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目的:运用小干扰RNA下调果蝇zeste基因增强子人类同源物(enhancer ofzeste homolog 2,EZH2)在肾癌细胞系769-P中的表达,明确其对肾癌细胞增殖的影响。方法:将处于对数生长期769-P细胞分为实验组(experiment group)、阴性对照组(negative group)、空白对照组(blank group),合成靶向EZH2基因的小干扰RNA片段(EZH2-siRNA)和无效序列片段后,通过脂质体介导分别转染至实理组和阴性对照组,空白对照组未做任何处理。以qRealtime-PCR检测EZH2基因mRNA水平的变化情况,以MTT法检测各组细胞增殖变化;流式细胞术(FCM)检测转染后细胞周期变化情况。结果:实理组中EZH2在mRNA表达水平明显受抑制;MTT实验中第4天始,实验组中769-P细胞的增殖能力开始受抑制,第5天时实验组细胞抑制更明显,与阴性对照组和空白组比较差异有统计学意义(P<0.05)。siRNA转染后实验组中G0/G1期细胞比例明显增多(81.32±3.14)%,与阴性对照组(44.13±1.52)%和空白对照组(45.71±2.32)%差异有统计学意义。结论:EZH2-siRNA可有效下调并抑制肾癌细胞769-P的增殖,EZH2在肾癌的发生、发展中发挥了重要作用,为下一步研究肾癌基因治疗提供了理论支持。
OBJECTIVE: To investigate the effect of enhancer ofzeste homolog 2 (EZH2) on the expression of 769-P in renal cell carcinoma cell line 769-P by using small interfering RNA (RNAi) to determine its effect on the proliferation of renal cell carcinoma. Methods: The 769-P cells in the logarithmic growth phase were divided into experiment group, negative group, blank group and EZH2- siRNA) and invalid sequence fragments were transfected by Lipofectamine into the real control group and the negative control group, but the blank control group did not do any treatment. The changes of mRNA level of EZH2 gene were detected by qRealtime-PCR. The proliferation of EZH2 gene was detected by MTT assay. The cell cycle was detected by flow cytometry (FCM). Results: The mRNA expression of EZH2 was significantly inhibited in the experimental group. On day 4 of MTT experiment, the proliferation of 769-P cells in the experimental group began to be inhibited. On the fifth day, the cell inhibition was more obvious in the experimental group than in the negative control The difference between the two groups was statistically significant (P <0.05). The percentage of cells in G0 / G1 phase increased significantly (81.32 ± 3.14)% after siRNA transfection compared with the control group (44.13 ± 1.52%) and the blank control group (45.71 ± 2.32)%. Conclusion: EZH2-siRNA can effectively down-regulate and inhibit the proliferation of renal carcinoma cell line 769-P. EZH2 plays an important role in the occurrence and development of renal cell carcinoma, which provides theoretical support for further study of gene therapy of renal cell carcinoma.