Atlas of the Immune Cell Repertoire in Mouse Atherosclerosis Defined by Single-Cell RNA-Sequencing and Mass Cytometry
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Rationale: Atherosclerosis is a chronic inflammatory disease that is driven by the interplay of pro- and anti-inflammatory leukocytes in the aorta. Yet, the phenotypic and transcriptional diversity of aortic leukocytes is poorly understood. Objective: We characterized leukocytes from healthy and atherosclerotic mouse aortas in-depth by single-cell RNA-sequencing and mass cytometry (cytometry by time of flight) to define an atlas of the immune cell landscape in atherosclerosis. Methods and Results: Using single-cell RNA-sequencing of aortic leukocytes from chow diet– and Western diet–fed Apoe −/− and Ldlr −/− mice, we detected 11 principal leukocyte clusters with distinct phenotypic and spatial characteristics while the cellular repertoire in healthy aortas was less diverse. Gene set enrichment analysis on the single-cell level established that multiple pathways, such as for lipid metabolism, proliferation, and cytokine secretion, were confined to particular leukocyte clusters. Leukocyte populations were differentially regulated in atherosclerotic Apoe −/− and Ldlr −/− mice. We confirmed the phenotypic diversity of these clusters with a novel mass cytometry 35-marker panel with metal-labeled antibodies and conventional flow cytometry. Cell populations retrieved by these protein-based approaches were highly correlated to transcriptionally defined clusters. In an integrated screening strategy of single-cell RNA-sequencing, mass cytometry, and fluorescence-activated cell sorting, we detected 3 principal B-cell subsets with alterations in surface markers, functional pathways, and in vitro cytokine secretion. Leukocyte cluster gene signatures revealed leukocyte frequencies in 126 human plaques by a genetic deconvolution strategy. This approach revealed that human carotid plaques and microdissected mouse plaques were mostly populated by macrophages, T-cells, and monocytes. In addition, the frequency of genetically defined leukocyte populations in carotid plaques predicted cardiovascular events in patients. Conclusions: The definition of leukocyte diversity by high-dimensional analyses enables a fine-grained analysis of aortic leukocyte subsets, reveals new immunologic mechanisms and cell-type–specific pathways, and establishes a functional relevance for lesional leukocytes in human atherosclerosis.
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scRNA-seq has been widely used to reveal the characteristics of immunity in different fields.
“It is notable that scRNA-seq has been widely used to reveal the characteristics of immunity in different fields. 6-10”
From Single-Cell RNA Sequencing to Dissect the Immunological Network of Autoimmune Myocarditis. (2020) · cites this paper as [7]
The Results section ('Cell-type specific genetic pathways in aortic leukocytes'; page unavailable) uses scRNA-seq to characterize immune-cell populations and pathways in mouse atherosclerosis. This supports an example of the method revealing immune characteristics, but the paper does not establish that it is widely used across different fields. (LLM, conf=0.83)
Experimental — accuracy not yet measuredHuman and mouse atherosclerotic plaques contain mostly macrophages.
“Both human and mouse atherosclerotic plaques are populated mostly by macrophages134,135.”
From Inflammation and its resolution in atherosclerosis: mediators and therapeutic opportunities (2019) · cites this paper as [135]
Single-cell analyses of unique macrophage subsets such as TREM2+ macrophages have revealed extensive phenotypic heterogeneity of macrophages and monocyte-derived dendritic cells in atherosclerotic lesions and produced a novel immune cell atlas of atherosclerosis.
“Indeed, findings on unique macrophage subsets (such as TREM2+ macrophages) on the basis of single-cell analyses have uncovered unprecedented phenotypic heterogeneity of macrophages and monocyte-derived dendritic cells in atherosclerotic lesions and have defined a novel atlas of the immune cell repertoire in atherosclerosis135,154.”
From Inflammation and its resolution in atherosclerosis: mediators and therapeutic opportunities (2019) · cites this paper as [135]
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Self-citation
- Authors
- Winkels, H., Ehinger, E., Vassallo, M., Buscher, K., Dinh, H. Q., Kobiyama, K., Hamers, A. A. J., Cochain, C., Vafadarnejad, E., Saliba, A.-E., Zernecke, A., Pramod, A. B., Ghosh, A. K., Anto Michel, N., Hoppe, N., Hilgendorf, I., Zirlik, A., Hedrick, C. C., Ley, K., & Wolf, D.
- Affiliations
- La Jolla Institute for Immunology (US)
- Universitätsklinikum Würzburg (DE)
- Helmholtz Institute for RNA-based Infection Research (DE)
- Department of Cardiology and Angiology I, University Heart Center Freiburg, Germany (N.A.M., N.H., I.H., A.Z., D.W.)
- University of Freiburg (DE)
- Corresponding author
- Winkels, Holger
- Ehinger, Erik
- Ley, Klaus
- ORCID
- Venue
- Circulation Research
- Published
- 2018-06-08
- Volume
- 122
- Issue
- 12
- Pages
- 1675-1688
- ISSN
- 0009-7330 (ISSN-L)
- 1524-4571
- PMC
- PMC5993603
- OpenAlex
- W2791581835
- Cited by
- 547
- References
- 40
- Citations by year
- 2026: 42 · 2025: 66 · 2024: 77 · 2023: 72 · 2022: 79 · 2021: 74 · 2020: 64 · 2019: 51 · 2018: 20
- FWCI
- 22.45
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Locations
- Circulation Research (publishedVersion)
- PubMed (publishedVersion)
- Europe PMC (PubMed Central) (acceptedVersion)
- PubMed Central (acceptedVersion)
- HZI (Helmholtz Centre for Infection Research) (submittedVersion, cc-by-nc-sa)
- Funders
- Deutsche Forschungsgemeinschaft
- National Institutes of Health (1s10od018499-01)
- National Heart, Lung, and Blood Institute (R01HL134236, R01HL121697, R01HL115232, P01HL088093, P01HL136275)
- National Center for Research Resources (S10RR027366)
- Topics
- Life Sciences › Immunology and Microbiology › Immunology › Atherosclerosis and Cardiovascular Diseases
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Single-cell and spatial transcriptomics
- Health Sciences › Medicine › Surgery › Cholesterol and Lipid Metabolism
- Keywords
- atherosclerosis, leukocyte heterogeneity, B cell subsets, macrophages, T cells, monocytes, carotid plaque, lipid metabolism, cytokine production, ApoE-deficient mice, Ldlr-deficient mice, Western diet, cardiovascular events
- MeSH terms
- Animals, Aortic Diseases, Apolipoproteins E, B-Lymphocytes, Flow Cytometry, Humans, Leukocytes, Macrophages, Medical Illustration, Monocytes, Phenotype, Receptors, LDL, T-Lymphocytes, Sequence Analysis, RNA, Atherosclerosis, Mice, Single-Cell Analysis, Transcriptome
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