Eg5基因在神经母细胞瘤细胞增殖和凋亡中的作用

Role of Eg5 gene in proliferation and apoptosis of neuroblastoma cells

目的探讨Eg5基因对神经母细胞瘤(NB)细胞增殖和凋亡的影响。方法取2株NB细胞株SHSY-5Y和NBL-S,分别转染针对Eg5基因的小干扰RNA(siRNA),分为细胞对照组(用等体积的水代替siRNA,只含有脂质体,未转染细胞)、siRNA对照组(转染siRNA对照)和siRNA干扰组(转染Eg5 siRNA),每组6孔,每孔1×10^6个细胞。使用实时定量聚合酶链式反应(real-time PCR)检测Eg5 mRNA表达水平的变化;使用5-乙炔基-2’-脱氧尿苷(EdU)掺入实验和克隆形成实验检测下调Eg5表达对NB细胞增殖的影响;使用流式细胞术检测下调Eg5表达对NB细胞凋亡和细胞周期的影响。结果荧光定量PCR检测结果显示,siRNA干扰组Eg5 mRNA表达水平与细胞对照组和siRNA对照组相比显著减少,差异有统计学意义(P<0.05);EdU掺入实验和克隆形成结果表明,siRNA干扰组细胞中DNA的合成量和细胞克隆数量低于细胞对照和siRNA对照组,差异有统计学意义(P<0.05);流式细胞术检测结果显示,细胞对照组和siRNA对照组细胞的凋亡比率低于siRNA干扰组细胞的凋亡比率,差异有统计学意义(P<0.05);细胞周期的检测结果显示,细胞对照组和siRNA对照组细胞中G0/G1期所占比率<50%,而siRNA干扰组细胞中G0/G1期所占比率>70%,差异有统计学意义(P<0.05)。结论在NB细胞中使用siRNA下调Eg5表达,能够抑制NB细胞增殖,促进细胞凋亡。

Objective To investigate the effect of Eg5 gene on the proliferation and apoptosis of neuroblastoma cells.Methods The siRNA targeting Eg5 gene was transfected into NB cells SHSY-5Y and NBL-S,and the down-regulation level of Eg5 gene was detected by fluorescence real-time PCR.The changes of cell proliferation after down-regulation of Eg5 expression.were detected by EdU incorporation assay and clone formation assay.And the apoptotic level and cell cycle were assessed by flow cytometry.Results After transfection of siRNA targeting Eg5 gene into NB cells,real-time PCR showed that the expression level of Eg5 gene was significantly lower than that of cell control group and siRNA control group.EdU incorporation experiment and clone formation showed that the down-regulation of Eg5 expression in NB cells could inhibit DNA synthesis and proliferation.The results of flow cytometry showed that the down-regulation of Eg5 expression in NB cells could promote cell apoptosis,block cell cycle at G0/G1 phase and inhibit cell DNA synthesis and proliferation.Conclu sion The down-regulation of Eg5 expression by siRNA in NB cells can inhibit the proliferation of NB cells and promote cell apoptosis.