Use of four new human-derived liver-cell lines for the detection of genotoxic compounds in the single-cell gel electrophoresis (SCGE) assay.
One of the main problems of in vitro genotoxicity assays is that the lack of adequate representation of drug-metabolising enzymes in indicator cell lines that are currently used in routine testing may lead to false results. In the present study, we investigated the ability of four new human-derived livercell lines to detect the DNA-damaging effects of representatives of different classes of genotoxic carcinogens that require metabolic activation, namely the nitrosamine N-nitrosodimethylamine (NDMA), the heterocyclic aromatic amines 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1), the polycyclic aromatic hydrocarbon benzo(a)pyrene (B(a)P) and the mycotoxin aflatoxin B1 (AFB1). Hydrogen peroxide (H2O2) was used in all experimental series as a positive control and parallel experiments were carried out with human HepG2 cells, which have been used in earlier studies. DNA damage was monitored in single-cell gel electrophoresis (SCGE) assays. Furthermore, RT-PCR experiments were carried out to study the expression of genes encoding for a panel of different phase-I and phase-II enzymes, which are involved in the activation/detoxification of genotoxic carcinogens. With one of the newly isolated hepatocellular lines, HCC1.2, positive results were obtained with all model compounds, two other new lines (HCC2 and HCC3), HepG2 and the virally immortalized line NKNT-3 were less sensitive and/or failed to detect some of the genotoxins. PCR analyses showed that all cell lines express genes coding for a variety of xenobiotic drug-metabolising enzymes. The highest levels were found in general in HCC1.2, while in NKNT-3 cells some genes were not transcribed. Overall, our results indicate that the line HCC1.2 may be useful for the development of improved in vitro genotoxicity test systems.
Winter
Heike K
HK
Institute of Cancer Research, Medical University of Vienna, Borschkegasse 8a, A-1090 Vienna, Austria.
Ehrlich
Veronika A
VA
Grusch
Michael
M
Lackner
Andreas
A
Schulte-Hermann
Rolf
R
Grasl-Kraupp
Bettina
B
Mikulits
Wolfgang
W
Knasmüller
Siegfried
S
D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism; D002455:Cell Division; D002460:Cell Line; D045744:Cell Line, Tumor; D020552:Comet Assay; D006801:Humans; D008099:Liver / Q000166:cytology*; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism; D009153:Mutagens / Q000633:toxicity*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D009153:Mutagens