause a decrease in DNA-damaging intermediates produced from the two indirect mutagens. The absolute amounts of [3H]thymidine incorporated by the solvent control cells of the 52-h groups was consistently found to be twice the level measured in the corresponding 4-h control groups. The incorporation of deoxy[3H]cytidine was not changed when tested in parallel. The increase in [3H]thymidine incorporation can therefore be explained by the assumption of a diminution of the thymidine pool during cell cultivation.
Beales
S
S
Sandoz Ltd, Drug Toxicology Department, Basle, Switzerland.
Suter
W
W
D015073:2-Acetylaminofluorene / Q000494:pharmacology; D000818:Animals; D001564:Benzo(a)pyrene / Q000494:pharmacology; D001711:Biotransformation / Q000187:drug effects / Q000528:radiation effects; D002478:Cells, Cultured; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D004260:DNA Repair / Q000187:drug effects / Q000528:radiation effects*; D004307:Dose-Response Relationship, Radiation; D004795:Enzyme Stability; D006861:Hydrogen Peroxide / Q000494:pharmacology; D008099:Liver / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D008297:Male; D009153:Mutagens; D051381:Rats; D011919:Rats, Inbred Strains; D014466:Ultraviolet Rays
D009153:Mutagens; D001564:Benzo(a)pyrene; D003577:Cytochrome P-450 Enzyme System; D015073:2-Acetylaminofluorene; D006861:Hydrogen Peroxide