BCL6, DUSP3, and IL6R Are Identified as Shared Druggable Immune-Regulatory Axis in Atrial Fibrillation and...

Atrial fibrillation (AF) and atherosclerosis (ATH) are increasingly recognized as interconnected cardiovascular conditions with shared immune and inflammatory underpinnings. However, the molecular mechanisms linking their pathogenesis remain poorly defined.

A multiplatform transcriptomic analysis was conducted using publicly available microarray datasets for AF and ATH. Differentially expressed genes (DEGs) were identified using linear modeling and batch correction. A total of 29 overlapping DEGs were found between AF and ATH, from which five immune-related DEGs were identified using the ImmPort database. LASSO regression selected three genes, that is, BCL6, DUSP3, and IL6R, as optimal immune-regulatory hub genes. Functional enrichment, drug-target interaction profiling, transcriptional regulatory network modeling, immune infiltration estimation, and single-cell RNA-seq analysis were conducted using R-based pipelines, DGIdb, iRegulon, CIBERSORTx, ImmuCellAI, and CellChat. To experimentally validate their regulatory role in AF, in vitro assays were performed using angiotensin II-treated mouse cardiac fibroblasts (MCFs). Gene-specific knockdown was achieved via siRNA transfection, followed by RT-qPCR, western blotting, colony formation, wound healing, and proliferation assays using Thermo Fisher-validated kits and reagents.

Transcriptomic pathway enrichment revealed strong involvement of MAPK signaling, phosphatase regulation, and T cell immunomodulatory pathways. Drug-gene interaction analysis identified four immune DEGs as druggable targets. Transcription factor regulatory modeling identified nine TFs converging on BCL6, DUSP3, and IL6R. Immune deconvolution analysis revealed macrophage and dendritic cell enrichment in both conditions, with broader immune remodeling in AF. Single-cell RNA-seq localized BCL6, DUSP3, and IL6R to T cells, macrophages, and fibroblasts, with divergent intercellular signaling inferred via CellChat. In vitro, siRNA-mediated knockdown of BCL6, DUSP3, and IL6R in Ang II-stimulated cardiac fibroblasts significantly suppressed their expression and attenuated fibroblast activation, while CUL4A knockdown showed supporting effects showing the pathogenic relevance of the core immune-regulatory axis in AF-ATH.

This integrative study identifies BCL6, DUSP3, and IL6R as shared immune-regulatory genes in AF and ATH, with transcriptomic and in vitro evidence supporting their pathogenic role and potential as dual-disease therapeutic targets.