Defining mast cell differentiation and heterogeneity through single-cell transcriptomics analysis

Mast cells (MCs) are widely recognized as central effector cells during type 2 inflammatory reactions and thought to also play a role in innate immune responses, wound healing, and potentially cancer. Circulating progenitor cells mature to MCs in peripheral tissues, where they exhibit phenotypic and functional heterogeneity. This diversity likely originates from differences in MC development imprinted by microenvironmental signals. The advent of single-cell transcriptomics reveals MC diversity beyond differences in proteases that were classically used to identify MC phenotypes. Here, we provide an overview of the current knowledge on MC progenitor differentiation and characteristics, and MC heterogeneity seen in health versus disease, that are drastically advanced through single-cell profiling technologies. This powerful approach can provide detailed cellular maps of tissues to decipher the complex cellular functions and interactions that may lead to identifying candidate factors to target in therapies. Mast cells (MCs) are widely recognized as central effector cells during type 2 inflammatory reactions and thought to also play a role in innate immune responses, wound healing, and potentially cancer. Circulating progenitor cells mature to MCs in peripheral tissues, where they exhibit phenotypic and functional heterogeneity. This diversity likely originates from differences in MC development imprinted by microenvironmental signals. The advent of single-cell transcriptomics reveals MC diversity beyond differences in proteases that were classically used to identify MC phenotypes. Here, we provide an overview of the current knowledge on MC progenitor differentiation and characteristics, and MC heterogeneity seen in health versus disease, that are drastically advanced through single-cell profiling technologies. This powerful approach can provide detailed cellular maps of tissues to decipher the complex cellular functions and interactions that may lead to identifying candidate factors to target in therapies. Mast cells (MCs) are histochemically distinctive tissue-resident effector cells derived from the hematopoietic system. Although best known as principal effector cells involved in IgE-driven type 1 hypersensitivity that underlies many allergic disorders, MCs are an ancient component of the immune system, with MC-like cells detected in tunicates, predating adaptive immunity by hundreds of millions of years.1Cavalcante M.C. de Andrade L.R. Du Bocage Santos-Pinto C. Straus A.H. Takahashi H.K. Allodi S. et al.Colocalization of heparin and histamine in the intracellular granules of test cells from the invertebrate Styela plicata (Chordata-Tunicata).J Struct Biol. 2002; 137: 313-321Crossref PubMed Scopus (50) Google Scholar,2Voehringer D. Protective and pathological roles of mast cells and basophils.Nat Rev Immunol. 2013; 13: 362-375Crossref PubMed Scopus (294) Google Scholar MCs almost certainly serve diverse additional functions in inflammation, innate and adaptive host defense, wound healing, regulation of vasomotor tone, and cancer. Such diverse functions in turn require the capacity for context-specific modifications of MC effector properties. Although MC functional diversification has long been suspected on the basis of histochemical markers, the exclusive tissue residence of mature MCs presents significant challenges to understanding the true range of MC diversity and how this diversity is regulated on the basis of functional studies. The recent application of single-cell genomics to the MC field has begun to overcome some of these limitations and reveals a much broader range of diversification than predicted by histochemistry alone. Moreover, these studies have permitted the identification of receptors and transcriptional systems that are promising targets for the development of therapeutics for diseases associated with or driven by MC-dependent effector systems. Here, we provide an overview of insights gained on MC progenitor (MCp) characteristics and MC heterogeneity across tissue and diseases using single-cell transcriptomic analysis and discuss the findings and remaining questions to unravel the underlying drivers of their infiltration and diversity in health and disease. MCs arise from circulating hematopoietic MCps that mature following recruitment to peripheral tissues.3Gurish M.F. Austen K.F. Developmental origin and functional specialization of mast cell subsets.Immunity. 2012; 37: 25-33Abstract Full Text Full Text PDF PubMed Scopus (237) Google Scholar Recent murine studies have indicated that MC populations found in peripheral connective tissues arise from a series of developmentally discrete waves of progenitor cells during and following embryogenesis. In the first wave, early erythro-myeloid progenitor (EMP) cells from yolk sac give rise to MCs expressing the stem cell factor receptor CD117 and the IL-33 receptor T1/ST2 and containing avidin-binding granules. These early MCs are then replaced by a second wave of late EMP-derived MCps that give rise to transcriptionally distinct connective tissue MCs (CTMCs) that are maintained independently of bone marrow (BM) through adulthood, indicating that progenitor origin and developmental stage of the host likely play roles in shaping MC identity.4Li Z. Liu S. Xu J. Zhang X. Han D. Liu J. et al.Adult connective tissue-resident mast cells originate from late erythro-myeloid progenitors.Immunity. 2018; 49: 640-653.e5Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar,5Gentek R. Ghigo C. Hoeffel G. Bulle M.J. Msallam R. Gautier G. et al.Hemogenic endothelial fate mapping reveals dual developmental origin of mast cells.Immunity. 2018; 48: 1160-1171.e5Abstract Full Text Full Text PDF PubMed Scopus (187) Google Scholar In contrast, MCs in the intestinal mucosa arise from MCps derived from hematopoietic stem cells, first from aorta-gonad-mesonephros hemogenic endothelium and later from the fetal liver and BM.4Li Z. Liu S. Xu J. Zhang X. Han D. Liu J. et al.Adult connective tissue-resident mast cells originate from late erythro-myeloid progenitors.Immunity. 2018; 49: 640-653.e5Abstract Full Text Full Text PDF PubMed Scopus (114) Google Scholar,6Derakhshan T. Samuchiwal S.K. Hallen N. Bankova L.G. Boyce J.A. Barrett N.A. et al.Lineage-specific regulation of inducible and constitutive mast cells in allergic airway inflammation.J Exp Med. 2020; 218e20200321Google Scholar,7Kitamura Y. Shimada M. Hatanaka K. Miyano Y. Development of mast cells from grafted bone marrow cells in irradiated mice.Nature. 1977; 268: 442-443Crossref PubMed Scopus (246) Google Scholar MCs across peripheral connective tissues share a core transcriptional signature, consistent with a shared developmental origin, yet also show tissue-specific differences in transcriptional profile and cell surface protein expression, indicating a role for their tissue microenvironment in shaping MC identity.8Dwyer D.F. Barrett N.A. Austen K.F. Expression profiling of constitutive mast cells reveals a unique identity within the immune system.Nat Immunol. 2016; 17: 878-887Crossref PubMed Scopus (251) Google Scholar A recent study from our laboratory examined the transcriptional distinction between constitutive and inflammation-induced MCs within murine lung tissue.6Derakhshan T. Samuchiwal S.K. Hallen N. Bankova L.G. Boyce J.A. Barrett N.A. et al.Lineage-specific regulation of inducible and constitutive mast cells in allergic airway inflammation.J Exp Med. 2020; 218e20200321Google Scholar Through this approach, we found that the resident lung CTMCs were significantly enriched for a transcriptional signature shared by CTMCs across 5 tissues in naive mice relative to recruited BM-derived MCs, even following allergic lung inflammation.6Derakhshan T. Samuchiwal S.K. Hallen N. Bankova L.G. Boyce J.A. Barrett N.A. et al.Lineage-specific regulation of inducible and constitutive mast cells in allergic airway inflammation.J Exp Med. 2020; 218e20200321Google Scholar,8Dwyer D.F. Barrett N.A. Austen K.F. Expression profiling of constitutive mast cells reveals a unique identity within the immune system.Nat Immunol. 2016; 17: 878-887Crossref PubMed Scopus (251) Google Scholar Notably, the BM-derived MCs infiltrating the pulmonary tissue during inflammation exhibit enrichment for TGF-β signature genes along with inflammation-associated superimposed signals.6Derakhshan T. Samuchiwal S.K. Hallen N. Bankova L.G. Boyce J.A. Barrett N.A. et al.Lineage-specific regulation of inducible and constitutive mast cells in allergic airway inflammation.J Exp Med. 2020; 218e20200321Google Scholar These studies suggest that MCp origin, microenvironmentally derived tissue signals, and tissue inflammatory state all influence the ultimate phenotype of MC subsets. Although peripheral CTMCs have minimal transcriptional overlap with basophils beyond components of the high-affinity IgE receptor FcɛR1 and histamine biosynthetic components, numerous studies have suggested the existence of a bipotent basophil/MC progenitor within the mouse BM and spleen.9Arinobu Y. Iwasaki H. Gurish M.F. Mizuno S.-I. Shigematsu H. Ozawa H. et al.Developmental checkpoints of the basophil/mast cell lineages in adult murine hematopoiesis.Proc Natl Acad Sci. 2005; 102: 18105-18110Crossref PubMed Scopus (281) Google Scholar, 10Qi X. Hong J. Chaves L. Zhuang Y. Chen Y. Wang D. et al.Antagonistic regulation by the transcription factors C/EBPα and MITF specifies basophil and mast cell fates.Immunity. 2013; 39: 97-110Abstract Full Text Full Text PDF PubMed Scopus (103) Google Scholar, 11Iwasaki H. Mizuno S. Arinobu Y. Ozawa H. Mori Y. Shigematsu H. et al.The order of expression of transcription factors directs hierarchical specification of hematopoietic lineages.Genes Dev. 2006; 20: 3010-3021Crossref PubMed Scopus (242) Google Scholar Supporting this concept, we determined that Mcpt8, previously identified as a basophil-specific transcript, is expressed by BM-derived MCs in vivo, a finding also reported by Hamey et al.6Derakhshan T. Samuchiwal S.K. Hallen N. Bankova L.G. Boyce J.A. Barrett N.A. et al.Lineage-specific regulation of inducible and constitutive mast cells in allergic airway inflammation.J Exp Med. 2020; 218e20200321Google Scholar,12Hamey F.K. Lau W.W.Y. Kucinski I. Wang X. Diamanti E. Wilson N.K. et al.Single-cell molecular profiling provides a high-resolution map of basophil and mast cell development.Allergy. 2021; 76: 1731-1742Crossref PubMed Scopus (28) Google Scholar Although these findings are consistent with the derivation of MCs and basophils from a common progenitor, there has been substantial debate within the field as to where exactly the MC and basophil progenitor fits within broader hematopoiesis. Historically, hematopoietic progenitors have been identified on the basis of surface marker expression and characterized on the basis of their progeny in clonal expansion assays and following transplantation.13Akashi K. Traver D. Miyamoto T. Weissman I.L. A clonogenic common myeloid progenitor that gives rise to all myeloid lineages.Nature. 2000; 404: 193-197Crossref PubMed Scopus (1961) Google Scholar,14Osawa M. Hanada K.-I. Hamada H. Nakauchi H. Long-term lymphohematopoietic reconstitution by a single CD34-low/negative hematopoietic stem cell.Science. 1996; 273: 242-245Crossref PubMed Scopus (1752) Google Scholar Classical model of hematopoiesis proposes that multipotent progenitors (MPPs) give rise to common myeloid progenitors (CMPs) with an immunophenotype distinct from that of common lymphoid progenitors. In this model, CMPs give rise to both granulocyte-monocyte progenitors (GMPs) and megakaryocyte-erythroid progenitors (MEPs).13Akashi K. Traver D. Miyamoto T. Weissman I.L. A clonogenic common myeloid progenitor that gives rise to all myeloid lineages.Nature. 2000; 404: 193-197Crossref PubMed Scopus (1961) Google Scholar,15Reya T. Morrison S.J. Clarke M.F. Weissman I.L. Stem cells, cancer, and cancer stem cells.Nature. 2001; 414: 105-111Crossref PubMed Scopus (8084) Google Scholar Adolfsson et al16Adolfsson J. Månsson R. Buza-Vidas N. Hultquist A. Liuba K. Jensen C.T. et al.Identification of Flt3+ lympho-myeloid stem cells lacking erythro-megakaryocytic potential: a revised road map for adult blood lineage commitment.Cell. 2005; 121: 295-306Abstract Full Text Full Text PDF PubMed Scopus (947) Google Scholar identified lymphoid-primed multipotent progenitors (LMPPs) with the capacity to give rise to both lymphocytes and granulocyte/macrophages but not the megakaryocyte-erythroid lineage, establishing that MPPs branch into CMPs and LMPPs, with both possessing the potential to develop to GMPs. Studies based on clonal assays and transplantation suggested that GMPs, MEPs, and MPPs all may have the potential to differentiate into MCs.9Arinobu Y. Iwasaki H. Gurish M.F. Mizuno S.-I. Shigematsu H. Ozawa H. et al.Developmental checkpoints of the basophil/mast cell lineages in adult murine hematopoiesis.Proc Natl Acad Sci. 2005; 102: 18105-18110Crossref PubMed Scopus (281) Google Scholar,17Suda T. Suda J. Ogawa M. 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Distinguishing mast cell and granulocyte differentiation at the single-cell level.Cell Stem Cell. 2010; 6: 361-368Abstract Full Text Full Text PDF PubMed Scopus (86) Google Scholar were among the first to combine single-cell cloning– and transcriptomics-based approaches to understand mouse granulocyte differentiation, finding that MCps are distinct from the GMP population that gives rise to monocytes and neutrophils, instead sharing expression of a set of transcription factors (Gata2, Gata1) and surface receptors (T1/ST2 and erythropoietin receptor) with the MEP pool. Drissen et al21Drissen R. Buza-Vidas N. Woll P. Thongjuea S. Gambardella A. Giustacchini A. et al.Distinct myeloid progenitor–differentiation pathways identified through single-cell RNA sequencing.Nat Immunol. 2016; 17: 666-676Crossref PubMed Scopus (162) Google Scholar similarly identified Gata1 expression as an early point of lineage divergence through single-cell RNA-sequencing (scRNA-seq), generating a reporter mouse and discovering that Gata1+ cells in the immunophenotypically defined pre-GMP and GMP pools had megakaryocyte, erythrocyte, MC, and eosinophil differentiation capacity but minimal monocyte and neutrophil potential. Although LMPP, Gata1− GMPs, and pre-GMPs were able to give rise to monocytes and neutrophils, they were unable to differentiate into MCs. These findings were consistent with previous lineage-tracing approaches supporting early bifurcation of myeloid and erythroid lineages within a heterogeneous MPP pool.22Perié L. Duffy K.R. Kok L. de Boer R.J. Schumacher T.N. The branching point in erythro-myeloid differentiation.Cell. 2015; 163: 1655-1662Abstract Full Text Full Text PDF PubMed Scopus (121) Google Scholar Subsequent studies using scRNA-seq supported this bifurcation and further found that the MPP trajectory toward defined lineages progresses in a continuum rather than through distinct subpopulations.23Paul F. Arkin Y. Giladi A. Jaitin D.A. Kenigsberg E. Keren-Shaul H. et al.Transcriptional heterogeneity and lineage commitment in myeloid progenitors.Cell. 2015; 163: 1663-1677Abstract Full Text Full Text PDF PubMed Scopus (645) Google Scholar,24Nestorowa S. Hamey F.K. Pijuan Sala B. Diamanti E. Shepherd M. Laurenti E. et al.A single-cell resolution map of mouse hematopoietic stem and progenitor cell differentiation.Blood. 2016; 128: e20-e31Crossref PubMed Scopus (412) Google Scholar Tusi et al25Tusi B.K. Wolock S.L. Weinreb C. Hwang Y. Hidalgo D. Zilionis R. et al.Population snapshots predict early haematopoietic and erythroid hierarchies.Nature. 2018; 555: 54-60Crossref PubMed Scopus (215) Google Scholar similarly characterized a linked developmental trajectory for MCs and basophils that was closely associated with the megakaryocyte and erythroid lineages and separate from the neutrophil and monocyte. This linked trajectory was confirmed by Dahlin et al,26Dahlin J.S. Hamey F.K. Pijuan-Sala B. Shepherd M. Lau W.W.Y. Nestorowa S. et al.A single-cell hematopoietic landscape resolves 8 lineage trajectories and defects in Kit mutant mice.Blood. 2018; 131: e1-e11Crossref PubMed Scopus (116) Google Scholar who used a massively parallel scRNA-seq–based approach to analyze the BM of wild-type and W41/W41 Kit mutant mice. Although Kit mutant mice had intact MEP and basophil differentiation trajectories, the MC progenitor branch from the MC/basophil bipotent progenitor was notably absent, indicating a requirement for Kit signaling for MCp development in vivo. Although eosinophils were identified through sequencing in this study, their developmental relationship to MCs and basophils was unclear. The same data set was later used by Wanet et al27Wanet A. Bassal M.A. Patel S.B. Marchi F. Mariani S.A. Ahmed N. et al.E-cadherin is regulated by GATA-2 and marks the early commitment of mouse hematopoietic progenitors to the basophil and mast cell fates.Sci Immunol. 2021; 6eaba0178Crossref PubMed Google Scholar to identify E-cadherin as a surface marker capable of identifying both basophil and MC progenitors within the BM, not only confirming a link between these cells, eosinophils, and the MEP lineage but also identifying rare clones capable of giving rise to monocytes or neutrophils in addition to basophils and/or eosinophils. In humans, progenitor cells from both the aorta-gonad-mesonephros and the yolk sac contribute to the fetal liver progenitor pool that seeds peripheral tissues before the emergence of BM-derived progenitors.28Holt P.G. Jones C.A. The development of the immune system during pregnancy and early life.Allergy. 2000; 55: 688-697Crossref PubMed Scopus (397) Google Scholar MCs develop in yolk sac before the emergence of hematopoietic stem cells in nonlymphoid tissues, suggesting the potential for multiple waves of MCs in humans as in mouse, although this has not been experimentally confirmed.29Popescu D.-M. Botting R.A. Stephenson E. Green K. Webb S. Jardine L. et al.Decoding human fetal liver haematopoiesis.Nature. 2019; 574: 365-371Crossref PubMed Scopus (281) Google Scholar Several studies suggest that MC differentiation in BM is also similar between mouse and human. As in mouse, the classical model of hematopoiesis in human was challenged due to the potential of both LMPPs and CMPs to give rise to myeloid cells and granulocytes.30Doulatov S. Notta F. Eppert K. Nguyen L.T. Ohashi P.S. Dick J.E. Revised map of the human progenitor hierarchy shows the origin of macrophages and dendritic cells in early lymphoid development.Nat Immunol. 2010; 11: 585-593Crossref PubMed Scopus (383) Google Scholar,31Goardon N. Marchi E. Atzberger A. Quek L. Schuh A. Soneji S. et al.Coexistence of LMPP-like and GMP-like leukemia stem cells in acute myeloid leukemia.Cancer Cell. 2011; 19: 138-152Abstract Full Text Full Text PDF PubMed Scopus (486) Google Scholar Using a sort-based approach, Görgens et al32Görgens A. Radtke S. Möllmann M. Cross M. Dürig J. Horn P.A. et al.Revision of the human hematopoietic tree: granulocyte subtypes derive from distinct hematopoietic lineages.Cell Rep. 2013; 3: 1539-1552Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar found that the eosinophil and basophil lineages originated from a common EMP that segregated from GMPs based on surface expression of CD133, whereas neutrophils arose from GMPs. Macrophages also developed from EMP-cultured cells, albeit within colonies forming a mixed population of erythroid and myeloid cells (CFU-MIX) that were assumed to be transient CD133low CMPs that lacked the potential to develop into neutrophils.32Görgens A. Radtke S. Möllmann M. Cross M. Dürig J. Horn P.A. et al.Revision of the human hematopoietic tree: granulocyte subtypes derive from distinct hematopoietic lineages.Cell Rep. 2013; 3: 1539-1552Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar Their findings challenged the existence of a true CMP and proposed a model in which MPPs branched to LMPPs and EMPs, the latter lacking GMP potential. The advent of transcriptomics contributed to resolving the relationship between the progenitors and dynamic of progression from primitive progenitor cells to more defined lineages. Velten et al33Velten L. Haas S.F. Raffel S. Blaszkiewicz S. Islam S. Hennig B.P. et al.Human haematopoietic stem cell lineage commitment is a continuous process.Nat Cell Biol. 2017; 19: 271-281Crossref PubMed Scopus (524) Google Scholar integrated cell indexing and scRNA-seq approaches to illustrate that early stem and progenitor cells gradually progress in a continuum into more defined lineages expressing CD38. This finding indicated that traditional use of a limited number of surface markers creates subpopulations that, although immunophenotypically distinct, exhibit transcriptional overlap and potentially share differentiation trajectories. The progression to the basophil/eosinophil/MC lineages coincided with simultaneous induction and suppression of shared modules of transcripts where these progenitors displayed an immunophenotype that was similar to the MEP trajectory but distinct from classic GMP commitment. A similar relationship was observed in umbilical cord blood by Zheng et al,34Zheng S. Papalexi E. Butler A. Stephenson W. Satija R. Molecular transitions in early progenitors during human cord blood hematopoiesis.Mol Syst Biol. 2018; 14e8041Crossref PubMed Scopus (95) Google Scholar who identified a close link between the basophil/eosinophil/MC lineages and MEP, and determined that human MCps strongly express the transferrin receptor CD71 (TFRC), a classical marker of erythroid precursors. They further identified expression of MC-associated transcripts in previous bulk transcriptomics studies of hematopoiesis that used CD71 as a marker for the MEP lineage. Drissen et al35Drissen R. Thongjuea S. Theilgaard-Mönch K. Nerlov C. Identification of two distinct pathways of human myelopoiesis.Sci Immunol. 2019; 4: eaau7148Crossref PubMed Scopus (51) Google Scholar similarly used transcriptomic analysis to show that differential expression of CD131 and CD114 separates the Bas/Eos/MC lineages from neutrophil/monocyte progenitors, respectively. Thus, scRNA-seq–based approaches have proven highly useful in identifying markers and modulators of each lineage. These important studies clarified the lineage relationship between MCs, basophils, and eosinophils in humans, suggesting that all 3 major granulocytes involved in type 2 (T2) inflammation may have evolved from a common precursor. Within peripheral blood, Dahlin et al36Dahlin J.S. Malinovschi A. Öhrvik H. Sandelin M. Janson C. Alving K. et al.Lin− CD34hi CD117int/hi FcεRI+ cells in human blood constitute a rare population of mast cell progenitors.Blood. 2016; 127: 383-391Crossref PubMed Scopus (87) Google Scholar used flow cytometry to identify the existence of a Lin− CD34+ CD117+ FcɛR1+-committed MCp. Ex vivo culture assays suggested that these MCps lack the multilineage capacity of MC/basophil/eosinophil progenitors within the BM or cord blood. More recently, Wu et al37Wu C. Boey D. Bril O. Grootens J. Vijayabaskar M.S. Sorini C. et al.Single-cell transcriptomics reveals the identity and regulators of human mast cell progenitors.Blood Adv. 2022; 6: 4439-4449Crossref PubMed Scopus (4) Google Scholar used scRNA-seq to define MCps within the context of other circulating CD34+ CD117+ cells, finding that although some MC-associated transcripts (TPSAB1, HDC) were tightly linked to the MCp, others associated with FcɛR1α were more broadly expressed. They further found that IL-3 drove the proliferation of the MCp population, whereas IL-5 supported its survival. However, identification of human MCps within peripheral tissue has proven more elusive, in part because CD34 is inconsistently detected on the surface of MCs in inflamed tissue.38Dwyer D.F. Ordovas-Montanes J. Allon S.J. Buchheit K.M. Vukovic M. Derakhshan T. et al.Human airway mast cells proliferate and acquire distinct inflammation-driven phenotypes during type 2 inflammation.Sci Immunol. 2021; 6: eabb7221Crossref PubMed Google Scholar,39Andersson C.K. Shikhagaie M. Mori M. Al-Garawi A. Reed J.L. Humbles A.A. et al.Distal respiratory tract viral infections in young children trigger a marked increase in alveolar mast cells.ERJ Open Res. 2018; 4: 00038-2018Crossref PubMed Scopus (6) Google Scholar The differentiation potential of the progenitor cells into various lineages as evaluated in vitro using a cocktail of cytokines and a layer of feeder cells may not recapitulate their actual fate potential in vivo due to a lack of critical microenvironmental signals or the simultaneous administration of cytokines that may not be present within the same niche in vivo. Transcriptomic studies suggest that hematopoietic stem cells progress through intermediate progenitor stages, gradually losing their potential to develop into some lineages while increasing their propensity to become others following fate decision points along the pathway (Fig 1). It is therefore likely that some intermediate populations retain the capacity to develop into more than 1 cell type depending on the growth factors in the microenvironment and the degree to which they have progressed down a differentiation pathway to a particular lineage. An example is the derivation of macrophages from eosinophil, basophil common progenitor before commitment to the latter cell types.32Görgens A. Radtke S. Möllmann M. Cross M. Dürig J. Horn P.A. et al.Revision of the human hematopoietic tree: granulocyte subtypes derive from distinct hematopoietic lineages.Cell Rep. 2013; 3: 1539-1552Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar Moreover, although scRNA-seq approaches have substantially increased our understanding of how MCs fit within the broader hematopoietic landscape, several important questions remain. In particular, whether and how MCp characteristics may change in disease and the potential contribution of other less committed progenitors to driving MC expansion in health and disease are yet to be explored. As noted previously, both the scarcity of MCs and their exclusive tissue residence present challenges to their study in humans. Traditionally, MCs in human tissues have therefore been studied using histologic approaches. Collectively, these studies have contributed substantially to our understanding of the correlation of MC burden in tissues to disease outcomes, such as in allergic rhinitis and severe asthma, and identified disease-associated differences in MC protease expression, such as the induced expression of carboxypeptidase A3 by intraepithelial MCs arising in several T2 inflammatory diseases.40Bentley A.M. Jacobson M.R. Cumberworth V. Barkans J.R. Moqbel R. Schwartz L.B. et al.Immunohistology of the nasal mucosa in seasonal allergic rhinitis: increases in activated eosinophils and epithelial mast cells.J Allergy Clin Immunol. 1992; 89: 877-883Abstract Full Text PDF PubMed Scopus (250) Google Scholar, 41Abonia J.P. Blanchard C. Butz B.B. Rainey H.F. Collins M.H. Stringer K. et al.Involvement of mast cells in eosinophilic esophagitis.J Allergy Clin Immunol. 2010; 126: 140-149Abstract Full Text Full Text PDF PubMed Scopus (234) Google Scholar, 42Takabayashi T. Kato A. Peters A.T. Suh L.A. Carter R. Norton J. et al.Glandular mast cells with distinct phenotype are highly elevated in chronic rhinosinusitis with nasal polyps.J Allergy Clin Immunol. 2012; 130: 410-420.e5Abstract Full Text Full Text PDF PubMed Scopus (117) Google Scholar, 43Balzar S. Fajt M.L. Comhair S.A. Erzurum S.C. Bleecker E. Busse W.W. et al.Mast cell phenotype, location, and activation in severe asthma. Data from the Severe Asthma Research Program.Am J Respir Crit Care Med. 2011; 183: 299-309Crossref PubMed Scopus (242) Google Scholar Although functional studies performed on MCs derived by in vitro culture techniques have identified the capacity of MCs to generate a broad range of proinflammatory mediators, the relationship between what MCs have the capacity to make in vitro and what they actually make in vivo remains unclear.44Mukai K. Tsai M. Saito H. Galli S.J. Mast cells as sources of cytokines, chemokines, and growth factors.Immunol Rev. 2018; 282: 121-150Crossref PubMed Scopus (439) Google Scholar Whole-tissue transcriptomic studies have long hinted at a prominent role for MCs in T2 disease, with MC-specific transcripts appearing among the most differentially regulated transcripts between health and disease in eosinophilic esophagitis (EoE) and asthma.41Abonia J.P. Blanchard C. Butz B.B. Rainey H.F. Collins M.H. Stringer K. et al.Involvement of mast cells in eosinophilic esophagitis.J Allergy Clin Immunol. 2010; 126: 140-149Abstract Full Text Full Text PDF PubMed Scopus (234) Google Scholar,45Woodruff P.G. Boushey H.A. Dolganov G.M. Barker C.S. Yang Y.H. Donnelly S. et al.Genome-wide profiling identifies epithelial cell genes associated with asthma and with treatment response to corticosteroids.Proc Natl Acad Sci U S A. 2007; 104: 15858-15863Crossref PubMed Scopus (679) Google Scholar However, we still have little understanding of how and whether these mediators differ among MC subsets, an