Prediction of Functional Genes in Primary Varicose Great Saphenous Veins Using the lncRNA-miRNA-mRNA Network

Abstract

Background: Long noncoding RNAs (lncRNAs) have been widely suggested to bind with the microRNA (miRNA) sites and play roles of competing endogenous RNAs (ceRNAs), which can thus affect and regulate target gene and mRNA expression. Such lncRNA-related ceRNAs are identified to exert vital parts in vascular disease. Nonetheless, it remains unknown about how the lncRNA-miRNA-mRNA network functions in the varicose great saphenous veins.

Methods: This study acquired the lncRNA and mRNA expression patterns from the GEO database and identifies the differentially expressed mRNAs and lncRNAs by adopting the R software "limma" package. Then, miRcode, miRDB, miRTarbase, and TargetScan were used to establish the miRNA-mRNA pairs and lncRNA-miRNA pairs. In addition, the lncRNA-miRNA-mRNA ceRNA network was constructed by using Cytoscape. Protein-protein interaction, Gene Ontology functional annotations, and Kyoto Encyclopedia of Genes and Genomes enrichment were carried out to examine the candidate hub genes, the functions of genes, and the corresponding pathways.

Results: In line with the preset theory, we constructed ceRNA network comprising 12 lncRNAs, 38 miRNAs, and 149 mRNAs. Kyoto Encyclopedia of Genes and Genomes analysis indicated that the PI3K/Akt signaling pathway played a vital part in the development of varicose great saphenous veins. AC114730, AC002127, and AC073342 were significant biomarkers. At the same time, we predicted the potential miRNA, which may exert a significant influence on the varicose great saphenous veins, namely, miR-17-5p, miR-129-5p, miR-1297, miR-20b-5p, and miR-33a-3p.

Conclusion: By performing ceRNA network analysis, our study detects new lncRNAs, miRNAs, and mRNAs, which can be applied as underlying biomarkers of varicose great saphenous veins and as therapeutic targets for the treatment of varicose great saphenous veins.

Figure 1

Acquisition of differential genes. (a) Volcano plot showing RNAs with differential expression. Genes with upregulation and downregulation are denoted as light red and light green, respectively. (b) lncRNAs with differential expression within GSVs. (c) mRNAs with differential expression within GSVs. Red and blue boxes stand for genes with upregulation and downregulation, separately. Six replicates were set for every group. ∣log2(fold change, FC) | >1 and adjusted P < 0.05 were the thresholds for selection.

Figure 2

The ceRNA network for lncRNAs-miRNAs-mRNAs for GSVs. Rectangles, ellipses, and diamonds stand for lncRNAs, mRNAs, and miRNAs denoted as green, blue, and red colors, separately.

Figure 3

GO analysis as well as KEGG analysis. (a) GO: biological process terms. (b) GO: cellular component terms. (c) GO: molecular function terms. (d) The enriched KEGG pathways.

Figure 4

PPI network and hub genes. (a) The PPI network showing DEGs. Node size indicates degree level, with greater nodes indicating the greater degree. (b) Selection of critical mRNAs in PPI network based on DEM connection degree using R.

Figure 5

Critical subnetwork of lncRNAs-miRNAs-mRNAs. (a) The network of lncRNA AC114730-miRNAs-mRNAs. (b) The network of lncRNA AC002127-miRNAs-mRNAs. (c) The network of lncRNA AC073342-miRNAs-mRNAs. lncRNAs, miRNAs, and mRNAs are denoted in green, red, and blue colors, separately.

Figure 6

Critical subnetworks of lncRNAs-miRNAs-mRNAs. (a) lncRNA-miR-17-5p-mRNA subnetwork. (b) lncRNA-miR-1297-mRNA subnetwork. (c) lncRNA-miR-129-5p-mRNA subnetwork. (d) lncRNA-miR-33a-3p-mRNA subnetwork. (e) lncRNA-miR-20b-5p-mRNA subnetwork. Dark green, light green, and blue nodes represent lncRNAs, miRNAs, and mRNAs, respectively.