EGCG对乳癌细胞RAFFS1A基因表达及甲基化的影响

Effect of EGCG on the gene expression of breast cancer cell RAFFS1A and methylation

目的:探讨表没食子儿茶素没食子酸酯(epigallocatechin gallate,EGCG)对乳腺癌细胞RASSF1A甲基化的影响。方法:体外培养乳腺癌细胞系MCF-7,用10μmol/L和20μmol/L EGCG处理后,采用Western blot和实时定量PCR分别检测RASSF1A基因蛋白和m RNA表达;高效液相色谱-电喷雾质谱检测RASSF1A甲基化,同时采用MTT检测MCF-1细胞增殖。结果:MCF-7细胞基础状态下RASSF1A表达水平较低,而经10μmol/L和30μmol/L EGCG处理后,能显著增强RASSF1A蛋白和m RNA的表达。高效液相色谱-电喷雾质谱结果显示,30μmol/L EGCG处理MCF-7细胞后,与对照组相比,RASSF1A启动子甲基化水平降低至36%,全基因组甲基化水平降低44%。此外,10μmol/L和20μmol/L EGCG作用后,MCF-7细胞的存活率降低至39%和23%。结论:EGCG能上调RASSF1A基因表达,降低细胞内甲基化水平,从而可能发挥对MCF-7细胞生长抑制作用。

To investigate the effect ofepigallocatechin gallate on the gene expression of breast cancer cell RAFFS1A and methylation. Methods: Breast cancer cells MCF-7 were in vitro cultured and respectively detected on expression of RASSF1A and mRNA by Western blot and real time quantitative PCR, after treated by 10μmol/L and 20μmol/L EGCG. RAFFS1A and methylation was detected by high performance liquid chromatography and electrospray ionization mass spectrometry, meanwhile the proliferation of MCF-1 cells was detected by MTT. Results: The expression level of RASSF1A under the basal conditions of MCF-7 cells was lower, but after treated by 10μmol/L and 30μmol/L EGCG, the expressionof RASSF1A and mRNA was improved. High performance liquid chromatography and electrospray ionization mass spectrometry showed that after MCF-7 cells were treated by 30μmol/L EGCG and compared to the control group, the level of RASSF1A promoter methylation decreased to 36%,the whole genome methylation level reduced to 44%. In addition, after treated by 10μ mol/L and 20μ mol/L EGCG, the survival rate of MCF-7 cell decreased to 39% and 23%. Conclusion: EGCG can improve the gene expression of RASSF1A and reduce methylation level in order to exert growth inhibitory effects on MCF-7 cells.