Growth-inhibiting Effects Of Cinobufacini Combined With RRM2 SiRNA On Endometrial Cancer Ishikawa Cells

Objective: To investigate the effect of cinobufacini associated with RRM2 siRNA on the growth of the endometrial cancer Ishikawa cells cultured in vitro.Methods: Small interfering RNA (siRNA) targeting RRM2 gene was designed, then ds-siRNA was transfected in to Ishikawa cells with Lipofectamine2000 . human endometrial cancer cell lines Ishikawa were divided into four different treatment groups: control, RRM2 siRNA, cinobufacini and RRM2 siRNA+ cinobufacini. The effect on cellular proliferation was analyzed by MTT assay and the transfection efficiency, cell cycle and apoptosis were measured by flow cytometry(FCM).The expressions of RRM2 on mRNA and protein level were analyzed by RT-PCR and Wesern blotting respectively.Ability of invasion was measured by transwell chamber model.Results: The siRNA targeting human RRM2 gene effectively inhibited the proliferation of Ishikawa cells, and the inhibitory action was increased when RRM2 siRNA associated with cinobufacini. 2mg/ml cinobufacini was the best inhibiting concentration. RRM2 siRNA associated with cinobufacini moderately increased apoptosis rate compared to either treatment alone, the apoptosis rates of the RRM2 siRNA , 2mg/ml cinobufacini and RRM2 siRNA+2mg/ml cinobufacini were 44.3%,42.0%,71.1% , respectively. Treatment with RRM2 siRNA and cinobufacini also produced an increase in the G₁ phase and a decrease in the G₂ and S phase ,decreased the expression of RRM2 on both mRNA and protein level and reduced cell invasion.But these effects did not appear in scrambled siRNA (N-siRNA) control group.Conclusion: The siRNA targeting human RRM2 gene could reduce RRM2 expression, inhibit cellular proliferation and invasion ,induce apoptosis and produce growth retardation of Ishikawa cells in vitro.