肺结核患者外周血单核细胞中差异表达miRNA的筛选

Investigation of differentially expressed miRNA in peripheral blood mononuclear cell from the patients with pulmonary tuberculosis

目的研究肺结核患者外周血单个核细胞中miRNA的差异表达。方法选择10例肺结核患者和10例健康人。每人抽取5ml静脉血,提取外周血单个核细胞(PBMC),分别将肺结核患者组和健康对照组的PBMC混合,提取总RNA和miRNA,进行miRNA荧光标记和纯化,用芯片筛选差异表达的miRNA,以校正后的两个样本间杂交信号强度的比值,来判断芯片的结果。以实时定量PCR验证。结果筛选出26个差异表达的miRNA,其中13个上调的miRNA和13个下调的miRNA。通过比较肺结核患者与健康对照者实时定量PCR结果证实,hsa-miR-1(分别为1.085±0.005,1.018±0.011)表达上调(t=4.334,P<0.01),hsa-miR-146a(分别为0.948±0.008,1.036±0.005)(t=4.460,P<0.01)与hsa-let-7e(分别为1.020±0.007,1.091±0.004)(t=4.149,P<0.01)表达下调。结论多种miRNA在肺结核患者外周血单个核细胞中有异常表达。

Objective To investigate the differentially expressed miRNAs in peripheral blood mononuelear cell from the patients with pulmonary tubereulosis. Methods Ten patients with pulmonary tuberculosis and 10 healthy controls were selected as the participants. 5ml of venous blood was collected, and then peripheral blood mononuclear cells （PBMC） were isolated and mixed from these 2 groups, respectively. The total RNA and miRNA were extracted. The miRNA was labeled and purified, and the endogenous differentially expressed miRNAs were sereened by high-throughput miRNA mieroarray. The result of miRNA microarray was analyzed by the corrected ra- tio of hybridization signal intensity in 2 group. The validation was performed by real-time quantitative PCR. Results Twenty-six differentially expressed miRNAs were obtained in 2 groups, including 13 overexpressed miRNAs and 13 downexpressed miRNAs. The results of comparation between the lung tuberculosis patients and healthy controls by real-time quantitative PCR approved that hsa-miR-l（1. 085 ± 0. 005,1. 018 ± 0. 011）（t= 4. 334, P〈0. 01） was overexpressed, and hsa-miR-146（0. 9484±0. 008,1. 0364±0. 005）（t=4. 460,P〈0. 01） and hsa-let-Te （1.0204±0. 007,1.0914±0. 004）（t=4. 149,P〈0. 01） were downexpressed. Conclusion There are many differen- tially expressed miRNA between the patients with pulmonary tuberculosis and healthy controls.