稳定转染ERβ基因对MCF-7乳腺癌细胞系生长特性的影响

Effects of exogenous ER β expression on the cell growth properties of MCF-7 breast cancer cell line

目的研究在不同处理因子作用下,外源基因ERβ的表达对MCF-7乳腺癌细胞系生长特性的影响。方法利用lipofectamine2000将ERβ真核表达载体pCDNA3-ERβ导入MCF-7乳腺癌细胞系。采用含雌激素应答元件(ERE)的荧光素酶报告基因及Western blot方法,检测转染细胞中ERβ的转录活性和蛋白表达水平,筛选阳性克隆。以亲本细胞MCF-7及转染空载体质粒pCDNA3的MCF-7细胞为对照,在雌激素E2和雌激素受体拮抗剂4-OHT作用下观察细胞的生长特点。结果在转染ERβ基因的MCF-7细胞系中,ERβ的转录激活活性明显升高;Western blot检测证实,ERβ的蛋白表达水平显著增高。在无处理因子情况下,外源基因ERβ在MCF-7细胞系中的表达对细胞的形态及生长速度无明显影响。与亲本细胞MCF-7及转染空载体质粒的MCF-7细胞相比,稳定转染ERβ的MCF-7细胞对雌激素的敏感性下降,但对4-OHT处理的敏感性无明显减少。结论外源性ERβ基因在MCF-7乳腺癌细胞中的稳定表达不增加对4-OHT的耐药性,但使之对雌激素的敏感性下降。

Objective To study the effects of exogenous ERβ on the growth of breast cancer MCF-7 cells under different treatment. Methods An eukaryotic expression vector containing 1.6 kb of human entire coding sequence of ER 13 （pCDNA3-ERβ） was transfected into human breast cancer MCF-7 cells using lipofectamine 2000. The biological activity of ERβ was detected with the luciferase reporter containing estrogen responsive element （ERE） and the expression of ERβ protein by Western blot. The growth properties of MCF-7, pCDNA 3-transfected MCF-7 and pCDNA 3-ER β-transfected MCF-7 cells under different treatment, including E2 （ 17beta-estradiol ） and 4-OHT （ 4-hydroxytamoxifen ） , were observed. Results A stronger activation of the reporter by ERβ in the presence of E2 was observed in the pCDNA 3-ER β-transfected MCF-7 cells than in the pCDNA 3-transfected MCF-7 and in MCF-7 cells. Western blot analysis showed that the protein level of ERβ in the pCDNA 3-ER β-transfected MCF-7 cells was markedly increased. Exogenous ERβ expression did not change the growth properties and the morphology of MCF-7 cells under normal condition. The pCDNA 3-ER β-transfected MCF-7 cells proliferated at the same rate as naive cells in the presence of 4-OHT, whereas a strong inhibition of the proliferation of the pCDNA 3-ER β- transfected MCF-7 cells in the presence of E2 was observed. Conclusion Exogenous ERβ expression does not increase the resistance to 4-OHT, and a strong inhibition of the proliferation may occurre in the presence of E2.