Abstract
The acute administration of ethanol to intestinal epithelial cells causes increased intestinal permeability and the translocation of endotoxins. The changes caused by ethanol in intestinal cells may be related to oxidative stress and DNA damage. However, DNA damage and repair-related molecules which act against stresses, including ethanol, have not been fully investigated in intestinal cells. Heat shock proteins (Hsps) are involved in the recovery and protection from cell damage and may be associated with DNA repair. Therefore, the aim of our study was to investigate cytotoxicity, DNA damage and the expression of DNA repair-related molecules, antioxidant proteins and Hsps in intestinal cells exposed to ethanol. Human intestinal Caco-2 cells were incubated with 1-8% ethanol for 1 h. Cell viability and DNA damage were determined using the MTT and comet assays, respectively. We measured DNA repair-related molecules, including DNA polymerase β, apurinic/apyrimidinic endonuclease/redox factor-1 (APE/Ref-1), growth arrest and DNA damage 45α (GADD45α) and proliferating cell nuclear antigen (PCNA), in Caco-2 cells using western blot analysis. We also measured glutathione peroxidase-1 (GPx-1), peroxiredoxin-1 (PRX-1), superoxide dismutase-2 (SOD-2), Hsp10, Hsp27, Hsp60, heat shock cognate (Hsc)70, Hsp70 and Hsp90. The viability of the Caco-2 cells exposed to ethanol decreased at concentrations ≥ 7% (P<0.05). The Olive tail moment, indicating DNA damage, increased dose dependently in ≥ 3% ethanol (P<0.05). Among the DNA repair proteins, the expression of PCNA and APE/Ref-1 increased significantly at 1% ethanol. Antioxidant enzymes, including GPx-1, PRX-1 and SOD-2, had an increased expression at 1% ethanol. Hsp10, Hsp27 and Hsp70 expression also increased significantly at 1% ethanol. In conclusion, the expression of DNA repair molecules, antioxidants and Hsps increased in intestinal Caco-2 cells exposed to low concentrations of ethanol. In particular, PCNA, APE/Ref-1, Hsp10, Hsp27 and Hsp70 were sensitive to low ethanol concentrations, indicating that they may be useful in evaluating the DNA repair and cytoprotective effects of the drug against stress in intestinal cells.