Effects of pEGFP-Bubl-shRNA plasmid on cell cycle and paclitaxel-sensitivity in human ovarian cancer cell line A2780 / 中国癌症杂志

ABSTRACT

Background and purpose:Previous studies have shown that Bubl was a critical component of the spindle checkpoint.Paclitaxel sensitivity was considered to be dependent on the functionality of this spindle checkpoint.This study investigated the effects of pEGFP-Bubl-shRNA plasmid stably transfected into the cell cycle and its sensitivity in human ovarian cancer cell line A2780.Methods:After the pEGFP-Bubl-shRNA plasmid and empty plasmids were constructed.they were transfected into A2780 cells by the Lipofectamine 2000~(TM).The nontransfected cells were the control.RT-PCR and Western blotting were used to determine the target gene and protein expression.The rate of proliferation inhibition was tested by an MTT assay,apoptosis and cell cycles were determined by flow cytometry,and the mitotic index was determined bv Hoechst33342 dye.Results:RT-PCR and Western blotting showed that pEGFP-Bubl-shRNA/A2780 group displayed a low expression of Bubl compared to the A2780 and pEGFP-Cl/A2780 group(P＜0.05).The sensitivity of the pEGFP-Bubl-shRNA/A2780 group was significantly lower than the non-transfccted and pEGFP-Cl/A2780 cells(P＜0.05).Flow cytometry revealed that the rates of G2/M phase and apoptosis were significantly lower in the pEGFP-Bubl-shRNA/A2780 group than those in the control group (P＜0.05).Conclusion:Bubl plays an important role in the paclitaxel treatment.A down-regulation of Bubl could reduce the drug sensitivity and rate of G_2/M cells in human ovarian cancer cell line A2780.