RESUMEN
Hematopoietic stem and progenitor cells can differentiate into all types of blood cells. Regulatory mechanisms underlying pluripotency in progenitors, such as the ability of lymphoid progenitor cells to differentiate into T-lineage, remain unclear. We have previously reported that LIM domain only 2 (Lmo2), a bridging factor in large transcriptional complexes, is essential to retain the ability of lymphoid progenitors to differentiate into T-lineage. However, biochemical characterization of Lmo2 protein complexes in physiological hematopoietic progenitors remains obscure. Here, we identified approximately 600 Lmo2-interacting molecules in a lymphoid progenitor cell line by two-step affinity purification with LC-MS/MS analysis. Zinc finger and BTB domain containing 1 (Zbtb1) and CBFA2/RUNX1 partner transcriptional corepressor 3 (Cbfa2t3) were found to be the functionally important binding partners of Lmo2. We determined CRISPR/Cas9-mediated acute disruption of Zbtb1 or Cbfa2t3 in the lymphoid progenitor or bone marrow-derived primary hematopoietic progenitor cells causes significant defects in the initiation of T-cell development when Notch signaling is activated. Our transcriptome analysis of Zbtb1- or Cbfa2t3-deficient lymphoid progenitors revealed that Tcf7 was a common target for both factors. Additionally, ChIP-seq analysis showed that Lmo2, Zbtb1, and Cbfa2t3 cobind to the Tcf7 upstream enhancer region, which is occupied by the Notch intracellular domain/RBPJ transcriptional complex after Notch stimulation, in lymphoid progenitors. Moreover, transduction with Tcf7 restored the defect in the T-lineage potential of Zbtb1-deficient lymphoid progenitors. Thus, in lymphoid progenitors, the Lmo2/Zbtb1/Cbfa2t3 complex directly binds to the Tcf7 locus and maintains responsiveness to the Notch-mediated inductive signaling to facilitate T-lineage differentiation.
Asunto(s)
Células Progenitoras Linfoides , Factores de Transcripción , Células Progenitoras Linfoides/fisiología , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Cromatografía Liquida , Espectrometría de Masas en Tándem , Células Madre Hematopoyéticas/metabolismo , Diferenciación Celular , Proteínas con Dominio LIM/genética , Proteínas con Dominio LIM/metabolismoRESUMEN
Innate lymphoid cells (ILCs) include cytotoxic natural killer cells and distinct groups of cytokine-producing innate helper cells which participate in immune defense and promote tissue homeostasis. Circulating human ILC precursors (ILCP) able to generate all canonical ILC subsets via multi-potent or uni-potent intermediates according to our previous work. Here we show potential cooperative roles for the Notch and IL-23 signaling pathways for human ILC differentiation from blood ILCP using single cell cloning analyses and validate these findings in patient samples with rare genetic deficiencies in IL12RB1 and RORC. Mechanistically, Notch signaling promotes upregulation of the transcription factor RORC, enabling acquisition of Group 1 (IFN-γ) and Group 3 (IL-17A, IL-22) effector functions in multi-potent and uni-potent ILCP. Interfering with RORC or signaling through its target IL-23R compromises ILC3 effector functions but also generally suppresses ILC production from multi-potent ILCP. Our results identify a Notch->RORC- > IL-23R pathway which operates during human ILC differentiation. These observations may help guide protocols to expand functional ILC subsets in vitro with an aim towards novel ILC therapies for human disease.
Asunto(s)
Inmunidad Innata , Linfocitos , Diferenciación Celular , Humanos , Interleucina-23 , Células Asesinas Naturales , Células Progenitoras Linfoides/fisiología , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/genética , Receptores Notch/metabolismoRESUMEN
T cell factor 1 (Tcf1) is known as a critical mediator for natural killer (NK) cell development and terminal maturation. However, its essential targets and precise mechanisms involved in early NK progenitors (NKP) are not well clarified. To investigate the role of Tcf1 in NK cells at distinct developmental phases, we employed three kinds of genetic mouse models, namely, Tcf7fl/flVavCre/+, Tcf7fl/flCD122Cre/+ and Tcf7fl/flNcr1Cre/+ mice, respectively. Similar to Tcf1 germline knockout mice, we found notably diminished cell number and defective development in BM NK cells from all strains. In contrast, Tcf7fl/flNcr1Cre/+ mice exhibited modest defects in splenic NK cells compared with those in the other two strains. By analyzing the published ATAC-seq and ChIP-seq data, we found that Tcf1 directly targeted 110 NK cell-related genes which displayed differential accessibility in the absence of Tcf1. Along with this clue, we further confirmed that a series of essential regulators were expressed aberrantly in distinct BM NK subsets with conditional ablating Tcf1 at NKP stage. Eomes, Ets1, Gata3, Ikzf1, Ikzf2, Nfil3, Runx3, Sh2d1a, Slamf6, Tbx21, Tox, and Zeb2 were downregulated, whereas Spi1 and Gzmb were upregulated in distinct NK subsets due to Tcf1 deficiency. The dysregulation of these genes jointly caused severe defects in NK cells lacking Tcf1. Thus, our study identified essential targets of Tcf1 in NK cells, providing new insights into Tcf1-dependent regulatory programs in step-wise governing NK cell development.
Asunto(s)
Factor Nuclear 1-alfa del Hepatocito/metabolismo , Células Asesinas Naturales/fisiología , Subgrupos Linfocitarios/fisiología , Células Progenitoras Linfoides/fisiología , Animales , Antígenos Ly/genética , Antígenos Ly/metabolismo , Diferenciación Celular , Células Cultivadas , Regulación de la Expresión Génica , Granzimas/genética , Granzimas/metabolismo , Factor Nuclear 1-alfa del Hepatocito/genética , Activación de Linfocitos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptor 1 Gatillante de la Citotoxidad Natural/genética , Receptor 1 Gatillante de la Citotoxidad Natural/metabolismo , Proteínas de Dominio T Box/genética , Proteínas de Dominio T Box/metabolismoRESUMEN
Our mathematical model of integration site data in clinical gene therapy supported the existence of long-term lymphoid progenitors capable of surviving independently from hematopoietic stem cells. To date, no experimental setting has been available to validate this prediction. We here report evidence of a population of lymphoid progenitors capable of independently maintaining T and NK cell production for 15 years in humans. The gene therapy patients of this study lack vector-positive myeloid/B cells indicating absence of engineered stem cells but retain gene marking in both T and NK. Decades after treatment, we can still detect and analyse transduced naïve T cells whose production is likely maintained by a population of long-term lymphoid progenitors. By tracking insertional clonal markers overtime, we suggest that these progenitors can support both T and NK cell production. Identification of these long-term lymphoid progenitors could be utilised for the development of next generation gene- and cancer-immunotherapies.
Asunto(s)
Células Asesinas Naturales/fisiología , Linfocitos/fisiología , Células Progenitoras Linfoides/fisiología , Linfocitos T/fisiología , Linfocitos B , Terapia Genética/métodos , Células Madre Hematopoyéticas , Humanos , Interferón gamma/metabolismo , Mutagénesis , Células Mieloides/fisiología , Proto-Oncogenes/genética , Proto-Oncogenes/fisiologíaRESUMEN
Innate lymphoid cells (ILCs) develop from common lymphoid progenitors (CLPs), which further differentiate into the common ILC progenitor (CILP) that can give rise to both ILCs and natural killer (NK) cells. Murine ILC intermediates have recently been characterized, but the human counterparts and their developmental trajectories have not yet been identified, largely due to the lack of homologous surface receptors in both organisms. Here, we show that human CILPs (CD34+CD117+α4ß7+Lin-) acquire CD48 and CD52, which define NK progenitors (NKPs) and ILC precursors (ILCPs). Two distinct NK cell subsets were generated in vitro from CD34+CD117+α4ß7+Lin-CD48-CD52+ and CD34+CD117+α4ß7+Lin-CD48+CD52+ NKPs, respectively. Independent of NKPs, ILCPs exist in the CD34+CD117+α4ß7+Lin-CD48+CD52+ subset and give rise to ILC1s, ILC2s, and NCR+ ILC3s, whereas CD34+CD117+α4ß7+Lin-CD48+CD52- ILCPs give rise to a distinct subset of ILC3s that have lymphoid tissue inducer (LTi)-like properties. In addition, CD48-expressing CD34+CD117+α4ß7+Lin- precursors give rise to tissue-associated ILCs in vivo. We also observed that the interaction of 2B4 with CD48 induced differentiation of ILC2s, and together, these findings show that expression of CD48 by human ILCPs modulates ILC differentiation.
Asunto(s)
Antígeno CD48/metabolismo , Diferenciación Celular/inmunología , Células Asesinas Naturales/fisiología , Células Progenitoras Linfoides/fisiología , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/metabolismo , Animales , Antígeno CD52/metabolismo , Separación Celular , Células Cultivadas , Citometría de Flujo , Técnicas de Inactivación de Genes , Humanos , Inmunidad Innata , Ratones , Cultivo Primario de Células , RNA-Seq , Transducción de Señal/genética , Transducción de Señal/inmunología , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/genética , Análisis de la Célula Individual , Especificidad de la EspecieRESUMEN
Plasmacytoid dendritic cells (pDCs) are an immune subset specialized in the production of Type I Interferons (IFNs). They are characterized by co-expression of myeloid and lymphoid markers. Their developmental origin has been studied since their discovery and the identification of a myeloid progenitor capable of generating all dendritic cell (DC) subsets, including pDCs, led to their classification within the myeloid compartment. However, recent findings challenge this hypothesis and provide evidence for a lymphoid origin for the majority of pDCs 46-48. In this review we discuss and present the original myeloid and the newer lymphoid developmental trajectories of pDCs.
Asunto(s)
Diferenciación Celular/inmunología , Linaje de la Célula/inmunología , Linaje de la Célula/fisiología , Células Dendríticas/fisiología , Células Progenitoras Linfoides/fisiología , Animales , Humanos , Células Progenitoras MieloidesRESUMEN
Tumor progression in the host leads to severe impairment of intrathymic T-cell differentiation/maturation, leading to the paralysis of cellular anti-tumor immunity. Such suppression manifests the erosion of CD4+CD8+ double-positive (DP) immature thymocytes and a gradual increase in CD4-CD8- double negative (DN) early T-cell progenitors. The impact of such changes on the T-cell progenitor pool in the context of cancer remains poorly investigated. Here, we show that tumor progression blocks the transition of Lin-Thy1.2+CD25+CD44+c-KitlowDN2b to Lin-Thy1.2+CD25+CD44-c-Kit-DN3 in T-cell maturation, instead leading to DN2-T-cell differentiation into dendritic cells (DC). We observed that thymic IL-10 expression is upregulated, particularly at cortico-medullary junctions (CMJ), under conditions of progressive disease, resulting in the termination of IL-10Rhigh DN2-T-cell maturation due to dysregulated expression of Notch1 and its target, CCR7 (thus restricting these cells to the CMJ). Intrathymic differentiation of T-cell precursors in IL-10-/- mice and in vitro fetal thymic organ cultures revealed that IL-10 promotes the interaction between thymic stromal cells and Notch1low DN2-T cells, thus facilitating these DN2-T cells to differentiate toward CD45+CD11c+MHC-II+ thymic DCs as a consequence of activating the Ikaros/IRF8 signaling axis. We conclude that a novel function of thymically-expressed IL-10 in the tumor-bearing host diverts T-cell differentiation toward a DC pathway, thus limiting the protective adaptive immune repertoire.
Asunto(s)
Células Dendríticas/fisiología , Factor de Transcripción Ikaros/metabolismo , Células Progenitoras Linfoides/fisiología , Receptor Notch1/metabolismo , Sarcoma/inmunología , Neoplasias Cutáneas/inmunología , Linfocitos T/fisiología , Timo/citología , Animales , Puntos de Control del Ciclo Celular , Diferenciación Celular , Línea Celular Tumoral , Proliferación Celular , Regulación Neoplásica de la Expresión Génica , Humanos , Factor de Transcripción Ikaros/genética , Interleucina-10/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptor Notch1/genética , Transducción de SeñalRESUMEN
CD8+ T cell exhaustion is a state of dysfunction acquired in chronic viral infection and cancer, characterized by the formation of Slamf6+ progenitor exhausted and Tim-3+ terminally exhausted subpopulations through unknown mechanisms. Here we establish the phosphatase PTPN2 as a new regulator of the differentiation of the terminally exhausted subpopulation that functions by attenuating type 1 interferon signaling. Deletion of Ptpn2 in CD8+ T cells increased the generation, proliferative capacity and cytotoxicity of Tim-3+ cells without altering Slamf6+ numbers during lymphocytic choriomeningitis virus clone 13 infection. Likewise, Ptpn2 deletion in CD8+ T cells enhanced Tim-3+ anti-tumor responses and improved tumor control. Deletion of Ptpn2 throughout the immune system resulted in MC38 tumor clearance and improved programmed cell death-1 checkpoint blockade responses to B16 tumors. Our results indicate that increasing the number of cytotoxic Tim-3+CD8+ T cells can promote effective anti-tumor immunity and implicate PTPN2 in immune cells as an attractive cancer immunotherapy target.
Asunto(s)
Adenocarcinoma/inmunología , Linfocitos T CD8-positivos/fisiología , Neoplasias del Colon/inmunología , Inmunoterapia/métodos , Coriomeningitis Linfocítica/inmunología , Virus de la Coriomeningitis Linfocítica/fisiología , Células Progenitoras Linfoides/fisiología , Melanoma/inmunología , Proteína Tirosina Fosfatasa no Receptora Tipo 2/metabolismo , Neoplasias Cutáneas/inmunología , Animales , Senescencia Celular , Citotoxicidad Inmunológica , Femenino , Receptor 2 Celular del Virus de la Hepatitis A/metabolismo , Tolerancia Inmunológica , Interferón Tipo I/metabolismo , Masculino , Melanoma Experimental , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteína Tirosina Fosfatasa no Receptora Tipo 2/genética , Transducción de Señal , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/metabolismoRESUMEN
Development of lymphoid progenitors requires a coordinated regulation of gene expression, DNA replication, and gene rearrangement. Chromatin-remodeling activities directed by SWI/SNF2 superfamily complexes play important roles in these processes. In this study, we used a conditional knockout mouse model to investigate the role of Smarca5, a member of the ISWI subfamily of such complexes, in early lymphocyte development. Smarca5 deficiency results in a developmental block at the DN3 stage of αß thymocytes and pro-B stage of early B cells at which the rearrangement of Ag receptor loci occurs. It also disturbs the development of committed (CD73+) γδ thymocytes. The αß thymocyte block is accompanied by massive apoptotic depletion of ß-selected double-negative DN3 cells and premitotic arrest of CD4/CD8 double-positive cells. Although Smarca5-deficient αß T cell precursors that survived apoptosis were able to undergo a successful TCRß rearrangement, they exhibited a highly abnormal mRNA profile, including the persistent expression of CD44 and CD25 markers characteristic of immature cells. We also observed that the p53 pathway became activated in these cells and that a deficiency of p53 partially rescued the defect in thymus cellularity (in contrast to early B cells) of Smarca5-deficient mice. However, the activation of p53 was not primarily responsible for the thymocyte developmental defects observed in the Smarca5 mutants. Our results indicate that Smarca5 plays a key role in the development of thymocytes undergoing ß-selection, γδ thymocytes, and also B cell progenitors by regulating the transcription of early differentiation programs.
Asunto(s)
Adenosina Trifosfatasas/metabolismo , Linfocitos B/fisiología , Proteínas Cromosómicas no Histona/metabolismo , Células Progenitoras Linfoides/fisiología , Linfocitos T/fisiología , Timocitos/fisiología , Adenosina Trifosfatasas/genética , Animales , Diferenciación Celular , Células Cultivadas , Proteínas Cromosómicas no Histona/genética , Selección Clonal Mediada por Antígenos , Reordenamiento Génico , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de Antígenos de Linfocitos B/genética , Receptores de Antígenos de Linfocitos B/metabolismo , Receptores de Antígenos de Linfocitos T alfa-beta/genética , Receptores de Antígenos de Linfocitos T alfa-beta/metabolismo , Receptores de Antígenos de Linfocitos T gamma-delta/genética , Receptores de Antígenos de Linfocitos T gamma-delta/metabolismo , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
Innate lymphoid cells (ILCs) play strategic roles in tissue homeostasis and immunity. ILCs arise from lymphoid progenitors undergoing lineage restriction and the development of specialized ILC subsets. We generated "5x polychromILC" transcription factor reporter mice to delineate ILC precursor states by revealing the multifaceted expression of key ILC-associated transcription factors (Id2, Bcl11b, Gata3, RORγt, and RORα) during ILC development in the bone marrow. This approach allowed previously unattained enrichment of rare progenitor subsets and revealed hitherto unappreciated ILC precursor heterogeneity. In vivo and in vitro assays identified precursors with potential to generate all ILC subsets and natural killer (NK) cells, and also permitted discrimination of elusive ILC3 bone marrow antecedents. Single-cell gene expression analysis identified a discrete ILC2-committed population and delineated transition states between early progenitors and a highly heterogeneous ILC1, ILC3, and NK precursor cell cluster. This diversity might facilitate greater lineage potential upon progenitor recruitment to peripheral tissues.
Asunto(s)
Médula Ósea/inmunología , Subgrupos Linfocitarios/fisiología , Linfocitos/fisiología , Células Progenitoras Linfoides/fisiología , Factores de Transcripción/metabolismo , Animales , Diferenciación Celular , Línea Celular , Linaje de la Célula , Regulación del Desarrollo de la Expresión Génica , Genes Reporteros , Inmunidad Innata , Células Asesinas Naturales/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Análisis de la Célula Individual , Factores de Transcripción/genéticaRESUMEN
During development innate lymphoid cells and specialized lymphocyte subsets colonize peripheral tissues, where they contribute to organogenesis and later constitute the first line of protection while maintaining tissue homeostasis. A few of these subsets are produced only during embryonic development and remain in the tissues throughout life. They are generated through a unique developmental program initiated in lympho-myeloid-primed progenitors, which lose myeloid and B cell potential. They either differentiate into innate lymphoid cells or migrate to the thymus to give rise to embryonic T cell receptor-invariant T cells. At later developmental stages, adaptive T lymphocytes are derived from lympho-myeloid progenitors that colonize the thymus, while lymphoid progenitors become specialized in the production of B cells. This sequence of events highlights the requirement for stratification in the establishment of immune functions that determine efficient seeding of peripheral tissues by a limited number of cells.
Asunto(s)
Linfocitos B/inmunología , Linfocitos/fisiología , Células Progenitoras Linfoides/fisiología , Células T Asesinas Naturales/inmunología , Timo/inmunología , Animales , Diferenciación Celular , Linaje de la Célula , Microambiente Celular , Citocinas/metabolismo , Humanos , Inmunidad Innata , Activación de Linfocitos , Comunicación Paracrina , TranscriptomaRESUMEN
The developmental programs that generate a broad repertoire of regulatory T cells (Treg cells) able to respond to both self antigens and non-self antigens remain unclear. Here we found that mature Treg cells were generated through two distinct developmental programs involving CD25+ Treg cell progenitors (CD25+ TregP cells) and Foxp3lo Treg cell progenitors (Foxp3lo TregP cells). CD25+ TregP cells showed higher rates of apoptosis and interacted with thymic self antigens with higher affinity than did Foxp3lo TregP cells, and had a T cell antigen receptor repertoire and transcriptome distinct from that of Foxp3lo TregP cells. The development of both CD25+ TregP cells and Foxp3lo TregP cells was controlled by distinct signaling pathways and enhancers. Transcriptomics and histocytometric data suggested that CD25+ TregP cells and Foxp3lo TregP cells arose by coopting negative-selection programs and positive-selection programs, respectively. Treg cells derived from CD25+ TregP cells, but not those derived from Foxp3lo TregP cells, prevented experimental autoimmune encephalitis. Our findings indicate that Treg cells arise through two distinct developmental programs that are both required for a comprehensive Treg cell repertoire capable of establishing immunotolerance.
Asunto(s)
Diferenciación Celular/inmunología , Encefalomielitis Autoinmune Experimental/inmunología , Células Progenitoras Linfoides/fisiología , Linfocitos T Reguladores/fisiología , Timo/crecimiento & desarrollo , Animales , Autoantígenos/inmunología , Colitis/inmunología , Modelos Animales de Enfermedad , Factores de Transcripción Forkhead/genética , Factores de Transcripción Forkhead/metabolismo , Adyuvante de Freund/administración & dosificación , Adyuvante de Freund/inmunología , Humanos , Tolerancia Inmunológica/inmunología , Subunidad alfa del Receptor de Interleucina-2/metabolismo , Células Progenitoras Linfoides/trasplante , Ratones , Ratones Transgénicos , Mycobacterium tuberculosis/inmunología , Glicoproteína Mielina-Oligodendrócito/administración & dosificación , Glicoproteína Mielina-Oligodendrócito/inmunología , Fragmentos de Péptidos/administración & dosificación , Fragmentos de Péptidos/inmunología , Transducción de Señal , Organismos Libres de Patógenos Específicos , Timo/citología , Timo/inmunologíaRESUMEN
Innate lymphoid cells (ILCs) guard epithelial tissue integrity during homeostasis, but can be potent immune effector cells during inflammation. Precursors to all ILC subsets (ILC precursors [ILCP]) have been identified in human peripheral blood (PB). We found that during homeostasis, ILCP in PB of mouse and human expressed homing receptors for secondary lymphoid organs, mainly CD62L. These ILCP entered mouse lymph nodes in a CD62L-dependent way and relied on S1P receptors for their exit. Importantly, CD62L expression was absent on human ILCs expressing NKp44 in tonsils and PB of Crohn disease patients, and relatively fewer CD62L+ ILCP were present in PB of Crohn disease patients. These data are in agreement with selective expression of CD62L on nonactivated ILCP. As such, we conclude that CD62L not only serves as a functional marker of ILCP, but has potential to be used in the clinic as a diagnostic marker in inflammatory disorders.
Asunto(s)
Células Sanguíneas/inmunología , Enfermedad de Crohn/inmunología , Selectina L/metabolismo , Ganglios Linfáticos/inmunología , Linfocitos/inmunología , Células Progenitoras Linfoides/fisiología , Animales , Células Cultivadas , Femenino , Homeostasis , Humanos , Inmunidad Innata , Selectina L/genética , Activación de Linfocitos , Masculino , Ratones , Ratones Endogámicos C57BL , Receptor 2 Gatillante de la Citotoxidad Natural/metabolismo , Receptores de Lisoesfingolípidos/metabolismoRESUMEN
Innate lymphoid cells (ILCs) are critical to effective immune surveillance against pathogens, have malignant counterparts, and contribute to disease. Thus, it is important to understand ILC development. All ILCs are derived from the common lymphoid progenitor cell; however, the exact mechanisms and signals that initiate their divergence from T cells, B cells and one and other are incompletely understood. Evidence now supports a stepwise developmental process that includes distinct cellular intermediates, progressively narrowed differentiation, and some plasticity. While the current models of human and murine ILC development share many similarities, they also include some distinct differences. Together these findings have established a working dynamic model of ILC development.
Asunto(s)
Plasticidad de la Célula , Inmunidad Innata , Linfocitos/fisiología , Células Progenitoras Linfoides/fisiología , Linfopoyesis , Animales , Diferenciación Celular , Humanos , Ratones , Modelos Inmunológicos , Transducción de SeñalRESUMEN
Within the hematopoietic system, the Notch pathway is critical for promoting thymic T cell development and suppressing the B and myeloid lineage fates; however, its impact on NK lymphopoiesis is less understood. To study the role of Notch during NK cell development in vivo, we investigated different NK cell compartments and function in Rbp-Jkfl/flVav-Cretg/+ mice, in which Rbp-Jk, the major transcriptional effector of canonical Notch signaling, was specifically deleted in all hematopoietic cells. Peripheral conventional cytotoxic NK cells in Rbp-Jk-deleted mice were significantly reduced and had an activated phenotype. Furthermore, the pool of early NK cell progenitors in the bone marrow was decreased, whereas immature NK cells were increased, leading to a block in NK cell maturation. These changes were cell intrinsic as the hematopoietic chimeras generated after transplantation of Rbp-Jk-deficient bone marrow cells had the same NK cell phenotype as the Rbp-Jk-deleted donor mice, whereas the wild-type competitors did not. The expression of several crucial NK cell regulatory pathways was significantly altered after Rbp-Jk deletion. Together, these results demonstrate the involvement of canonical Notch signaling in regulation of multiple stages of NK cell development.
Asunto(s)
Proteína de Unión a la Señal Recombinante J de las Inmunoglobulinas/genética , Células Asesinas Naturales/fisiología , Células Progenitoras Linfoides/fisiología , Linfopoyesis , Receptores Notch/metabolismo , Animales , Diferenciación Celular , Células Cultivadas , Quimera , Citotoxicidad Inmunológica , Activación de Linfocitos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Transducción de SeñalRESUMEN
De novo generation of T cells depends on continual colonization of the thymus by bone marrow-derived progenitors. Thymus seeding progenitors (TSPs) constitute a heterogeneous population comprising multipotent and lineage-restricted cell types. Entry into the thymic microenvironment is tightly controlled and recent quantitative studies have revealed that the adult murine thymus only contains approximately 160 niches to accommodate TSPs. Of these niches only about 6% are open for seeding on average at steady-state. Here, I review the state of understanding of colonization of the adult murine thymus with a particular focus on past and current controversies in the field. Improving thymus colonization and/or maintaining intact TSP niches during the course of pre-conditioning regimens are likely to be critical for efficient T-cell regeneration after hematopoietic stem cell transplantation.
Asunto(s)
Células de la Médula Ósea/fisiología , Células Progenitoras Linfoides/fisiología , Nicho de Células Madre/fisiología , Linfocitos T/fisiología , Timo/fisiología , Animales , Diferenciación Celular , Linaje de la Célula , Movimiento Celular , Trasplante de Células Madre Hematopoyéticas , Homeostasis , Humanos , Ratones , Acondicionamiento PretrasplanteRESUMEN
Single-cell RNA sequencing is a powerful technology for assessing heterogeneity within defined cell populations. Here, we describe the heterogeneity of a B220+CD117intCD19-NK1.1- uncommitted hematopoietic progenitor having combined lymphoid and myeloid potential. Phenotypic and functional assays revealed four subpopulations within the progenitor with distinct lineage developmental potentials. Among them, the Ly6D+SiglecH-CD11c- fraction was lymphoid-restricted exhibiting strong B-cell potential, whereas the Ly6D-SiglecH-CD11c- fraction showed mixed lympho-myeloid potential. Single-cell RNA sequencing of these subsets revealed that the latter population comprised a mixture of cells with distinct lymphoid and myeloid transcriptional signatures and identified a subgroup as the potential precursor of Ly6D+SiglecH-CD11c- Subsequent functional assays confirmed that B220+CD117intCD19-NK1.1- single cells are, with rare exceptions, not bipotent for lymphoid and myeloid lineages. A B-cell priming gradient was observed within the Ly6D+SiglecH-CD11c- subset and we propose a herein newly identified subgroup as the direct precursor of the first B-cell committed stage. Therefore, the apparent multipotency of B220+CD117intCD19-NK1.1- progenitors results from underlying heterogeneity at the single-cell level and highlights the validity of single-cell transcriptomics for resolving cellular heterogeneity and developmental relationships among hematopoietic progenitors.
Asunto(s)
Células Madre Hematopoyéticas/fisiología , Análisis de Secuencia de ARN/métodos , Animales , Linfocitos B/citología , Linfocitos B/fisiología , Diferenciación Celular , Linaje de la Célula , Femenino , Perfilación de la Expresión Génica , Heterogeneidad Genética , Células Madre Hematopoyéticas/citología , Secuenciación de Nucleótidos de Alto Rendimiento , Células Progenitoras Linfoides/citología , Células Progenitoras Linfoides/fisiología , Masculino , Ratones Endogámicos C57BL , Células Progenitoras Mieloides/citología , Células Progenitoras Mieloides/fisiología , Análisis de la Célula IndividualRESUMEN
Total body irradiation (TBI) damages hematopoietic cells in the bone marrow and thymus; however, the long-term effects of irradiation with aging remain unclear. In this study, we found that the impact of radiation on thymopoiesis in mice varied by sex and dose but, overall, thymopoiesis remained suppressed for ≥12 mo after a single exposure. Male and female mice showed a long-term dose-dependent reduction in thymic cKit+ lymphoid progenitors that was maintained throughout life. Damage to hematopoietic stem cells (HSCs) in the bone marrow was dose dependent, with as little as 0.5 Gy causing a significant long-term reduction. In addition, the potential for T lineage commitment was radiation sensitive with aging. Overall, the impact of irradiation on the hematopoietic lineage was more severe in females. In contrast, the rate of decline in thymic epithelial cell numbers with age was radiation-sensitive only in males, and other characteristics including Ccl25 transcription were unaffected. Taken together, these data suggest that long-term suppression of thymopoiesis after sublethal irradiation was primarily due to fewer progenitors in the BM combined with reduced potential for T lineage commitment. A single irradiation dose also caused synchronization of thymopoiesis, with a periodic thymocyte differentiation profile persisting for at least 12 mo postirradiation. This study suggests that the number and capability of HSCs for T cell production can be dramatically and permanently damaged after a single relatively low TBI dose, accelerating aging-associated thymic involution. Our findings may impact evaluation and therapeutic intervention of human TBI events.
Asunto(s)
Células de la Médula Ósea/fisiología , Hematopoyesis/efectos de la radiación , Síndromes de Inmunodeficiencia/inmunología , Células Progenitoras Linfoides/fisiología , Linfocitos T/fisiología , Timo/efectos de la radiación , Envejecimiento , Animales , Diferenciación Celular , Linaje de la Célula , Células Cultivadas , Femenino , Síndromes de Inmunodeficiencia/etiología , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas Proto-Oncogénicas c-kit/metabolismo , Timo/inmunología , Irradiación Corporal Total/efectos adversosRESUMEN
Lineage-Sca-1+c-Kit- (LSK-) cells are a lymphoid progenitor population that expands in the spleen and preferentially differentiates into mature B cells in response to Plasmodium yoelii infection in mice. Furthermore, LSK- derived B cells can subsequently contribute to the ongoing immune response through the generation of parasite-specific Ab-secreting cells, as well as germinal center and memory B cells. However, the factors that promote their differentiation into B cells in the spleen postinfection are not defined. In this article, we show that LSK- cells produce the cytokine IL-17 in response to Plasmodium infection. Using Il-17ra-/- mice, IL-17R signaling in cells other than LSK- cells was found to support their differentiation into B cells. Moreover, primary splenic stromal cells grown in the presence of IL-17 enhanced the production of CXCL12, a chemokine associated with B cell development in the bone marrow, by a population of IL-17RA-expressing podoplanin+CD31- stromal cells, a profile associated with fibroblastic reticular cells. Subsequent blockade of CXCL12 in vitro reduced differentiation of LSK- cells into B cells, supporting a direct role for this chemokine in this process. Immunofluorescence indicated that podoplanin+ stromal cells in the red pulp were the primary producers of CXCL12 after P. yoelii infection. Furthermore, podoplanin staining on stromal cells was more diffuse, and CXCL12 staining was dramatically reduced in Il-17ra-/- mice postinfection. Together, these results identify a distinct pathway that supports lymphoid development in the spleen during acute Plasmodium infection.
Asunto(s)
Células Productoras de Anticuerpos/fisiología , Linfocitos B/fisiología , Interleucina-17/metabolismo , Células Progenitoras Linfoides/fisiología , Malaria/inmunología , Plasmodium yoelii/inmunología , Bazo/inmunología , Animales , Anticuerpos Antiprotozoarios/metabolismo , Células Productoras de Anticuerpos/parasitología , Linfocitos B/parasitología , Diferenciación Celular , Células Cultivadas , Quimiocina CXCL12/metabolismo , Femenino , Humanos , Memoria Inmunológica , Células Progenitoras Linfoides/parasitología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas Proto-Oncogénicas pp60(c-src)/metabolismo , Receptores de Interleucina-17/genéticaRESUMEN
Innate lymphoid cells (ILCs) are lineage- and antigen receptor-negative lymphocytes including natural killer (NK) cells and at least three distinguishable cell subsets (ILC1, ILC2, ILC3) that rapidly produce cytokines (IFN-γ, IL-5, IL-13, IL-17A, IL-22) upon activation. As such, ILCs can act as first-line defenders in the context of infection, inflammation and cancer. Because of the strong conservation between the expression of key transcription factors that can drive signature cytokine outputs in ILCs and differentiated helper T cells, it has been proposed that ILCs represent innate counterparts of the latter. Several distinct ILC precursors (ILCP) with pan-ILC (giving rise to all ILCs) or subset-restricted potentials have been described in both mouse and man. How and where these different ILCP give rise to more mature tissue-resident ILCs remains unclear. Recently, environmental signals have been shown to epigenetically influence canonical ILC differentiation pathways, generating substantial functional plasticity. These new results suggest that while ILC differentiation may be 'fixed' in principle, it remains 'flexible' in practice. A more comprehensive knowledge in the molecular mechanisms that regulate ILC development and effector functions may allow for therapeutic manipulation of ILCs for diverse disease conditions.