Human Blood Monocyte Subsets

Abstract

Objective Human monocyte subsets are defined as classical (CD14++CD16−), intermediate (CD14++CD16+), and nonclassical (CD14+CD16+). Alterations in monocyte subset frequencies are associated with clinical outcomes, including cardiovascular disease, in which circulating intermediate monocytes independently predict cardiovascular events. However, delineating mechanisms of monocyte function is hampered by inconsistent results among studies. Approach and Results We utilize CyTOF mass cytometry to profile human monocytes using a panel of 36 cell surface markers. Using the dimensionality reduction approach viSNE, we define monocytes by incorporating all cell surface markers simultaneously. Using viSNE, we find that although classical monocytes are defined with high purity using CD14 and CD16, intermediate and nonclassical monocytes defined using CD14 and CD16 alone are frequently contaminated, with average intermediate and nonclassical monocyte purity of approximately 86.0% and 87.2% respectively. To improve the monocyte purity, we devised a new gating scheme that takes advantage of the shared coexpression of cell surface markers on each subset. In addition to CD14 and CD16, CCR2, CD36, HLA-DR and CD11c are the most informative markers that discriminate among the three monocyte populations. Using these additional markers as filters, our revised gating scheme increases the purity of both intermediate and nonclassical monocyte subsets to 98.8% and 99.1% respectively. We demonstrate the utility of this new gating scheme using conventional flow cytometry of PBMCs from subjects with cardiovascular disease. Conclusions Using CyTOF mass cytometry we have identified a small panel of surface markers that can significantly improve monocyte subset identification and purity in flow cytometry. Such a revised gating scheme will be useful for clinical studies of monocyte function in human cardiovascular disease.

Differential Interaction of Platelet-Derived Extracellular Vesicles with Leukocyte Subsets in Human Whole Blood

“…4a and c ), as well as by their expression patterns of the chemokine receptors CCR2, CX 3 CR1, and CCR5 (Fig. 4b and c ) 48 – 50 . Complementing their analysis in whole blood, we characterized monocyte subsets after isolation of monocytes from peripheral blood mononuclear cells (PBMCs) by negative depletion with antibody-coated magnetic beads.…”

“…In addition to CD14 and CD16 surface expression, monocyte subsets were characterized via the exposure of chemokine receptors CCR2, CX 3 CR1, CCR5 as summarized in Fig. 4c 48 , 49 (n = 3).…”

Intracapillary immune complexes recruit and activate slan-expressing CD16+ monocytes in human lupus nephritis

“…CD16 + monocytes are a heterogeneous cell population, as precisely revealed by recent transcriptomic and phenotypic studies (65)(66). We are just beginning to understand their role in immunity.…”

The MHC class II antigen presentation pathway in human monocytes differs by subset and is regulated by cytokines

“…Perturbations in the frequency of intermediate as well as non-classical monocytes have been observed in a variety of conditions, including autoimmune diseases, bacterial and viral infections and several inflammatory conditions such as asthma, coronary artery disease, periodontitis and stroke [ 6 , 43 ]. Further characterization of monocytes subsets, including improved ways to discriminate among the monocyte subsets [ 44 ], will help understand the role of these cells in health as well as in disease.…”