The autoradiographic test for unscheduled DNA synthesis: a sensitive assay for the detection of DNA repair in the HepG2 cell line.
We assessed the DNA-repair capacity of HepG2 cells, which were derived from a human hepatoma, by the unscheduled DNA synthesis assay, using the autoradiography protocol (UDS-AR). We evaluated DNA repair following exposure to direct mutagens (4-nitroquinoline-N-oxide (4-NQO), methyl methanesulfonate (MMS)), to mutagens requiring metabolic activation (benzo[a]pyrene (B[a]P), 2-acetylaminofluorene (2-AAF), N-dimethylnitrosoamine (NDMA)) or to structurally related non-mutagens such as pyrene and 4-acetylaminofluorene (4-AAF). All positive compounds tested induced UDS in HepG2 cells. With 4-NQO and MMS, a concentration-dependent increase in net nuclear grains per cell was observed, with 73 and 90% of cells, respectively, in repair at the highest concentration. B[a]P, 2-AAF and NDMA displayed similar dose-dependent UDS responses, but the percentage of cells in repair was lower (about 45%) than that for 4-NQO and MMS. We assessed the genotoxicity of the compounds tested by determining IC(5NNG): the concentration required to induce 5NNG. The compounds studied were ranked in order of IC(5NNG) as follows: 4-NQO = B[a]P > 2-AAF > MMS > NDMA. The UDS assay discriminated between mutagens and non-mutagens, as pyrene and 4-AAF failed to induce DNA repair. The present study demonstrates that UDS can be used as an endpoint for the detection of DNA damage in HepG2 cells.
Valentin-Severin
Isabelle
I
UMR (1234), Toxicologie Alimentaire, INRA/ENSBANA, 1 Esplanade Erasme, 21000 Dijon, France.
Thybaud
Véronique
V
Le Bon
Anne-Marie
AM
Lhuguenot
Jean-Claude
JC
Chagnon
Marie-Christine
MC
D000704:Analysis of Variance; D001345:Autoradiography / Q000379:methods; D004260:DNA Repair / Q000187:drug effects* / Q000235:genetics; D004261:DNA Replication / Q000187:drug effects* / Q000235:genetics; D023381:Endpoint Determination / Q000379:methods*; D006801:Humans; D009153:Mutagens / Q000633:toxicity*; D012044:Regression Analysis; D014407:Tumor Cells, Cultured
D009153:Mutagens