AT1-R siRNA对雌激素诱导的Ishikawa细胞生物学行为的影响

Effect of AT1R Knockdown on Ishikawa Cell Proliferation Induced by Estrogen

目的:通过转染小干扰RNA(siRNA)沉默血管紧张素受体1(angiotensin receptor 1,AT1R)的表达来研究其对雌激素诱导的Ishikawa细胞增殖和细胞周期及凋亡的影响。方法:免疫荧光检测AT1R在子宫内膜癌Ishikawa细胞中的表达,转染AT1-R siRNA,Western blot检测转染前后Ishikawa细胞中AT1R蛋白的表达。MTT检测雌激素诱导Ishikawa细胞的增殖及雌激素诱导的雌激素受体抑制剂作用下转染前后Ishikawa细胞的增殖,Western blot检测细胞外调节蛋白激酶(extracellular signal-regulatedkinases 1/2,ERK1/2)表达。结果:免疫荧光检测AT1R表达于Ishikawa细胞,转染AT1-R siRNA 72 h后AT1R蛋白表达降低最显著,降低82.40%(P<0.05),为此检测转染72h后细胞增殖周期及凋亡,MTT结果显示,雌激素能诱导Ishikawa细胞增殖,封闭雌激素受体后雌激素诱导的Ishikawa细胞增殖受到抑制,封闭雌激素受体后雌激素诱导的转染组细胞增殖较未转染组受到明显抑制;流式细胞周期结果显示,雌激素受体封闭后雌激素诱导的转染组细胞较未转染组S期减少,凋亡增多(P<0.05),其ERK1/2表达较未转染组明显降低(P<0.05)。结论:AT1R可促进雌激素诱导的Ishikawa细胞增殖,其机制可能与ERK1/2表达下降有关。

Objective：To investigate the role of angiotensinⅡtype 1 in the estrogen-induced proliferation, cell cycle, and apop -tosis in endometrial carcinoma cell line Ishikawa by transfection of siRNA-AT 1R. Methods：The expression of Angiotensin Receptor1 （AT 1-R） was identified using immunofluorescence assay. The expression of AT 1-R protein was examined by Western blotting before and after transfection. The effect of AT 1-R silence on 17β-E2-induced proliferation of cell line Ishikawa was measured by MTT assay, whereas the expression of ERK 1/2 protein was examined by Western blotting. Results：After transfection with AT 1-R siRNA plasmid for 72hours, the expression of AT1-R protein significantly decreased by 82.40% （P〈0.05）. AT1-R blocking inhibited the proliferation of Ishikawa cells treated with17β-E2, whereas AT 1-R silencing further inhibited the proliferation. AT 1-R silence reversed the promo-tion of 17β-E2 on the cell cycle transformation of Ishikawa and decreased the number of S-phase cells （ P〈0.05）. The apoptotic cells in -creased. ERK 1/2 expression decreased in the transfection group. Conclusion：AT1-R plays an important role in 17β-E2-induced prolif-eration of endometrial carcinoma cell line Ishikawa and could be related to the decrease in the expression of ERK 1/2.