From the Division of Inflammation Biology, La Jolla Institute for Allergy and Immunology, CA (H.W., E.E., M.V., K.B., H.Q.D., K.K., A.A.J.H., A.B.P., A.K.G., C.C.H., K.L., D.W.).
Institute of Experimental Biomedicine, University Hospital Würzburg, Germany (C.C., A.Z.).
Circ Res. 2018 Jun 8;122(12):1675-1688. doi: 10.1161/CIRCRESAHA.117.312513. Epub 2018 Mar 15.
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.
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.
Using single-cell RNA-sequencing of aortic leukocytes from chow diet- and Western diet-fed and 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 and 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.
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.
动脉粥样硬化是一种慢性炎症性疾病，其发生是由主动脉中促炎和抗炎白细胞的相互作用驱动的。然而，人们对主动脉白细胞的表型和转录多样性知之甚少。
我们通过单细胞 RNA 测序和液质流式细胞术（飞行时间质谱流式细胞术）对来自健康和动脉粥样硬化小鼠主动脉的白细胞进行了深入研究，以定义动脉粥样硬化免疫细胞图谱。
通过对喂食标准饮食和西方饮食的 和 小鼠主动脉白细胞进行单细胞 RNA 测序，我们检测到 11 个主要的白细胞簇，它们具有不同的表型和空间特征，而健康主动脉中的细胞组成则相对较少。在单细胞水平上进行基因集富集分析确定，多个通路，如脂质代谢、增殖和细胞因子分泌等，局限于特定的白细胞簇。在动脉粥样硬化的 和 小鼠中，白细胞群体存在差异调节。我们使用一种新型的基于金属标记抗体的 35 标志物组质谱流式细胞术面板和常规流式细胞术，对这些簇的表型多样性进行了验证。这些基于蛋白质的方法获取的细胞群体与转录定义的簇高度相关。在单细胞 RNA 测序、液质流式细胞术和荧光激活细胞分选的综合筛选策略中，我们检测到 3 个主要的 B 细胞亚群，其表面标志物、功能途径和体外细胞因子分泌均发生改变。白细胞簇基因特征通过遗传去卷积策略揭示了 126 个人斑块中的白细胞频率。这种方法表明，人类颈动脉斑块和微解剖的小鼠斑块主要由巨噬细胞、T 细胞和单核细胞组成。此外，颈动脉斑块中遗传定义的白细胞群体的频率可预测患者的心血管事件。
通过高维分析定义白细胞多样性可以对主动脉白细胞亚群进行精细分析，揭示新的免疫机制和细胞类型特异性途径，并确定人类动脉粥样硬化中病灶白细胞的功能相关性。
