Abstract:

Abstract:Objective
To investigate the inhibitory effect of siRNA interfering E2F3 gene  on prolifera
tion of bladder cancer 5637 cells,and clarify  the biomechanism of inhibitory effect of   E2F3 gene  silencing  on the proliferation of  5637 cells.Methods There pairs of  nucleotide chains coding short RNA sequence and  targeting E2F3 gene were chemically synthesized and cloned into pRNAT-U6.1/Neo digested  with BamHⅠand Hind Ⅲ． RNAi plasmid(pRNAT-U6.1-E2F3/Neo) was constructed．The reconstructed RNAi plasmids were identified by electrophoresis  and  confirmed by sequencing analysis. The recombinant plasmids were transformed into strain DH5α, and the recombinant pRNAT-U6.1/Neo was transfected into 5637 cells. Before and after transfection, the levels of E2F3 mRNA and E2F3 protein expression were detected by RT-PCR and Western blotting,the  cell cycle was assessed by flow cytometry, the  apoptotic rate was detected by Annexin-V/PS kit. Results Compared with control group,the  E2F3 gene expression was markedly down-regulated in 5637 cells following RNA interference treatment( P<0.001), the percentage of cells at G1 phase and the apoptotic rate were up-regulated in transfection cells group,there were significant difference between RNAi cells groups and control group（P<0.001）.  Conclusion siRNA can significantly  inhibit the E2F3 gene expression in bladder cancer 5637 cells, decrease cell proliferation , arrest cell cycle and increase apoptotic rate.

Key words: RNA interference, small interfering, E2F3;bladder neoplasms, suppression of cell proliferation