雄激素受体阳性乳腺癌干细胞的富集及其特性鉴定  

Enrichment and identification of androgen receptor-positive breast cancer stem cells

作　　者：刘珊[1] 张巍[1] 刘夏[1] 秦颖[1] 牛昀[1] 

机构地区：[1]天津医科大学肿瘤医院乳腺病理研究室国家肿瘤临床医学研究中心乳腺癌防治教育部重点实验室天津市肿瘤防治重点实验室,300060

出　　处：《中华实验外科杂志》2014年第1期67-69,共3页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金资助项目（81172532）;教育部长江学者和创新团队计划资助项目（TRT0743）

摘　　要：目的 从雄激素受体阳性（AR＋）乳腺癌细胞株中分离出乳腺微球体（MSs）进行培养和鉴定.方法 将3种AR＋乳腺癌细胞株（MCF-7、MDA-MB-231及T47D）接种在添加生长因子的DMEM/F12培养基中,观察MSs的形成及生长；细胞免疫荧光检测MCF-7、MDA-MB-231及T47D中雄激素受体（AR）的表达；流式细胞术（FCM）检测贴壁细胞及MSs中CD44＋ CD24-/low细胞的含量；单克隆形成实验检测MSs细胞的自我更新能力.结果 乳腺癌细胞株在无血清培养基中呈悬浮生长,7-10d可形成MSs;细胞免疫荧光显示MCF-7、T47D中AR主要在细胞质中表达、MDA-MB-231中AR在细胞核及细胞质中均有表达；流式细胞术检测MCF-7、MDA-MB-231及T47D贴壁细胞中CD44＋ CD24-/low细胞含量分别为50.0％、86.4％、3.2％,悬浮细胞中CD44＋ CD24-/low细胞含量分为87.4％、98.8％、38.4％；克隆形成实验显示随着时间的推移形成的克隆球体积不断增大.结论 悬浮生长的MDA-MB-231及MCF-7细胞可作为研究AR阳性乳腺癌干细胞的良好模型.Objective To isolate the mammospheres （MSs） from the androgen receptor positive（AR ＋） breast cancer cell lines for culture and identification to screen AR ＋ breast cancer stem cell model for further study.Methods The three types of AR ＋ breast cancer cell lines were seeded in culture medium DMEM/F12 supplemented with growth factors.The formation and growth of MSs were observed.Immunofluorescence was used to detect the expression of AR in MCF-7,MDA-MB-231 and T47D cell lines.Flow cytometry （FCM） was used to detect CD44 ＋ CD24-/low cell content in adherent cells and mammospheres.Clonogenic assay was used to detect the self-renew capcity of every single cell of the MSs.Results Breast cancer cell lines in serum-free medium showed suspension growth,and MSs formed during 7-10 days.Immunofluorescence showed MCF-7 and T47D cells expressed AR mainly in the cytoplasm,and MDA-MB-231 cells in both the nucleus and cytoplasm.FCM revealed that CD44＋ CD24-/low cell levels in MCF-7,MDA-MB-231 and T47D adherent cells were 50.0%,86.4% and 3.2%,and those in suspended cells were 87.4%,98.8% and 38.4%,respectively.Colony formation assay demonstrated that the volume of MSs was gradually increased with time.Conclusion Suspension growth of MDA-MB-231 and MCF-7 cells can be used as an optimal model for the study of AR-positive breast cancer stem cells.

关 键 词：乳腺癌干细胞 雄激素受体 

分 类 号：R737.9[医药卫生—肿瘤]