Study of N-nitrosomorpholine-induced DNA strand breaks in Caco-2 cells by the classical and modified comet assay: influence of vitamins E and C.
We have studied the genotoxic effects of the well-known heterocyclic liver carcinogen N-nitrosomorpholine (NMOR), an N-nitroso compound, which was prepared in our laboratory by nitrosation of the secondary amine morpholine with NaNO2. NMOR induced DNA strand breaks in human colon carcinoma Caco-2 cells. The concentration-dependent DNA-damaging effects of NMOR were proved by the comet assay. We further characterized DNA strand breaks induced by NMOR as follows: 1) We pretreated cells with vitamins E and C, which are able to eliminate oxidative DNA damage. 2) We varied the pH of the comet assay (12.1 and 13). In general, alkali-labile sites are stable until pH is raised to 12.5. 3) We used the site-specific repair enzymes exonuclease III and formamidopyrimidine-DNA glycosylase in the modified comet assay. Results showed that NMOR-induced DNA strand breaks have their origin exclusively in alkali-labile sites. Nevertheless, vitamins E and C decreased the level of DNA strand breaks. These results showed that antioxidants may have biological activities other than free radical scavenging that relate to their cancer-prevention properties. Our conceptions about reduction of NMOR-induced DNA lesions in Caco-2 cells by vitamins E and C are presented in this work.
Robichová
S
S
Department of Mutagenesis and Carcinogenesis, Cancer Research Institute, Slovak Academy of Sciences, 833 91 Bratislava, Slovak Republic. exonroso@savba.sk
Slamenová
D
D
D016588:Anticarcinogenic Agents / Q000494:pharmacology*; D000975:Antioxidants / Q000494:pharmacology; D001205:Ascorbic Acid / Q000494:pharmacology*; D018938:Caco-2 Cells; D002273:Carcinogens / Q000494:pharmacology*; D020552:Comet Assay; D004249:DNA Damage / Q000187:drug effects*; D004260:DNA Repair; D005090:Exodeoxyribonucleases / Q000378:metabolism; D016166:Free Radical Scavengers; D006801:Humans; D006863:Hydrogen-Ion Concentration; D009602:Nitrosamines / Q000494:pharmacology*; D010084:Oxidation-Reduction; D014810:Vitamin E / Q000494:pharmacology*
D016588:Anticarcinogenic Agents; D000975:Antioxidants; D002273:Carcinogens; D016166:Free Radical Scavengers; D009602:Nitrosamines; D014810:Vitamin E; C002741:N-nitrosomorpholine; D005090:Exodeoxyribonucleases; C020133:exodeoxyribonuclease III; D001205:Ascorbic Acid