Abstract
Dendritic cells (DCs) were initially defined as mononuclear phagocytes with a dendritic morphology and an exquisite efficiency for naïve T-cell activation. DC encompass several subsets initially identified by their expression of specific cell surface molecules and later shown to excel in distinct functions and to develop under the instruction of different transcription factors or cytokines. Very few cell surface molecules are expressed in a specific manner on any immune cell type. Hence, to identify cell types, the sole use of a small number of cell surface markers in classical flow cytometry can be deceiving. Moreover, the markers currently used to define mononuclear phagocyte subsets vary depending on the tissue and animal species studied and even between laboratories. This has led to confusion in the definition of DC subset identity and in their attribution of specific functions. There is a strong need to identify a rigorous and consensus way to define mononuclear phagocyte subsets, with precise guidelines potentially applicable throughout tissues and species. We will discuss the advantages, drawbacks, and complementarities of different methodologies: cell surface phenotyping, ontogeny, functional characterization, and molecular profiling. We will advocate that gene expression profiling is a very rigorous, largely unbiased and accessible method to define the identity of mononuclear phagocyte subsets, which strengthens and refines surface phenotyping. It is uniquely powerful to yield new, experimentally testable, hypotheses on the ontogeny or functions of mononuclear phagocyte subsets, their molecular regulation, and their evolutionary conservation. We propose defining cell populations based on a combination of cell surface phenotyping, expression analysis of hallmark genes, and robust functional assays, in order to reach a consensus and integrate faster the huge but scattered knowledge accumulated by different laboratories on different cell types, organs, and species.
Highlights
The immune system includes a large variety of myeloid and lymphoid cell types which develop through distinct ontogenic pathways, express specific phenotypes, and exert specialized functions.Universal identification of Dendritic cells (DCs) subsetsThe mononuclear phagocytes form a complex group of myeloid cells that encompass three major cell types, i.e., monocytes, macrophages, and dendritic cells (DC), together with their proximal progenitors
We found the Xcr1 gene among genes expressed in mouse spleen CD8α+ DC when compared to a number of other immune cell types [see Supplementary Material “Additional file 5; gb-2008-9-1-r17-s5.xls” from Robbins et al [39], in the “CD8a_DC_gene_signature” established from our microarray data and confirmed from our own re-analysis of the microarray dataset independently generated by Dudziak et al [87]]
We showed that, upon many types of in vivo or in vitro stimulation, human and murine plasmacytoid DC (pDC) and conventional DC (cDC) undergo a remodeling of their gene expression program related to their plasticity, including induction of NFκB and IFN target genes, but still keep the canonical gene expression associated to their subset identity [41, 97]
Summary
The immune system includes a large variety of myeloid and lymphoid cell types which develop through distinct ontogenic pathways, express specific phenotypes, and exert specialized functions. It has been proposed that the study of their developmental pathway, in other words ontogeny, was the best way to classify mononuclear phagocyte cell types, at least in the mouse model where the knowledge in DC subset properties is the most advanced In this model, genetically modified animals unambiguously permit to track the development of cell types and to dissect their phenotypes and functions, in different contexts in vivo. Ontogeny Ontogeny studies in mice, in particular studies on the dependence of DC subset development on transcription factors, have been instrumental in identifying the homologies between lymphoid tissue-resident CD8α+ cDC and the CD103+CD11b− cDC present in non-lymphoid tissues and migrating into the draining lymph nodes once activated [58] These studies, together with gene expression profiling analyses [9, 40], allowed grouping mouse CD8α+ cDC and CD103+CD11b− cDC together under the umbrella of the XCR1+ cDC subset [38, 40, 59, 60]. The establishment of the concept that mouse bona fide DC constitute a separate hematopoietic lineage, and the discrimination between mouse CD11b+ cDC and MoDC, were confirmed using mutant animals allowing to track natural precursor–progeny relationships in vivo through irreversible fluorescent tagging of all daughter cells of a given type of hematopoietic progenitor, based on Cre-mediated conditional activation
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