The Killing Effects Of HSV-tk Gene Mediated By ApoE Modified Liposome On Hepatocellular Carcinoma Cells

Objectives:1. To evaluate the selective affinity of the cationic liposome modified by apo E on hepatocyte and hepatocellular carcinoma cells.2. The plasmid p Survivin-TK-PBI-CMV2 containing suicide gene HSVtk was constructed. Apo E-lipoplex carrying p Survivin-TK-PBI-CMV2 plasmid was used to transfect hepatocyte and hepatocellular carcinoma cells, the HSVtk gene expression was detected.3. To detect the selective killing effects of apo E-lipoplex/HSVtk/GCV system on hepatocellular carcinoma cells.Methods:1. Apo E-lipoplex carrying the plasmid p Genesil-1 was transfected into the hepatocyte cell HL-7702, hepatocellular carcinoma cells Hep G2 and HUh-7, cervical cancer cell Hela.Transfection efficiency was detected by flow cytometry.2. p Survivin-TK-PBI-CMV2 plasmid carried by apo E-lipoplex was transfected into Hep G2, Huh-7, HL-7702 respectively. The m RNA expression of HSVtk gene was detected by RT-PCR, and the HSVtk protein expression was detected by western blot.3. Following the transfection of p Survivin-TK-PBI-CMV2 into Hep G2, Huh-7 and HL-7702 cells, GCV was added into the culture medium. Two days later the cell apoptosis rates were examined by FCM, cell proliferaton was examined by MTT, cell morphology was observed using fluorescence microscope after Hoechst staining.Results:1. Compared with the Hela cell group, hepatocyte cell and hepatocellular carcinoma cells groups had higher transfection rates by the laboratory synthesized apo E-lipoplex(P <0.05).2. p Survivin-TK-PBI-CMV2 plasmid carried by apo E-lipoplex was transfected into Hep G2, Huh-7, HL-7702, the HSVtk m RNA and protein were found specially expressed in the hepatocellular carcinoma cells Hep G2 and Huh-7, but not in the hepatocyte HL-7702.3. The FCM results showed that the transfected hepatocellular carcinoma cells Hep G2 and Huh-7 had higher apoptosis rates(P < 0.05). The MTT results showed that the transfected Hep G2 and Huh-7 cells had slower cell proliferation(P < 0.05). Hoechst staining under fluorescence microscope showed characteristic morphology of apoptosis in transfected Hep G2 and Huh-7 cells.Conclusions:1. The apo E-lipoplex had higher transfection efficiency toward hepatocyte and hepatocellular carcinoma cells, which showed the targeted transfection ability of apo E-lipoplex.2. HSVtk gene driven by survivin promoter was specially expressed in hepatocellular carcinoma cells Hep G2 and Huh-7.3. apo E-lipoplex+ p Survivin-TK-PBI-CMV2+GCV system had targeted and evident killing effects on hepatocellular carcinoma cells Hep G2 and Huh-7,whereas not on the hepatocyte HL-7702.