脑神经胶质瘤替莫唑胺耐药 circRNA-miRNA-mRNA调控网络的构建

目的 筛选脑神经胶质瘤替莫唑胺耐药细胞株U87/TR与亲本细胞株U87的环状RNA(circRNA)差异表达谱以及构建关键环状RNA(circRNA)-微小RNA(miRNA)-信使RNA(mRNA)调控网络.方法 2020年1—6月,应用circRNAs芯片检测神经胶质瘤替莫唑胺耐药细胞circRNAs差异表达谱,并进行RT-PCR验证.用miRanda数据库预测circRNA-miRNA靶向结合关系,通过miRanda v5、TargetScan、miBase数据库预测靶基因,Cytoscape软件构建circRNA-miRNA-mRNA调控网络.结果 circRNA芯片结果显示,与U87细胞比较,U87/TR细胞有313个差异有统计学意义表达的circRNAs(P<0.05),其中145个显著上调,168个明显下调;RT-qPCR验证结果与芯片结果相符.miRNA靶位点预测结果显示,关键上调人环状RNAhsa_circRNA_009054,hsa_circRNA_104338(n=3,t=12.09、9.52,P=0.007、0.011)以及下调hsa_circRNA_016459,hsa_circRNA_101975(n=3,t=13.86、7.75,P=0.005、0.016)可与hsa-miR-33a-5p,hsa-miR-15b-3p以及hsa-miR-193b-5p,hsa-miR-23a-5p等多个miRNAs靶向结合,关键circRNA-miRNA-mRNA调控网络包括4个关键circRNAs,20个关键miRNAs,以及278个关键mRNAs.结论 cir?cRNA在脑神经胶质瘤替莫唑胺耐药细胞中异常表达,hsa_circRNA_009054,hsa_circRNA_104338以及hsa_circRNA_016459,hsa_circRNA_101975等关键circRNA可能通过circRNA-miRNA-mRNA调控网络参与脑神经胶质瘤替莫唑胺的耐药过程.

Construction of circRNA-miRNA-mRNA regulatory network in glioma with temozolomide resistance

Objective To screen the differential expression profile of Circular RNA (circRNA) in brain glioma temozolomide-resistant cell line U87/TR and parental cell line U87, and to construct a key Circular RNA (circRNA) -microRNA (miRNA) -messenger RNA(mRNA) regulatory network.Methods From January to June 2020, circRNAs chip was used to detect the differential expression profile of circRNAs in glioma-resistant temozolomide cells and verified by RT-PCR. miRanda database was used to predict the circRNA-miRNA target binding relationship. miRanda v5, TargetScan, and miBase databases were used to predict the target genes. Cytoscape software was used to construct the circRNA-miRNA-mRNA regulatory network.Results The results of circRNA microarrayshowed that U87/TR cells had 313 significantly differentially expressed circRNAs compared with U87 cells (P<0.05), of which 145 were significantly up-regulated and 168 were significantly down-regulated, RT-PCR verification results were consistent with the results of the chip. The prediction results of miRNA target sites showed that the key up-regulation of hsa_circRNA_009054, hsa_circRNA_ 104338 (n=3, t=12.09, 9.52, P=0.007, 0.011) and down-regulation of hsa_circRNA_016459, hsa_circRNA_101975 (n=3, t=13.86, 7.75, P=0.005, 0.016) can target multiple miRNAs such as hsa-miR-33a-5p, hsa-miR-15b-3phsa-miR-193b-5p and hsa-miR-23a-5p, the key circRNA-miRNA-mRNA regulatory network included 4 key circRNAs, 20 key miRNAs, and 278 key mRNAs.Conclusion circRNA is abnormally expressed in temozolomide-resistant cells of cerebral glioma. Key circRNAs such as hsa_circRNA_009054,hsa_circRNA_104338 and hsa_circRNA_016459, hsa_circRNA_101975 and other key circRNAs may participate in the drug resistance process of temozolomide in brain glioma through the circRNA-miRNA-mRNA regulatory network.