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目的通过RNA干扰阻断T24细胞中Eg5基因的表达,研究Eg5基因对T24细胞增殖和迁移的影响。方法 T24细胞株分为3组:干扰组、空白对照组、阴性对照组。化学合成针对Eg5的小干扰RNA,通过脂质体转染至T24细胞中,应用Western blot法检测转染后T24细胞中Eg5蛋白表达,采用台盼蓝拒染实验检测细胞死亡率,用噻唑蓝(MTT)比色分析法和平板克隆形成实验检测细胞增殖抑制率,通过细胞划痕实验检测细胞的迁移能力。结果干扰组Eg5蛋白表达显著低于空白对照组和阴性对照组;干扰组细胞死亡率显著升高,与空白对照组及阴性对照组比较差异有统计学意义(P<0.05);干扰组的克隆形成率下降,与空白对照组和阴性对照组比较差异有统计学意义(P<0.05);MTT结果提示,干扰Eg5基因表达后,T24细胞的增殖率明显下降(P<0.05);划痕实验结果显示干扰组细胞迁移能力受到明显抑制(P<0.05)。结论 Eg5基因在膀胱移行细胞癌起重要作用,干扰Eg5基因能有效抑制T24细胞的增殖与迁移,促进细胞死亡,以Eg5为靶点有望成为膀胱癌基因治疗的新基因靶点。
Objective To investigate the effect of Eg5 gene on the proliferation and migration of T24 cells by blocking the expression of Eg5 gene in T24 cells by RNA interference. Methods T24 cell lines were divided into three groups: interference group, blank control group, negative control group. Small interfering RNA against Eg5 was chemically synthesized and transfected into T24 cells by liposome. The expression of Eg5 in transfected T24 cells was detected by Western blot. The cell death rate was detected by trypan blue exclusion assay. (MTT) colorimetric assay and plate clone formation assay to detect cell proliferation inhibition rate, cell migration assay by cell scratch assay. Results The expression of Eg5 in the interference group was significantly lower than that in the blank control group and the negative control group. The cell death rate in the interference group was significantly higher than that in the blank control group and the negative control group (P <0.05) (P <0.05). The results of MTT showed that the proliferation rate of T24 cells was significantly decreased (P <0.05) after interference with Eg5 gene expression; the scratch test The results showed that the interference ability of cell migration was significantly inhibited (P <0.05). Conclusion Eg5 plays an important role in transitional cell carcinoma of the bladder. Interruption of Eg5 can effectively inhibit the proliferation and migration of T24 cells and promote cell death. Eg5 targeting Eg5 is expected to be a new gene target for gene therapy of bladder cancer.