乳腺癌细胞与乳腺上皮细胞MicroRNAs表达谱的差异性分析

Differential expression profiles of microRNAs between breast cancer cells and mammary epithelial cells

背景与目的:已有研究表明,MicroRNAs(miRNAs)的异常表达参与了乳腺癌的发生和发展,常起到抑癌基因或癌基因作用。本研究旨在探讨乳腺癌细胞与乳腺上皮细胞miRNAs的差异性表达,并预测异常表达的miRNAs可能调控的靶基因。方法:培养乳腺癌细胞株MCF-7和正常乳腺上皮细胞株HBL-100,分别提取细胞总RNA,并进行质检;利用miRNAs芯片技术,检测乳腺癌细胞和正常乳腺上皮细胞miRNAs的表达,对其表达谱进行差异性分析;采用实时定量RT-PCR进行验证;运用MiRanda、TargetScan、PicTar、DIANA-microT软件预测miRNAs可能调控的靶基因。结果:从MCF-7细胞与HBL-100细胞提取到的总RNA纯度较高,其A260/A280为1.90,A260/A230为2.0;琼脂糖凝胶实验结果显示总RNA完整性好。通过miRNAs表达谱的差异性分析,获得173个与乳腺癌相关的miRNAs,其中113个表达上调,60个表达下调;实时定量PCR检测到mir-18a和mir-195在MCF-7细胞中高表达;对乳腺癌细胞显著异常表达的mir-200b进行信息学分析,共筛选出68个靶基因。结论:获得了乳腺癌细胞与乳腺上皮细胞miRNAs的差异表达谱,其中部分miRNAs可能通过调控其靶基因而参与乳腺癌的发病机制。

Background and Objective： Aberrant expression of microRNAs （miRNAs） is involved in the development of breast cancer. This study was to explore differential expression of miRNAs between breast cancer cells and mammary epithelial cells, and to predict potential targets of the diferentially expressed miRNAs. Methods. Human breast cancer MCF-7 cells and normal human mammary epithelial HBL-100 cells were cultured. The total RNA was isolated and examined. The expression profiles of miRNAs in MCF-7 and HBL-100 cells were detected with a miRNA microarray chip and compared. Array results were confirmed by real-time polymerase chain reaction （PCR）. Potential mRNA targets were predicted by MiRanda, TargetScan, PicTar, and DIANA-microT software. Results= The purification of total RNA from MCF-7 and HBL-100 cells was relatively high. According to a microarray- based screening, 173 breast cancer-related miRNAs were identified. Of the 173 miRNAs, 113 were up-regulated and 60 were down-regulated in MCF-7 cells （P〈0.01）. Real-time PCR showed that mir-18a and mir-195 were highly expressed in MCF-7 cells. Sixty-eight potential target genes of mir-200b were predicted by informatics analysis. Conclusions： The miRNA expression profiles of breast cancer cells and mammary epithelial cells are obtained. Some miRNAs may be involved in the pathogensis of breast cancer by regulating their target genes.