AT1-R在雌激素诱导子宫内膜癌细胞增殖中的作用

The Role of Angiotensin Ⅱ Type 1 Receptor in Estrogen-induced Proliferation of Endometrial Carcinoma Cell Line HEC-1A

目的:探讨血管紧张素受体1(angiotensin receptor 1,AT1-R)在雌激素诱导子宫内膜癌细胞HEC-1A增殖、细胞周期转化中的作用及其对细胞外调节蛋白激酶(extracellular signal-regulated kinases 1/2,ERK1/2)表达的影响。方法:免疫荧光技术检测AT1-R在HEC-1A细胞中的表达;脂质体介导AT1-R-siRNA质粒转染人子宫内膜癌细胞HEC-1A,经G418选择培养,采用Western blot检测转染前后HEC-1A细胞中AT1-R蛋白的表达。MTT法检测无雌激素诱导及雌激素诱导10min转染前后细胞的增殖;流式细胞技术检测细胞周期;Western blot检测ERK1/2蛋白及ER蛋白表达水平。结果:转染AT1-R-siRNA-GFP后,与未转染组相比AT1-R蛋白表达降低41.79%(P<0.01);沉默AT1-R能够抑制17β-E2对HEC-1A细胞的促增殖作用。17β-E2处理10 min后,HEC-1A细胞G_1期细胞减少,S期细胞增多,细胞增殖明显(P<0.05);AT1-R沉默后,能够抑制17β-E2诱导的HEC-1A细胞周期转化,使G_1期细胞增加,S期细胞减少(P<0.05);雌激素处理10 min后,HEC-1A细胞ERK1/2蛋白表达水平升高,沉默AT1-R后ERK1/2蛋白表达水平又明显降低。17β-E2诱导及AT1-R沉默对HEC-1A细胞ER蛋白表达无明显影响。结论:AT1-R在雌激素诱导子宫内膜癌细胞HEC-1A增殖,细胞周期转化中具有重要作用,其机制可能与ERK1/2表达降低有关,与ER表达无关。

To investigate the role of angiotensin II type 1 （AT1-R） in estrogen-induced proliferation, cell cycle progression and expression of ERK1/2 in endometrial carcinoma cell line HEC-1A. Methods： The expression of AT1-R was assessed by immunofluorescence. AT1-R siRNA was transfected into human endometdal carcinoma cell line HEC-1A via Lipofectamine 2000. After screening the cultures with G418, the expression of AT1-R protein was examined by Western blot before and after transfection. The effect of AT1-R silencing on 17[3-E2-induced proliferation of HEC-1A cells was measured by MTT assay, the cell cycle was analyzed by flow cytometry, and the expression of EtRK1/2 protein was examined by Western blot. Results： After transfection with AT1-R siRNA plasmid, the expression of AT1-R protein was decreased by 41.79% （ P 〈 0.01）. AT1-R silencing inhibited the proliferation of HEC-1A cells treated with 17β-E2. HEC-1A cells treated with 10^-8mol/L 17β-E2 for 10 min had a decreased number of cells in G1 phase and an increased number of cells in S phase （ P 〈 0.05）. AT1-R silencing reversed the promoting effect of 17β-E2 on the cell cycle, increasing the number of cells in G1 phase and decreasing the number of cells in S phase （ P 〈 0.05）. After treatment with 10^-8mol/L 17β-E2 for 10 min, the expression of ERK1/2 protein in the HEC-1A cell line was increased and AT1-R silencing decreased the expression of ERK1/2 protein. Conclusion： AT1-R plays an important role in 17β-E2-induced proliferation and cell cycle progression in endometrial carcinoma cell line HEC-1A, which may be related to decreased expression of ERK1/2.