Background: Atrial fibrillation (AF) is the most common persistent arrhythmia. Valvular heart disease (VHD) and AF frequently coexist. In our study, from performing bioinformatics analysis, we sought to identify immune-related genes (IRGs) and explore the role of immune cell infiltration in AF-VHD in depth, aiming at investigating the potential molecular mechanism and developing new therapeutic targets for AF, including AF-VHD. Methods: The gene expression of the GSE41177 and GSE79768 datasets were downloaded from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) were analyzed via the limma package in Bioconductor with R software. Differentially expressed immune-related genes (DEIRGs) were selected via combination ImmPort database with DEGs, and the enrichment function and pathway analysis were explored. A protein-protein interaction (PPI) network was built with a Search Tool for the Retrieval of Interacting Genes/Proteins plugin in Cytoscape. The CIBERSORT algorithm was used to evaluate immune infiltration in the left atrial (LA) tissues between AF-VHD and sinus rhythm (SR) patients. Finally, a correlation analysis between key DEIRGs and infiltrating immune cells was performed.Results: A total of 130 DEIRGs were detected. Enrichment function of DEIRGs demonstrated that they are significant in immune and inflammatory responses. The key DEIRGs assessed by the PPI network and involved in both the immune and inflammatory responses were the C-X-C motif chemokine ligand (CXCL) 1, pro-platelet basic protein (PPBP), CXCL12, and C-C motif chemokine ligand 4 (CCL4). The immune infiltration findings indicated that, compared with the LA tissues from SR patients, the tissues from AF-VHD patients contained a higher proportion of gamma delta T cells, but a lower proportion of CD8 and regulatory T cells. The results of correlation analysis demonstrated that CXCL1 was positively correlated with activated mast cells and significantly negatively correlated with resting mast cells. PPBP, CXCL12, and CCL4 were positively correlated with the infiltration of various immune cells, such as neutrophils, plasma cells, and resting dendritic cells. Conclusions: The key immune-related genes and the differences in immune infiltration in LA tissues play an essential role in the occurrence and progression of AF-VHD.

Several studies have indicated that long noncoding RNAs (lncRNAs)-HOX transcript antisense RNA (HOTAIR) is involved in some cardiovascular diseases by regulating gene expression as a competitive endogenous RNA (ceRNA). GJA1 encoding Cx43 is one potential target gene of microRNA-613 (miR-613). Meanwhile, there is a potential target regulatory relationship between HOTAIR and miR-613. The present study is aimed at investigating whether HOTAIR functions as a ceRNA to regulate the Cx43 expression in atrial fibrillation (AF) by sponging miR-613. The expressions of HOTAIR, miR-613, and Cx43 were detected in the right atrial appendages of 45 patients with heart valve disease, including 23 patients with chronic AF. The HOTAIR overexpressed and underexpressed HL-1 cell model were constructed to confirm the effect of HOTAIR on Cx43. Then, the Cx43 expression was detected to testify the interplay between HOTAIR and miR-613 after cotransfecting HOTAIR and miR-613. Furthermore, luciferase assays were performed to verify that HOTAIR could regulate Cx43 remolding as a ceRNA by sponging miR-613. The expression of HOTAIR and Cx43 was significantly downregulated in chronic AF group. HOTAIR regulated positively the Cx43 expression in HL-1 cells. The upregulated effect of HOTAIR on the Cx43 expression could be remarkably attenuated by miR-613. Moreover, the inhibitory effect of miR-613 on the Cx43 expression could be obviously mitigated by HOTAIR. At last, luciferase assays confirmed HOTAIR functioned as a ceRNA in the Cx43 expression by sponging miR-613. Our study suggests that HOTAIR, functioning as a ceRNA by sponging miR-613, is an important contributor to Cx43 remolding in AF.

Atrial fibrillation (AF) is one of the most common forms of cardiac arrhythmia. Novel evidence has indicated that a competing endogenous RNA (ceRNA) mechanism may occur in AF. The present study aimed to identify differentially expressed microRNAs (miRNAs/miRs) in AF and predict their targeting long non-coding RNAs (lncRNAs) to identify a potential ceRNA network involved in AF using bioinformatics analysis. The GSE68475 microarray dataset was downloaded from the Gene Expression Omnibus database and differentially expressed miRNAs in AF were obtained. In addition, right atrial appendage (RAA) tissues from patients with AF were collected to determine the expression levels of the miRNAs identified following bioinformatics analysis using reverse transcription-quantitative PCR (n=8 per group). Subsequently, Gene Ontology (GO) functional term and Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathway enrichment analyses of the target genes of differentially expressed miRNAs of interest were performed. The potential upstream lncRNAs targeting the identified miRNAs were predicted using bioinformatics analysis. A dual luciferase reporter assay was used to verify the existence of a targeted relationship between the differentially expressed miRNA and lncRNA of interest. The results identified 43 differentially expressed miRNAs, including 23 upregulated miRNAs. The trends in the expression levels of miR-223-3p were inconsistent between the microarray data and those recorded in the RAA tissues from patients with persistent AF. Therefore, miR-223-3p was selected as the miRNA of interest for further investigations. The target gene of miR-233-3p was found to be enriched in 57 GO terms and 21 KEGG signaling pathways. According to the bioinformatics prediction, 69 lncRNAs targeting miR-223-3p were identified, including the lncRNA growth arrest-specific transcript 5, lncRNA KCNQ1 opposite strand/antisense transcript 1 (KCNQ1OT1) and lncRNA MYC-induced long non-coding RNA. The results from dual luciferase assay confirmed that miR-223-3p was a direct target of KCNQ1OT1. A ceRNA regulatory relationship may exist between KCNQ1OT1 and miR-223-3p in AF, providing therefore a novel potential research target for further studies.