SMC-derived foam cells regulate atherosclerotic plaque stability via the crosstalk between ECM remodeling and WNT signaling

Abstract Purpose The study investigates atherosclerotic plaque instability via multi-omics integration, focusing on Smooth Muscle Cell (SMC)-to-foam cell transition, ECM remodeling, and WNT signaling. while analyzing their epigenetic basis to develop biomarkers and therapies. Method Bulk RNA sequencing (24 human atherosclerotic plaques) and single-cell RNA-seq (3 human atherosclerotic plaques) are analyzed to dissect plaque instability. Bulk RNA analysis included PCA for transcriptional heterogeneity assessment and DESeq2 for identifying differentially expressed genes (DEGs). Reactome pathway enrichment of upregulated DEGs revealed WNT signaling activation in unstable plaques. For scRNA-seq, Seurat and UMAP resolved 10 cell clusters, including SMCs and foam cells. Diffusion mapping (DM) and pseudotime analysis delineated SMC-to-foam cell transitions, with Reactome enrichment implicating extracellular matrix (ECM)-related pathway in phenotypic regulation. Then we quantify the gene expression dynamics of key pathways (e.g., ECM remodeling, Wnt signaling) across pseudotemporal trajectories. hdWGCNA identified foamification-associated gene modules, and gene regulatory networks (GRNs) were analyzed for transcription factor binding sites (TFBS) with TCF4 identified as a key regulatory hub. Using differential methylation analysis confirmed epigenetic regulation drives foam cell formation. Results SMC foamification drives atherosclerotic plaque instability through synergistic ECM degradation and WNT signaling dysregulation. Early-phase upregulation of ECM remodeling genes (SPARC, FN1, VCAN) drives collagen breakdown and inflammation, destabilizing the fibrous cap. ECM degradation products further activate the WNT antagonist SOST, suppressing β-catenin signaling and amplifying ECM destruction via a self-reinforcing loop. In later stages, overexpression of the WNT agonist RSPO2 enhances β-catenin activity, driving SMC transition to a pro-inflammatory phenotype while silencing contractile markers (ACTA2, MYH11) weakens fibrous cap structure, accelerating plaque vulnerability. TCF4 functions as a dual regulatory hub for ECM degradation and WNT signaling dysregulation during SMC foamification, TCF4 is markedly upregulated in unstable plaques, correlating with enhanced collagen breakdown and WNT pathway suppression. Plaque stability analyses further validate these findings, demonstrating that TCF4 overexpression aligns with fibrous cap thinning, calcification, and inflammatory activation. TCF4's dual role in ECM-WNT crosstalk and epigenetic regulation establishes it as a therapeutic target for stabilizing vulnerable plaques. Genome-wide methylation analysis links hypomethylated ECM-receptor/WNT pathway genes in unstable plaques to plaque rupture via epigenetic dysregulation. Conclusion TCF4 mediates ECM organization and WNT signaling to promote unstable atherosclerotic plaque formation, offering novel diagnostic and therapeutic targets for atherosclerosis.