Overexpression of bcl-2 protects hepatoma cell line HCC-9204 from ethanol-i nduced apoptosis

Objective To investigate the effect of overexpression of bcl-2 on ethanol-induced apopto sis of primary hepatocellular carcinoma (HCC) cells. Methods The retrovirus expression vector pDOR-SB containing human bcl-2 cDNA was intro duced into a human HCC cell line HCC-9204 by liposome-mediated transfection. pDOR-transfected and non-transfected HCC-9204 cells were used as controls. T he expression of Bcl-2 protein by transfected HCC-9204 cells was detected by t he immunohistochemical method. Then the cells were cloned with the limited dilu tion method continually until a monoclonal cell strain whose positive rate of Bc l-2 protein was 100% detected by flow cytometry was obtained. The killing rat es of cells were detected by Methabenzthiazuron assay after the treatment of 6% ethanol for 6 h. The extent of apoptosis was analyzed by transferase-mediate d dUTP nick end labeling (TUNEL) staining and flow cytometry. Results Most of the pDOR-SB-transfected cells demonstrated Bcl-2 positive signals, wh ile no signal was found in the controls. The positive rate of Bcl-2 protein de tected by flow cytometry in the obtained monoclonal cell strain, which was named HCC-bcl2, was 100% after the cells had been cloned 3 times continually. The k illing rate, TUNEL index and the scale of sub-G1 apoptotic peak in HCC-bcl2 cells were all significantly lower than those in the control cells. Conclusion Overexpression of Bcl-2 protein suppresses ethanol-induced apoptosis of the HC C cell line HCC-9204.