5-Aza-CdR对肾细胞癌Caki-1细胞生长及E-钙黏蛋白基因表达的影响

Effects of 5-Aza-CdR on proliferation of renal clear cell carcinoma line Caki-1 and expression of E-cadherin gene

目的:研究甲基化抑制剂5-氮杂-2′-脱氧胞苷(5-Aza-CdR)对肾细胞癌细胞株Caki-1增殖抑制作用、对E-钙黏蛋白(E-cadherin,E-cad)甲基化状态及E-cad蛋白状态的影响。方法:不同浓度5-Aza-CdR处理肾细胞癌细胞株Caki-1后,四甲基偶氮唑盐(MTT)比色实验观察细胞经药物处理前后的生长活性;甲基化特异性PCR(methylationspecial PCR,MSP)检测细胞处理前后E-cad基因的甲基化状态;蛋白质印迹法检测5-Aza-CdR处理细胞前后E-cad蛋白的表达。结果:5-Aza-CdR能抑制肾细胞癌细胞株Caki-1的增殖;未经药物处理组的基因高甲基化,经10-6mol/L5-Aza-CdR处理72 h,E-cad基因启动子区域高甲基化得到逆转,同时E-cad蛋白表达也得到增强。结论:5-Aza-CdR能够有效逆转Caki-1细胞E-cad基因的异常甲基化,恢复E-cad蛋白表达。

Objective： To investigate the effects of methylation inhibitor 5-Aza-2′-deoxycytidine on the growth of renal clear cell carcinoma cell line Caki-1 and the expression of the E-cadherin（E-cad） gene and protein.Methods： Renal clear cell carcinoma cell line Caki-1 were treated with different concentration of 5-Aza-CdR respectively.Then the growth rate of the cells was detected by MTT assay.The methylation and demethylation status of E-cad gene were detected by methylation special PCR（MSP）.The western blot was used to detect E-cad protein levels in the cell line before and after treatment with 5-Aza-CdR.Results： 5-Aza-CdR can inhibit the growth of renal clear cell carcinoma Caki-1.The high methylation status of E-cad gene promoter region was reversed with 10-6 mol/L 5-Aza-CdR treated for 72 h,while E-cad protein expression was also enhanced.Conclusion： 5-Aza-CdR effectively caused the demethylation of E-cad gene,and recovered the E-cad protein expression.