Survivin在NF-κB圈套寡核苷酸促进肝癌细胞HepG2凋亡中的作用

Effects of Survivin on the NF-κB Decoy Oligonucleotides Inducing Apoptosis in Hepatic Cancer Cells HepG2

目的观察核转录因子-κB(NF-κB)圈套寡核苷酸对NF-κB活性的影响,研究Survivin在其诱导肝癌细胞HepG2凋亡中的作用。方法将异硫氰酸荧光素(FITC)标记的NF-κB圈套寡核苷酸转染至HepG2细胞中,荧光倒置显微镜和共聚焦显微镜进行核内定位观察,凝胶迁移率实验(electrophoretic mobility shift assay,EMSA)检测转染前后NF-κB活性变化;流式细胞仪和TUNEL检测细胞凋亡率;Western blot检测凋亡抑制蛋白Survivin的改变。结果NF-κB圈套寡核苷酸转染后定位于细胞核内,EMSA显示转染后NF-κB活性明显下降。与未处理组、转染错义组相比,转染NF-κB圈套寡核苷酸组的HepG2细胞凋亡率增加,Survivin蛋白表达下调[(39.8±1.9)、(42.7±2.5)vs(20.1±3.1)]。结论NF-κB圈套寡核苷酸具有促进肝癌细胞HepG2凋亡的作用,其机制可能与下调Survivin的表达有关。

Objective To investigate the effects of Survivin on the NF-κB decoy oligodeoxynucleotides（ODNs） inducing apoptosis in hepatic cancer cells HepG2.Methods FITC-labeled NF-κB decoy ODNs were transfected into HepG2 with LipofectamineTM2000.The activity of NF-κB before and after transfection was measured by electrophoretic mobility shift assay（EMSA）.The apoptosis of cells was detected by flow cytometry and TdT mediated dUTP-biotin nick end labeling（TUNEL）,and the expression of Survivin by Western blot.Results 1 h after FITC-labeled decoy ODNs were transfected,the decoy ODNs were detected in the nuclei of HepG2 cells.EMSA revealed the significant reduction of NF-κB expression.As compared with control group,apoptosis rate in NF-κB decoy ODNs group was increased in HepG2 cells,and the expression of Survivin protein was down-regulated.Conclusion The NF-κB decoy ODNs can accelerate the apoptosis of hepatic cancer cells HepG2 by down-regulating the expression of Survivin.