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目的 :探讨AURKA基因表达下调对骨肉瘤细胞增殖和细胞周期分布的影响。方法:脂质体瞬时转染法介导AURKA小干扰RNA(small interfering RNA,siRNA)处理骨肉瘤U2-OS细胞,采用实时荧光定量PCR及Western blot方法检测转染前后AURKA mRNA和蛋白表达,CCK8实验检测细胞活力变化,流式细胞术检测细胞周期分布变化,Western blot检测周期相关蛋白的表达,5-溴脱氧尿嘧啶核苷(Brd U)细胞增殖实验检测细胞增殖情况。结果:AURKA siRNA作用U2-OS细胞后,AURKA mRNA和蛋白的表达水平较正常对照组及阴性对照组显著降低,且差异有统计学意义(P<0.05),两对照组间AURKA表达无明显差异;下调AURKA表达后,细胞活力降低,且作用72 h后抑制率最高,约为(36.63±2.38)%;细胞周期中S期细胞比例下降,G2/M期细胞比例增加,与正常对照组及阴性对照组相比差异有统计学意义(P<0.05),提示存在G2/M期阻滞,S期细胞增殖率下降(P<0.05),周期相关蛋白D1(Cyclin D1)表达水平下降(P<0.05),周期相关蛋白B1(Cyclin B1)表达水平明显增加(P<0.05)。结论:下调AURKA表达可抑制骨肉瘤细胞U2-OS的增殖,同时导致细胞阻滞于G2/M期。
Objective: To investigate the effect of down-regulation of AURKA gene on the proliferation and cell cycle distribution of osteosarcoma cells. Methods: Transient transfection of liposomes mediated the expression of AURKA mRNA and protein in osteosarcoma U2-OS cells by small interfering RNA (siRNA), and the expression of AURKA mRNA and protein were detected by real-time fluorescence quantitative PCR and Western blot. The expressions of CCK8 Cell viability was detected by flow cytometry. Cell cycle distribution was analyzed by flow cytometry. Western blot was used to detect the expression of related proteins. BrdU proliferation assay was used to detect cell proliferation. Results: The expression of AURKA mRNA and protein in U2-OS cells after AURKA siRNA treatment was significantly lower than that in normal control group and negative control group (P <0.05). There was no significant difference in AURKA expression between the two control groups (P <0.05). After AURKA expression was down-regulated, the cell viability decreased and the highest inhibitory rate was reached at 72 h (36.63 ± 2.38)%. The percentage of cells in S phase decreased and the percentage of G2 / M phase increased in cell cycle. Compared with normal control group (P <0.05), suggesting that there was G2 / M arrest, decreased cell proliferation in S phase (P <0.05) and decreased expression of Cyclin D1 (P <0.05) <0.05), and the expression of cyclin B1 was significantly increased (P <0.05). Conclusion: Down-regulation of AURKA can inhibit the proliferation of osteosarcoma cells U2-OS, and lead to cell arrest in G2 / M phase.