南方医科大学学报

南方医科大学学报 ›› 2019, Vol. 39 ›› Issue (07): 784-.doi: 10.12122/j.issn.1673-4254.2019.07.06

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胃癌中miRNA-340 的体内外增殖能力检测及生物信息分析

王健，陈文静，林慧娟，张江宇   

  • 出版日期:2019-07-20 发布日期:2019-07-20

Role of miRNA-340 in modulating gastric cancer cell proliferation and bioinformatic analysis

  • Online:2019-07-20 Published:2019-07-20

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摘要： 目的了解miRNA-340在胃癌发生中的可能机制，在线预测与其相互作用的circRNAs及其下游靶基因和参与的信号通 路。方法利用芯片筛选胃癌细胞株中差异表达miRNA；利用Real-time RT-PCR，检测miRNA-340在21对胃癌组织与癌旁正 常组织中的表达水平；利用MTT及EdU细胞增殖实验，检测miRNA-340对正常胃上皮细胞HFE145与胃癌细胞BGC-823的体 外增殖能力；利用裸鼠成瘤实验检测miRNA-340在体内成瘤的能力。随后，通过在线软件Targetscan、David数据库及Starbase 分别对miRNA-340 的下游的靶基因、相关信号通路及相互作用的circRNAs 进行生物信息分析。结果与正常胃上皮细胞 HFE145相比较，miRNA-340在胃癌细胞株中的表达水平显著下调；Real-time RT-PCR结果显示，miRNA-340在胃癌组织中的 表达明显低于正常粘膜组织（P<0.048）；体内体外实验表明miRNA-340对细胞的体外增殖能力具有一定的抑制作用；生物信息 分析显示，预测的miRNA-340相互作用的下游靶基因中，筛选出21个具有3个及以上的保守结合部位的靶基因，其中多个靶基 因均参与肿瘤的发生及侵袭；相互作用的circRNAs中筛选出分值前10个circRNAs作为与miRNA-340可能作用最强的sponge circRNAs。结论miRNA-340 在肿瘤发生发展中可能起到了抑癌基因的作用，对胃癌细胞的增殖具有一定的抑制作用； miRNA-340与多个circRNAs可能存在相互作用，共同调控下游靶基因参与胃癌的发生发展。

Abstract: Objective To investigate the mechanism of miRNA-340 for regulating the proliferation of gastric cancer (GC) cells and predict its interacting circular RNAs (circRNAs), its downstream target genes and the involved signaling pathways. Methods The differentially expressed miRNAs in GC cell lines were analyzed and screened using miRNA microarrays. The expression level of miRNA-340 in 21 pairs of GC tissues and adjacent normal tissues was detected using real-time PCR. MTT and EdU assays were performed to examine the effect of miRNA-340 on the proliferation ability of HFE145 and BGC-823 cells. We also tested the effect of miRNA-340 inhibition on subcutaneous tumorigenesis of GC cells in a nude mouse model. The downstream target genes of miRNA-340 and the probable signal pathways were predicted online using Targetscan and DAVID database, respectively. The interacting circRNAs of miRNA-340 were analyzed using starBase platform. Results Among the differentially expressed miRNAs, miRNA-340 was significantly down-regulated in GC cell lines. Real-time PCR results showed that the expression of miRNA-340 was significantly lower in GC tissues than in the adjacent tissues (P<0.05). MTT and EdU cell proliferation assays showed that miRNA-340 overexpression inhibited the proliferation of GC cells in vitro. In the nude mouse models, the proliferation of GC cells transfected with miRNA-340 inhibitor was obviously enhanced. Bioinformatics analysis suggested that miRNA-340 had 21 target genes with 3 or more conserved sites, and these genes were involved in tumorigenesis and invasion. The top 10 circRNAs were selected as the most powerful sponge circRNAs interacting with miRNA-340. Conclusion miRNA-340 may play the role of a tumor suppressor in tumorigenesis and progression. Overexpression of miRNA-340 suppress the proliferation of GC cells, suggesting its involvement in the development of GC along with multiple circRNAs.