Integrative single-cell meta-analysis reveals disease-relevant vascular cell states and markers in human atherosclerosis

Abstract

Atherosclerosis is a complex inflammatory process driven by plaque formation in the major elastic arteries and often leads to reduced blood flow, coronary artery disease (CAD), myocardial infarction and stroke. CAD progression involves complex interactions and phenotypic plasticity within and between distinct vascular and immune cell lineages. Several single-cell RNA-seq (scRNA-seq) studies have highlighted lineage-specific transcriptomic signatures however there remains variability on the reported cell phenotypes in humans. In this study we meta-analyzed scRNA-seq datasets across four publications to create a comprehensive map of human atherosclerosis cell diversity. We applied standardized QC, processing, and integration benchmarking to harmonize 118,578 high-quality cells for this atlas. Beyond characterizing vascular and immune cell diversity, we derived insights into smooth muscle cell (SMC) phenotypic modulation through pseudotime, transcription factor activity inference and cell-cell communication analyses. We also integrated genome-wide association study (GWAS) data to identify etiologic cell types for GWAS diseases and traits, which uncovered a critical role for modulated SMC phenotypes in CAD and coronary artery calcification. Finally, we identified candidate markers (e.g., CRTAC1) of synthetic and osteochondrogenic SMCs that may serve as proxies of atherosclerosis progression. Together, this represents an important step towards creating a unified cellular map of atherosclerosis to inform cell state-specific mechanistic and translational studies of cardiovascular diseases.

Multi-ancestry genetic analysis of gene regulation in coronary arteries prioritizes disease risk loci

“…Fewer than 5% of shared associations had the same lead eQTL, but among shared lead eQTLs, we observed 98% directional consistency (64% of all replicated lead eQTLs, Figures 2 D and S2 ). Nearly one in five eGenes exhibited cell-type-specific expression in a coronary artery single-cell RNA sequencing reference dataset (cell specificity expression score ≥0.7, Figure S2 ; Table S8 ), 30 and functional enrichment analysis revealed several pathways for cell adhesion and inflammation ( Table S9 ). 31 …”

Integrative multi-ancestry genetic analysis of gene regulation in coronary arteries prioritizes disease risk loci

“…Fewer than 5% of shared associations had the same lead eQTL, but among shared lead eQTLs we observed 98% directional consistency (64% of all replicated lead eQTLs, Figure 2d, Figure S2 ). Nearly one in five eGenes exhibited cell-type-specific expression in a coronary artery single-cell RNA sequencing reference dataset (cell-specificity expression score ≥0.7, Figure S2, Table S8 ), 30 and functional enrichment analysis revealed several pathways for cell adhesion and inflammation ( Table S9 ). 31…”

“…Coronary artery single-cell RNA and Assay for Transposase-Accessible Chromatin sequencing (scRNA-seq, snATAC-seq) reference datasets show that TARID and TCF21 are both expressed more highly and more accessible in fibroblasts and SMCs (including SMC-derived fibromyocytes resulting from SMC phenotypic switching during atherosclerosis, Figures 3d,e, S3 ). 30,37…”

“…The ARHGAP42 eQTL overlapped with GTEx AOR and COR associations, and colocalized with both BP traits and CAD GWAS associations ( Table S12 , Figure S5c,d ). ARHGAP42 (also known as GRAF3) regulates vascular tone via expression predominantly in mural cells and fibroblasts ( Figure S5e,f ), 30,37,39 and insufficiency causes hypertension. 40,41 rs2455569 is located in a pericyte-specific chromatin-accessibility peak in the first intron of ARHGAP42 , with fine-mapping highlighting proximal SNPs encompassing a region accessible in multiple vascular cell types ( Figure 4a-c ).…”

“…Furthermore, eGenes colocalizing to one GWAS trait can be prioritized for functional characterization, particularly those exhibiting cell-type specificity– e.g., Heparanase 2 ( HPSE2 ), is enriched in SMCs in coronary artery and for which our eQTL colocalizes with SBP and DBP. 30 HPSE2 would not be a strong candidate gene in a QTL study focused on disease outcomes rather than intermediate phenotypes, but recent characterization of this extracellular matrix protein in endothelial maintenance suggests a contribution to vascular function during inflammation. 55,56…”