Synergistic effect of IFN-γ gene on LIGHT-induced apoptosis in HepG2 cells via down regulation of Bcl-2.
To detect the expression of anti-apoptotic factor Bcl-2 and Survivin in transferred HepG2 cells and evaluate the synergistic effect of IFN-γ gene on LIGHT-induced apoptosis signal transduction pathways, the full-length ORF of LIGHT and IFN-γ gene were cloned into pcDNA4 and verified by DNA sequencing. After being optimized by EGFP, recombinant LIGHT and IFN-γ were transferred into the HepG2 cells mediated by a cationic liposome in vitro. The expression of LIGHT and IFN-γ was identified in the supernatants by ELISA. The HepG2 cells were divided into three groups: the control, LIGHT gene transfection alone, and simultaneous transfection of LIGHT and IFN-γ genes. The cell apoptosis and expression of Bcl-2 and Survivin in cell lysate were detected through FCM. After transfection, the apoptosis rate of HepG2 cells was increased with the prolonged time, and the apoptosis rate of LIGHT group was higher than the control group, while the LIGHT/IFN-γ group was higher than the LIGHT group P < 0.01). The expression of Bcl-2 and Survivin in LIGHT group and LIGHT/IFN-γ group decreased dramatically compared with the control group. LIGHT gene alone can result in significant inhibition of HepG2 cells proliferation. INF-γ can synergistically precede LIGHT-induced apoptotic processes through down-regulation of Bcl-2 expression, but not survivin expression.
Department of Hepatobiliary Surgery, Affiliated Hospital of Medical College, Qingdao University, Qingdao, Shandong, China.
Bing
B
Han
Li-Qun
LQ
Wu
Xiang
X
Ma
Zheng-Hua
ZH
Wang
Jin-Peng
JP
Li
Chong-Yao
CY
Bi
Sun
S
Yong
D017209:Apoptosis / Q000235:genetics; D015536:Down-Regulation; D056945:Hep G2 Cells; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000378:metabolism; D007371:Interferon-gamma / Q000235:genetics* / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D000077022:Survivin; D014162:Transfection; D053326:Tumor Necrosis Factor Ligand Superfamily Member 14 / Q000378:metabolism*
C107489:BIRC5 protein, human; D051034:Inhibitor of Apoptosis Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D000077022:Survivin; D053326:Tumor Necrosis Factor Ligand Superfamily Member 14; D007371:Interferon-gamma