基于微RNA-mRNA调控网络对草酸钙肾结石形成关键分子的筛选和分析

Screening and analyses of core molecules in calcium oxalate nephrolithiasis based on microRNA-mRNA regulatory network

目的应用生物信息学技术构建草酸钙肾结石大鼠miRNA-mRNA调控网络并筛选出调控草酸钙肾结石形成的关键分子。方法从基因表达汇编(GEO)数据库中筛选出乙二醇喂养14 d诱导的草酸钙肾结石大鼠肾组织的miRNA和mRNA数据集,利用GEO2R在线工具分析得到差异表达miRNA(DEM)和差异表达基因(DEG),并用热图进行展示。利用在线靶基因预测工具miRWalker预测DEM的靶基因,根据miRNA负向调控mRNA原理应用R语言“venn.diagram”包与DEG取交集,得到参与调控草酸钙肾结石形成的基因,然后应用“clusterprofile”包对调控基因进行基因本体(GO)和京都基因与基因组百科全书(KEGG)富集分析,利用String数据库进行蛋白质-蛋白质相互作用(PPI)网络分析,利用Cytoscape软件实现对PPI和miRNA-mRNA调控网络可视化和关键基因的筛选。结果共分析得到38个DEM和364个DEG。将预测得到的DEM靶基因与DEG取交集后得到116个调控基因,这些基因富集在24个GO条目和22个KEGG信号通路中。成功构建了miRNA-mRNA调控网络,处于核心调控位置的miRNA为miRNA-6215、miRNA-758-5p和miRNA-214-3p。PPI网络分析筛选出5个关键基因,分别为叉头框转录因子M1(Foxm1)、核分裂周期蛋白80(Ndc80)、细胞周期蛋白依赖性激酶抑制因子2B(Cdkn2b)、Cdc10依赖性转录因子1(Cdt1)和B细胞编码κ轻链多肽抑制基因(Ikbkg)。结论通过构建miRNA-mRNA调控网络,筛选出在草酸钙肾结石形成过程中起关键调控作用的miRNA和基因,为草酸钙结石的防治提供了新的研究思路。

Objective To construct a microRNA(miRNA)-mRNA regulatory network in rats with calcium oxalate nephrolithiasis using bioinformatics technology,and to screen for the related core molecules regulating the formation of calcium oxalate nephrolithiasis.Methods The miRNA and mRNA datasets of rats with calcium oxalate nephrolithiasis induced by glycol feeding for 14 d were screened from the Gene Expression Omnibus(GEO)database.The differentially expressed miRNAs(DEMs)and differentially expressed genes(DEGs)were analyzed by GEO2R online tool,and displayed by heatmap diagrams.The target genes of DEMs were predicted by miRWalker and then were intersected with DEGs to identify the regulatory genes by“venn.diagram”package of R software according to the principle of negative regulation of miRNA to mRNA.“Clusterprofile”package was used to perform Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses of the regulatory genes,and the protein-protein interaction(PPI)analysis was performed by String database.Lastly,Cytoscape software was used to visualize the PPI and miRNA-mRNA regulatory network and screen the hub genes.Results A total of 38 DEMs and 364 DEGs were screened.Totally 116 regulatory genes were obtained through the intersection of DEMs’target genes and DEGs,and they were enriched in 24 GO items and 22 KEGG signaling pathways.The miRNA-mRNA regulatory network was successfully constructed,and the miRNAs at the core regulatory position were miRNA-6215,miRNA-758-5p,and miRNA-214-3p.Five hub genes were identified from PPI network,namely forkhead box M1(Foxm1),nuclear division cycle 80(Ndc80),cyclin dependent kinase inhibitor 2B(Cdkn2b),Cdc10-dependent transcript 1(Cdt1),and inhibitor of nuclear factorκB kinase regulatory subunit gamma(Ikbkg).Conclusion Through constructing miRNA-mRNA regulatory network,miRNAs and genes that play key regulatory roles in the formation of calcium oxalate nephrolithiasis are obtained,which provides new research ideas for the prevention and treatment of calcium oxalate nephrolithiasis.