Effect Of Silencing E2F-1 Gene With SiRNA On Expression Of Rb Protein In MGC803 Cells

Objective To investigate the effect of silencing E2F-1 gene expression with siRNA on the tumor cell line MGC803 and Rb gene protein expression.Methods Human gastric carcinoma cell line MGC803 was cultured in vitro and transfected with siRNA of E2F-1 by lipofectamine2000 transient-transfection assay.After 48 hours, the levels of E2F-1 mRNA expression before and after transfection were detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). The levels of E2F-1 protein expression before and after transfection were detected by western blot.The levels of total Rb protein and Rb phosphorylation protein expression before and after transfection were also detected by western blot. It could compute the dephosphorylation of Rb protein expression for the further analysis.Results After plasmid amplification, extraction, the level of E2F-1 mRNA in MGC803 cells was inhibited by the specific siRNA. The expression of E2F-1 mRNA at 48h after transfection was apparent difference from normal MGC803 cells and negative control cells. Western-blot indicated that the expression of E2F-1 protein was upregulated in MGC803 cells after interfered with siRNA (83.2%, 84.6%, p<0.05) while the dephosphorylation of Rb protein expression was upregulated (61.22%, 66.60%, p<0.05).Conclusion RNA interference can downregulate the level of E2F-1 mRNA and E2F-1 protein expression in MGC803 cells, downregulate the activity of the dephosphorylation of Rb protein. There may be some antagonistic effects between E2F-1 gene and Rb gene. E2F-1 may be used as a gene therapy target on gastric carcinoma .The mechanism of gastric carcinoma will be investigated furthermore.