RESUMO
Germinal centers (GCs) are microanatomical sites of B cell clonal expansion and antibody affinity maturation. Therein, B cells undergo the Darwinian process of somatic diversification and affinity-driven selection of immunoglobulins that produces the high-affinity antibodies essential for effective humoral immunity. Here, we review recent developments in the field of GC biology, primarily as it pertains to GCs induced by infection or immunization. First, we summarize the phenotype and function of the different cell types that compose the GC, focusing on GC B cells. Then, we review the cellular and molecular bases of affinity-dependent selection within the GC and the export of memory and plasma cells. Finally, we present an overview of the emerging field of GC clonal dynamics, focusing on how GC and post-GC selection shapes the diversity of antibodies secreted into serum.
Assuntos
Linfócitos B , Centro Germinativo , Animais , Anticorpos , Afinidade de Anticorpos , Humanos , Imunidade HumoralRESUMO
Germinal centers (GCs) form in secondary lymphoid organs in response to infection and immunization and are the source of affinity-matured B cells. The duration of GC reactions spans a wide range, and long-lasting GCs (LLGCs) are potentially a source of highly mutated B cells. We show that rather than consisting of continuously evolving B cell clones, LLGCs elicited by influenza virus or SARS-CoV-2 infection in mice are sustained by progressive replacement of founder clones by naive-derived invader B cells that do not detectably bind viral antigens. Rare founder clones that resist replacement for long periods are enriched in clones with heavily mutated immunoglobulins, including some with very high affinity for antigen, that can be recalled by boosting. Our findings reveal underappreciated aspects of the biology of LLGCs generated by respiratory virus infection and identify clonal replacement as a potential constraint on the development of highly mutated antibodies within these structures.
Assuntos
Linfócitos B , Centro Germinativo , Infecções por Vírus de RNA , Animais , Camundongos , Linfócitos B/citologia , Linfócitos B/imunologia , Células Clonais , COVID-19 , Centro Germinativo/citologia , Centro Germinativo/imunologia , SARS-CoV-2 , Influenza Humana , Infecções por Vírus de RNA/imunologia , Infecções por Vírus de RNA/patologia , Infecções por Vírus de RNA/virologiaRESUMO
Repeated exposure to pathogens or their antigens triggers anamnestic antibody responses that are higher in magnitude and affinity than the primary response. These involve reengagement of memory B cell (MBC) clones, the diversity and specificity of which determine the breadth and effectiveness of the ensuing antibody response. Using prime-boost models in mice, we find that secondary responses are characterized by a clonality bottleneck that restricts the engagement of the large diversity of MBC clones generated by priming. Rediversification of mutated MBCs is infrequent within secondary germinal centers (GCs), which instead consist predominantly of B cells without prior GC experience or detectable clonal expansion. Few MBC clones, generally derived from higher-affinity germline precursors, account for the majority of secondary antibody responses, while most primary-derived clonal diversity is not reengaged detectably by boosting. Understanding how to counter this bottleneck may improve our ability to elicit antibodies to non-immunodominant epitopes by vaccination.
Assuntos
Linfócitos B/imunologia , Centro Germinativo/imunologia , Memória Imunológica/imunologia , Imunidade Adaptativa/imunologia , Animais , Formação de Anticorpos/imunologia , Formação de Anticorpos/fisiologia , Antígenos/imunologia , Linfócitos B/metabolismo , Células CHO , Linhagem Celular , Cricetulus , Feminino , Centro Germinativo/metabolismo , Humanos , Memória Imunológica/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Modelos AnimaisRESUMO
Germinal centers (GCs) were described more than 125 years ago as compartments within secondary lymphoid organs that contained mitotic cells. Since then, it has become clear that this structure is the site of B cell clonal expansion, somatic hypermutation, and affinity-based selection, the combination of which results in the production of high-affinity antibodies. Decades of anatomical and functional studies have led to an overall model of how the GC reaction and affinity-based selection operate. More recently, the introduction of intravital imaging into the GC field has opened the door to direct investigation of certain key dynamic features of this microanatomic structure, sparking renewed interest in the relationship between cell movement and affinity maturation. We review these and other recent advances in our understanding of GCs, focusing on cellular dynamics and on the mechanism of selection of high-affinity B cells.
Assuntos
Centro Germinativo/imunologia , Animais , Linfócitos B/imunologia , Linfócitos B/metabolismo , Centro Germinativo/citologia , Centro Germinativo/metabolismo , Humanos , Tolerância a Antígenos Próprios/imunologiaRESUMO
A longitudinal study by Davis et al. followed the evolution of antibody responses in four survivors of the 2014 Ebola outbreak treated in the United States and provides insight into the emergence of neutralizing antibodies long after convalescence.
Assuntos
Ebolavirus/imunologia , Doença pelo Vírus Ebola , Anticorpos Neutralizantes , Linfócitos B , Humanos , Estudos Longitudinais , Estados UnidosRESUMO
Re-exposure to an antigen generates abundant antibody responses and drives the formation of secondary germinal centers (GCs). Recall GCs in mice consist almost entirely of naïve B cells, whereas recall antibodies derive overwhelmingly from memory B cells. Here, we examine this division between cellular and serum compartments. After repeated immunization with the same antigen, tetramer analyses of recall GCs revealed a marked decrease in the ability of B cells in these structures to bind the antigen. Boosting with viral variant proteins restored antigen binding in recall GCs, as did genetic ablation of primary-derived antibody-secreting cells through conditional deletion of Prdm1, demonstrating suppression of GC recall responses by pre-existing antibodies. In hapten-carrier experiments in which B and T cell specificities were uncoupled, memory T cell help allowed B cells with undetectable antigen binding to access GCs. Thus, antibody-mediated feedback steers recall GC B cells away from previously targeted epitopes and enables specific targeting of variant epitopes, with implications for vaccination protocols.
Assuntos
Linfócitos B , Centro Germinativo , Memória Imunológica , Centro Germinativo/imunologia , Animais , Camundongos , Memória Imunológica/imunologia , Linfócitos B/imunologia , Células T de Memória/imunologia , Camundongos Endogâmicos C57BL , Fator 1 de Ligação ao Domínio I Regulador Positivo/imunologia , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Formação de Anticorpos/imunologia , Células B de Memória/imunologia , Camundongos KnockoutRESUMO
T cell activation and function require a structured engagement of antigen-presenting cells. These cell contacts are characterized by two distinct dynamics in vivo: transient contacts resulting from promigratory junctions called immunological kinapses or prolonged contacts from stable junctions called immunological synapses. Kinapses operate in the steady state to allow referencing to self-peptide-MHC (pMHC) and searching for pathogen-derived pMHC. Synapses are induced by T cell receptor (TCR) interactions with agonist pMHC under specific conditions and correlate with robust immune responses that generate effector and memory T cells. High-resolution imaging has revealed that the synapse is highly coordinated, integrating cell adhesion, TCR recognition of pMHC complexes, and an array of activating and inhibitory ligands to promote or prevent T cell signaling. In this review, we examine the molecular components, geometry, and timing underlying kinapses and synapses. We integrate recent molecular and physiological data to provide a synthesis and suggest ways forward.
Assuntos
Sinapses Imunológicas/imunologia , Ativação Linfocitária , Linfócitos T/imunologia , Animais , Comunicação Celular , Humanos , Sinapses Imunológicas/metabolismo , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais , Linfócitos T/citologia , Linfócitos T/metabolismoRESUMO
Intestinal intraepithelial lymphocytes (IELs) are located at the critical interface between the intestinal lumen, which is chronically exposed to food and microbes, and the core of the body. Using high-resolution microscopy techniques and intersectional genetic tools, we investigated the nature of IEL responses to luminal microbes. We observed that TCRγδ IELs exhibit unique microbiota-dependent location and movement patterns in the epithelial compartment. This behavioral pattern quickly changes upon exposure to different enteric pathogens, resulting in increased interepithelial cell (EC) scanning, expression of antimicrobial genes, and glycolysis. Both dynamic and metabolic changes to γδ IEL depend on pathogen sensing by ECs. Direct modulation of glycolysis is sufficient to change γδ IEL behavior and susceptibility to early pathogen invasion. Our results uncover a coordinated EC-IEL response to enteric infections that modulates lymphocyte energy utilization and dynamics and supports maintenance of the intestinal epithelial barrier. VIDEO ABSTRACT.
Assuntos
Intestinos/citologia , Intestinos/imunologia , Infecções por Salmonella/imunologia , Linfócitos T/imunologia , Animais , Células Epiteliais/metabolismo , Vigilância Imunológica , Mucosa Intestinal/imunologia , Camundongos , Infecções por Salmonella/microbiologia , Salmonella typhimurium/fisiologiaRESUMO
Germinal centers (GCs) are the primary sites of clonal B cell expansion and affinity maturation, directing the production of high-affinity antibodies. This response is a central driver of pathogenesis in autoimmune diseases, such as systemic lupus erythematosus (SLE), but the natural history of autoreactive GCs remains unclear. Here, we present a novel mouse model where the presence of a single autoreactive B cell clone drives the TLR7-dependent activation, expansion, and differentiation of other autoreactive B cells in spontaneous GCs. Once tolerance was broken for one self-antigen, autoreactive GCs generated B cells targeting other self-antigens. GCs became independent of the initial clone and evolved toward dominance of individual clonal lineages, indicating affinity maturation. This process produced serum autoantibodies to a breadth of self-antigens, leading to antibody deposition in the kidneys. Our data provide insight into the maturation of the self-reactive B cell response, contextualizing the epitope spreading observed in autoimmune disease.
Assuntos
Linfócitos B/imunologia , Evolução Clonal , Centro Germinativo/citologia , Centro Germinativo/imunologia , Tolerância Imunológica , Animais , Autoanticorpos/imunologia , Autoantígenos/imunologia , Doenças Autoimunes/imunologia , Linfócitos B/citologia , Quimera/imunologia , Epitopos/imunologia , Rim/imunologia , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Upon viral infection, natural killer (NK) cells expressing certain germline-encoded receptors are selected, expanded, and maintained in an adaptive-like manner. Currently, these are thought to differentiate along a common pathway. However, by fate mapping of single NK cells upon murine cytomegalovirus (MCMV) infection, we identified two distinct NK cell lineages that contributed to adaptive-like responses. One was equivalent to conventional NK (cNK) cells while the other was transcriptionally similar to type 1 innate lymphoid cells (ILC1s). ILC1-like NK cells showed splenic residency and strong cytokine production but also recognized and killed MCMV-infected cells, guided by activating receptor Ly49H. Moreover, they induced clustering of conventional type 1 dendritic cells and facilitated antigen-specific T cell priming early during MCMV infection, which depended on Ly49H and the NK cell-intrinsic expression of transcription factor Batf3. Thereby, ILC1-like NK cells bridge innate and adaptive viral recognition and unite critical features of cNK cells and ILC1s.
Assuntos
Imunidade Adaptativa/imunologia , Linhagem da Célula/imunologia , Infecções por Herpesviridae/imunologia , Imunidade Inata/imunologia , Células Matadoras Naturais/imunologia , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , MuromegalovirusRESUMO
In this Minireview, we discuss basic aspects of germinal center biology in the context of immunity to influenza infection and speculate on how the simultaneous evolutionary races of virus and antibody may impact our efforts to design a universal influenza vaccine.
Assuntos
Centro Germinativo/imunologia , Vacinas contra Influenza/imunologia , Influenza Humana/imunologia , Animais , Formação de Anticorpos , Humanos , Influenza Humana/prevenção & controleRESUMO
Cancer cells frequently alter their lipids to grow and adapt to their environment1-3. Despite the critical functions of lipid metabolism in membrane physiology, signalling and energy production, how specific lipids contribute to tumorigenesis remains incompletely understood. Here, using functional genomics and lipidomic approaches, we identified de novo sphingolipid synthesis as an essential pathway for cancer immune evasion. Synthesis of sphingolipids is surprisingly dispensable for cancer cell proliferation in culture or in immunodeficient mice but required for tumour growth in multiple syngeneic models. Blocking sphingolipid production in cancer cells enhances the anti-proliferative effects of natural killer and CD8+ T cells partly via interferon-γ (IFNγ) signalling. Mechanistically, depletion of glycosphingolipids increases surface levels of IFNγ receptor subunit 1 (IFNGR1), which mediates IFNγ-induced growth arrest and pro-inflammatory signalling. Finally, pharmacological inhibition of glycosphingolipid synthesis synergizes with checkpoint blockade therapy to enhance anti-tumour immune response. Altogether, our work identifies glycosphingolipids as necessary and limiting metabolites for cancer immune evasion.
RESUMO
Immune cells rely on transient physical interactions with other immune and non-immune populations to regulate their function1. To study these 'kiss-and-run' interactions directly in vivo, we previously developed LIPSTIC (labelling immune partnerships by SorTagging intercellular contacts)2, an approach that uses enzymatic transfer of a labelled substrate between the molecular partners CD40L and CD40 to label interacting cells. Reliance on this pathway limited the use of LIPSTIC to measuring interactions between CD4+ T helper cells and antigen-presenting cells, however. Here we report the development of a universal version of LIPSTIC (uLIPSTIC), which can record physical interactions both among immune cells and between immune and non-immune populations irrespective of the receptors and ligands involved. We show that uLIPSTIC can be used, among other things, to monitor the priming of CD8+ T cells by dendritic cells, reveal the steady-state cellular partners of regulatory T cells and identify germinal centre-resident T follicular helper cells on the basis of their ability to interact cognately with germinal centre B cells. By coupling uLIPSTIC with single-cell transcriptomics, we build a catalogue of the immune populations that physically interact with intestinal epithelial cells at the steady state and profile the evolution of the interactome of lymphocytic choriomeningitis virus-specific CD8+ T cells in multiple organs following systemic infection. Thus, uLIPSTIC provides a broadly useful technology for measuring and understanding cell-cell interactions across multiple biological systems.
Assuntos
Linfócitos B , Linfócitos T CD8-Positivos , Comunicação Celular , Células Dendríticas , Células Epiteliais , Células T Auxiliares Foliculares , Linfócitos T Reguladores , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Comunicação Celular/imunologia , Células Dendríticas/citologia , Células Dendríticas/imunologia , Ligantes , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/imunologia , Células T Auxiliares Foliculares/citologia , Células T Auxiliares Foliculares/imunologia , Linfócitos B/citologia , Linfócitos B/imunologia , Centro Germinativo/citologia , Análise da Expressão Gênica de Célula Única , Células Epiteliais/citologia , Células Epiteliais/imunologia , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Vírus da Coriomeningite Linfocítica/imunologia , Coriomeningite Linfocítica/imunologia , Coriomeningite Linfocítica/virologia , Especificidade de ÓrgãosRESUMO
The gut epithelium is populated by intraepithelial lymphocytes (IELs), a heterogeneous T cell population with cytotoxic and regulatory properties, which can be acquired at the epithelial layer. However, the role of T cell receptor (TCR) in this process remains unclear. Single-cell transcriptomic analyses revealed distinct clonal expansions between cell states, with CD4+CD8αα+ IELs being one of the least diverse populations. Conditional deletion of TCR on differentiating CD4+ T cells or of major histocompatibility complex (MHC) class II on intestinal epithelial cells prevented CD4+CD8αα+ IEL differentiation. However, TCR ablation on differentiated CD4+CD8αα+ IELs or long-term cognate antigen withdraw did not affect their maintenance. TCR re-engagement of antigen-specific CD4+CD8αα+ IELs by Listeria monocytogenes did not alter their state but correlated with reduced bacterial invasion. Thus, local antigen recognition is an essential signal for differentiation of CD4+ T cells at the epithelium, yet differentiated IELs are able to preserve an effector program in the absence of TCR signaling.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Linfócitos Intraepiteliais/imunologia , Linfócitos Intraepiteliais/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Animais , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Evolução Clonal/genética , Evolução Clonal/imunologia , Antígenos de Histocompatibilidade Classe II/genética , Antígenos de Histocompatibilidade Classe II/imunologia , Imunofenotipagem , Camundongos , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Transdução de Sinais , Análise de Célula Única , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismoRESUMO
To produce high-affinity antibodies, B cells must undergo iterative cycles of targeted mutagenesis and affinity-based selection. These cycles take place in a structure known as the germinal center, which forms in secondary lymphoid organs upon exposure to antigenic stimuli. Here, we summarize our current understanding of the germinal center reaction.
Assuntos
Anticorpos/imunologia , Centro Germinativo/metabolismo , Animais , Antígenos/imunologia , Linfócitos B/imunologia , Linfócitos T/imunologiaRESUMO
The protective efficacy of serum antibodies results from the interplay of antigen-specific B cell clones of different affinities and specificities. These cellular dynamics underlie serum-level phenomena such as original antigenic sin (OAS)-a proposed propensity of the immune system to rely repeatedly on the first cohort of B cells engaged by an antigenic stimulus when encountering related antigens, in detriment to the induction of de novo responses1-5. OAS-type suppression of new, variant-specific antibodies may pose a barrier to vaccination against rapidly evolving viruses such as influenza and SARS-CoV-26,7. Precise measurement of OAS-type suppression is challenging because cellular and temporal origins cannot readily be ascribed to antibodies in circulation; its effect on subsequent antibody responses therefore remains unclear5,8. Here we introduce a molecular fate-mapping approach with which serum antibodies derived from specific cohorts of B cells can be differentially detected. We show that serum responses to sequential homologous boosting derive overwhelmingly from primary cohort B cells, while later induction of new antibody responses from naive B cells is strongly suppressed. Such 'primary addiction' decreases sharply as a function of antigenic distance, allowing reimmunization with divergent viral glycoproteins to produce de novo antibody responses targeting epitopes that are absent from the priming variant. Our findings have implications for the understanding of OAS and for the design and testing of vaccines against evolving pathogens.
Assuntos
Formação de Anticorpos , Linfócitos B , Imunização Secundária , Humanos , Anticorpos Antivirais/biossíntese , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Antígenos Virais/imunologia , Vacinas contra Influenza/imunologia , SARS-CoV-2/imunologia , Vacinação , Linfócitos B/imunologia , Vacinas Virais/imunologiaRESUMO
Class-switch recombination (CSR) is a DNA recombination process that replaces the immunoglobulin (Ig) constant region for the isotype that can best protect against the pathogen. Dysregulation of CSR can cause self-reactive BCRs and B cell lymphomas; understanding the timing and location of CSR is therefore important. Although CSR commences upon T cell priming, it is generally considered a hallmark of germinal centers (GCs). Here, we have used multiple approaches to show that CSR is triggered prior to differentiation into GC B cells or plasmablasts and is greatly diminished in GCs. Despite finding a small percentage of GC B cells expressing germline transcripts, phylogenetic trees of GC BCRs from secondary lymphoid organs revealed that the vast majority of CSR events occurred prior to the onset of somatic hypermutation. As such, we have demonstrated the existence of IgM-dominated GCs, which are unlikely to occur under the assumption of ongoing switching.
Assuntos
Linfócitos B/imunologia , Centro Germinativo/imunologia , Switching de Imunoglobulina , Plasmócitos/imunologia , Linfoma Plasmablástico/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Diferenciação Celular , Células Cultivadas , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Filogenia , Receptores de Antígenos de Linfócitos B/metabolismoRESUMO
CD4(+) follicular helper T cells (T(FH) cells) are essential for germinal center (GC) responses and long-lived antibody responses. Here we report that naive CD4(+) T cells deficient in the transcription factor Foxp1 'preferentially' differentiated into T(FH) cells, which resulted in substantially enhanced GC and antibody responses. We found that Foxp1 used both constitutive Foxp1A and Foxp1D induced by stimulation of the T cell antigen receptor (TCR) to inhibit the generation of T(FH) cells. Mechanistically, Foxp1 directly and negatively regulated interleukin 21 (IL-21); Foxp1 also dampened expression of the costimulatory molecule ICOS and its downstream signaling at early stages of T cell activation, which rendered Foxp1-deficient CD4(+) T cells partially resistant to blockade of the ICOS ligand (ICOSL) during T(FH) cell development. Our findings demonstrate that Foxp1 is a critical negative regulator of T(FH) cell differentiation.
Assuntos
Diferenciação Celular , Fatores de Transcrição Forkhead/fisiologia , Proteínas Repressoras/fisiologia , Linfócitos T Auxiliares-Indutores/citologia , Animais , Linfócitos T CD4-Positivos/imunologia , Proteína Coestimuladora de Linfócitos T Induzíveis/genética , Interleucinas/genética , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Antígenos de Linfócitos T/fisiologiaRESUMO
During antibody affinity maturation, germinal center (GC) B cells cycle between affinity-driven selection in the light zone (LZ) and proliferation and somatic hypermutation in the dark zone (DZ). Although selection of GC B cells is triggered by antigen-dependent signals delivered in the LZ, DZ proliferation occurs in the absence of such signals. We show that positive selection triggered by T cell help activates the mechanistic target of rapamycin complex 1 (mTORC1), which promotes the anabolic program that supports DZ proliferation. Blocking mTORC1 prior to growth prevented clonal expansion, whereas blockade after cells reached peak size had little to no effect. Conversely, constitutively active mTORC1 led to DZ enrichment but loss of competitiveness and impaired affinity maturation. Thus, mTORC1 activation is required for fueling B cells prior to DZ proliferation rather than for allowing cell-cycle progression itself and must be regulated dynamically during cyclic re-entry to ensure efficient affinity-based selection.