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[目的]检测并验证正常成年小鼠肺内支气管肺泡干细胞(BASCs)的miRNAs表达情况。[方法]流式细胞仪分选小鼠BASCs和其对照细胞(CD45-CD31-Sca-1-CD34-细胞);将分选出的细胞常规提取RNA后,经YM-100(Millipore)微离心过滤柱抽提小于300核苷酸长度的小RNA;微阵列法检测BASCs和其对照细胞的miRNAs表达谱,筛选出差异miRNAs;构建miRNAs特异性TaqManMGB探针,qRT-PCR法验证所选差异miRNAs在两种细胞的表达。[结果]微阵列法检测显示BASCs和对照细胞有116个miRNAs的表达有显著性差异(P<0.01),其中有56个miRNAs在BASCs高表达,60个miRNAs在BASCs低表达。在这些差异miRNAs中选取了10个与细胞周期、干细胞分化、肿瘤发生相关的miRNAs,通过qRT-PCR法检测其在BASCs和对照细胞中的表达,qRT-PCR的检测结果与miRNAs芯片检测结果一致。[结论]研究通过微阵列法鉴定了小鼠BASCs的miRNAs表达谱,与对照细胞相比,发现116个miRNAs的表达具有显著性差异(56个高表达,60个低表达)。
[Objective] To detect and validate the expression of miRNAs in bronchoalveolar stem cells (BASCs) of normal adult mice. [Methods] Mouse BASCs and their control cells (CD45-CD31-Sca-1-CD34- cells) were sorted by flow cytometry. RNA was isolated from the sorted cells and then centrifuged by YM-100 (Millipore) MicroRNAs were used to detect miRNAs expression in BASCs and their control cells, and miRNAs were screened out. The miRNAs-specific TaqManMGB probes were constructed by qRT-PCR, and the selected miRNAs were verified by qRT-PCR Expression in both cells. [Results] The results of microarray assay showed that the expression of 116 miRNAs in BASCs and control cells was significantly different (P <0.01). Among them, 56 miRNAs were highly expressed in BASCs and 60 miRNAs were low in BASCs. Ten miRNAs related to cell cycle, stem cell differentiation and tumorigenesis were selected from these miRNAs, and their expression in BASCs and control cells was detected by qRT-PCR. The result of qRT-PCR was consistent with that of miRNAs . [Conclusion] The miRNAs expression profiles of mouse BASCs were identified by microarray. Compared with the control cells, 116 miRNAs showed significant differences (56 high expression and 60 low expression).