Abstract Body: Background: Atherosclerosis is a multicellular pathology involving dynamic interactions of infiltrated and resident cells in the arterial wall. To understand the contribution and phenotypic plasticity of adventitial fibroblasts to atherogenesis, the spatiotemporal dynamics of fibroblasts were determined during progression of atherosclerosis.
Methods: First, fibroblast clusters were analyzed in human coronary artery single-cell RNA-sequencing (scRNA-seq) from datasets (GSE131778). Second, atherosclerosis was induced in male C57BL/6J mice by administration of Ldlr antisense oligonucleotides (ASO) in combination with Western diet feeding and angiotensin II infusion for 4 or 12 weeks to generate early and progressive stages of lesions, respectively. These results were compared to data from mice that did not receive Ldlr ASO, Western diet, or angiotensin II. Aortic roots were harvested for integrated scRNA-seq and spatial transcriptomics analyses to map the phenotypic evolution and spatial distribution of fibroblasts. Since decorin (DCN) is highly abundant in fibroblasts, fibroblast clusters were defined based on the presence of DCN and five other major fibroblast markers (PI16, LUM, DPP4, LY6A, and LY6C1).
Results: DCN was highly enriched in fibroblasts present in human coronary arteries, consistent with its use as a fibroblast marker. scRNA-seq of mouse tissue revealed that Dcn⁺ cells increased progressively during atherosclerotic development. The largest expansion of Dcn⁺ fibroblast clusters was characterized by increased mRNA abundance of extracellular matrix genes (Col1a1, Fn1) but reduced contractile markers (Myh11, Cnn1) as lesions progressed. Spatial transcriptomics revealed progressive redistribution with migration of adventitia to intima. While Dcn⁺ cells were confined to the adventitia in the non-atherosclerotic state, this cluster infiltrated the medial layers and accumulated within the intima during progression of atherosclerosis, accompanied by a parallel accumulation of Lyz2⁺ cells in atherosclerotic lesions.
Conclusion: Dcn⁺ cells exhibit phenotypic transition and migrate from the adventitia to the intima during progression of atherosclerosis. We will validate these findings using in vivo Dcn-lineage tracing and complementary in vitro systems.
Key Words: Decorin, Atherosclerosis, Fibroblasts, Single-cell, Spatial Transcriptomics