肺癌患者血浆外泌体microRNAs标志物的筛选

Screening of plasma exosome microRNAs in patients with lung cancer

目的:筛选肺癌患者血浆外泌体中的异常microRNA(miRNA),以期为肺癌筛查及辅助诊断提供新的标志物和方向。方法:选择6名肺癌患者(腺癌3例、鳞癌2例,小细胞肺癌1例)和4名健康对照者血浆样本,采用差速离心法提取血浆外泌体,采用Agilent Bioanalyzer对抽提的RNA进行质量检验,对血浆外泌体miRNA进行高通量测序分析,采用TargetScan、PITA和miRNAorg数据库进行靶基因预测,应用R软件进行靶基因的KEGG和GO分析。结果:miRNA表达分析发现,与正常对照者比较,肺癌患者血浆外泌体中有142种miRNAs表达异常,其中62种miRNAs表达上调,80种miRNAs表达下调。GO分析显示,这些miRNAs的靶基因主要参与了以DNA为模板的转录、转录调控及信号转导等。KEGG分析显示,这些miRNAs的靶基因主要参与轴突导向信号通路、钙信号通路、肌动蛋白细胞骨架调节等。结论:通过对肺癌患者和健康对照者血浆外泌体miRNAs进行高通量测序,初步筛选出差异表达miRNA,经生物信息学分析,推测其对肺癌转移具有良好预测潜力,有望成为肺癌筛查和辅助诊断的候选生物标志物。

OBJECTIVE:Existence of miRNAs in plasma exosomes of lung cancer patients were screened to develop new methods for the screening and auxiliary diagnosis of lung cancers.METHODS:Plasma samples were collected from 6 lung cancer patients(3 adenocarcinomas,2 squamous carcinomas and 1 small cell lung cancer)and 4 healthy controls.Agilent Bioanalyzer was used for qualitative inspection of the extracted RNA.Plasma exosomal miRNA was analyzed by high-throughput sequencing.Target gene prediction was performed by TargetScan,PITA and miRNAorg database,and R software was applied for KEGG and GO analysis of target genes.RESULTS:The exosomal differential miRNA expression analyses showed that 142 miRNAs were expressed abnormally in the lung cancer group compared with to the normal group,with 62 miRNAs up-regulated and 80 miRNAs down-regulated.GO analysis showed that these abnormally-expressed(target)genes were mainly involved in DNA-based template transcription,transcriptional regulation and signal transduction,etc.The target KEGG analysis indicated that these target genes were mainly involved in axon guidance signaling pathway,calcium signaling pathway,and actin cytoskeleton regulation.CONCLUSION:Our data suggest that differentially expressed miRNAs may have good predictive potential for lung cancer metastasis.In addition,they may be candidate biomarkers for the screening and diagnosis of lung cancers.