大气细颗粒物暴露后小鼠肺组织microRNA差异表达谱芯片分析

Microarray-based Analysis of Differentially Expressed MicroRNAs in Mouse Lung Tissue Induced by Particulate Matter

microRNAs作为临床疾病早期诊断的新型生物标记物越来越受到重视,为了进一步探究其在大气污染暴露后引起疾病的分子染毒机制。本研究通过建立大气污染小鼠染毒模型,利用Agilent芯片筛查小鼠肺组织中microRNAs差异表达谱,并通过实时荧光定量PCR方法验证芯片结果,使用Target Scan,PITA,microRNAorg数据对差异mi RNA进行靶基因预测,进行靶基因富集的基因功能(GO)和信号通路(KEGG)分析。结果显示,大气细颗粒物暴露2周后小鼠肺组织microRNAs有显著差异表达谱,高剂量暴露组与对照组比较有4个mi RNAs上调,低剂量暴露组与对照组比较有2个mi RNAs上调,高剂量组与低剂量组比较,有4个mi RNAs上调(标准为fold change值>=2.0且P值<=0.05),挑选差异明显的mi RNAs进行q RT-PCR验证,mi R-139-5p、mi R-691及mi R-340-3p变化趋势与芯片一致,生物信息学结果显示,差异表达的mi RNAs所调控的靶基因明显富集于34个GO通路(包括RNA转录酶II启动子的转录,RNA拼接,DNA模板,蛋白质结合和核酸结合)和32个KEGG通路(主要集中轴突导向通路和癌症通路)。综上所述,大气细颗粒物暴露染毒可诱导小鼠肺组织中mi R-139-5p、mi R-691及mi R-340-3p明显上调,且生物信息学分析提示中枢神经系统发育及癌症通路可能作为PM2.5暴露相关差异表达mi RNAs调控靶基因介导的主要致病通路。

MicroRNAs can be regarded as new biomarkers for the early diagnosis of clinical diseases, which has been paid more and more attentions. In order to investigate the pathogenic mechanism of air pollution, BALB/cmice were treated with noninvasive tracheal instillation of PM2.5 suspension to construct experimental animal model.Differentially expressed microRNAs in mouse lung tissues were screened by Agilent chip, and the results were verified by real-time quantitative PCR. Target genes prediction of differential mi RNAs was performed using Target Scan, PITA, and micro RNAorg. Then, target gene enrichment, gene function（GO） and signal pathway（KEGG）analysis were performed, and differentially expressed mi RNAs were detected after exposed to particulate matter for2 weeks. 4 mi RNAs were up-regulated in high dose exposure group compared to control group, 2 mi RNAs were up-regulated in low dose exposure group compared to control group, and 4 mi RNAs were up-regulated in high dose exposure group compared to low dose exposure group（standard value = 2 and the P value = 0.05）. The expression of mi R-139-5 p, mi R-691 and mi R-340-3 p detected by q RT-PCR exhibited similar patterns of up-regulation to those shown in microarray results. Bioinformatics analysis revealed that the target genes of mi RNAs were significantly enriched in 34 GOs（including transcription from RNA polymerase II promoter, RNA splicing, DNAtemplated, protein binding, RNA binding, DNA binding） and 32 signal pathways（including mainly axon guidance and pathways in cancer）. In conclusion, the expression levels of mi R-139-5 p, mi R-691 and mi R-340-3 p in mice lung tissues were up-regulated remarkably after exposure to particulate matter. Bioinformatics analysis suggested that central nervous system development and cancer pathway may be the major pathogenic pathway mediated by differentially expressed mi RNAs related to target genes.