Effects Of RNAi Targeted To STAT3 On Apoptosis Induction And On STAT3 Signaling In Human Hepatocellular Carcinoma Cells

Objective:To investigate the eukaryotic expression of STAT3-targeting siRNAs (STAT3-siRNAs) and their effects on apoptosis in liver cancer cell line SMMC7721, in order to provide theoretical and experimental data for siRNA therapy of liver cancer.Methods:1:Three eukaryotic expression plasmids of STAT3-siRNA, STAT3-siRNA1, STAT3-siRNA2, and STAT3-siRNA3, were constructed to selectively silence the STAT3 gene in SMMC7721 cells. Laser confocal microscopy was used to observe the sub-cellular organel changes after transfection of the STAT3-siRNA expression plasmids into SMMC7721 cells.2:The expression of STAT3 protein in SMMC7721 cells was determined by Western bloting. The best STAT3-siRNA expressing plasmid was selected for the use of further study. 3:Cytoflowmetry was used to analyze the effects of STAT3-siRNA on the apoptosis of SMMC7721 cells.4:The morphological changes in SMMC7721 cells after transfection with STAT3-siRNA expression plasmids were observed by using transmission electron microscope.5:Expression levels of apoptosis-associated genes, survivin and bcl-2, were determined by using semi-quantitative RT-PCR, after transfection of the STAT3-siRNA expression plasmids into SMMC7721 cells.Results:1.48hours after STAT3-siRNA eukaryotic expression plasmids were transfected into SMMC7721 cells, expression of the green fluorescence protein was found by laser confocal microscopy in 67% of the SMMC7721 cells.2. Western bloting found that STAT3-siRNA significantly inhibited the expression of STAT3 proteins.3. After the STAT3-siRNA eukaryotic expression plasmid STAT3-siRNA1 was transfected into SMMC7721 cells, expression of the STAT3 mRNAs, including survivin and bcl-2, was found by semi-quantitative RT-PCR to be decreased when compared to the control group (P<0.01), and the apoptosis rate was significantly increased in the STAT3-siRNA1 transfected group(P<0.01).Conclusion:STAT3-siRNA eukaryotic expression plasmids were successfully transfected into SMMC7721 cells and the STAT3-siRNAs were significantly expressed in 48 hours. Our results suggested that STAT3-siRNA technology can effectively silence the STAT3 gene expression in SMMC7721 cells, resulting in apoptosis through down-regulateing the expression of bcl-2 and survivin genes.