In-Depth Proteomic Characterization of Classical and Non-Classical Monocyte Subsets

Segura, Víctor; Valero, M. Luz; Cantero, Laura; Muñoz, Javier; Zarzuela, Eduardo; Garcı́a, Fernando; Aloria, Kerman; Beaskoetxea, Javier; Arizmendi, Jesús M.; Navajas, Rosana; Paradela, Alberto; Díez, Paula; Dégano, Rosa Ma; Fuentes, Manuel; Órfão, Alberto; Montero, Andrés; Garin-Muga, Alba; Corrales, Fernando J.; Pino, Manuel M. Sánchez del

doi:10.3390/proteomes6010008

Cited by 14 publications

(17 citation statements)

References 49 publications

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“…However, they secrete TNF-alpha, IL-1beta, and CCL3 in response to virus and immune complex containing nucleic acids via the MyD88–MEK pathway ( 26 ). In depth proteome analysis further supports the established functions of classical and non-classical monocyte subsets ( 35 ).…”

Section: Functional Role Of Monocyte Subsets Defined By Transcriptomimentioning

confidence: 52%

Monocyte Subsets: Phenotypes and Function in Tuberculosis Infection

et al. 2018

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Monocytes are critical defense components that play an important role in the primary innate immune response. The heterogeneous nature of monocytes and their ability to differentiate into either monocyte-derived macrophages or monocyte-derived dendritic cells allows them to serve as a bridge between the innate and adaptive immune responses. Current studies of monocytes based on immunofluorescence, single-cell RNA sequencing and whole mass spectrometry finger printing reveals different classification systems for monocyte subsets. In humans, three circulating monocyte subsets are classified based on relative expression levels of CD14 and CD16 surface proteins, namely classical, intermediate and non-classical subsets. Transcriptomic analyses of these subsets help to define their distinct functional properties. Tuberculosis (TB) is a disease instigated by the deadly pathogen Mycobacterium tuberculosis. Current research on monocytes in TB has indicated that there are alterations in the frequency of intermediate and non-classical subsets suggesting their impact in bacterial persistence. In this review, we will focus on these monocyte subsets, including their classification, frequency distribution, cytokine profiles, role as a biomarker and will comment on future directions for understanding the salient phenotypic and functional properties relevant to TB pathogenesis.

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Section: Functional Role Of Monocyte Subsets Defined By Transcriptomimentioning

confidence: 52%

Monocyte Subsets: Phenotypes and Function in Tuberculosis Infection

et al. 2018

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“…Although variability among donors in the expression of multiple genes in human monocytes has been previously described by cytofluorometry and transcriptome analysis 6,32,33 , clustering of individuals in distinct profiles of expression has not been achieved. The mechanisms leading to shared interindividual variations in monocyte phenotypes described here are unknown.…”

Section: Discussionmentioning

confidence: 99%

Identification of Novel Human Monocyte Subsets and Evidence for Phenotypic Groups Defined by Interindividual Variations of Expression of Adhesion Molecules

Merah-Mourah

Cohen

Charron

et al. 2020

Sci Rep

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Monocytes contribute to immune responses as a source for subsets of dendritic cells and macrophages. Human blood monocytes are classified as classical, non-classical and intermediate cells. However, the particular functions of these subsets have been hard to define, with conflicting results and significant overlaps. One likely reason for these ambiguities is in the heterogeneity of these monocyte subsets regrouping cells with divergent functions. To better define monocyte populations, we have analysed expression of 17 markers by multicolour flow cytometry in samples obtained from 28 control donors. Data acquisition was tailored to detect populations present at low frequencies. Our results reveal the existence of novel monocyte subsets detected as larger CD14 + cells that were CD16 + or CD16 neg. 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. 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. Distinct inflammatory responses to TLR agonists were found in small and large monocytes. Overall, refining the definition of monocyte subsets should lead to the identification of populations with specific functions. 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. This may be due in part to the high number and complexity of phenotypes present in monocyte populations. A 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. 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. Further analysis identified CD14 + CD16 + monocytes with expression of CCR5 and intermediate expression of receptors divergently expressed in the two other subsets (e.g. CCR2 and CX3CR1 4. This additional subset was identified as "intermediate" 5. Evidence for this third subset was confirmed in transcriptome analysis 6-10. Despite progress in phenotypic analysis, immune functions associated with monocyte subpopulations in the steady state remained ill defined. 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 11-13. Thus, intermediate monocytes were described as the major source of pro-inflammatory cytokines upon stimulation 14,15. In contrast, non-classical monocytes were also described as the most inflammatory monocytes 3,13,16. Similarly, anti-inflammatory cytokine secretion was alternatively found high in intermediate 13 or in classical monoc...

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“…Although two previous reports have quantified the whole cell proteome of monocyte subsets 34,35 , ours represents the first selective study of the cell surface proteome of these primary cell types. Furthermore, one of the previous studies 34 pooled non-classical and intermediate phenotype cells prior to analysis, and the other did not examine intermediate phenotype cells 35 . We compared our data to both studies.…”

Section: Discussionmentioning

confidence: 99%

“…There are a number of possible explanations for this finding, particularly including a small sample size of differentially expressed proteins that were also quantified in our study in each case. Additionally, correspondence between cell surface and whole-cell protein abundances can in any case be poor, and there may have been some confounding in one of the prior studies from isolation and measurement of protein samples in five different locations, using four different types of mass spectrometer 35 . Neither prior study examined intermediate-phenotype cells.…”

Section: Discussionmentioning

confidence: 99%

Comprehensive cell surface proteomics defines markers of classical, intermediate and non-classical monocytes

Ravenhill

Soday

Houghton

et al. 2020

Sci Rep

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Monocytes are a critical component of the cellular innate immune system, and can be subdivided into classical, intermediate and non-classical subsets on the basis of surface CD14 and CD16 expression. classical monocytes play the canonical role of phagocytosis, and account for the majority of circulating cells. Intermediate and non-classical cells are known to exhibit varying levels of phagocytosis and cytokine secretion, and are differentially expanded in certain pathological states. Characterisation of cell surface proteins expressed by each subset is informative not only to improve understanding of phenotype, but may also provide biological insights into function. Here we use highly multiplexed Tandem-Mass-Tag (TMT)-based mass spectrometry with selective cell surface biotinylation to characterise the classical monocyte surface proteome, then interrogate the phenotypic differences between each monocyte subset to identify novel protein markers. Monocytes play critical roles in the response to infection and inflammation and antigen presentation. Key effector functions are mediated by differentiated cells, including macrophages and myeloid dendritic cells. Monocytes can be divided into subsets based on the presence of the cell surface lipopolysaccharide (LPS) co-receptor CD14, and Fc-Gamma Receptor III (CD16). CD14++ CD16− classical monocytes account for ~85% of circulating monocytes, intermediate cells (CD14++ CD16+) for ~5% and non-classical cells (CD14+ CD16++) for ~10% of circulating monocytes 1,2. It has become increasingly well established that each subset plays divergent roles in different diseases, as well as differing in the ability to secrete cytokines and respond to pathogen associated molecular patterns (PAMPs). Classical monocytes are phagocytic and readily secrete inflammatory cytokines. Conversely, the CD16 positive non-classical cells are poorly phagocytic and are suggested to secrete TNFα in response to some stimuli, but less of other pro-inflammatory molecules 2-4. Intermediate monocytes are increased in diseases such as severe asthma, rheumatoid arthritis and sarcoidosis, and there is some evidence for expansion of classical monocytes in atherosclerosis 5-8. It is still unclear whether intermediate monocytes represent a truly distinct monocyte subset, or merely a transitional stage between classical and non-classical cells 9. Cells of the innate and adaptive immune systems can be categorised on the basis of microscopic appearance and expression of plasma membrane (PM) proteins, enabling separation by fluorescence activated cell sorting (FACS). As such, systematic evaluation of the entire cell surface proteome expressed by a given immune cell population is a powerful tool to characterise cellular function and distinguish cell types. Previous studies of monocytes subsets have examined transcriptional differences and offer varying depths of information about subset markers, suitability of individual proteins for discriminating subsets by cell surface flow cytometry and the importance of each protein...

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In-Depth Proteomic Characterization of Classical and Non-Classical Monocyte Subsets

Cited by 14 publications

References 49 publications

Monocyte Subsets: Phenotypes and Function in Tuberculosis Infection

Monocyte Subsets: Phenotypes and Function in Tuberculosis Infection

Identification of Novel Human Monocyte Subsets and Evidence for Phenotypic Groups Defined by Interindividual Variations of Expression of Adhesion Molecules

Comprehensive cell surface proteomics defines markers of classical, intermediate and non-classical monocytes

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