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目的细胞增殖是影响肾细胞癌进展的重要因素,本研究旨在探讨组蛋白赖氨酸甲基转移酶SET8在调节肾透明细胞癌786-O细胞增殖中的作用。方法将体外培养肾透明细胞癌786-O细胞随机分为空白对照组、空质粒组和SET8-shRNA组3组。采用四甲基偶氮唑蓝(MTT)法检测3组细胞的增殖能力;RT-PCR和蛋白质印迹法检测3组细胞Survivin和Caspase-3的表达水平,分析干扰SET8对肾透明细胞癌786-O细胞增殖及相关基因和蛋白表达的影响。结果 shRNA-SET8可成功抑制786-O细胞中SET8蛋白的表达。MTT检测结果显示,0、12、24、36和48 h SET8-shRNA组增殖抑制率分别为(10.97±2.68)%、(20.38±7.12)%、(25.03±7.60)%、(30.35±1.97)%和(31.33±1.04)%,SET8-shRNA组786-O细胞的增殖能力较空白对照组和空质粒组明显降低,P<0.05。RT-PCR和蛋白质印迹法结果显示,SET8-shRNA组Survivin mRNA相对表达量为0.257±0.024,较空白对照组(1.046±0.041)和空质粒组(0.994±0.018)明显下降(P<0.05),其蛋白的相对表达量为0.834±0.072,较空白对照组(0.951±0.047)和空质粒组(1.203±0.092)明显下降,P<0.05;而Caspase-3mRNA的相对表达量为0.921±0.072,较空白对照组(0.421±0.022)和空质粒组(0.439±0.007)明显上升(P<0.05),蛋白的相对表达量为1.188±0.022,较空白对照组(0.541±0.060)和空质粒组(0.617±0.028)明显上升,P<0.05。结论干扰SET8可以有效抑制肾透明细胞癌786-O细胞SET8蛋白表达,从而抑制Survivin和上调Caspase-3的表达而抑制肾透明细胞癌786-O细胞增殖能力。
The purpose of cell proliferation is an important factor affecting the progression of renal cell carcinoma, the purpose of this study is to investigate the role of SET8 in the regulation of proliferation of renal clear cell carcinoma 786-O cells. Methods 786-O cells of renal clear cell carcinoma were randomly divided into blank control group, empty plasmid group and SET8-shRNA group. The proliferation of three groups of cells was detected by MTT method. The expression of Survivin and Caspase-3 was detected by RT-PCR and Western blotting. The expression of Survivin and Caspase- O cells proliferation and related gene and protein expression. Results shRNA-SET8 successfully inhibited the expression of SET8 protein in 786-O cells. The results of MTT assay showed that the inhibitory rates of SET8-shRNA group were (10.97 ± 2.68)%, (20.38 ± 7.12)%, (25.03 ± 7.60)% and (30.35 ± 1.97) at 0,12,24,36 and 48 h % And (31.33 ± 1.04)% respectively. The proliferation of 786-O cells in SET8-shRNA group was significantly lower than that in blank control group and empty plasmid group (P <0.05). The results of RT-PCR and Western blot showed that the relative expression of Survivin mRNA in SET8-shRNA group was 0.257 ± 0.024, significantly lower than that in blank control group (1.046 ± 0.041) and empty plasmid group (0.994 ± 0.018) (P <0.05) The relative expression of protein was 0.834 ± 0.072, which was significantly lower than that of blank control group (0.951 ± 0.047) and empty plasmid group (1.203 ± 0.092) (P <0.05), while the relative expression of Caspase-3 mRNA was 0.921 ± 0.072 Compared with control group (0.421 ± 0.022) and empty plasmid group (0.439 ± 0.007), the relative expression level of protein was 1.188 ± 0.022 (0.541 ± 0.060) and empty plasmid group ± 0.028) increased significantly, P <0.05. Conclusion Interference of SET8 can inhibit the expression of SET8 protein in 786-O cells of clear cell renal carcinoma, thus inhibiting the expression of Survivin and up-regulating Caspase-3 and inhibiting the proliferation of 786-O cells.