摘要
目的:应用第二代DNA测序技术及生物信息学法分析肺腺癌组织中环状RNA(circular RNA,circRNA)差异表达的情况及可能的作用,以期为肺腺癌患者的精确诊断和治疗提供有价值的分子标志物。方法:收集3例经病理及免疫组织化学检测确诊为肺腺癌患者的肺腺癌及癌旁肺组织,采用第二代DNA测序法检测腺癌及癌旁肺组织中circRNA的表达情况。利用生物信息学的方法,对差异表达的circRNA进行筛选及分层聚类分析,并通过软件预测结合人工搜索公共数据库的方式,构建相应的circRNA-miRNA-mRNA互作关系网。最后随机选取5种差异表达的circRNA(hsa_circ_0072309、hsa_circ_0008234、hsa_circ_0000199、hsa_circ_0008870和hsa_circ_0000097),采用实时荧光定量PCR法检测它们在30对肺腺癌及癌旁肺组织中的表达水平,以验证测序结果的准确性。结果:3对肺腺癌及癌旁肺组织经第二代DNA测序,共检测到38018种circRNA;通过火山图和聚类分析共得到482种具有显著差异的circRNA,其中70种表达水平显著上调,412种表达水平显著下调。通过绘制circRNA-miRNA-mRNA互作图后发现,差异最明显的4种circRNA对应的miRNA或靶基因(如miR-211和RAB22A)已被证实在肺癌等多种癌症的发生和发展中发挥重要作用。实时荧光定量PCR检测结果显示,hsa_circ_0072309、hsa_circ_0008234、hsa_circ_0000199、hsa_circ_0008870和hsa_circ_0000097在30例肺腺癌组织中的表达水平均明显下调(P值均<0.05),与测序结果一致,说明测序数据是准确可信的。结论:通过二代测序结合生物信息学分析的方法,从肺腺癌组织中筛选出了大量差异表达的circRNA,这为进一步研究circRNA对肺腺癌发生和发展的作用机制奠定了基础。
Objective:To investigate the differential expression of circular RNA(circRNA)in lung adenocarcinoma and its possible role by the second-generation DNA sequencing technology and bioinformatics,so as to provide valuable molecular markers for the accurate diagnosis and treatment of lung adenocarcinoma.Methods:Lung adenocarcinoma tissues and the adjacent lung tissues were collected from 3 patients with lung adenocarcinoma diagnosed by pathological and immunohistochemical detection.The expression of circRNAs in 3 pairs of adenocarcinoma and adjacent lung tissues was detected by the second-generation DNA sequencing.The differentially expressed circRNAs were screened and stratified by bioinformatics,and the corresponding circRNA-miRNA-mRNA interaction network was constructed through software prediction combined with manual search of the public database.Finally,five differentially expressed circRNAs(hsa_circ_0072309,hsa_circ_0008234,hsa_circ_0000199,hsa_circ_0008870 and hsa_circ_0000097)were randomly selected,and their expression levels in 30 pairs of lung adenocarcinoma tissues and adjacent lung tissues were detected by real-time fluorescent quantitative PCR to verify the accuracy of sequencing results.Results:Second generation sequencing showed that the three pairs of cancer tissues and adjacent lung tissues had a total of 38018 circRNAs.Volcano map and cluster analysis showed a total of 482 circRNAs with significant differences between the cancer tissues and adjacent lung tissues,of which 70 were significantly up-regulated and 412 were significantly down-regulated in lung adenocarcinoma tissues.By plotting the circRNA-miRNA-mRNA interaction map,it was found that the microRNAs(miRs)or target genes(such as miR-211 and RAB22A)corresponding to the four circRNAs with the most significant differences played an important role in the occurrence and development of various cancers,including lung cancer.The real-time fluorescent quantitative PCR showed that the expression levels of hsa_circ_0072309,hsa_circ_0008234,hsa_circ_0000
作者
王阳
关键
王耀林
WANG Yang;GUAN Jian;WANG Yaolin(Medical College of Shihezi University,Shihezi 832000,Xinjiang Uygur Autonomous Region,China;Department of Respiratory and Critical Care Medicine,People’s Hospital of Suzhou New District,Suzhou 215010,Jiangsu Province,China)
出处
《肿瘤》
CAS
CSCD
北大核心
2020年第1期41-51,共11页
Tumor
基金
苏州高新区医疗卫生科技计划项目(编号:2018Z001)
苏州高新区人民医院科学创新基金项目(编号:SGY2018C01).
关键词
癌
非小细胞肺
RNA
环状
高通量核苷酸序列分析
计算生物学
Carcinoma,non-small-cell lung
RNA,circular
High-throughput nucleotide sequencing
Computational biology
