靶向抑制UBE2C基因对SMMC-7721细胞生物学行为的影响

Effects of UBE2C inhibition on the biological behaviour of SMMC-7721 cell line

目的将构建好的UBE2C干扰质粒转入SMMC-7721细胞中,观察其对SMMC-7721细胞生物学行为的影响。方法采用脂质体法将构建好的UBE2C干扰质粒转入SMMC-7721细胞株中,潮霉素筛选4周,并经Western blot鉴定,选出干扰效果最佳的低表达稳定株(shUBE2C组)用于后续实验。用MTT法检测细胞增殖,流式细胞仪检测细胞周期,软琼脂克隆形成实验检测克隆形成能力。结果 Western blot检测显示,转shUBE2C组UBE2C的蛋白表达水平低于转阴性对照shNC组(P<0.05)。shUBE2C组的SMMC-7721细胞生长曲线明显右移,显示靶向抑制UBE2C可降低SMMC-7721细胞的增殖能力。与shNC组相比,shUBE2C组细胞的G1期细胞比例明显增高(P﹤0.05),而S期的比例明显降低(P﹤0.05),提示其延缓细胞周期进程。shUBE2C组的克隆形成率明显低于shNC组(P﹤0.05),提示其降低克隆形成能力。结论 UBE2C低表达可改变SMMC-7721细胞的生物学行为。

To observe the effects of UBE2C inhibition on the biological behaviour of SMMC-7721 cell line, a UBE2C interference plasmid was constructed and transfected into SMMC-7721 cell line by lipofeetamine 2000. After screened with G418, the expression of UBE2C in protein level was detected by Western blot. Stable and bestinterfered cell line with low UBE2C expression （shUBE2C group） was picked out for follow-up experiments. We employed MTT assay to detect cell proliferation, flow cytometry to detect cell cycle, and soft agar colony formation assay to detect colony forming ability. Western blot showed that the protein expression of UBE2C in shUBE2C group was lower than that of negative control shNC group （P 〈 0.05）. SMMC-7721 cell growth curve in shUBE2C group was significantly shifted to the right, which showed shUBE2C could reduce the proliferation of SMMC-7721 cell line. Compared with shNC group, the percentage of SMMC-7721 cells in G1 phase of shUBE2C group was significantly higher （P 〈 0.05）, while the percentage of S phase was significantly lower （P 〈 0.05）, suggesting that shUBE2C could delay cell cycle progression. The colony formation rate of shUBE2C group was significantly lower than that of shNC group （P 〈 0.05）, suggesting that shUBE2C could reduce the colony-forming ability. All result indicated that low expression of UBE2C may change the biological behavior of SMMC-7721 cell line.