中国医科大学学报
中國醫科大學學報
중국의과대학학보
JOURNAL OF CHINA MEDICAL UNIVERSITY
2015年
5期
394-399
,共6页
孟琳%王天一%李晓曦%马萍
孟琳%王天一%李曉晞%馬萍
맹림%왕천일%리효희%마평
microRNA%乳腺癌%肿瘤干细胞%miR-206%miR-1
microRNA%乳腺癌%腫瘤榦細胞%miR-206%miR-1
microRNA%유선암%종류간세포%miR-206%miR-1
microRNA%breast cancer%tumor stem cell%miR-206%miR-1
目的：探讨上调microRNA?206（miR?206）和microRNA?1（miR?1）表达对乳腺癌干细胞增殖的影响以及其作用机制。方法采用流式细胞分选技术从乳腺癌细胞株MCF?7中分离出乳腺癌干细胞。实验分为空白对照组、阴性对照组、miR?206转染组、miR?1转染组。除空白对照组外，其他各组分别转染阴性对照mimic、hsa?miR?206 mimic、hsa?miR?1 mimic。应用实时定量PCR检测miR?206、miR?1以及转录因子EVI?1基因的表达水平；Western blot法检测EVI?1蛋白的表达；应用MTT法检测miR?206、miR?1对乳腺癌干细胞增殖能力的影响。结果从MCF?7细胞系中分选出CD44+/CD24-/low乳腺癌干细胞，经无血清培养后可以成功传代，用于后续试验；转染hsa?miR?206 mimic、hsa?miR?1 mimic 48 h后miR?206、miR?1相对表达水平提高，EVI?1 mRNA表达水平明显下降；Western blot、MTT结果显示，上调miR?206和miR?1表达水平后显著降低EVI?1蛋白的表达，抑制了乳腺癌干细胞增殖能力。乳腺癌干细胞中miR?206、miR?1表达水平以及EVI?1蛋白表达水平的差异有统计学意义（P<0.05）。结论上调miR?206和miR?1表达能够抑制乳腺癌干细胞增殖能力，其机制可能与下调EVI?1的表达有关。
目的：探討上調microRNA?206（miR?206）和microRNA?1（miR?1）錶達對乳腺癌榦細胞增殖的影響以及其作用機製。方法採用流式細胞分選技術從乳腺癌細胞株MCF?7中分離齣乳腺癌榦細胞。實驗分為空白對照組、陰性對照組、miR?206轉染組、miR?1轉染組。除空白對照組外，其他各組分彆轉染陰性對照mimic、hsa?miR?206 mimic、hsa?miR?1 mimic。應用實時定量PCR檢測miR?206、miR?1以及轉錄因子EVI?1基因的錶達水平；Western blot法檢測EVI?1蛋白的錶達；應用MTT法檢測miR?206、miR?1對乳腺癌榦細胞增殖能力的影響。結果從MCF?7細胞繫中分選齣CD44+/CD24-/low乳腺癌榦細胞，經無血清培養後可以成功傳代，用于後續試驗；轉染hsa?miR?206 mimic、hsa?miR?1 mimic 48 h後miR?206、miR?1相對錶達水平提高，EVI?1 mRNA錶達水平明顯下降；Western blot、MTT結果顯示，上調miR?206和miR?1錶達水平後顯著降低EVI?1蛋白的錶達，抑製瞭乳腺癌榦細胞增殖能力。乳腺癌榦細胞中miR?206、miR?1錶達水平以及EVI?1蛋白錶達水平的差異有統計學意義（P<0.05）。結論上調miR?206和miR?1錶達能夠抑製乳腺癌榦細胞增殖能力，其機製可能與下調EVI?1的錶達有關。
목적：탐토상조microRNA?206（miR?206）화microRNA?1（miR?1）표체대유선암간세포증식적영향이급기작용궤제。방법채용류식세포분선기술종유선암세포주MCF?7중분리출유선암간세포。실험분위공백대조조、음성대조조、miR?206전염조、miR?1전염조。제공백대조조외，기타각조분별전염음성대조mimic、hsa?miR?206 mimic、hsa?miR?1 mimic。응용실시정량PCR검측miR?206、miR?1이급전록인자EVI?1기인적표체수평；Western blot법검측EVI?1단백적표체；응용MTT법검측miR?206、miR?1대유선암간세포증식능력적영향。결과종MCF?7세포계중분선출CD44+/CD24-/low유선암간세포，경무혈청배양후가이성공전대，용우후속시험；전염hsa?miR?206 mimic、hsa?miR?1 mimic 48 h후miR?206、miR?1상대표체수평제고，EVI?1 mRNA표체수평명현하강；Western blot、MTT결과현시，상조miR?206화miR?1표체수평후현저강저EVI?1단백적표체，억제료유선암간세포증식능력。유선암간세포중miR?206、miR?1표체수평이급EVI?1단백표체수평적차이유통계학의의（P<0.05）。결론상조miR?206화miR?1표체능구억제유선암간세포증식능력，기궤제가능여하조EVI?1적표체유관。
Objective To investigate the effects of up?regulated miR?206/miR?1 on the proliferation of breast cancer stem cells and the effect mech?anism. Methods Breast cancer stem cells(BCSCs)were isolated from breast cancer cell line MCF?7 by fluorescence?activated cell sorting. Cells in the experiment were divided into the blank control group,the negative control group,the miR?206 group and the miR?1 group. The BCSCs were transfected by negative control mimic,hsa?miR?206mimic and hsa?miR?1mimic in all groups except the blank control group. MiR?206and miR?1 expression levels as well as the transcription factor EVI?1 gene were detected by real time PCR. The expression levels of the transcription factor EVI?1 protein were detected by Western blot. MTT method was used to detect the effects of miR?206 and miR?1 on the proliferation of BCSCs. Results The BCSCs(CD44+/CD24-/low cells)isolated from MCF?7 cell lines were successfully cultured in serum?free medium for subsequent studies. After transfection of hsa?miR?206mimic and hsa?miR?1mimic for 48 hours,miR?206and miR?1relative expression levels increased. EVI?1mRNA ex?pression levels significantly decreased. The results of Western blot and MTT showed that up?regulated expression levels of miR?206 and miR?1 could significantly reduce the expression of EVI?1 protein and inhibited the proliferation of BCSCs. The differences in levels of miR?206,miR?1 and EVI?1 protein were statistically significant(P<0.05). Conclusion Up?regulated miR?206 and miR?1 expression can inhibit the proliferation ability of BCSCs,which may be related to the down?regulation of EVI?1.
