沉默Sema6A基因对骨肉瘤细胞U-2OS增殖和转移的影响

Effects of siRNA-mediated Semaphorin 6A silencing on proliferation and migration of human osteosaroma U-2OS cells

目的检测信号素6A(Sema6A)基因在骨肉瘤细胞系中的表达情况,探讨RNA干扰Sema6A基因对骨肉瘤U-2OS细胞增殖、侵袭及迁移的影响。方法实时定量逆转录聚合酶链反应(qRT-PCR)检测骨肉瘤细胞株U-2OS、Saos-2及MG-63中Sema6A的mRNA表达水平。设计并合成针对Sema6A的小干扰RNA(siRNA)3条及阴性对照siRNA,在Lipofectamine RNAi MAX介导下转染U-2OS细胞,通过qRT-PCR检测Sema6A mRNA水平表达及蛋白质印迹法检测Sema6A蛋白水平,进而筛选出转染效率高的siRNA,CCK-8检测各组细胞活力,Transwell细胞侵袭和迁移实验检测各组细胞的侵袭及迁移能力。结果骨肉瘤细胞株中,Sema6A mRNA在U-2OS中表达最高。瞬时转染Sema6A siRNA的U-2OS细胞中Sema6A的mRNA及蛋白水平较阴性对照组均下降,与阴性对照组相比,实验组细胞的增殖、侵袭及迁移能力均增强,差异有统计学意义(P<0.05)。结论 RNA干扰沉默Sema6A基因可以增强U-2OS细胞的增殖、侵袭及迁移能力。

Objective To detect the expression of Semaphorin 6A gene in osteosaroma cell lines,and to explore the effect of RNA interference Sema6 A gene on proliferation,invasion and migration of osteosaroma U-2OS cells. Methods Small inference RNA targeting different regions of Sema6 A gene were transfected into U-2OS cells by Lipofectamine RNAimax. The most effective siRNA in knockdown Sema 6A was screened by quantitative PCR and Western blotting. The changes of cell proliferation,migration and invasion ability after Sema 6A knockdown were examined by CCK-8,Transwell cell invasion and migration assays. Results In osteosaroma cell lines,mRNA Sema6 A expressed the highest in U-2OS. The mRNA and protein levels of Sema6 A in siRNA U-2OS cells by Transient transfection were significantly lower than the negative control group（ P〈 0. 05）. Compared with the negative control group,proliferation,invasion and metastasis of cells in the experimental group increased significantly（ P〈 0. 05）. Conclusion Down-regulation of Sema6 A by siRNA can significantly enhance the proliferation and migration of U-2OS cells.