目的:研究雌激素受体β(ERβ)对人乳腺癌细胞株MDA-MB-435生物学行为的影响。
方法:通过稳定转染将人ERβcDNA导入MDA-MB-435细胞。MTT法检测ERβ表达对细胞增殖和侵袭的影响,流式细胞术和transwell技术。细胞周期蛋白A,细胞周期蛋白E,细胞周期蛋白D1,p21,MMPs,通过RT-PCR和/或Westernblot或明胶酶谱检测Ets-1,VEGF和b-FGF。
结果:显示ERβ能够以不依赖雌二醇的方式显着增加MDA-MB-435细胞的增殖和侵袭。ERβ过表达的细胞的S期分布为46.8%,显著高于野生型(29.9%)和mock转染细胞(27.6%)(P=0.01)。在ERβ转染的细胞中,p21的表达在mRNA水平上下降了33.3%(P=0.03),在蛋白水平上下降了47.4%(P=0.02),分别。MMP-9在mRNA水平上的表达增加了91.3%(P<0.01),其活性上调了67.3%(P=0.02)。此外,Ets-1mRNA和蛋白水平分别增加62.2%(P=0.01)和51.0%(P=0.01),分别。细胞周期蛋白A的mRNA水平没有观察到显着差异,细胞周期蛋白E,这些细胞中的细胞周期蛋白D1、MMP-1、MMP-2、MMP-7、VEGF和b-FGF。
结论:ERβ能增强乳腺癌细胞的增殖和侵袭能力。p21的下调和MMP-9和Ets-1的上调可能参与其机制。
OBJECTIVE: To
study effects of estrogen receptor beta (ER beta) on the biological behavior of a human breast cancer cell line MDA-MB-435.
METHODS: Human ER beta cDNA was introduced into MDA-MB-435 cells by stable transfection. Effects of ER beta expression on cell proliferation and invasion were investigated by MTT, flow cytometry and transwell techniques. Cyclin A, cyclin E, cyclin D1, p21, MMPs, Ets-1, VEGF and b-FGF were detected by RT-PCR and/or Western blot or gelatin zymography.
RESULTS: ER beta was shown to be able to significantly increase the proliferation and invasion of MDA-MB-435 cells in an estradiol-independent manner. The S phase distribution of the cells with ER beta overexpression was 46.8%, significantly higher than that of wild type (29.9%) and mock transfected cells (27.6%) (P = 0.01). In ER beta transfected cells, the expression of p21 decreased by 33.3% at mRNA level (P = 0.03) and by 47.4% at protein level (P = 0.02), respectively. The expression of MMP-9 increased by 91.3% at mRNA level (P < 0.01) and its activity was up-regulated by 67.3% (P = 0.02). Furthermore, the mRNA and protein levels of Ets-1 increased 62.2% (P = 0.01) and 51.0% (P = 0.01), respectively. No significant difference was observed in the mRNA levels of cyclin A, cyclin E, cyclin D1, MMP-1, MMP-2, MMP-7, VEGF and b-FGF among these cells.
CONCLUSIONS: ER beta can enhance proliferation and invasion of breast cancer cells. Down-regulation of p21 and up-regulation of MMP-9 and Ets-1 may be involved in its mechanisms.