Integrated analysis of differential lncRNA and mRNA expression in cholelithiasis based on high-throughput sequencing and bioinformatics

Abstract Background: The etiology of GSD (gallstone disease)has not been fully elucidated. Thus, the current study was designed to analyze the expression profiles of lncRNAs and mRNAs to unveil the potential mechanisms and investigate the role of lncRNA in GSD.Methods:Four gallstone and four control samples were collected and used for RNA sequencing. Differentially expressed lncRNAs (DElncRNAs) and mRNAs (DEmRNAs) were identified between gallstone and control samples by limma R package. The biological function of DElncRNAs and DEmRNAs were analyzed by clusterProfiler R package and IPA software. The Pearson correlation between DElncRNAs and DEmRNAs were calculated to construct the co-expression network of DElncRNAs and DEmRNAs. Moreover, to explore the role of lncRNA in gallstone, the cis- and trans-regulatory network of selected lncRNAs were constructed and visualized by Cytoscape software. In addition, we also constructed a ceRNA regulatory network in gallstone disease. To validate the RNA-sequencing data, we performed RT-qPCR to determine the expressions of top 4 DEmRNAs and DElncRNAs in gallstone and control samples.Results:A total of 934 DEmRNAs and 304 DElncRNAs were identified between gallstone and control samples, respectively. Functional enrichment analysis revealed that both DElncRNAs and DEmRNAs were involved in metabolism-related biological functions. The results of correlation analysis showed that the expressions of 597 DEmRNAs had extremely strong relationship with the expressions of 194 DElncRNAs. A cis-lncRNA-mRNA and trans-lncRNA-TF-mRNA regulatory network were constructed. In addition, a ceRNA composed of 24 DElncRNAs, 201 DEmRNAs and 120 predicted miRNAs was constructed by Cytoscape software. Moreover, by RT-qPCR, we found that the expressions of AC004692.4, HECW1-IT1, SFRP4 and COMP were significantly higher, whereas the expressions of LINC01564, SLC26A3, RP1-27K12.2 and GSTA2 were remarkedly lower in gallstone samples compared with those in control ones, which were consistent with the sequencing results.Conclusion: Our study investigated the role of DElncRNAs and DEmRNAs in gallstone, which may help us understand the etiology of gallstone on the genetic level.