TGF-β信号通路调控Twist高表达型乳腺肿瘤细胞微球体的形成及迁移  

TGF-β signaling pathway regulates the mammosphere formation and migration of highly Twist-expressed breast cancer cells

作　　者：郎磊[1] 周明莉[1,2] 徐丽云[1] 杜燕娥[1] 文思阳 柳满然[1] 

机构地区：[1]重庆医科大学检验医学院临床检验诊断学教育部重点实验室,重庆400016 [2]成都市第三人民医院检验科,四川成都610031

出　　处：《肿瘤》2016年第10期1083-1089,1097,共8页Tumor

基　　金：国家自然科学基金面上项目(编号:31171336)~~

摘　　要：目的 :探讨转化生长因子β(transforming growth factor-beta,TGF-β)信号通路对Twist高表达型乳腺癌细胞微球体形成及微球体细胞迁移的影响。方法:应用蛋白质印迹法和微球体形成实验分别检测正常乳腺上皮MCF10A细胞及乳腺癌MCF7和BT549细胞中Twist的表达水平及微球体形成能力。蛋白质印迹法检测MCF10A、MCF7和BT549微球体细胞中TGF-β、SMAD3和磷酸化SMAD3(phospho-SMAD3,p-SMAD3)的表达水平。在微球体形成实验的基础上,加入TGF-β信号通路特异性抑制剂LY2109761,应用微球体形成实验和蛋白质印迹法分别检测乳腺癌BT549微球体细胞的形成能力和p-SMAD3蛋白、肿瘤干细胞相关标志物性别决定区Y框蛋白2(sex determining region Y-box 2,SOX2)和八聚体绑定转录因子4(octamer-binding transcription factor 4,OCT4)的表达水平,Transwell法检测乳腺癌BT549来源的微球体细胞的迁移能力。结果:乳腺癌BT549细胞中Twist的表达水平明显高于MCF10A细胞和MCF7细胞(P值均<0.05),BT549细胞微球体形成能力显著高于MCF10A和MCF7细胞(P值均<0.05)。BT549微球体细胞中TGF-β、p-SMAD3和SMAD3蛋白的表达水平均显著高于MCF10A和MCF7微球体细胞(P值均<0.05)。LY2109761可显著抑制乳腺癌BT549细胞的微球体形成能力(P<0.05),下调乳腺癌BT549微球体细胞中p-SMAD3及干细胞相关蛋白SOX2和OCT4的表达(P值均<0.05),抑制乳腺癌BT549微球体细胞的迁移(P<0.05)。结论 :TGF-β信号通路特异性抑制剂LY2109761可抑制Twist高表达乳腺癌细胞微球体的形成及微球体细胞的迁移。Objective： To investigate the effects of transforming growth factor-beta（TGF-β） signaling pathway on the mammosphere formation and migration of highly Twist-expressed breast cancer cells.Methods： The expression level of Twist and mammosphere formation efficiency（MFE） in normal breast epithelial cell line MCF10 A and breast cancer MCF7 and BT549 cells were detected by Western blotting and mammosphere formation assay, respectively. The expression levels of TGF-β, SMAD3 and phospho-SMAD3（p-SMAD3） in mammosphere cells derived from normal breast epithelial MCF10 A cells and the breast cancer MCF7 and BT549 cells were detected by Western blotting. Based on the foundation of mammosphere formation assay, the mammosphere formation efficiency and the expressions of p-SMAD3 and cancer stem cell markers sex determining region Y-box 2（SOX2） and octamer-binding transcription factor 4（OCT4） in mammosphere cells derived from breast cancer BT549 cells after treatment with LY2109761, a specific inhibitor of TGF-β signaling pathways, were detected by mammosphere formation assay and Western blotting, respectively. The ability of migration of mammosphere cells derived from breast cancer BT549 cells was detected by Transwell assay.Results： The expression level of Twist protein in breast cancer BT549 cells was higher than those in MCF10 A and MCF7 cells（both P〈0.05）. The mammosphere formation efficiency of breast cancer BT549 cells was higher than those of MCF10 A and MCF7 cells（both P〈0.05）. The expression levels of TGF-β, p-SMAD3 and SMAD3 proteins in mammosphere cells derived from breast cancer BT549 cells were higher than those in mammosphere cells derived from MCF10 A and MCF7 cells（all P〈0.05）. The ability of mammosphere formation of breast cancer BT549 cells was inhibited by LY2109761（P〈0.05）. The expression levels of p-SMAD3 and cancer stem cell markers SOX2 and OCT4 in the mammosphere cells derived from breast cancer BT549 cells after treatment with LY2109761 were down-regu

关 键 词：乳腺肿瘤 转化生长因子 细胞迁移分析 微球体 信号通路 Twsit 

分 类 号：R737.9[医药卫生—肿瘤]