Ethanol enhanced the genotoxicity of acrylamide in human, metabolically competent HepG2 cells by CYP2E1 induction and glutathione depletion.
In the present study, the genotoxicity of acrylamide (AA) was investigated in HepG2 cells using SCGE. Additionally, the influence of ethanol on the modulation of AA-induced DNA-migration caused by CYP2E1-upregulation and/or GSH-depletion was examined in the same cell line. For the ethanol/AA combination assays, the cells were treated with ethanol for 24h prior to exposure to 5mM AA for another 24h. 1.25 to 10mM AA-induced DNA migration (OTM) in HepG2 cells in a concentration-dependent manner, e.g., exposure to 10mM AA, resulted in an 8-fold increase of DNA migration compared to the negative control. Treatment with 120mM ethanol prior to exposure to 5mM AA increased the level of DNA migration more than 2-fold as compared to cells treated with 5mM AA alone. Immunoblotting showed a clear ethanol-induced increase of CYP2E1, which plays a pivotal role in AA toxification. Additionally, intracellular GSH levels were significantly reduced after ethanol or AA treatment. In the ethanol/AA combination experiments, GSH depletion was comparable to the additive effect of the single compounds. No induction of apoptosis (ssDNA assay), but necrosis was identified as responsible for the reduction of viability with increasing compound concentration. The data clearly show a higher genotoxic potential of ethanol/AA combination treatment compared to AA treatment alone. In conclusion, both the ethanol-mediated induction of CYP2E1 and the depletion of GSH provide a mechanistic explanation for the over-additive effects of ethanol and AA. Even though the concentrations used in this study were rather high, consequences for the dietary intake of AA-containing food and alcoholic beverages should be discussed.
Lamy
Evelyn
E
Institute of Indoor and Environmental Toxicology, University of Giessen, Aulweg 123, 35385 Giessen, Germany. evelyn.lamy@uniklinikum-giessen.de
Völkel
Yvonne
Y
Roos
Peter H
PH
Kassie
Fekadu
F
Mersch-Sundermann
Volker
V
D020106:Acrylamide / Q000633:toxicity*; D000428:Alcohol Drinking / Q000009:adverse effects*; D017209:Apoptosis / Q000187:drug effects; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D020552:Comet Assay; D019392:Cytochrome P-450 CYP2E1 / Q000187:drug effects / Q000378:metabolism*; D004249:DNA Damage; D004785:Environmental Pollutants / Q000633:toxicity*; D000431:Ethanol / Q000633:toxicity*; D005978:Glutathione / Q000187:drug effects / Q000378:metabolism*; D006801:Humans
D004785:Environmental Pollutants; D020106:Acrylamide; D000431:Ethanol; D019392:Cytochrome P-450 CYP2E1; D005978:Glutathione