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1.
Nat Immunol ; 17(12): 1467-1478, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27776108

RESUMO

Tissue-resident memory T cells (TRM cells) in the airways mediate protection against respiratory infection. We characterized TRM cells expressing integrin αE (CD103) that reside within the epithelial barrier of human lungs. These cells had specialized profiles of chemokine receptors and adhesion molecules, consistent with their unique localization. Lung TRM cells were poised for rapid responsiveness by constitutive expression of deployment-ready mRNA encoding effector molecules, but they also expressed many inhibitory regulators, suggestive of programmed restraint. A distinct set of transcription factors was active in CD103+ TRM cells, including Notch. Genetic and pharmacological experiments with mice revealed that Notch activity was required for the maintenance of CD103+ TRM cells. We have thus identified specialized programs underlying the residence, persistence, vigilance and tight control of human lung TRM cells.


Assuntos
Linfócitos T CD8-Positivos/fisiologia , Memória Imunológica , Vírus da Influenza A Subtipo H3N2/imunologia , Pulmão/imunologia , Infecções por Orthomyxoviridae/imunologia , Receptor Notch1/metabolismo , Receptor Notch2/metabolismo , Infecções Respiratórias/imunologia , Animais , Antígenos CD/metabolismo , Células Cultivadas , Feminino , Humanos , Cadeias alfa de Integrinas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Pessoa de Meia-Idade , Receptor Notch1/genética , Receptor Notch2/genética
2.
Nat Immunol ; 15(12): 1143-51, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25344724

RESUMO

Activated CD8(+) T cells choose between terminal effector cell (TEC) or memory precursor cell (MPC) fates. We found that the signaling receptor Notch controls this 'choice'. Notch promoted the differentiation of immediately protective TECs and was correspondingly required for the clearance of acute infection with influenza virus. Notch activated a major portion of the TEC-specific gene-expression program and suppressed the MPC-specific program. Expression of Notch was induced on naive CD8(+) T cells by inflammatory mediators and interleukin 2 (IL-2) via pathways dependent on the metabolic checkpoint kinase mTOR and the transcription factor T-bet. These pathways were subsequently amplified downstream of Notch, creating a positive feedback loop. Notch thus functions as a central hub where information from different sources converges to match effector T cell differentiation to the demands of an infection.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Diferenciação Celular/imunologia , Receptores Notch/imunologia , Subpopulações de Linfócitos T/imunologia , Imunidade Adaptativa/imunologia , Transferência Adotiva , Animais , Linfócitos T CD8-Positivos/citologia , Separação Celular , Citometria de Fluxo , Vírus da Influenza A , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Infecções por Orthomyxoviridae/imunologia , Reação em Cadeia da Polimerase em Tempo Real , Subpopulações de Linfócitos T/citologia , Transcriptoma , Transdução Genética
3.
Eur J Immunol ; 53(3): e2149548, 2023 03.
Artigo em Inglês | MEDLINE | ID: mdl-36642930

RESUMO

To specifically tailor immune responses to a given pathogenic threat, dendritic cells (DC) are highly heterogeneous and comprise many specialized subtypes, including conventional DC (cDC) and monocyte-derived DC (MoDC), each with distinct developmental and functional characteristics. However, the functional relationship between cDC and MoDC is not fully understood, as the overlapping phenotypes of certain type 2 cDC (cDC2) subsets and MoDC do not allow satisfactory distinction of these cells in the tissue, particularly during inflammation. However, precise cDC2 and MoDC classification is required for studies addressing how these diverse cell types control immune responses and is therefore currently one of the major interests in the field of cDC research. This review will revise murine cDC2 and MoDC biology in the steady state and under inflammatory conditions and discusses the commonalities and differences between ESAMlo cDC2, inflammatory cDC2, and MoDC and their relative contribution to the initiation, propagation, and regulation of immune responses.


Assuntos
Células Dendríticas , Monócitos , Animais , Camundongos , Fenótipo
4.
Eur J Immunol ; 53(11): e2249819, 2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-36512638

RESUMO

This article is part of the Dendritic Cell Guidelines article series, which provides a collection of state-of-the-art protocols for the preparation, phenotype analysis by flow cytometry, generation, fluorescence microscopy and functional characterization of mouse and human dendritic cells (DC) from lymphoid organs and various nonlymphoid tissues. DC are sentinels of the immune system present in almost every mammalian organ. Since they represent a rare cell population, DC need to be extracted from organs with protocols that are specifically developed for each tissue. This article provides detailed protocols for the preparation of single-cell suspensions from various mouse nonlymphoid tissues, including skin, intestine, lung, kidney, mammary glands, oral mucosa and transplantable tumors. Furthermore, our guidelines include comprehensive protocols for multiplex flow cytometry analysis of DC subsets and feature top tricks for their proper discrimination from other myeloid cells. With this collection, we provide guidelines for in-depth analysis of DC subsets that will advance our understanding of their respective roles in healthy and diseased tissues. While all protocols were written by experienced scientists who routinely use them in their work, this article was also peer-reviewed by leading experts and approved by all coauthors, making it an essential resource for basic and clinical DC immunologists.


Assuntos
Células Dendríticas , Pele , Animais , Humanos , Citometria de Fluxo , Células Mieloides , Rim , Mamíferos
6.
Proc Natl Acad Sci U S A ; 118(38)2021 09 21.
Artigo em Inglês | MEDLINE | ID: mdl-34526403

RESUMO

The spleen contains phenotypically and functionally distinct conventional dendritic cell (cDC) subpopulations, termed cDC1 and cDC2, which each can be divided into several smaller and less well-characterized subsets. Despite advances in understanding the complexity of cDC ontogeny by transcriptional programming, the significance of posttranslational modifications in controlling tissue-specific cDC subset immunobiology remains elusive. Here, we identified the cell-surface-expressed A-disintegrin-and-metalloproteinase 10 (ADAM10) as an essential regulator of cDC1 and cDC2 homeostasis in the splenic marginal zone (MZ). Mice with a CD11c-specific deletion of ADAM10 (ADAM10ΔCD11c) exhibited a complete loss of splenic ESAMhi cDC2A because ADAM10 regulated the commitment, differentiation, and survival of these cells. The major pathways controlled by ADAM10 in ESAMhi cDC2A are Notch, signaling pathways involved in cell proliferation and survival (e.g., mTOR, PI3K/AKT, and EIF2 signaling), and EBI2-mediated localization within the MZ. In addition, we discovered that ADAM10 is a molecular switch regulating cDC2 subset heterogeneity in the spleen, as the disappearance of ESAMhi cDC2A in ADAM10ΔCD11c mice was compensated for by the emergence of a Clec12a+ cDC2B subset closely resembling cDC2 generally found in peripheral lymph nodes. Moreover, in ADAM10ΔCD11c mice, terminal differentiation of cDC1 was abrogated, resulting in severely reduced splenic Langerin+ cDC1 numbers. Next to the disturbed splenic cDC compartment, ADAM10 deficiency on CD11c+ cells led to an increase in marginal metallophilic macrophage (MMM) numbers. In conclusion, our data identify ADAM10 as a molecular hub on both cDC and MMM regulating their transcriptional programming, turnover, homeostasis, and ability to shape the anatomical niche of the MZ.


Assuntos
Proteína ADAM10/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Células Dendríticas/metabolismo , Proteínas de Membrana/metabolismo , Proteína ADAM10/fisiologia , Secretases da Proteína Precursora do Amiloide/fisiologia , Animais , Células Apresentadoras de Antígenos/metabolismo , Antígeno CD11c/metabolismo , Diferenciação Celular , Proliferação de Células , Feminino , Homeostase , Tecido Linfoide/metabolismo , Macrófagos/metabolismo , Masculino , Proteínas de Membrana/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Células Mieloides/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Processamento de Proteína Pós-Traducional/genética , Processamento de Proteína Pós-Traducional/fisiologia , Transdução de Sinais , Baço/citologia , Baço/metabolismo
7.
Eur J Immunol ; 2022 Dec 23.
Artigo em Inglês | MEDLINE | ID: mdl-36563125

RESUMO

This article is part of the Dendritic Cell Guidelines article series, which provides a collection of state-of-the-art protocols for the preparation, phenotype analysis by flow cytometry, generation, fluorescence microscopy, and functional characterization of mouse and human DC from lymphoid organs, and various non-lymphoid tissues. Within this chapter, detailed protocols are presented that allow for the generation of single-cell suspensions from mouse lymphohematopoietic tissues including spleen, peripheral lymph nodes, and thymus, with a focus on the subsequent analysis of DC by flow cytometry. However, prepared single-cell suspensions can be subjected to other applications including sorting and cellular enrichment procedures, RNA sequencing, Western blotting, and many more. While all protocols were written by experienced scientists who routinely use them in their work, this article was also peer-reviewed by leading experts and approved by all co-authors, making it an essential resource for basic and clinical DC immunologists.

8.
Eur J Immunol ; 2022 Dec 23.
Artigo em Inglês | MEDLINE | ID: mdl-36563126

RESUMO

This article is part of the Dendritic Cell Guidelines article series, which provides a collection of state-of-the-art protocols for the preparation, phenotype analysis by flow cytometry, generation, fluorescence microscopy, and functional characterization of mouse and human dendritic cells (DC) from lymphoid organs and various non-lymphoid tissues. Recent studies have provided evidence for an increasing number of phenotypically distinct conventional DC (cDC) subsets that on one hand exhibit a certain functional plasticity, but on the other hand are characterized by their tissue- and context-dependent functional specialization. Here, we describe a selection of assays for the functional characterization of mouse and human cDC. The first two protocols illustrate analysis of cDC endocytosis and metabolism, followed by guidelines for transcriptomic and proteomic characterization of cDC populations. Then, a larger group of assays describes the characterization of cDC migration in vitro, ex vivo, and in vivo. The final guidelines measure cDC inflammasome and antigen (cross)-presentation activity. While all protocols were written by experienced scientists who routinely use them in their work, this article was also peer-reviewed by leading experts and approved by all co-authors, making it an essential resource for basic and clinical DC immunologists.

9.
J Immunol ; 206(8): 1681-1689, 2021 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-33820829

RESUMO

The original concept stated that immature dendritic cells (DC) act tolerogenically whereas mature DC behave strictly immunogenically. Meanwhile, it is also accepted that phenotypically mature stages of all conventional DC subsets can promote tolerance as steady-state migratory DC by transporting self-antigens to lymph nodes to exert unique functions on regulatory T cells. We propose that in vivo 1) there is little evidence for a tolerogenic function of immature DC during steady state such as CD4 T cell anergy induction, 2) all tolerance as steady-state migratory DC undergo common as well as subset-specific molecular changes, and 3) these changes differ by quantitative and qualitative markers from immunogenic DC, which allows one to clearly distinguish tolerogenic from immunogenic migratory DC.


Assuntos
Células Dendríticas/imunologia , Tolerância Imunológica/imunologia , Linfócitos T Reguladores/imunologia , Animais , Autoimunidade , Diferenciação Celular , Movimento Celular , Humanos , Imunidade Celular , Modelos Imunológicos
10.
Trends Immunol ; 36(12): 802-814, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26617322

RESUMO

Differentiation of naïve T cells into effector cells is required for optimal protection against different classes of microbial pathogen and for the development of immune memory. Recent findings have revealed important roles for the Notch signaling pathway in T cell differentiation into all known effector subsets, raising the question of how this pathway controls such diverse differentiation programs. Studies in preclinical models support the therapeutic potential of manipulating the Notch pathway to alleviate immune pathology, highlighting the importance of understanding the mechanisms through which Notch regulates T cell differentiation and function. We review these findings here, and outline both unifying principles involved in Notch-mediated T cell fate decisions and cell type- and context-specific differences that may present the most suitable points for therapeutic intervention.


Assuntos
Diferenciação Celular/imunologia , Receptores Notch/imunologia , Linfócitos T/imunologia , Animais , Humanos , Ativação Linfocitária/imunologia , Transdução de Sinais/imunologia
11.
Proc Natl Acad Sci U S A ; 109(23): 9041-6, 2012 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-22615412

RESUMO

Generation of effective immune responses requires expansion of rare antigen-specific CD4(+) T cells. The magnitude of the responding population is ultimately determined by proliferation and survival. Both processes are tightly controlled to limit responses to innocuous antigens. Sustained expansion occurs only when innate immune sensors are activated by microbial stimuli or by adjuvants, which has important implications for vaccination. The molecular identity of the signals controlling sustained T-cell responses is not fully clear. Here, we describe a prominent role for the Notch pathway in this process. Coactivation of Notch allows accumulation of far greater numbers of activated CD4(+) T cells than stimulation via T-cell receptor and classic costimulation alone. Notch does not overtly affect cell cycle entry or progression of CD4(+) T cells. Instead, Notch protects activated CD4(+) T cells against apoptosis after an initial phase of clonal expansion. Notch induces a broad antiapoptotic gene expression program that protects against intrinsic, as well as extrinsic, apoptosis pathways. Both Notch1 and Notch2 receptors and the canonical effector RBPJ (recombination signal binding protein for immunoglobulin kappa J region) are involved in this process. Correspondingly, CD4(+) T-cell responses to immunization with protein antigen are strongly reduced in mice lacking these components of the Notch pathway. Our findings, therefore, show that Notch controls the magnitude of CD4(+) T-cell responses by promoting cellular longevity.


Assuntos
Apoptose/imunologia , Receptores Notch/metabolismo , Transdução de Sinais/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Apoptose/genética , Sobrevivência Celular/imunologia , Sobrevivência Celular/fisiologia , Citometria de Fluxo , Hemocianinas , Imunização , Camundongos , Camundongos Endogâmicos C57BL , Análise em Microsséries , Reação em Cadeia da Polimerase em Tempo Real , Receptores Notch/genética
12.
Adv Exp Med Biol ; 785: 107-20, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23456843

RESUMO

A fundamental property of the adaptive immune system is the ability to generate antigen-specific memory, which protects against repeated infections with the same pathogens and determines the success of vaccination. Immune memory is built up alongside a response providing direct protection during the course of a primary immune response. For CD8 T cells, this involves the generation of two distinct types of effector cells. Short lived effector cells (SLECs) confer immediate protection, but contribute little to the memory repertoire. Memory precursor effector cells (MPECs) have the ability to respond to survival signals and develop into memory cells. These two types of cells can be distinguished on the basis of surface markers and express distinct genetic programs. A single naive CD8 T cell can give rise to both MPEC and SLEC daughter cells. This may involve an initial asymmetric division or depend on later instructive signals acting on equipotent daughter cells. Strong inflammatory signals favor the generation of SLECs and weaker inflammation favors the generation of MPECs. A distinguishing feature of MPECs is their ability to persist when most effector cells die. This survival depends on signals from the IL-7 receptor, which induce expression of anti-apoptotic factors. MPECs are therefore characterized by expression of the IL-7 receptor as well as the CCR7 chemokine receptor, which allows homing to areas in lymphoid organs where IL-7 is produced. Critical for persistence of MPECs is further their responsiveness to myeloid cell derived IL-15, which instructs these cells to switch their metabolic programs from glycolysis associated with rapid proliferation to fatty acid oxidation required during a more resting state. As the mechanisms determining generation of immunological memory are unraveled, opportunities will emerge for the improvement of vaccination strategies.


Assuntos
Imunidade Adaptativa , Linfócitos T CD8-Positivos/imunologia , Diferenciação Celular/imunologia , Memória Imunológica , Células Precursoras de Linfócitos T/imunologia , Subpopulações de Linfócitos T/imunologia , Animais , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/metabolismo , Sobrevivência Celular , Expressão Gênica , Humanos , Interleucina-15/genética , Interleucina-15/imunologia , Camundongos , Células Precursoras de Linfócitos T/citologia , Células Precursoras de Linfócitos T/metabolismo , Receptores CCR7/genética , Receptores CCR7/imunologia , Receptores de Interleucina-7/genética , Receptores de Interleucina-7/imunologia , Transdução de Sinais , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/metabolismo
13.
J Invest Dermatol ; 140(1): 132-142.e3, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31260672

RESUMO

The cell adhesion molecule E-cadherin is a major component of adherens junctions and marks Langerhans cells (LC), the only dendritic cell (DC) population of the epidermis. LC form a dense network and attach themselves to the surrounding keratinocytes via homophilic E-cadherin binding. LC activation, mobilization, and migration require a reduction in LC E-cadherin expression. To determine whether E-cadherin plays a role in regulating LC homeostasis and function, we generated CD11c-specific E-cadherin knockout mice (CD11c-Ecaddel). In the absence of E-cadherin-mediated cell adhesion, LC numbers remained stable and similar as in control mice, even in aged animals. Intriguingly, E-cadherin-deficient LC displayed a dramatically changed morphology characterized by a more rounded cell body and fewer dendrites than wild-type cells. Nevertheless, maturation and migration of LC lacking E-cadherin was not altered, neither under steady-state nor inflammatory conditions. Accordingly, CD11c-Ecaddel and control mice developed comparable contact hypersensitivity reactions and imiquimod-triggered psoriatic skin inflammation, indicating that E-cadherin on LC does not influence their ability to orchestrate T cell-mediated immunity. In conclusion, our data demonstrate that E-cadherin is dispensable to maintain LC in the epidermis and does not regulate LC maturation, migration, and function.


Assuntos
Caderinas/metabolismo , Dermatite de Contato/imunologia , Epiderme/fisiologia , Células de Langerhans/fisiologia , Psoríase/imunologia , Animais , Antígeno CD11c/genética , Antígeno CD11c/metabolismo , Caderinas/genética , Diferenciação Celular , Movimento Celular , Forma Celular , Células Cultivadas , Dermatite de Contato/genética , Modelos Animais de Doenças , Homeostase , Humanos , Imiquimode , Camundongos , Camundongos Knockout , Psoríase/genética
14.
Front Immunol ; 10: 741, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31031751

RESUMO

Dendritic cells (DC) fulfill an essential sentinel function within the immune system, acting at the interface of innate and adaptive immunity. The DC family, both in mouse and man, shows high functional heterogeneity in order to orchestrate immune responses toward the immense variety of pathogens and other immunological threats. In this review, we focus on the Langerin+CD8+ DC subpopulation in the spleen. Langerin+CD8+ DC exhibit a high ability to take up apoptotic/dying cells, and therefore they are essential to prime and shape CD8+ T cell responses. Next to the induction of immunity toward blood-borne pathogens, i.e., viruses, these DC are important for the regulation of tolerance toward cell-associated self-antigens. The ontogeny and differentiation pathways of CD8+CD103+ DC should be further explored to better understand the immunological role of these cells as a prerequisite of their therapeutic application.


Assuntos
Antígenos CD/metabolismo , Antígenos CD8/metabolismo , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Lectinas Tipo C/metabolismo , Lectinas de Ligação a Manose/metabolismo , Baço/imunologia , Baço/metabolismo , Animais , Apresentação de Antígeno/imunologia , Biomarcadores , Comunicação Celular , Apresentação Cruzada/imunologia , Suscetibilidade a Doenças , Regulação da Expressão Gênica , Humanos , Imunofenotipagem , Imunoterapia/métodos , Macrófagos/imunologia , Macrófagos/metabolismo , Fenótipo , Baço/citologia
15.
Adv Immunol ; 137: 43-82, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29455847

RESUMO

CD8+ T cells clear primary infections with intracellular pathogens and provide long-term immunity against reinfection. Two different types of CD8+ T cells are responsible for these functions: short-lived effector T cells and memory T cells. The cellular relationship between these two types of CD8+ T cells has been subject to much investigation. Both cell types can derive from a single naïve CD8+ T cell precursor. Their generation requires a fate choice early during a T cell response. As a result, two populations of T cells emerge. One of these consists of terminally differentiated short-lived effector T cells. The other contains cells able to develop into long-lived memory T cells. A foundation for development of these two populations may be laid during the first division of an activated naïve T cell precursor, as a consequence of asymmetric segregation of fate-determining factors into the daughter cells. Nonetheless, the binary choice between the two lineages is strongly influenced by signals, which ensure that the differentiation process is matched with the needs posed by the infection. Here, we will discuss the genetic and metabolic programs governing differentiation of these two lineages as well as the processes leading to their induction and consolidation to create bistability. These processes involve extensive lateral inhibition between the programs as well as positive feedback between the genetic programs and the signaling pathways responsible for their induction. These features will be highlighted by discussing the role of the Notch signaling pathway in guiding the decision between the two lineages.


Assuntos
Diferenciação Celular , Homeostase , Memória Imunológica , Subpopulações de Linfócitos T/fisiologia , Linfócitos T/fisiologia , Animais , Linhagem da Célula , Retroalimentação Fisiológica , Humanos , Imunidade Celular , Transdução de Sinais
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