11/23/2023 0 Comments Define cytometryCD14 expression was considered a necessary condition for inclusion in monocyte populations (Fig. Settings, which were chosen to achieve a comprehensive study of monocyte populations, included an optimized procedure to reduce non-specific labelling, the acquisition of 1 × 10 6 events to analyse rare populations, and the exclusion of non-myeloid cell lineages (CD3 +, CD19 +, and NKp46 + (CD335) cells) as initial step (Fig. In order to further define human blood monocyte populations, we have analysed labelled PBMC from 28 healthy individuals by flow cytometry (Supplementary Table S1). Identification of populations of monocytes in human healthy donors Although monocyte populations had heterogeneous phenotypes among healthy donors they could nonetheless be resolved into phenotypic groups based on interindividual variations of expression. Results identified novel populations of monocytes with unique morphologic and phenotypic characteristics, and with distinct inflammatory responses to TLR agonists. To look for conserved phenotypes, the analysis was extended to 28 healthy Caucasian donors. In this study, we stringently assessed the phenotypic heterogeneity of human blood monocytes by multicolour cell surface labelling, flow cytometry analysis, and unsupervised detection of clusters and analysis of their phenotypes. Therefore, production of pro-inflammatory and anti-inflammatory cytokines upon activation, a hallmark property of monocytes, remains hard to ascribe unambiguously to given subsets. Similarly, anti-inflammatory cytokine secretion was alternatively found high in intermediate 13 or in classical monocytes 6, 11, 17. In contrast, non-classical monocytes were also described as the most inflammatory monocytes 3, 13, 16. Thus, intermediate monocytes were described as the major source of pro-inflammatory cytokines upon stimulation 14, 15. Marked functional redundancies between the sub-populations were found, and contradicting results in the literature added to the puzzling difficulties in assigning functions to specific populations 11, 12, 13. Evidence for this third subset was confirmed in transcriptome analysis 6, 7, 8, 9, 10.ĭespite progress in phenotypic analysis, immune functions associated with monocyte subpopulations in the steady state remained ill defined. This additional subset was identified as “intermediate” 5. Further analysis identified CD14 +CD16 + monocytes with expression of CCR5 and intermediate expression of receptors divergently expressed in the two other subsets (e.g. It consisted initially of two populations described as “classical” monocytes, which express CD14 but no CD16, and “nonclassical” monocytes with low CD14 and strong CD16 expression. This may be due in part to the high number and complexity of phenotypes present in monocyte populations.Ī classification of human blood monocyte subsets based on the expression of CD14 and CD16 cell surface receptors was proposed 1 and refined over the years 2, 3. Monocytes, which are mostly precursors of some macrophage and dendritic cell populations, have been hard to divide into populations with clear-cut inflammatory and immune functions. Overall, refining the definition of monocyte subsets should lead to the identification of populations with specific functions. Distinct inflammatory responses to TLR agonists were found in small and large monocytes. Unsupervised multidimensional analysis confirmed the existence of large monocytes and revealed interindividual variations that were grouped according to unique patterns of expression of adhesion molecules CD62L, CD49d, and CD43. These large monocytes differed from regular, smaller monocytes with respect to expression of various cell surface molecules, such as FcR, chemokine receptors, and adhesion molecules. Our results reveal the existence of novel monocyte subsets detected as larger CD14 + cells that were CD16 + or CD16 neg. Data acquisition was tailored to detect populations present at low frequencies. To better define monocyte populations, we have analysed expression of 17 markers by multicolour flow cytometry in samples obtained from 28 control donors. One likely reason for these ambiguities is in the heterogeneity of these monocyte subsets regrouping cells with divergent functions. However, the particular functions of these subsets have been hard to define, with conflicting results and significant overlaps. Human blood monocytes are classified as classical, non-classical and intermediate cells. Monocytes contribute to immune responses as a source for subsets of dendritic cells and macrophages.
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