Construction of atrial fibrillation-related circRNA/lncRNA-miRNA-mRNA regulatory network and analysis of potential biomarkers

Abstract

Background: The specific pathogenesis of atrial fibrillation (AF) remains unclear. In this study, we examined the expression of differential messenger RNAs (mRNAs), circular RNAs (circRNAs), and long-stranded noncoding RNAs (lncRNAs) from human peripheral blood mononuclear cells to initially construct a circRNA/lncRNA-miRNA-mRNA ceRNA regulatory network to explore the pathogenesis of AF and to screen for potential biomarkers.

Methods: A total of four pairs of AF cases and healthy subjects were selected to detect differentially expressed mRNAs, circRNAs, and lncRNAs in peripheral blood mononuclear cells by microarray analysis. And 20 pairs of peripheral blood from AF patients and healthy subjects were selected for validation of mRNA, circRNA, and lncRNA by quantitative real-time PCR (qRT-PCR).The relevant ceRNA networks were constructed by GO and KEGG and correlation analysis.

Results: The results showed that compared with healthy subjects, there were 813 differentially expressed mRNAs (DEmRNAs) in peripheral blood monocytes of AF, including 445 upregulated genes and 368 downregulated genes, 120 differentially expressed circRNAs (DEcircRNAs), including 65 upregulated and 55 downregulated, 912 differentially expressed lncRNAs (DElncRNAs), including 531 upregulated and 381 downregulated lncRNAs. GO and KEGG analysis of DERNA revealed the biological processes and pathways involved in AF. Based on microarray data and predicted miRNAs, a ceRNA network containing 34 mRNAs, 212 circRNAs, 108 lncRNAs, and 38 miRNAs was constructed.

Conclusion: We revealed a novel ceRNA network in AF and showed that downregulated XIST, circRNA_2773, and CADM1 were negatively correlated with miR-486-5p expression and had a potential targeting relationship with miR-486-5p.

Keywords: atrial fibrillation; biomarkers; ceRNA network; circRNA; lncRNA; mRNA.

FIGURE 1

mRNA expression profiles. (A) Volcano map of DEmRNA, red and green indicate up‐ and downregulation, respectively. (B) Hierarchical clustering analysis heat map analysis of DEmRNA. Blue: low expression; red: high expression. “T” represents AF samples and “C” represents healthy controls. (C) GO analysis of DEmRNA. (D) KEGG analysis of DEmRNA.

FIGURE 2

lncRNA expression profiles. (A) Volcano plot of DElncRNA, red and green indicate up‐ and downregulation, respectively. (B) Hierarchical clustering analysis heat map analysis of DElncRNA. Blue: low expression; red: high expression. “T” represents AF samples and “C” represents healthy controls. (C) GO analysis of DElncRNA. (D) KEGG analysis of DElncRNA. (E) Gene length. (F) lncRNA number of exons.

FIGURE 3

Establishment of circRNA/lncRNA‐miRNA‐mRNA network. (A) circRNA‐miRNA‐mRNA competitive endogenous RNA network. Triangle pink indicates circRNA, V‐shaped purple indicates miRNA, and parallelogram yellow indicates mRNA. (B) lncRNA‐miRNA‐mRNA competitive endogenous RNA network. Diamond green denotes lncRNA, V‐shaped purple denotes miRNA, and parallelogram yellow denotes mRNA. (C) circRNA/lncRNA‐miRNA‐mRNA competitive endogenous RNA network. (D) Venn diagram of circRNA‐miRNA, lncRNA‐miRNA, and mRNA‐miRNA intersections.

FIGURE 4

Establishment of circRNA/lncRNA‐miRNA‐hub mRNA network. (A) Scores of the top 10 hub genes. (B) Key modules. (C) The circRNA/lncRNA‐miRNA‐mRNA competitive endogenous RNA network constructed based on hubmRNA. triangle pink indicates circRNA, diamond green indicates lncRNA, V purple indicates miRNA, and parallelogram yellow indicates mRNA. (D) The interaction of crucial gene and atrial fibrillation.

FIGURE 5

Protein and protein interaction network of differentially expressed genes

FIGURE 6

Expression of hub genes in microarrays

FIGURE 7

Differential expression of ceRNA‐qRT‐PCR validation

FIGURE 8

Correlation of ceRNA expression levels with differential expression