Survivin基因干扰RNA对骨肉瘤MG-63细胞系增殖和凋亡的影响

Effect of a short hairpin RNA targeting Survivin on the proliferation and apoptosis in osteosarcoma cell line MG-63

[目的]观察Survivin基因siRNA表达载体对骨肉瘤细胞增殖和凋亡的影响。[方法]体外构建SurvivinshRNA表达载体pS ilence 2.1-neo-Survivin,转染骨肉瘤细胞系MG-63,筛选稳定表达的克隆,倒置显微镜观察各组细胞形态学变化,RT-PCR、W eston-b lot方法检测转染后Survivin mRNA及蛋白的表达,噻唑蓝(MTT)法、克隆形成实验检测细胞的增殖情况,吖啶橙荧光染色法观察细胞凋亡情况。[结果]转染后MG-63细胞Survivin基因mRNA水平和蛋白表达显著下降,其抑制率分别为85.08%和81.14%;细胞增殖受到显著抑制,培养48 h后与空白组比较,抑制率达63.41%;吖啶橙染色显示转染组细胞出现明显核碎裂等凋亡改变,转染组凋亡率为24.54%。[结论]靶向Survivin基因的siRNA表达载体可以显著抑制骨肉瘤细胞增殖,促进细胞凋亡。

[ Objective] To construct an expression plasmid of a short hairpin RNA targeting Survivin, and investigate its effect on the expression of Survivin, the proliferation and the apeptosis of human osteosarcoma cell line MG-63 in vitro. [ Method ] A plasmid of a short hairpin RNA targeting Survivin was constructed, and it was transfered into MG-63 cell line by dosper lipesomal method. Morphological change of tumor cells was observed under invert microscope. The expression of Survivin mRNA and protein were examined by RT-PCR and Weston-blot method respectively. The inhibition of the cell proliferation was estimated by MTT method and the colony forming test. The apeptotic cells were stained by acridine orange. [ Result] The number of drift cells was increased after transfection, Survivin shRNA could significantly reduce the expressions of Survivin mRNA and protein The expression of Survivin mRNA and protein after transfection was respectively inhibited by 85.08% and 81.14%, Survivin shRNA also inhibit the growth of the cell by MTT method and the inhibition rate reached 63.41% at 48 h, the apoptosis rate of the cells in the transfected group was 24. 54%. [ Conclusion] Survivin shRNA could significantly reduce the expressions of Survivin mRNA and protein and inhibit the proliferation of the MG-63 cell, and increase the apeptosis of the cells.