结肠癌组织中差异表达circRNA的筛选与验证

Screening and verification of differentially expressed circRNA in colon cancer tissues

目的筛选结肠癌组织中差异表达的环状RNA(circRNA),分析其在结肠癌发生、发展中的作用。方法选取该院2018年1月至2019年12月收治的3例结肠癌患者的肿瘤组织标本及配对癌旁正常组织标本,用Arraystar Human circRNA Arrays V2芯片检测、筛选差异表达的circRNA;采用聚类分析筛选出表达水平有明显差异的circRNA。另选取同期9例结肠癌肿瘤组织标本及配对癌旁组织标本用实时荧光定量PCR(qPCR)检测进行初步验证。筛选到的差异表达circRNA进行GO基因功能富集分析(GO分析)及KEGG信号通路富集分析(KEGG分析)。结果结肠癌组织与癌旁组织比较,按照P<0.05且差异表达倍数(FC)>1.5筛选标准,筛选出114个差异表达circRNA,其中62个circRNA上调表达,52个circRNA下调表达,经过聚类分析筛选出在结肠癌中显著下调的8个circRNA:hsa_circRNA_404686、hsa_circRNA_102293、hsa_circRNA_000367、hsa_circRNA_104270、hsa_circRNA_001729、hsa_circRNA_100790、hsa_circRNA_105039、hsa_circRNA_102049。qPCR检测结果显示,这8个circRNA在结肠癌组织中表达水平降低。GO分析显示,被富集的基因主要参与转录调节、RNA结合、蛋白质结合、细胞周期调控等。KEGG分析显示,主要参与调节干细胞多能性的信号通路、FOXO信号通路、p53信号通路等。结论在结肠癌组织筛选到8个显著下调的circRNA,经基因功能注释分析其参与肿瘤的发生、发展,可能会成为结肠癌诊断及预后评估的生物标志物。

Objective To screen out the differentially expressed circular RNA(circRNA)in colon cancer tissues and analyze their possible role in the development of colon cancer.Methods The cancer tissues and matched adjacent tissues of 3 patients with colon cancer were collected from January 2018 to December 2019 in the hospital.Differentially expressed circRNAs were detected and screened by using Arraystar Human circRNA Arrays V2 chip.Cluster analysis was performed to screen out circRNAs with significantly different expression levels.Tissues and matched adjacent tissues from 9 patients with colon cancer admitted from January 2018 to December 2019 were collected to preliminary verify the differentially expressed circRNA by using real-time quantitative PCR(qPCR)method.The screened differentially expressed circRNAs were analyzed by using annotation of circRNA,GO pathway analysis and KEGG pathway analysis.Results A total of 114 differentially expressed circRNAs were detected in colon cancer tissues with>1.5 fold change(FC)in their matched adjacent tissues(P<0.05);62 circRNAs were up-regulated and 52 circRNAs were down-regulated.After cluster analysis,8 circRNAs that were significantly down-regulated in colon cancer were screened out:hsa_circRNA_404686,hsa_circRNA_102293,hsa_circRNA_000367,hsa_circRNA_104270,hsa_circRNA_001729,hsa_circRNA_100790,hsa_circRNA_105039,hsa_circRNA_102049.Compared with adjacent tissues,qPCR test showed that these 8 circRNAs were significantly down-regulated in colon cancer tissues.GO analysis revealed that numerous target genes were involved in transcription regulation,RNA binding,protein binding,cell cycle.KEGG analysis revealed that numerous target genes were involved in signaling pathways regulating pluripotency of stem cell,FOXO signaling pathways,p53 signaling pathways.Conclusion There are 8 circRNAs expression significantly down-regulated in colon cancer tissue.Gene function annotation analysis indicates that the 8 circRNAs may be related to the occurrence and development of tumors,and may become markers for the diagnosis and prognostic evaluation of colon cancer.