雄激素抑制雌激素受体阴性雄激素受体阳性乳腺癌细胞增殖相关微小RNA的筛选及其功能学研究  

MiRNAs expression related to androgen inhibition of human MDA-MB-453 cell growth and initial functional analysis

作　　者：吕淑华[1] 牛昀[1] 于琦[3] 王淑玲[2] 王亚红[1] 张静[1] 

机构地区：[1]天津医科大学附属肿瘤医院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 [2]天津市人民医院病理科 [3]天津静海县医院肿瘤科

出　　处：《中华实验外科杂志》2012年第8期1521-1524,共4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）;天津市自然科学基金资助项目（09JCYBJC10100）

摘　　要：目的筛选雌激素受体阴性（ER-）雄激素受体阳性（AR＋）乳腺癌细胞中AR相关的微小RNA（miRNA），并研究其细胞功能学。方法选择MDA—MB-453乳腺癌细胞，雄激素二氢睾酮作用后miRNA芯片杂交筛选出明显上调的miRNA，对其中的let-7a采用实时定量逆转录-聚合酶链反应（RT—qPCR）进行验证；通过转染let-7a特异性反义寡核苷酸使let-7a下调4倍，通过构建质粒和细胞转染实验使let-7a上调〉8倍；噻唑蓝（MTT）和流式细胞术检测转染后细胞增殖和细胞周期的变化。结果miRNA芯片筛选出明显上调的miRNA只有let-7a、b、c、d4个，RT—qPCR结果显示1et-7a上调近13倍；细胞转染后，在let-7a低表达组，MTT检测结果显示，细胞增殖活性增高，至第4天之后与对照组比较差异有统计学意义（P〈0．05），流式细胞术检测结果显示对照组和实验组细胞G，期分别为（72．30±3．16）％和（63．24±3．63）％，S期分别为（19．12±4．45）％和（31．00±4．56）％，G，期细胞比例减少，S期细胞比例增加，差异有统计学意义（P〈0．05），在let-7a高表达组，则出现相反的结果。结论let-7a能够抑制ER—AR＋乳腺癌细胞增殖、使细胞周期阻滞在G1-S期，可能在雄激素抑制ER-AR＋乳腺癌细胞生长过程中发挥主要作用。Objective To screen the androgen receptor-related microRNA （miRNA） in estrogen receptor negative but androgen receptor positive （ER - AR ＋ ） MDA-MB-453 breast cancer cell line and study the function. Methods The MDA-MB-453 breast cancer cells were cultured in vitro, and treated with dihydrotestosterone （DHT）. MiRNA array hybridization technique was used to screen the differentially expressed miRNAs. The significantly up-regulated miRNA, let-Ta was selected and checked by using real- time quantitative reverse transcriptase polymerase chain reaction （RT-qPCR）. The MDA-MB-453 cells were transiently transfected with specific antisense oligonucleotide or vector. Cell proliferation was deter- mined by MTr assay and cell cycle was analyzed by flow cytometry ＇after transfection. Results Let-7a was significantly up-regulated in DHT-treated cells as compared with control group. After the cells was trans- fected with oligonucleotide or vector, cells proliferation was inhibited （P 〈 0. 05 ） , and the number of ceils in the G1 phase was significantly increased [ （72. 30 ±3. 16）% vs （63.24±3.63）% ] and that in S phase was significantly decreased [ （ 19. 12 ± 4. 45 ） % vs （ 31.00± 4. 56 ） % ] in let-7a-overexpressed group （P 〈0. 05）. Meanwhile, in let-7a-blocked group, there were the opposite results. Conclusion Let-7a may play an important role in the process of DHT inhibiting ER-AR ＋ breast cancer growth probably by in- hibiting the cell proliferation and causing cell cycle arrest at the G1-S phases.

关 键 词：乳腺癌 let-7a 雄激素受体 增殖 

分 类 号：R737.9[医药卫生—肿瘤]