The study on apoptosis and its molecular mechanism in mouse insulinoma cells induced by ethanol

Objective To investigate the effect of ethanol on apoptosis in mouse insulinoma cells(NIT-1 cells) and to evaluate the pathway associated with the regulation of Bcl-2,Bax and Caspase-3.Methods After NIT-1 cells were treated with ethanol at concentrations of 0,50,100,200 and 400 mmol/L for 24 h,single cell gel electrophoresis(SCGE) and AnnexinV/PI methods were performed to detect the apoptosis.After NIT-1 cells were exposed to various concentrations of ethanol (same as above) for 6,12 and 24 h,the levels of Bcl-2,Bax and Caspase-3 mRNA expression were determined by RT-PCR method.Results SCGE results showed that the low concentration of ethanol induced no obviously DNA damage in NIT-1 cells,but the rate of DNA migration,degree of DNA damage and average length of DNA migration of 200 and 400 mmol/L ethanol-treated groups were significantly changed compared to those of untreated group.The results of Annexin V/PI assay indicated that the apoptosis rates of NIT-1 cells in 200 and 400 mmol/L ethanol-treated groups were higher than that of untreated group.At 6 h,the ration of Bcl-2/Bax mRNA expression was increased first,then decreased with the increasing of ethanol dosage.At 12 h and 24 h,the ration was decreased.By treatment at 400 mmol/L,the decrease was dramatically compared to control group.Ethanol increased the caspase-3 mRNA expression at high concentrations and long exposure time.At 24 h,the caspase3 mRNA expression was increased significantly by treatment at 400 mmol/L.Conclusion Ethanol induced apoptosis in NIT-1 cells and the potential molecular mechanism probably relates with the Bcl-2 family and Caspse-3.