Overexpression of bcl-2 protects hepatoma cell line HCC-9204 from ethanol-induced apoptosis.
OBJECTIVE
To investigate the effect of overexpression of bcl-2 on ethanol-induced apoptosis of primary hepatocellular carcinoma (HCC) cells.


METHODS
The retrovirus expression vector pDOR-SB containing human bcl-2 cDNA was introduced into a human HCC cell line HCC-9204 by liposome-mediated transfection. pDOR-transfected and non-transfected HCC-9204 cells were used as controls. The expression of Bcl-2 protein by transfected HCC-9204 cells was detected by the immunohistochemical method. Then the cells were cloned with the limited dilution method continually until a monoclonal cell strain whose positive rate of Bcl-2 protein was 100% detected by flow cytometry was obtained. The killing rates of cells were detected by Methabenzthiazuron assay after the treatment of 6% ethanol for 6 h. The extent of apoptosis was analyzed by transferase-mediated dUTP nick end labeling (TUNEL) staining and flow cytometry.


RESULTS
Most of the pDOR-SB-transfected cells demonstrated Bcl-2 positive signals, while no signal was found in the controls. The positive rate of Bcl-2 protein detected 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 killing 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.


CONCLUSIONS
Overexpression of Bcl-2 protein suppresses ethanol-induced apoptosis of the HCC cell line HCC-9204.
Yang
Lianjun
L
Department of Pathology, Fourth Military Medical University, Xi'an 710032, China. yangsi@sina.com
Si
Xiaohui
X
Wang
Wenliang
W
D017209:Apoptosis / Q000187:drug effects*; D006528:Carcinoma, Hepatocellular / Q000378:metabolism / Q000473:pathology*; D000431:Ethanol / Q000494:pharmacology*; D005434:Flow Cytometry; D006801:Humans; D020287:In Situ Nick-End Labeling; D008113:Liver Neoplasms / Q000378:metabolism / Q000473:pathology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000502:physiology*; D014162:Transfection; D014407:Tumor Cells, Cultured
D019253:Proto-Oncogene Proteins c-bcl-2; D000431:Ethanol