Evaluation of the single cell gel electrophoresis assay with human hepatoma (Hep G2) cells.
Human Hep G2 cells have retained the activities of phase I and phase II enzymes which are involved in the metabolism of environmental genotoxins. The present study describes the results of single cell gel electrophoresis (SCGE) assays with a panel of different model compounds with these cells. With genotoxic carcinogens such as aflatoxin B(1) (AFB(1)), benzo(a)pyrene (B(a)P), nitrosodimethylamine (NDMA) and cyclophosphamide (CP), statistically significant dose dependent induction of DNA migration was measured. With the two heterocyclic amines, 2-amino-3-methyl-3H-imidazo[4, 5-f]quinoline (IQ) and 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1), and also with rodent carcinogens such as safrole, hexamethylphosphoramide (HMPA) and the pyrrolizidine alkaloid isatidine, which give negative results in other in vitro genotoxicity tests, positive results were obtained in Hep G2/SCGE assays. Nitrosomethylurea (NMU) was the only directly acting compound tested in the study and was by far (ca. 10(3)-fold) more active than the corresponding nitrosamine. The exposure concentrations required to cause significant effects varied over a broad range. The most pronounced effect was seen with AFB(1) (0.008 microM) followed by HMPA (15 microM), B(a)P (25 microM), NMU (100 microM), isatidin (500 microM), CP (900 microM), IQ (1200 microM), safrol (4000 microM), and NDMA (90x10(3) microM). Numbers in parenthesis give the lowest concentrations, which caused a significant increase of DNA migration. With two compounds, namely, the non-carcinogen pyrene and the synthetic hormone tamoxifen (TF), negative results were obtained under all test conditions. These findings are in agreement with the results of recent investigations which indicated that human hepatocytes are unable to convert TF to DNA reactive metabolites, whereas it is activated by rat liver cells and causes DNA adducts in these cells. Comparisons of the present results with data from earlier experiments indicate that the Hep G2/SCGE assay enables the detection of genotoxins including compounds which give misleading results in other in vitro genotoxicity tests and appears to be an alternative to tests with primary liver cells from laboratory animals.
Uhl
M
M
Institute of Tumor Biology and Cancer Research, University of Vienna, Austria.
Helma
C
C
Knasmüller
S
S
D001711:Biotransformation; D002273:Carcinogens / Q000378:metabolism / Q000633:toxicity*; D006528:Carcinoma, Hepatocellular / Q000201:enzymology / Q000473:pathology*; D002648:Child; D020552:Comet Assay; D004249:DNA Damage; D004273:DNA, Neoplasm / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D006801:Humans; D008113:Liver Neoplasms / Q000201:enzymology / Q000473:pathology*; D008297:Male; D009153:Mutagens / Q000378:metabolism / Q000633:toxicity*; D011237:Predictive Value of Tests; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000201:enzymology
D002273:Carcinogens; D004273:DNA, Neoplasm; D009153:Mutagens