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Functional Responses and Costimulator Dependence of Memory CD4+ T Cells
Abstract
To examine the functional characteristics of memory CD4+ T cells, we used an adoptive transfer system to generate a stable population of Ag-specific memory cells in vivo and compared their responses to Ag with those of a similar population of Ag-specific naive cells. Memory cells localized to the spleen and lymph nodes of mice and exhibited extremely rapid recall responses to Ag in vivo, leaving the spleen within 3-5 days of Ag encounter. Unlike their naive counterparts, memory cells produced effector cytokines (IFN-gamma, IL-4, IL-5) within 12-24 h of Ag exposure and did not require multiple cycles of cell division to do so. Memory cells proliferated at lower Ag concentrations than did naive cells, were less dependent on costimulation by B7 molecules, and independent of costimulation by CD40. Furthermore, effector cytokine production by memory cells also occurred in the absence of either B7 or CD40 costimulation. Lastly, memory cells were resistant to tolerance induction. Together, these findings suggest that the threshold for activation of memory CD4+ cells is lower than that of naive cells. This would permit memory cells to rapidly express their effector functions in vivo earlier in the course of a secondary immune response, when the levels of Ag and the availability of costimulation may be relatively low.
... PB ILCs fail to induce T-cell antigen recall responses. In contrast to naive T helper cells, memory CD4 + T cells might not necessarily require co-stimulation 29,30 . We therefore examined the possibility that ILCs could induce memory CD4 + T-cell responses to the human cytomegalovirus (CMV) immunedominant protein pp65. ...
... In line with this, PB ILC2 and ILC3-like cells upregulated HLA-DR expression following 5-day stimulation with IL-2 alone ( Supplementary Fig. 17a). Under these conditions, however, the cells did not express any co-stimulatory molecules (Fig. 4), underlining that costimulation may not be essential to induce cytokine response by antigen-specific memory T cells as reported by others 29,30 . Moreover, the magnitude of the observed T-cell response correlated with the HLA-DR expression levels on the antigen-presenting ILC populations (Fig. 6e). ...
... Nonetheless, it remains unclear whether co-stimulation is needed for activation of memory CD4 + T cells 29,30 . We observed an increased antigen-presentation capacity of PB ILC2 and ILC3like cells cultured with IL-2 alone for 5 days. ...
ILCs and T helper cells have been shown to exert bi-directional regulation in mice. However, how crosstalk between ILCs and CD4+ T cells influences immune function in humans is unknown. Here we show that human intestinal ILCs co-localize with T cells in healthy and colorectal cancer tissue and display elevated HLA-DR expression in tumor and tumor-adjacent areas. Although mostly lacking co-stimulatory molecules ex vivo, intestinal and peripheral blood (PB) ILCs acquire antigen-presenting characteristics triggered by inflammasome-associated cytokines IL-1β and IL-18. IL-1β drives the expression of HLA-DR and co-stimulatory molecules on PB ILCs in an NF-κB-dependent manner, priming them as efficient inducers of cytomegalovirus-specific memory CD4+ T-cell responses. This effect is strongly inhibited by the anti-inflammatory cytokine TGF-β. Our results suggest that circulating and tissue-resident ILCs have the intrinsic capacity to respond to the immediate cytokine milieu and regulate local CD4+ T-cell responses, with potential implications for anti-tumor immunity and inflammation. Murine ILCs can modulate T cell responses in MHCII-dependent manner. Here the authors show that human ILCs process and present antigens and induce T-cell responses upon exposure to IL-1-family cytokines; along with the article by Lehmann et al, this work elucidates how cytokines set context specificity of ILC-T cell crosstalk by regulating ILC antigen presentation.
... Memory CD4 T cells can respond more quickly to a secondary challenge. This is, in part, because they are less reliant on a heightened level of costimulatory signals, [12][13][14] although memory CD4 T-cell cytokine responses do require the presence of costimulatory molecules. 14,15 While this contributes to rapid pathogen control, this presents significant hurdles for treatments that aim to induce antigen-specific tolerance in autoimmunity, allergy or transplantation. ...
... Memory CD4 T cells respond to low doses of antigen and costimulatory signals, suggesting they will be refractory to tolerance induction. [12][13][14]44 This presents significant hurdles for therapies that aim to induce antigen-specific Tcell tolerance. 6,17,45 Our previous 18 and current data demonstrate that some but not all functions of memory CD4 T cells are altered following the exposure of memory CD4 T cells to antigen delivered in the absence of adjuvant. ...
Memory T cells respond rapidly in part because they are less reliant on heightened levels of costimulatory molecules. This enables rapid control of secondary infecting pathogens but presents challenges to efforts to control or silence memory CD4 T cells, for example in antigen specific tolerance strategies for autoimmunity. We have examined the transcriptional and functional consequences of re-activating memory CD4 T cells in the absence of an adjuvant. We find that memory CD4 T cells generated by infection or immunisation survive secondary activation with antigen delivered without adjuvant, regardless of their location in secondary lymphoid organs or peripheral tissues. These cells were, however, functionally altered following a tertiary immunisation with antigen and adjuvant, proliferating poorly but maintaining their ability to produce inflammatory cytokines. Transcriptional and cell cycle analysis of these memory CD4 T cells suggest they are unable to commit fully to cell division potentially because of low expression of DNA repair enzymes. In contrast, these memory CD4 T cells could proliferate following tertiary reactivation by viral re-infection. These data indicate that antigen specific tolerogenic strategies must examine multiple parameters of T cell function, and provide insight into the molecular mechanisms that may lead to deletional tolerance of memory CD4 T cells.
... Nevertheless, it remains controversial whether CD28 engagement differentially impacts 55 activation of naive and memory T cells. Previous studies have suggested that memory T cells have lower costimulation thresholds or conversely, that naive cells have a greater requirement for costimulatory signals (Croft et al., 1994;Dubey et al., 1995;London et al., 2000;Luqman and Bottomly, 1992). This has given rise to a widely perceived notion that, in contrast to naive T cells, previously activated or memory T cells do not require CD28 60 costimulation. ...
... However, the relative quantities of both signals are likely to be highly variable depending on 420 the setting of T cell activation. There is evidence to suggest that naive and memory cells have different requirements for CD28 costimulation, and a widely held view is that CD28 costimulation is less required for activation of memory than naive T cells (Croft et al., 1994;Dubey et al., 1995;London et al., 2000;Luqman and Bottomly, 1992). Understanding the requirements for CD28 costimulation during immune responses is important for many 425 therapeutic approaches including immune suppression in autoimmunity and transplantation, as well as cancer immunotherapy. ...
T cell activation is a critical driver of immune response and if uncontrolled, it can result in failure to respond to infection or in excessive inflammation and autoimmunity. CD28 costimulatory pathway is an essential regulator of CD4 T cell responses. To deconvolute how T cell receptor (TCR) and CD28 orchestrate activation of human CD4 T cells we stimulated cells using varying intensities of TCR and CD28 signals followed by gene expression profiling. We demonstrate that T-helper differentiation and cytokine expression are controlled by CD28. Strikingly, cell cycle and cell division are sensitive to CD28 in memory cells, but under TCR control in naive cells, in contrast to the paradigm that memory cells are CD28-independent. Using a combination of chromatin accessibility and enhancer profiling, we observe that IRFs and Blimp-1 (PRDM1) motifs are enriched in naive and memory T cells in response to TCR. In contrast, memory cells initiate AP1 transcriptional regulation only when both TCR and CD28 are engaged, implicating CD28 as an amplifier of transcriptional programmes in memory cells. Lastly, we show that CD28-sensitive genes are enriched in autoimmune disease loci, pointing towards the role of memory cells and the regulation of T cell activation through CD28 in autoimmune disease development. This study provides important insights into the differential role of CD28 in naive and memory T cell responses and offers a new platform for design and interpretation of costimulatory based therapies.
One-sentence summary
Genomic profiling of CD4 T cell activation reveals a sensitivity switch from TCR in naive to CD28 in memory cells.
... Following pathogen clearance, antigen-specific T cells survive as memory T cells or effector T cells that either recirculate or reside permanently in the tissues 5 . In contrast to the three signal requirement for naive T cell priming 6 , it has long been presumed that MHC-TCR interactions alone are sufficient for memory CD4 T cell reactivation and effector function 7,8 . More recent work, however, has shown that co-stimulation via CD80/86 is also critical for reactivation of memory CD4 T cells [9][10][11] . ...
... DCs provide key signals for CD4 T cell effector function. It has long been understood that engagement of TCR and costimulatory molecules on antigen-experienced CD4 T cells (effector, effector memory and tissue resident memory cells) is sufficient to drive rapid production of effector cytokines 8,10 . This is based on the conceptual framework that once primed, CD4 T cells need not depend on additional innate immune signals as this would impose a degree of redundancy and might delay the effector response. ...
Innate cytokines are critical drivers of priming and differentiation of naive CD4 T cells, but their functions in memory T cell response are largely undefined. Here we show that IL-1 acts as a licensing signal to permit effector cytokine production by pre-committed Th1 (IFN-γ), Th2 (IL-13, IL-4, and IL-5) and Th17 (IL-17A, IL-17F, and IL-22) lineage cells. This licensing function of IL-1 is conserved across effector CD4 T cells generated by diverse immunological insults. IL-1R signaling stabilizes cytokine transcripts to enable productive and rapid effector functions. We also demonstrate that successful lineage commitment does not translate into productive effector functions in the absence of IL-1R signaling. Acute abrogation of IL-1R signaling in vivo results in reduced IL-17A production by intestinal Th17 cells. These results extend the role of innate cytokines beyond CD4 T cell priming and establish IL-1 as a licensing signal for memory CD4 T cell function.
... [71][72][73] Previously activated CD4 T cells are known to require lower epitope density to become reactivated and also to expand more rapidly than naive CD4 T cells. [74][75][76][77] Our later studies showed that the advantage that is afforded to memory CD4 T cells specific for internal virion proteins, at the expense of CD4 T cells specific for HA, could be overcome by an intermediate boost with HA-derived peptides derived from the second virus. This intermediate boosting allowed the CD4 T cell response to the subsequent infection to now include CD4 T cells specific for HA and with them, HA-specific antibody responses for the second virus. ...
... Recall of memory CD4 T cells has several advantages because they exist at a higher frequency than the naive CD4 T cell population, expand more rapidly and at lower epitope density than naive CD4 T cells. [74][75][76][77]175,176 For those healthy adults with a very limited repertoire of CD4 T cells that cross-react with avian HA proteins, prepandemic priming with H5 or H7 proteins or peptides might be a very useful strategy. 147,148,[177][178][179] By populating humans with avian HA-epitope-specific CD4 T memory cells that can be recalled, these hosts are more likely to produce neutralizing HA-specific antibody responses and/or to more vigorously respond to avian influenza derived vaccines, if avian influenza viruses gain the ability to transmit across human populations. ...
CD4 T cells convey a number of discrete functions to protective immunity to influenza, a complexity that distinguishes this arm of adaptive immunity from B cells and CD8 T cells. Although the most well recognized function of CD4 T cells is provision of help for antibody production, CD4 T cells are important in many aspects of protective immunity. Our studies have revealed that viral antigen specificity is a key determinant of CD4 T cell function, as illustrated both by mouse models of infection and human vaccine responses, a factor whose importance is due at least in part to events in viral antigen handling. We discuss research that has provided insight into the diverse viral epitope specificity of CD4 T cells elicited after infection, how this primary response is modified as CD4 T cells home to the lung, establish memory, and after challenge with a secondary and distinct influenza virus strain. Our studies in human subjects point out the challenges facing vaccine efforts to facilitate responses to novel and avian strains of influenza, as well as strategies that enhance the ability of CD4 T cells to promote protective antibody responses to both seasonal and potentially pandemic strains of influenza.
... However, many studies have highlighted that while the engagement of costimulatory molecules is often required for the development of robust T cell effectors when priming naive T cells, their engagement is redundant during the reactivation of already primed T cells, the latter of which is applicable to the model system presented here. [58][59][60][61] Moreover, the consistency of antigen-presenting cell activation across treatment groups demonstrates that the specificity of the T cell responses to Spike proteins and Spike EVs in vaccinated donors is driven by antigen recognition from the T cells, as opposed to a secondary effect driven by the differential activation of antigen-presenting cells following EV treatment. ...
Extracellular vesicles (EVs) are entering the clinical arena as novel biologics for infectious diseases, potentially serving as the immunogenic components of next generation vaccines. However, relevant human assays to evaluate the immunogenicity of EVs carrying viral antigens are lacking, contributing to challenges in translating rodent studies to human clinical trials. Here, we engineered EVs to carry SARS-CoV-2 Spike to evaluate the immunogenicity of antigen-carrying EVs using human peripheral blood mononuclear cells (PBMCs). Delivery of Spike EVs to PBMCs resulted in specific immune cell activation as assessed through T cell activation marker expression. Further, Spike EVs were taken up largely by antigen-presenting cells (monocytes, dendritic cells and B cells). Taken together, this human PBMC-based system models physiologically relevant pathways of antigen delivery, uptake and presentation. In summary, the current study highlights the suitability of using human PBMCs for evaluating the immunogenicity of EVs engineered to carry antigens for infectious disease therapeutics.
... Les cellules immunitaires mémoires ont des caractéristiques fonctionnelles qui leurs permettent d'apporter une réponse secondaire à l'Ag plus rapide et plus efficace (London, Lodge, and Abbas 2000). ...
L'anémie hémolytique auto-immune à auto-anticorps chaud (AIHA) est une maladie auto-immune rare et acquise, médiée par des anticorps ciblant les globules rouges. L'implication des lymphocytes T helper (Th) CD4+ dans la physiopathologie a été peu étudiée et la plupart des résultats sont extrapolés de modèles murins. Bien que cela soit encore débattu, l’AIHA pourrait être associée à une polarisation pro-inflammatoire de type Th17 et à un déficit quantitatif en lymphocytes T régulateurs (Treg).Au cours de ce travail, nous avons réalisé la première étude concomitante des LT effecteurs et des Treg circulants, ainsi qu’une étude fonctionnelle et transcriptomique des Treg de patients atteints d’AHAI au diagnostic.Nous avons observé une orientation des LT effecteurs vers une polarisation Th17 accompagnée d’une diminution des Treg circulants, principalement des Treg effecteurs. Ce déficit quantitatif est associé à une diminution de la capacité des Treg à inhiber la prolifération des LT effecteurs, ce qui semble favoriser d’avantage la polarisation Th17.Le déficit en Treg est associé à une diminution du niveau d’expression de la protéine Foxp3, le facteur de transcription dominant des Treg, qui assure la stabilité de leur phénotype et le maintien de leurs fonctions suppressives. Le profil transcriptomique des Treg des patients a révélé : |1. Un fort engagement des voies de signalisation du TNF dans les Treg, en accord avec une augmentation significative du TNF-α sérique chez les patients. |2. Un rôle possible de mécanismes post-traductionnels conduisant à la dégradation de Foxp3 à l’origine des dysfonctionnements des Treg.A travers ce travail nous rapportons deux observations inédites et importantes pour la compréhension de la physiopathologie de l’AHAI, l’augmentation du TNF-α sérique chez les patients AHAI au diagnostic ainsi que le déficit quantitatif et fonctionnel des Treg circulants. Bien que les aspects mécanistiques ou les liens entre ces deux observations restent à préciser, elles nous permettent d’ores et déjà d’envisager de nouveaux axes thérapeutiques pour les patients tels que le renforcement des populations Treg et/ou la neutralisation de la production du TNF-α.
... A key feature of memory/ effector T-cells is that they have a lower threshold for activation and are less dependent on costimulatory signals. Although current evidence predominantly suggests that although for memory/effector T cells costimulatory signals help to boost the recall response and are required for optimal T-cell activation and effector functions, multiple studies have shown that memory T-cells can be activated to express effector functions (including cytolytic function) with no or suboptimal costimulatory signals (Croft et al. 1994;Fuse et al. 2008;London et al. 2000;Suresh et al. 2001a). In the case of T-BsAb therapy, in vitro studies have shown that effector memory T-cells (TEM) constitute the majority of proliferating T cells, while naïve T-cells largely remain unchanged (Wong et al. 2013). ...
T-cell bispecific antibodies (T-BsAbs) are a new class of cancer immunotherapy drugs that can simultaneously bind to tumor-associated antigens on target cells and to the CD3 subunit of the T-cell receptor (TCR) on T cells. In the last decade, numerous T-BsAbs have been developed for the treatment of both hematological malignancies and solid tumors. Among them, blinatumomab has been successfully used to treat CD19 positive malignancies and has been approved by the FDA as standard care for acute lymphoblastic leukemia (ALL). However, in many clinical scenarios, the efficacy of T-BsAbs remains unsatisfactory. To further improve T-BsAb therapy, it will be crucial to better understand the factors affecting treatment efficacy and the nature of the T-BsAb-induced immune response. Herein, we first review the studies on the potential mechanisms by which T-BsAbs activate T-cells and how they elicit efficient target killing despite suboptimal costimulatory support. We focus on analyzing reports from clinical trials and preclinical studies, and summarize the factors that have been identified to impact the efficacy of T-BsAbs. Lastly, we review current and propose new approaches to improve the clinical efficacy of T-BsAbs.
... [135]. Conversely, the re-activation of memory T-cell is B7-independent {c.f., raft expression) [136]. ...
p>Agonistic monoclonal antibodies directed against murine CD40 can be used in experimental settings to induce anti-lymphoma T-cells that are able to affect primary tumour eradication in vivo . These anti-tumour CTL provide both protection against re-challenge with the original vaccinating tumour line and, furthermore, prevent relapse, thereby affecting long-term survival. Such therapeutic strategies are desirable in order to provide durable responses against human lymphomas without evolving adverse side-effects.
Here, we further investigate the mechanism by which anti-CD40 exerts its therapeutic activity and demonstrate that interactions via the TNFR family member 4-1BB contribute to the expansion of anti-tumour CTL and, hence, survival following immunotherapy of lymphoma; indeed, we show that all tumour-specific CTL generated during anti-CD40-induced immunotherapy express this molecule. Furthermore, we demonstrate for the first time that the murine lymphomas used in these studies invoke an immunological response prior to anti-CD40 administration which is characterised by the phenotypic maturation of DCs and the differentiation of CD8<sup>*</sup> T-cells, as well as the accumulation of these cells at the site of tumour. Importantly, the kinetics and magnitude of these responses are augmented by administration of anti-CD40 thereby suggesting that this mAb exerts its therapeutic activity by the boosting of a pre-existing ineffectual adaptive immune response. Data shown here suggest that enhanced DC maturation following administration of anti-CD40 to tumour-bearing animals may contribute to this effect and, hence, support the hypothesis that anti-CD40 operates at the axis of the professional APC in order to affect the rejection of both CD40-positive and CD40-negative tumours.
In addition, we show that both CD8<sup>*</sup> T-cell lines and immortalised hybridomas derived from tumour-reactive CD8<sup>*</sup> CTL can be established in vitro from animals in remission following anti-CD40-induced immunotherapy. These techniques are new to this laboratory and provide a basis for the screening of a lymphoma-derived cDNA library in order to identify the tumour rejection antigens operating in this system.</p
... Any process that increases the presentation of autoimmune epitopes has the potential to break this tolerance. Once tolerance of this nature has been broken it may be difficult to re-establish since memory T-cells with lower stimulation thresholds will have been generated (Hurst et al., 1999;Saparov et al., 1999;Bachmann et al., 1999;London et al., 2000). Presentation of endocytosed material might also break functional tolerance because the endocytic pathway may process self-proteins differently to the endogenous antigen pathway, thereby revealing cryptic epitopes. ...
Human immunodeficiency virus (HIV) infects about 40 million people worldwide. HIV is the causative agent of acquired immunodeficiency syndrome (AIDS). AIDS is characterised by a progressive decline in protective immunity that leads to opportunistic infection and eventually death. Although HIV-1 causes a decline in CD4' T-cell number and this undoubtedly contributes to the general immune deficit of AIDS, CD4+ T-cell loss does not completely explain the pathogenesis of AIDS. Death and anergy of uninfected T-cells is observed in AIDS, as are deficits in innate and specific immunity. Antigen presenting cells(APCs, including macrophages and dendritic cells, DCs, which are infectable by M-tropic strains of HIV-1 via the CCR-5 chemokine receptor) play a key role in orchestrating innate and adaptive immune responses and controlling T-cell activities including activation, anergy, deletion, tolerisation and memory by the provision of appropriate signals. APC dysregulation results in deficits of innate and adaptive immune responses. It is known that HIV-1 can cause APC dysregulation; this thesis examines some mechanisms by which this might occur. The HIV-1 envelope glycoprotein gpl20 mediates HIV-1 infection by binding to target cells via CD4 and CCR-5 and is focussed on throughout this work. Because gpl20 is found on the surface of HIV-1 and dissolved in the serum of HIV-1 infected patients, it has the ability to disrupt the function of both infected and uninfected APCs. Data in this thesis demonstrate that gp 120 causes a decline of cell-surface CD4 from human macrophages in vitro. A mechanism for this loss is proposed based on observations that it is significantly more substantial when CCR-5-binding gpl20, derived from M-tropicHIV-1 is used as opposed to CXCR-4-binding gpl20. CD4 loss is absent from macrophages that fail to express surface CCR-5 due to homozygosity for the naturally occurring ccr5A32mutation. It appears that CD4 loss by this novel CCR-5-dependent mechanism requires cross-linking of CCR-5, CD4 and gp 120 at the cell surface leading to receptor-mediated endocytosis of this protein complex. Confocal microscopy was used to visualise these endocytosed proteins inside macrophages and RT-PCR was used to investigate transcriptional regulation of CD4 and CCR-5 recovery. Endocytosis of the protein complex may change antigen presentation efficiencies. Possible implications for protective- and auto¬immunity are discussed. This thesis also presents evidence that pre-treatment with gp!20 leads to reduction in an APC's ability to stimulate antigen-specific proliferation of a T-cell line. Because this effect is not dependent on the tropism of the HIV-1 strain from which the gpl20 is derived, an alternative mechanism to CD4-loss was sought. The hypothesis that APC dysfunction is due to HIV-1 subversion of physiological mechanisms involving prostaglandin and the Notch signalling pathway, leading to inappropriate tolerance induction, was examined. Treatment of macrophages and DCs with gpl20 caused the transcriptional up-regulation of genes involved in the Notch pathway including Notch ligands, the presence of which on an APC has previously been shown to abrogate T-cell activation by the induction of an anergic phenotype. Preventing HIV-1 infection of APCs and the subsequent dysregulation of immune responses is a therapeutic goal. Branched, synthetic peptides based on discontinuous epitopes of gpl20and previously demonstrated to disrupt binding to CD4 and CCR-5 are shown to protect macrophages from infection with M-tropic HIV-1Bal- Possible refinements to peptide structure and their utility as anti-HIV-1 therapeutics or vaccines are discussed
(PDF) HIV-1 interactions with Antigen Presenting Cells. Available from: https://www.researchgate.net/publication/358769965_HIV-1_interactions_with_Antigen_Presenting_Cells#fullTextFileContent [accessed May 04 2023].
... CD4 + T M cells expand after antigen exposure and begin to make cytokines (which direct immune cell function), provide help in the B cell and CD8 + T cell responses, and directly exert effector functions (39,40). A major characteristic of CD4 + T M cells is their ability to respond to lower doses of antigen and/or to reduced levels of co-stimulation compared to naive CD4 + T cells (41,42). This enhanced antigenic sensitivity has been associated with the organization of the TCR into nanoclusters (5,6) and to the onset of a bioenergetic program in which OXPHOS dominates over glycolysis (43). ...
The inhibition of anabolic pathways, such as aerobic glycolysis, is a metabolic cornerstone of memory T cell differentiation and function. However, the signals that hamper these anabolic pathways are not completely known. Recent evidence pinpoints the chemokine receptor CCR5 as an important player in CD4+ T cell memory responses by regulating T cell antigen receptor (TCR) nanoclustering in an antigen-independent manner. This paper reports that CCR5 specifically restrains aerobic glycolysis in memory-like CD4+ T cells, but not in effector CD4+ T cells. CCR5-deficient memory CD4+ T cells thus show an abnormally high glycolytic/oxidative metabolism ratio. No CCR5-dependent change in glucose uptake nor in the expression of the main glucose transporters was detected in any of the examined cell types, although CCR5-deficient memory cells did show increased expression of the hexokinase 2 and pyruvate kinase M2 isoforms, plus the concomitant downregulation of Bcl-6, a transcriptional repressor of these key glycolytic enzymes. Further, the TCR nanoclustering defects observed in CCR5-deficient antigen-experienced CD4+ T cells were partially reversed by incubation with 2-deoxyglucose (2-DG), suggesting a link between inhibition of the glycolytic pathway and TCR nanoscopic organization. Indeed, the treatment of CCR5-deficient lymphoblasts with 2-DG enhanced IL-2 production after antigen re-stimulation. These results identify CCR5 as an important regulator of the metabolic fitness of memory CD4+ T cells, and reveal an unexpected link between T cell metabolism and TCR organization with potential influence on the response of memory T cells upon antigen re-encounter.
... The distinguishing attribute of this approach is the possibility of developing vaccines within the patients themselves such that the resource allocation requirement is minimized. To evoke a resilient anti-tumor memory immune response, a paragon in situ cancer vaccine should typically be capable of inducing immunogenic cancer cell apoptosis, facilitating the TAA release, enhancing uptake of antigen by APCs followed by their activation to effectuate tumor-directed T-cell responses that in turn engender systemic anti-tumor immunity [133][134][135]. Oncolytic virotherapy (OV) is progressively being recognized as a form of immunotherapy. ...
The tumor microenvironment is best described as the battleground for fighting cancer and the host immune system. Contrary to primitive beliefs that cancer growth and progression are solely dependent on cancer cells, recent studies suggest that the cancer cell mass environs also play a pivotal role. Active crosstalk between the tumor cells and their surrounding microenvironment permits their collusion to effectuate high cancer cell proliferation and metastasis. Tumors have been reported to actively recruit and alter immune cells' phenotypes and functions to either promote immune suppression or increase the tolerance towards tumor-associated antigens. Comprehending the part played by the tumor microenvironment in tumor progression and its mechanism of action paves the way for developing novel therapeutic approaches for a more personalized and efficient tumor microenvironment targeted anticancer treatment. This review elaborates on the nature and importance of the tumor microenvironment and the anti-cancer therapeutic strategies designed to target them. Furthermore, we discuss in-depth the employment of oncolytic viruses as nanomedicines for tumor microenvironment targeted anticancer therapy. This review also delineates the benefits of combining novel therapeutic approaches to existing treatment strategies to improve disease prognosis.
... La efficace lors de la rencontre ultérieure avec l'antigène (London, Lodge, et Abbas 2000). ...
Les réactivations à Polyomavirus, BK-virus (BKv) et JC-virus (JCv), sont des complications majeures en transplantation rénale, responsables de néphropathie à BKv (Nx BKv) et de leuco-encéphalopathie multifocale progressive (LEMP). Sans thérapeutique antivirale spécifique, ces infections virales menent à la perte du rein transplanté ou au décès du patient. Notre groupe a conduit une étude observationnelle incluant 100 patients transplantés rénaux avec différents niveaux de réactivation BKv (Etude MelTyK). Nous avons mis en évidence une altération progressive de la fonctionnalité des lymphocytes T spécifiques du BKv, associée à une corrélation inverse entre la polyfonctionnalité lymphocytaire ou le nombre d’incompatibilités HLA et la charge virale BKv plasmatique. Cette altération de la fonctionnalité suggérait un état d’épuisement des lymphocytes T spécifiques du BKv en fonction du niveau de réactivation BKv. Ces données nous ont conduit à élaborer une méthode biologique non-invasive d’évaluation du risque individuel de Nx BKv (brevet FR1855342). Cette méthode a pour objectif d’aider au diagnostic de Nx BKv sans avoir recours à la biopsie du greffon rénal et de stratifier le risque de développer cette complication. Par ailleurs, nous avons décrit un cas fatal de LEMP associé à un état d’anergie des lymphocytes T spécifiques du JCv. L’étude de la fonctionnalité des lymphocytes T spécifiques des Polyomavirus pourrait ouvrir de futures pistes diagnostiques et/ou thérapeutiques. Elle pourrait permettre de dépister les patients à risques de Nx BKv et pourrait contribuer au développement d’immunothérapies innovantes. La restauration de la fonctionalité des lymphocytes T spécifiques des Polyomavirus pourrait ainsi fournir une piste thérapeutique prometteuse afin de contrôler ces réactivations virales sans majorer le risque de rejet allogénique.
... There is an ongoing debate as to how naive and memory cells differ in their requirements for CD28 costimulation, and a prevalent view is that CD28 costimulation is less required for activation of memory than naive T cells [47][48][49][50]. More recently, studies have begun to question this conclusion and have shown CD28 impacts on memory T cells responses in mice [51,52]. ...
T-cell activation is a critical driver of immune responses. The CD28 costimulation is an essential regulator of CD4 T-cell responses, however, its relative importance in naive and memory T cells is not fully understood. Using different model systems, we observe that human memory T cells are more sensitive to CD28 costimulation than naive T cells. To deconvolute how the T-cell receptor (TCR) and CD28 orchestrate activation of human T cells, we stimulate cells using varying intensities of TCR and CD28 and profiled gene expression. We show that genes involved in cell cycle progression and division are CD28-driven in memory cells, but under TCR control in naive cells. We further demonstrate that T-helper differentiation and cytokine expression are controlled by CD28. Using chromatin accessibility profiling, we observe that AP1 transcriptional regulation is enriched when both TCR and CD28 are engaged, whereas open chromatin near CD28- sensitive genes is enriched for NF-kB motifs. Lastly, we show that CD28-sensitive genes are enriched in GWAS regions associated with immune diseases, implicating a role for CD28 in disease development. Our study provides important insights into the differential role of costimulation in naive and memory T-cell responses and disease susceptibility.
... Based on pilot studies in the field [33][34][35], it was generally accepted that one feature of immunological memory is that a memory T cell elicits a faster and stronger response to cognate antigens than a naïve T cell. However, recent evidence showed that, at least under certain conditions, the response of naïve T cells to an antigen is stronger than the response of memory T cells [36][37][38]. ...
Virtual memory T cells are foreign antigen-inexperienced T cells that have acquired memory-like phenotype and constitute for 10-20% of all peripheral CD8 ⁺ T cells in mice. Their origin, biological roles, and relationship to naïve and foreign antigen-experienced memory T cells are incompletely understood. By analyzing TCR repertoires and using retrogenic monoclonal T-cell populations, we show that virtual memory T cells originate exclusively from strongly self-reactive T cells. Moreover, we show that the stoichiometry of the CD8 interaction with Lck regulates the size of the virtual memory T-cell compartment via modulating the self-reactivity of individual T-cell clones. We propose a so far unappreciated peripheral T-cell fate decision checkpoint that eventually leads to the differentiation of highly self-reactive T cells into virtual memory T cells. This underlines the importance of the variable level of self-reactivity in polyclonal T cells for the generation of functional T-cell diversity. Although virtual memory T cells descend from the highly self-reactive clones and acquire a partial memory program, they do not show higher capacity to induce autoimmune diabetes than naïve T cells. Thus, virtual memory T cells are not generally more responsive than naïve T cells, because their activity highly depends on the immunological context.
Summary
We conclude that virtual memory T cells are formed from self-reactive CD8 ⁺ T cells in a process regulated by CD8-Lck stoichiometry. Despite their self-reactivity and partial memory differentiation program, virtual memory T cells did not show a strong autoimmune potential.
... While most analyses on miRNA expression in T cells are focusing on specific time points, only a few longitudinal studies analyzed a time window between one and several days after T cell activation (11). During the initial 24 h of T cell activation the cells undergo the transition from the resting to the proliferative stage, accompanied by pivotal changes of signaling pathways (18)(19)(20)(21). MiRNA expression profiles within the initial 24 h of T cell activation are, however, rarely described and limited to the analysis of individual time points (11,22). ...
T cells are central to the immune response against various pathogens and cancer cells. Complex networks of transcriptional and post-transcriptional regulators, including microRNAs (miRNAs), coordinate the T cell activation process. Available miRNA datasets, however, do not sufficiently dissolve the dynamic changes of miRNA controlled networks upon T cell activation. Here, we established a quantitative and time-resolved expression pattern for the entire miRNome over a period of 24 h upon human T-cell activation. Based on our time-resolved datasets, we identified central miRNAs and specified common miRNA expression profiles. We found the most prominent quantitative expression changes for miR-155-5p with a range from initially 40 molecules/cell to 1600 molecules/cell upon T-cell activation. We established a comprehensive dynamic regulatory network of both the up- and downstream regulation of miR-155. Upstream, we highlight IRF4 and its complexes with SPI1 and BATF as central for the transcriptional regulation of miR-155. Downstream of miR-155-5p, we verified 17 of its target genes by the time-resolved data recorded after T cell activation. Our data provide comprehensive insights into the range of stimulus induced miRNA abundance changes and lay the ground to identify efficient points of intervention for modifying the T cell response.
... The use of DNA microarrays showed that the expression of approximately 200 cDNA clones changed following T cell activation, these are expected to affect the different responses and properties of naïve, effector and memory CD4^ cells (Liu 2001). The enhanced response by memory cells is because the threshold of detection is lower for memory than naïve cells, and that memory cells have a decreased dependency on CD28 mediated costimulation (London 2000, Fujii 1992). ...
Responses of T cells to antigen depend on recognition of MHC-peptide complexes by cells with specific clonal T cell receptors. There is much data on the properties of individual clones responding to particular peptide-MHC complexes of infectious agents and also data analysing the TCR sequence variability of T cells responding to single peptide-MHC complexes. However, there is less data on global responses and the size and number of clones. Most studies to date have focussed on CD8+ T cells where large expanded clones have been detected in a number of cell populations, helping to define clonal responses to a range of infections. In contrast there is limited data in CD4+ cells where clones are much smaller and are present at a lower frequency. The data presented in this thesis examines the clonality of CD4+ T cells at a global level through heteroduplex analysis. Initial attempts to establish this technique for examination of clonality in mice failed; however the technique is proven in human studies. Human CD4+ cells were subdivided by their expression of CD45, CD27 and CD95 to determine which phenotypic markers distinguish naive, memory and effector cells. Phenotypic and clonal analysis have allowed a model of differentiation to be proposed suggesting that loss of CD27 and acquisition of CD95 defines antigen experienced cell populations. Data also suggests that a population of CD45RA+CD27' CD95+ cells might be a revertant population. The clonality of CD4+CD45R0+CD25+ regulatory cells has also been examined. There was a large overlap in clonality between CD25+ and CD25' cells suggesting that regulatory T cells may develop in the periphery from antigen experienced cells. Studies in humans are mostly performed on peripheral blood samples due to ethical and practical constraints of sampling tissues. The data presented here examines the overlap of clonality of cells in blood, secondary lymphoid tissues and non-lymphoid effector tissues, and shows that there is a high degree of overlap in clonality between blood and tissues. Collectively, these data provide further knowledge of the clonal responses of CD4+ cells.
... The strength of T EFF/MEM responses in both lymphoid and nonlymphoid tissues is a barrier to transplant tolerance. 45,46 The presence and number of alloantigen-specific T EFF/MEM cells in humans correlates with worse transplant outcomes. 47 Despite current immunosuppressive regimens capable of non-specific T cell depletion, certain T EFF/MEM subsets have been found to survive treatment with the commonly used depleting induction agents, Campath (anti-CD52 mAb) or anti-thymocyte globulin. ...
Organ transplantation is often lifesaving, but the long‐term deleterious effects of combinatorial immunosuppression regimens and allograft failure cause significant morbidity and mortality. Long‐term graft survival in the absence of continuing immunosuppression, defined as operational tolerance, has never been described in the context of multiple major histocompatibility complex (MHC) mismatches.
Here, we show that miR‐142 deficiency leads to indefinite allograft survival in a fully MHC mismatched murine cardiac transplant model in the absence of exogenous immunosuppression. We demonstrate that the cause of indefinite allograft survival in the absence of miR‐142 maps specifically to the T cell compartment. Of therapeutic relevance, temporal deletion of miR‐142 in adult mice prior to transplantation of a fully MHC mismatched skin allograft resulted in prolonged allograft survival. Mechanistically, miR‐142 directly targets Tgfbr1 for repression in regulatory T cells (TREG). This leads to increased TREG sensitivity to TGF‐β and promotes transplant tolerance via an augmented peripheral TREG response in the absence of miR‐142. These data identify manipulation of miR‐142 as a promising approach for the induction of tolerance in human transplantation.
... 47 Memory T cell responses usually do not require CD28-and CD40L-mediated costimulation; therefore, therapeutic strategies targeting these pathways are conventionally thought to be ineffective in sensitized recipients. 10,11,48,49 However, newer studies endorsing a beneficial effect of costimulation blockade have renewed interest in this approach in sensitized recipients. 14,50 In these studies, costimulation blockade-based regimen disrupts GC follicular T helper and B cell interaction, prevents memory B cell expansion and plasma cell generation, and prevents antibody-mediated rejection in allosensitized nonhuman primates. ...
Allo‐sensitization constitutes a major barrier in transplantation. Pre‐existing donor‐reactive memory T and B cells, and pre‐formed donor‐specific antibodies (DSAs) have all been implicated in accelerated allograft rejection in sensitized recipients. Here, we employ a sensitized murine model of islet transplantation to test strategies that promote long‐term immunosuppression‐free allograft survival. We demonstrate that donor‐specific memory T and B cells can be effectively inhibited by peri‐transplant infusions of donor apoptotic cells in combination with anti‐CD40L and rapamycin, and this treatment leads to significant prolongation of islet allograft survival in allo‐sensitized recipients. We further demonstrate that late graft rejection in recipients treated with this regimen is associated with a breakthrough of B cells and their aggressive graft infiltration. Consequently, additional post‐transplant B cell depletion effectively prevents late rejection and promotes permanent acceptance of islet allografts. In contrast, persistent low levels of DSAs do not seem to impair graft outcome in these recipients. We propose that B cells contribute to late rejection as antigen presenting cells for intragraft memory T cell expansion but not to alloantibody production, and that a therapeutic strategy combining donor apoptotic cells, anti‐CD40L and rapamycin effectively inhibits proinflammatory B cells and promotes long‐term islet allograft survival in such recipients.
... Naive MHC-restricted T cells are present at low frequency and require TCR ligation by their cognate antigen as well as additional stimulation by professional antigen presenting cells to become activated (56)(57)(58). These T cells will then undergo a rapid expansion, up to 25-fold in 72 h, and differentiate into effector T cells (59). ...
Diseases due to mycobacteria, including tuberculosis, leprosy, and Buruli ulcer, rank among the top causes of death and disability worldwide. Animal studies have revealed the importance of T cells in controlling these infections. However, the specific antigens recognized by T cells that confer protective immunity and their associated functions remain to be definitively established. T cells that respond to mycobacterial peptide antigens exhibit classical features of adaptive immunity and have been well-studied in humans and animal models. Recently, innate-like T cells that recognize lipid and metabolite antigens have also been implicated. Specifically, T cells that recognize mycobacterial glycolipid antigens (mycolipids) have been shown to confer protection to tuberculosis in animal models and share some biological characteristics with adaptive and innate-like T cells. Here, we review the existing data suggesting that mycolipid-specific T cells exist on a spectrum of “innateness,” which will influence how they can be leveraged to develop new diagnostics and vaccines for mycobacterial diseases.
... This contrasts markedly with older children and adults, in which there are preexisting populations of influenza-specific memory cells that can target conserved epitopes (Kreijtz et al. 2007;Lee et al. 2008;Koutsakos et al. 2019) and provide nonsterilizing immunity on reinfection (McMichael et al. 1983;Wilkinson et al. 2012;Sridhar et al. 2013). Because the responding memory cells have a significant advantage in terms of both precursor frequency and the threshold of activation required to induce a response (Croft et al. 1994;London et al. 2000;Rogers et al. 2000;von Essen et al. 2012), the induction of an adaptive immune response in young children is likely delayed. This may contribute to higher levels of viral replication and age-dependent differences in susceptibility to influenza virus infection. ...
Influenza poses a significant disease burden on children worldwide, with high rates of hospitalization and substantial morbidity and mortality. Although the clinical presentation of influenza in children has similarities to that seen in adults, there are unique aspects to how children present with infection that are important to recognize. In addition, children play a significant role in viral transmission within communities. Growing evidence supports the idea that early influenza infection can uniquely establish lasting immunologic memory, making an understanding of how viral immunity develops in this population critical to better protect children from infection and to facilitate efforts to develop a more universally protective influenza vaccine.
... Although memory cells primarily recognize pathogenrelated antigens, it has been demonstrated that cross-reactive memory cells can recognize alloantigens in a process called heterologous immunity, and therefore assume a critical relevance not only in donor-sensitized recipients (Adams, Williams et al. 2003). Memory T cells are characterized by a lower activation threshold, are less dependent on costimulation and are not efficiently inhibited by regulatory T cells (Lanzavecchia and Sallusto 2000;London, Lodge et al. 2000;Yang, Brook et al. 2007). As a result, the efficacy of costimulation blockers is limited in the context of a secondary immune response. ...
Die beste therapeutische Option für Patienten mit Organversagen ist häufig eine Transplantation. Um die immunologische Abstossung eines Organs aus einem genetisch nicht-identischen Spender zu verhindern, müssen transplantierte Patienten lebenslang eine immunsuppressive Therapie einnehmen. Derzeit verfügbare immunsuppressive Medikamente sind sehr wirksam gegen die akute Abstossung aber weniger effizient in der Prävention der chronischen Abstossung, und verursachen zum Teil schwerwiegende Nebenwirkungen. Neue Strategien zur Abstossungsprävention sind die Voraussetzung für eine verbesserte Lebenserwartung und Lebensqualität nach Organtransplantation. Die beste Lösung wäre die Induktion von immunologischer Toleranz, ein Zustand, in welchem das Immunsystem das Transplantat ohne Immunsuppression akzeptiert, aber auf pathogene Keime reagiert; verschiedene ungelöste Probleme verhindern aber eine breite klinische Anwendung von diesem Prinzip. Die Apoptose, eine Form von vorprogrammiertem Zelltod mit einer entscheidenden Funktion für die Gewebe-Homöostase in multizellulären Organismen, reguliert das Immunsystem und spielt eine kritische Rolle in der Erhaltung der Selbst-Toleranz. In dieser Studie untersuche ich die potentielle Rolle einer pharmakologischen Modulation der Apoptose als eine neue Strategie zur Abstossungsprävention und Toleranz-Induktion. Kapitel 1 ist eine Einführung zur Transplantationsimmunologie und zu den klinischen und experimentellen Optionen für die Abstossungsprävention. Kapitel 2 erklärt die Prinzipien der Apoptose, deren Rolle in der Regulation des Immunsystems und die Optionen für eine pharmakologische Modulation der Apoptose. Im Kapitel 3 werden die Resultate der Experimente über die immunsuppressive Wirkung der pro-apoptotischen Substanz ABT-737 in vitro und in einem Haut-Transplantationsmodell in der Maus vorgestellt. Die Wirkung von ABT-737 in vivo war begrenzt, aber verstärkt in Kombination mit dem Calcineurin Hemmer Cyclosporin A. Im Kapitel 4 werden der Synergismus mit Cyclosporin A und die immunomodulatorische Wirkung von ABT-737 weiter untersucht. Diese mechanistischen Studien waren die Basis für die Entwicklung eines neuen Protokolls zur Toleranzinduktion, wie beschrieben im Kapitel 5. Die Wirkung von ABT-737 auf Gedächtnis-Lymphozyten, ein wichtiges Hindernis für Toleranz in der Klinik wird im Kapitel 6 untersucht. Kapitel 7 fasst die immunmodulatorische Wirkung von ABT-737 und anderen pro-apoptotischen Substanzen zusammen. Im Kapitel 8 diskutiere ich die mögliche Applikation von diesem neuen Prinzip im Bereich der Transplantation. Transplantation is often the best therapeutic option for patients with end-stage organ failure. However, organ transplantation from a genetically not identical individual inevitably leads to immunological graft rejection, which must be prevented by lifelong immunosuppressive therapy. Currently available immunosuppressive drugs effectively prevent acute allograft rejection, but are less effective in the long-term and induce major side effects. Therefore, new strategies are required to improve survival and quality of life after transplantation. Induction of donor-specific tolerance, a state in which the immune system accepts the transplanted organ without immunosuppression and normally reacts to pathogens, would represent the ideal solution to this problem. However, several obstacles preclude the establishment of tolerance induction protocols clinical practice. Apoptosis, a form of programmed cell death with a fundamental role in the maintenance of tissue homeostasis in multicellular organisms, is critically involved in the regulation of the immune system and in the maintenance of self- tolerance. In this project, I investigated the potential role of a pharmacological modulation of the apoptosis pathway as a new strategy to prevent rejection and to induce tolerance after solid organ transplantation. Chapter 1 provides an introduction to transplantation immunology and to clinical and experimental approaches to prevent allograft rejection. Chapter 2 deals with apoptosis, its role as a regulator of the immune system and the available options to pharmacologically modulate it. Chapter 3 describes the immunosuppressive effect of the pro-apoptotic drug ABT-737 in vitro and in a skin transplantation model. The immunosuppressive effect of ABT-737 as a single agent was limited, but markedly increased in combination with the calcineurin inhibitor cyclosporine A. Chapter 4 analyzes more in details the synergistic effect of these two drugs and characterizes the immunomodulatory properties of ABT-737. The information obtained from these mechanistic studies was the basis for the development of a new therapeutic approach to induce immunological tolerance targeting the apoptosis pathway, as described in chapter 5. Finally, the effect of ABT-737 on memory lymphocytes, a major barrier to tolerance induction in humans, is described in chapter 6. In chapter 7 I give an overview about the immuno-modulatory properties of ABT-737 and other pro-apoptotic drugs. Chapter 8 discusses the potential role of these agents in transplantation.
... Studies utilizing human IEC lines (T84 and HT29) show that IFNγ-treated, protein antigen-pulsed IECs can stimulate antigenspecific immune responses in T cell hybridomas (107). T cell hybridomas do not need co-stimulation, which arguably mimics the reduced costimulatory requirements of the majority of T cells in the lamina propria, which are antigen-experienced memory cells (108,109). Follow-up work in IECs found that generation of specific MHC II-restricted peptide epitopes differed if antigen was taken up from the apical or basolateral IEC surface (78). During disease, inflammatory signals including IFNγ and TNFα in the gut increase epithelial permeability (Figure 2) (110)(111)(112). ...
As the primary barrier between an organism and its environment, epithelial cells are well-positioned to regulate tolerance while preserving immunity against pathogens. Class II major histocompatibility complex molecules (MHC class II) are highly expressed on the surface of epithelial cells (ECs) in both the lung and intestine, although the functional consequences of this expression are not fully understood. Here, we summarize current information regarding the interactions that regulate the expression of EC MHC class II in health and disease. We then evaluate the potential role of EC as non-professional antigen presenting cells. Finally, we explore future areas of study and the potential contribution of epithelial surfaces to gut-lung crosstalk.
... Generation of antibody after vaccination depends on follicular helper T cells (Tfh), a subset of CD4 + T cells that are specialized for providing help to B cells to support class switch recombination, affinity maturation, and B cell differentiation into long-lived plasma cells and memory cells in germinal centers (GC) in secondary lymphoid tissue. 38,39 A signature Tfh marker, CXCR5, is upregulated on naïve CD4 + T cells after antigen stimulation, permitting localization to the B cell zones of lymphoid tissue. 40,41 Mature Tfh cells expressing PD-1, ICOS and Bcl-6 provide signals for B cell survival and differentiation via CXCL13, IL-4 and IL-21 production as well as CD40L expression. ...
The delayed availability of vaccine during the 2009 H1N1 influenza pandemic created a sense of urgency to better prepare for the next influenza pandemic. Advancements in manufacturing technology, speed and capacity have been achieved but vaccine effectiveness remains a significant challenge. Here, we describe a novel vaccine design strategy called immune engineering in the context of H7N9 influenza vaccine development. The approach combines immunoinformatic and structure modeling methods to promote protective antibody responses against H7N9 hemagglutinin (HA) by engineering whole antigens to carry seasonal influenza HA memory CD4⁺ T cell epitopes – without perturbing native antigen structure – by galvanizing HA-specific memory helper T cells that support sustained antibody development against the native target HA. The premise for this vaccine concept rests on (i) the significance of CD4⁺ T cell memory to influenza immunity, (ii) the essential role CD4⁺ T cells play in development of neutralizing antibodies, (iii) linked specificity of hemagglutinin (HA)-derived CD4⁺ T cell epitopes to antibody responses, (iv) the structural plasticity of HA and (v) an illustration of improved antibody response to a prototype engineered recombinant H7-HA vaccine. Immune engineering can be applied to development of vaccines against pandemic concerns, including avian influenza, as well as other difficult targets.
... Their longevity and tissue localization allows T M cells to maintain effective immunity against bacteria, protozoa and viruses [1][2][3] . A key characteristic of both CD8 + and CD4 + T M cells is their capacity to produce substantial amounts of effector molecules within a few hours upon reinfection [4][5][6] . This rapid responsiveness limits pathogen spread and recruits innate immune cells to the site of infection 7,8 . ...
Memory T cells are critical for the immune response to recurring infections. Their instantaneous reactivity to pathogens is empowered by the persistent expression of cytokine-encoding mRNAs. How the translation of proteins from pre-formed cytokine-encoding mRNAs is prevented in the absence of infection has remained unclear. Here we found that protein production in memory T cells was blocked via a 3' untranslated region (3' UTR)-mediated process. Germline deletion of AU-rich elements (AREs) in the Ifng-3' UTR led to chronic cytokine production in memory T cells. This aberrant protein production did not result from increased expression and/or half-life of the mRNA. Instead, AREs blocked the recruitment of cytokine-encoding mRNA to ribosomes; this block depended on the ARE-binding protein ZFP36L2. Thus, AREs mediate repression of translation in mouse and human memory T cells by preventing undesirable protein production from pre-formed cytokine-encoding mRNAs in the absence of infection.
... We cannot exclude the possibility that VM T cells represent a major risk in other types of autoimmune diseases/conditions. Based on pilot studies in the field (Pihlgren et al, 1996;Curtsinger et al, 1998;London et al, 2000), it was generally accepted that one feature of immunological memory is that a memory T cell elicits a faster and stronger response to cognate antigens than a naïve T cell. However, recent evidence showed that, at least under certain conditions, the response of naïve T cells to an antigen is stronger than the response of memory T cells (Knudson et al, 2013;Mehlhop-Williams & Bevan, 2014;Cho et al, 2016). ...
Virtual memory T cells are foreign antigen-inexperienced T cells that have acquired memory-like phenotype and constitute 10-20% of all peripheral CD8+ T cells in mice. Their origin, biological roles, and relationship to naïve and foreign antigen-experienced memory T cells are incompletely understood. By analyzing T-cell receptor repertoires and using retrogenic monoclonal T-cell populations, we demonstrate that the virtual memory T-cell formation is a so far unappreciated cell fate decision checkpoint. We describe two molecular mechanisms driving the formation of virtual memory T cells. First, virtual memory T cells originate exclusively from strongly self-reactive T cells. Second, the stoichiometry of the CD8 interaction with Lck regulates the size of the virtual memory T-cell compartment via modulating the self-reactivity of individual T cells. Although virtual memory T cells descend from the highly self-reactive clones and acquire a partial memory program, they are not more potent in inducing experimental autoimmune diabetes than naïve T cells. These data underline the importance of the variable level of self-reactivity in polyclonal T cells for the generation of functional T-cell diversity.
... T cells expressing ICOS are phenotypically resting or T EM cells, 9,28,31,39 and these ICOS + T memory cells can undergo rapid expansion independent of CD28/B7 or CD40/CD154 ligation. 44 As predicted from ICOS-deficient human and mice, 28,39 in this study ICOS-Ig treatment demonstrated a decrease of CD4 + T EM cells in peripheral blood and mLNs during treatment. In contrast to rodent studies, 14,15 the decrease of T EM cells in our study did not affect allograft survival or acute or chronic rejection after transplantation, which may be due to intact cytotoxic CD8 + T cell responses of the . ...
Background. Inducible costimulator (ICOS) is rapidly upregulated with T-cell stimulation and may represent an escape pathway for T-cell costimulation in the setting of CD40/CD154 costimulation blockade. Induction treatment exhibited no efficacy in a primate renal allograft model, but rodent transplant models suggest that the addition of delayed ICOS/ICOS-L blockade may prolong allograft survival and prevent chronic rejection. Here, we ask whether ICOS-Ig treatment, timed to anticipate ICOS upregulation, prolongs NHP cardiac allograft survival or attenuates pathogenic alloimmunity.
Methods. Cynomolgus monkey heterotopic cardiac allograft recipients were treated with αCD40 (2C10R4, d0-90) either alone or with the addition of delayed ICOS-Ig (d63-110).
Results. Median allograft survival was similar between ICOS-Ig + αCD40 (120 days, 120-125 days) and αCD40 (124 days, 89-178 days) treated animals, and delayed ICOS-Ig treatment did not prevent allograft rejection in animals with complete CD40 receptor coverage. Although CD4+ TEM cells were decreased in peripheral blood (115 ± 24) and mLNs (49 ± 1.9%) during ICOS-Ig treatment compared with monotherapy (214 ± 27%, P = 0.01; 72 ± 9.9%, P = 0.01, respectively), acute and chronic rejection scores and kinetics of alloAb elaboration were similar between groups.
Conclusions. Delayed ICOS-Ig treatment with the reagent tested is probably ineffective in modulating pathogenic primate alloimmunity in this model.
... Memory T cells are different from naive T cells because they are longlasting cells, are antigenindependent persistent, and are capable of selfrenewal [183] . Further more, they are able to be activated more easily than naïve T cells because they are less dependent on TCR stimulation and on costimulatory molecules [184] . These cells can be CD4 + or CD8 + cells, with the CD8 + subtype much more frequent and commonly studied [185] . ...
The evolutionary emergence of an efficient immune system has a fundamental role in our survival against pathogenic attacks. Nevertheless, this same protective mechanism may also establish a negative consequence in the setting of disorders such as autoimmunity and transplant rejection. In light of the latter, although research has long uncovered main concepts of allogeneic recognition, immune rejection is still the main obstacle to long-term graft survival. Therefore, in order to define effective therapies that prolong graft viability, it is essential that we understand the underlying mediators and mechanisms that participate in transplant rejection. This multifaceted process is characterized by diverse cellular and humoral participants with innate and adaptive functions that can determine the type of rejection or promote graft acceptance. Although a number of mediators of graft recognition have been described in traditional immunology, recent studies indicate that defining rigid roles for certain immune cells and factors may be more complicated than originally conceived. Current research has also targeted specific cells and drugs that regulate immune activation and induce tolerance. This review will give a broad view of the most recent understanding of the allogeneic inflammatory/tolerogenic response and current insights into cellular and drug therapies that modulate immune activation that may prove to be useful in the induction of tolerance in the clinical setting.
... Once these genes have been primed in activated T cells, they remain primed when T cells revert to the quiescent state as memory T cells [64,70]. Memory T cells have the ability to re-activate genes faster, and in response to weaker stimuli than naïve T cells [58,64,[71][72][73][74][75][76]. ...
During ontogeny, cells progress through multiple alternate differentiation states by activating distinct gene regulatory networks. In this review we highlight the important role of chromatin priming in facilitating gene activation during lineage specification and in maintaining an epigenetic memory of previous gene activation. We describe that chromatin priming is part of a hugely diverse repertoire of regulatory mechanisms that genes use to ensure that they are expressed at the correct time, in the correct cell type, at the correct level, but also that they react to signals. We also emphasize how increasing our knowledge of these principles could inform our understanding of developmental failure and disease.
... Further, protein activation and phosphorylation events ultimately lead to calcium mobilization and activation of MAPK signaling pathways. While it is debated whether the threshold of TCR activation by antigen is lower for memory compared to na€ ıve T cells[5,7,[16][17][18], numerous studies have demonstrated that the two cell types have differences in the levels of activated signaling molecules downstream of the TCR[16,17,19]. Kersh et al. showed that, before the TCR is activated, the micro-domains within the plasma membrane of memory cells contain a higher concentration of the phosphoproteins required for signal transduction, such as phospho-LAT, and are therefore, more efficient at activating downstream pathways[16]. ...
We have identified a simple epigenetic mechanism underlying the establishment and maintenance of immunological memory in T cells. By studying the transcriptional regulation of inducible genes we found that a single cycle of activation of inducible factors is sufficient to initiate stable binding of pre-existing transcription factors to thousands of newly activated distal regulatory elements within inducible genes. These events lead to the creation of islands of active chromatin encompassing nearby enhancers, thereby supporting the accelerated activation of inducible genes, without changing steady state levels of transcription in memory T cells. These studies also highlighted the need for more sophisticated definitions of gene regulatory elements. The chromatin priming elements defined here are distinct from classical enhancers because they function by maintaining chromatin accessibility rather than directly activating transcription. We propose that these priming elements are members of a wider class of genomic elements that support correct developmentally regulated gene expression.
... As they are programmed to rapidly expand and generate large effector populations upon antigen re-encounter, they pose a significant barrier to tissue, cell, and protein replacement therapies in "primed" individuals. Due to their terminal differentiation and costimulation independence, memory T cells have long been considered to be resistant to tolerance induction (29,30). However, pioneering studies including our own, where antigen expression has been enforced in APCs, have shown that memory CD4 + or CD8 + T-cell responses can be silenced (8)(9)(10)(31)(32)(33). ...
Islet-specific memory T cells arise early in type 1 diabetes (T1D), persist for long periods, perpetuate disease and are rapidly reactivated by islet transplantation. As memory T cells are poorly controlled by 'conventional' therapies, memory T-cell mediated attack is a substantial challenge in islet transplantation and this will extend to application of personalized approaches using stem-cell derived replacement β cells. New approaches are required to limit memory autoimmune attack of transplanted islets or replacement β cells. Here we show that transfer of bone marrow encoding cognate antigen directed to dendritic cells, under mild, immune-preserving conditions inactivates established memory CD8(+) T-cell populations and generates a long-lived, antigen-specific tolerogenic environment. Consequently, CD8(+) memory T cell-mediated targeting of islet-expressed antigens is prevented and islet graft rejection alleviated. The immunological mechanisms of protection are mediated through deletion and induction of unresponsiveness in targeted memory T-cell populations. The data demonstrate that hematopoietic stem cell-mediated gene therapy effectively terminates antigen-specific memory T-cell responses and this can alleviate destruction of antigen-expressing islets. This addresses a key challenge facing islet transplantation and importantly, the clinical application of personalized β-cell replacement therapies using patient-derived stem cells.
p>Identification of the signals required for optimal differentiation of naive T cells into effector and memory cells is critical for the design of effective vaccines for use in immunotherapy. This thesis describes the generation and characterisation of soluble recombinant ligands for two members of the TNFR superfamily of cell surface receptors, CD27 and CD137 (4-1BB). These receptors provide co-stimulatory signals, which act in concert with antigen-driven T cell receptor signals to promote optimal T cell activation. The recombinant fusion proteins, which encompassed the extracellular domains of murine CD27 ligand (CD70) or 4-1BB ligand (4-1BBL) and the Fc domain of human IgG1, formed large multimeric complexes that proved to have potent signalling capacity. Stimulation of CD27 by soluble recombinant CD70 was found to enhance both the magnitude and quality of CD8<sup>+</sup> T cell responses. Triggering CD27 in the presence of antigen significantly enhanced the division and survival of CD8<sup>+</sup> T cells in vitro, as well as their ability to produce interleukin-2 and interferon-γ and up regulate expression of 4-1BB and CD25 (interleukin-2 receptor α-chain). In an in vivo model of T cell activation, administration of peptide antigen and soluble CD70 resulted in a massive (> 300-fold) expansion of antigen-specific CD8<sup>+</sup> T cells, due to the enhanced division and survival promoted by CD27 co-stimulation. In mice that received antigen and soluble CD70, CD8<sup>+</sup> T cells developed into effectors with direct ex vivo cytotoxicity and the capacity to eradicate syngeneic tumour cells expressing antigen in vivo . Furthermore, unlike immunization with peptide antigen alone, which resulted in a diminished secondary response after rechallenge, CD27 co-stimulation during primary T cell activation led to a strong secondary response being evoked upon rechallenge with the antigenic peptide. Thus, in addition to increasing the frequency of primed antigen-specific T cells, CD27 signalling during the primary response instils a programme of differentiation that allows CD8<sup>+</sup> T cells to maintain their responsiveness. Ligation of 4-1BB using soluble 4-1BBL exhibited similar co-stimulatory effects to those of CD27 when administered with a peptide antigen, causing enhanced primary CD8<sup>+</sup> T cell expansion and affecting the quality of the T cell response so that a population of reactive T cells was maintained after the primary response. However, closer comparison of the effects of CD27 versus 4-1BB co-stimulation using their soluble ligands demonstrated that while CD27 predominantly enhanced the initial burst of T cell expansion, 4-1BB altered the kinetics of the primary response so that the activated T cells survived for longer and the contraction phase was slowed.</p
The majority of JAK2 V617F -negative myeloproliferative neoplasms (MPNs) have disease-initiating frameshift mutations in calreticulin ( CALR ), resulting in a common carboxyl-terminal mutant fragment (CALR MUT ), representing an attractive source of neoantigens for cancer vaccines. However, studies have shown that CALR MUT -specific T cells are rare in patients with CALR MUT MPN for unknown reasons. We examined class I major histocompatibility complex (MHC-I) allele frequencies in patients with CALR MUT MPN from two independent cohorts. We observed that MHC-I alleles that present CALR MUT neoepitopes with high affinity are underrepresented in patients with CALR MUT MPN. We speculated that this was due to an increased chance of immune-mediated tumor rejection by individuals expressing one of these MHC-I alleles such that the disease never clinically manifested. As a consequence of this MHC-I allele restriction, we reasoned that patients with CALR MUT MPN would not efficiently respond to a CALR MUT fragment cancer vaccine but would when immunized with a modified CALR MUT heteroclitic peptide vaccine approach. We found that heteroclitic CALR MUT peptides specifically designed for the MHC-I alleles of patients with CALR MUT MPN efficiently elicited a CALR MUT cross-reactive CD8 ⁺ T cell response in human peripheral blood samples but not to the matched weakly immunogenic CALR MUT native peptides. We corroborated this effect in vivo in mice and observed that C57BL/6J mice can mount a CD8 ⁺ T cell response to the CALR MUT fragment upon immunization with a CALR MUT heteroclitic, but not native, peptide. Together, our data emphasize the therapeutic potential of heteroclitic peptide–based cancer vaccines in patients with CALR MUT MPN.
The ability of the innate and adaptive immune systems to communicate with each other is central to protective immune responses and maintenance of host health. Myeloid cells of the innate immune system are able to sense microbial ligands, perturbations in cellular homeostasis, and virulence factors, thereby allowing them to relay distinct pathogen-specific information to naïve T cells in the form of pathogen-derived peptides and a unique cytokine milieu. Once primed, effector T helper cells produce lineage-defining cytokines to help combat the original pathogen, and a subset of these cells persist as memory or effector-memory populations. These memory T cells then play a dual role in host protection by not only responding rapidly to reinfection, but by also directly instructing myeloid cells to express licensing cytokines. This means there is a bi-directional flow of information first from the innate to the adaptive immune system, and then from the adaptive back to innate immune system. Here, we focus on how signals, first from pathogens and then from primed effector and memory T cells, are integrated by myeloid cells and its consequences for protective immunity or systemic inflammation.
Why do we still have no cure for chronic inflammatory diseases? One reason could be that current therapies are based on the assumption that chronic inflammation is driven by persistent 'acute' immune reactions. Here we discuss a paradigm shift by suggesting that beyond these reactions, chronic inflammation is driven by imprinted, pathogenic 'memory' cells of the immune system. This rationale is based on the observation that in patients with chronic inflammatory rheumatic diseases refractory to conventional immunosuppressive therapies, therapy-free remission can be achieved by resetting the immune system; that is, by ablating immune cells and regenerating the immune system from stem cells. The success of this approach identifies antigen-experienced and imprinted immune cells as essential and sufficient drivers of inflammation. The 'dark side' of immunological memory primarily involves memory plasma cells secreting pathogenic antibodies and memory T lymphocytes secreting pathogenic cytokines and chemokines, but can also involve cells of innate immunity. New therapeutic strategies should address the persistence of these memory cells. Selective targeting of pathogenic immune memory cells could be based on their specificity, which is challenging, or on their lifestyle, which differs from that of protective immune memory cells, in particular for pathogenic T lymphocytes. The adaptations of such pathogenic memory cells to chronic inflammation offers entirely new therapeutic options for their selective ablation and the regeneration of immunological tolerance.
Memory T cells respond rapidly in part because they are less reliant on heightened levels of costimulatory molecules. This presents challenges to silencing memory T cells in tolerance strategies for autoimmunity or allergy. We find that memory CD4 T cells generated by infection or immunisation survive secondary activation with antigen delivered without adjuvant, regardless of their location in secondary lymphoid organs or peripheral tissues. These cells were, however, functionally altered following a tertiary immunisation with antigen and adjuvant, proliferating poorly but maintaining their ability to produce inflammatory cytokines. Transcriptional and cell cycle analysis of these memory CD4 T cells suggest they are unable to commit fully to cell division potentially because of low expression of DNA repair enzymes. In contrast, these memory CD4 T cells could proliferate following tertiary reactivation by viral re-infection. These data suggest that tolerance induction in memory CD4 T cells is partial and can be reversed.
Immunotherapy is revolutionizing the treatment of cancer, and the current immunotherapeutics have remarkably improved the outcomes for some cancer patients. However, we still need answers for patients with immunologically cold tumors that do not benefit from the current immunotherapy treatments. Here, we suggest a novel strategy that is based on using a very old and sophisticated system for cancer immunotherapy, namely "intrinsic cancer vaccination", which seeks to awaken our own immune system to activate tumor-specific T cells. To do this, we must take advantage of the genetic instability of cancer cells and the expression of cancer cell neoantigens to trigger immunity against cancer cells. It will be necessary to not only enhance the phagocytosis of cancer cells by antigen presenting cells but also induce immunogenic cancer cell death and the subsequent immunogenic clearance, cross-priming and generation of tumor-specific T cells. This strategy will allow us to avoid using known tumor-specific antigens, ex vivo manipulation or adoptive cell therapy; rather, we will efficiently present cancer cell neoantigens to our immune system and propagate the cancer-immunity cycle. This strategy simply follows the natural cycle of cancer-immunity from its very first step, and therefore could be combined with any other treatment modality to yield enhanced efficacy.
The concept of disease interception can be realistically applied in practice based on the knowledge of the at‐risk individual coupled with reliable markers identifying the earliest stages of dysregulation preceding onset of disease. Several therapeutic options can be pursued; however, antigen‐specific approaches may provide an opportunity to reset or reestablish tolerance. The ultimate value for disease interception is the successful application at a population level as a part of the standard health‐care practice. In the near future, integration of multitude datasets and measurements should enable personalized risk profiles for dominant autoimmune diseases. To this end, several relevant datasets could be considered: genome sequencing coupled with computational approaches to understand transcriptional regulation; tracking immune repertoire diversity and specificity from genesis to pathogenesis; and host microbiome signatures and environmental perturbations ‐ including infections, lifestyle modifications, etc., and their impact on above parameters.
Immunologische Gedächtnisreaktionen sind die Grundlage um wiederkehrende Erreger schnell und effizient zu bekämpfen und um einen Impfschutz zu generieren. Das zellvermittelte Gedächtnis wird unter anderem durch CD8 Gedächtnis-T-Zellen aufgebaut, welche vor allem im Kontext von Immunreaktionen gegen intrazellulärer Erreger vonnöten sind, um bei Reinfektion mit den Erregerstämmen einen schnellen Schutz zu gewährleisten. Ein detailliertes Wissen über die Generierung, Kontrolle und Reaktivierung der Gedächtniszellen ist nützlich, um Gedächtnisreaktionen verstehen und lenken zu können. Durch die Entdeckung des TZR und CD28 wurden Meilensteine für das Verständnis der T-Zellaktivierung gelegt und die Grundlage geschaffen, CD8 Gedächtnisreaktionen zu verstehen. Auch wenn für primäre Immunreaktionen die „2-Signal-Theorie“ lange als erwiesen gilt, so blieb die Rolle der Kostimulation für Gedächtnisreaktionen lange umstritten. In dieser Arbeit wurden verschiedene methodische Herangehensweisen verwendet, mit denen durchgehend die Bedeutung von CD28 vermittelter Kostimulation für immunologische CD8 T-Zell-Gedächtnisreaktionen nachgewiesen wurde. CD28 blockierende Antikörper und CD28 induzierbar deletierbare Mauslinien wurden im Modellinfektionssystem mit Ovalbumin produzierenden Listeria monocytogenes zur Analyse der Primär- und Sekundärantworten verwendet. Mit diesen Methoden konnte eine Beeinträchtigung der Expansion von CD8 Gedächtniszellen in Abwesenheit von CD28 bewiesen werden. Weiterhin werden Effektorfunktionen wie Degranulation und Produktion von IFN-γ während der Sekundärinfektion in Abwesenheit von Kostimulation eingeschränkt. Mit Hilfe von Experimenten, bei denen CD28 suffizienten Mäusen eine geringe Anzahl an naiven, antigenspezifischen, CD28 deletierbaren CD8 T-Zellen transferiert wurden, wurde die Bedeutung der Kostimulation für die Expansion von Gedächtniszellen bestätigt, jedoch konnte überraschenderweise auch ein Anstieg der Effektorfunktionen in Abwesenheit von CD28 sowohl während der Primär- als auch der Sekundärantwort dokumentiert werden. Diese zur globalen Blockade bzw. Deletion widersprüchlichen Ergebnisse lassen eine Beteiligung anderer CD28 abhängiger Zelltypen an der Induktion der Effektorfunktionen der CD8 T-Zellen plausibel erscheinen, wie zum Beispiel Einflüsse von T-Helferzellen, welche die Effektorfunktionen positiv verstärken, solange sie selbst Kostimulationssignale empfangen können. Weiterhin konnte gezeigt werden, dass sich Gedächtniszellen an den CD28 defizienten Phänotyp – eine CD28 intakte immunologische Umgebung vorausgesetzt – adaptieren können, wenn ausreichend Zeit nach Deletion und vor Sekundärinfektion verstreichen konnte.
T cell activation is a highly regulated process involving peptide–MHC engagement of the T cell receptor and positive costimulatory signals. Upon activation, coinhibitory 'checkpoints', including programmed cell death protein 1 (PD1), become induced to regulate T cells. PD1 has an essential role in balancing protective immunity and immunopathology, homeostasis and tolerance. However, during responses to chronic pathogens and tumours, PD1 expression can limit protective immunity. Recently developed PD1 pathway inhibitors have revolutionized cancer treatment for some patients, but the majority of patients do not show complete responses, and adverse events have been noted. This Review discusses the diverse roles of the PD1 pathway in regulating immune responses and how this knowledge can improve cancer immunotherapy as well as restore and/or maintain tolerance during autoimmunity and transplantation.
The failure of calcineurin inhibitor (CNI)-based regimens to improve long-term graft survival coupled with their known toxicities has prompted the development of alternative therapeutic agents, including monoclonal antibodies, fusion receptor proteins, and small molecules to target novel pathways. This chapter focuses on prominent pathways currently being targeted in transplantation, with an emphasis on pathways being developed for future use. Intracellular signal transduction pathways, B-cell therapeutic targets, including B-cell-activating factors (BAFF) and a proliferation-inducing ligand (APRIL), as well as co-stimulatory pathways, including the CD28/B7, CD40/CD154, LFA1/ICAM, and the CD2/LFA3 pathways, will be reviewed in this chapter. Ultimately, the goal of therapeutics in transplantation is to achieve operational, or preferably immunologic, tolerance to the allograft while maintaining an intact immune system for surveillance of other antigens, with agents devoid of the toxicities of CNIs. This chapter will focus on therapeutics aimed at achieving this goal.
Francisella tularensis subsp. tularensis strain SchuS4 (Ftt) is a highly virulent intracellular bacterium. Inhalation of 10 or fewer organisms results in an acute and potentially lethal disease called pneumonic tularemia. Ftt infections occur naturally in the U.S. and Ftt was developed as a bioweapon. Thus, there is a need for vaccines that protect against this deadly pathogen. Although a live vaccine strain of Francisella tularensis (LVS) exists, LVS fails to generate long-lived protective immunity against modest challenge doses of Ftt. We recently identified an important role for high avidity CD4(+) T cells in short-term protection and hypothesized that expanding this pool of cells would improve overall vaccine efficacy with regard to longevity and challenge dose. In support of our hypothesis, application of a prime/boost vaccination strategy increased the pool of high avidity CD4(+) T cells which correlated with improved survival following challenge with either increased doses of virulent Ftt or at late time points after vaccination. In summary, we demonstrate that both epitope selection and vaccination strategies that expand antigen-specific T cells correlate with superior immunity to Ftt as measured by survival.
Background
A number of food allergies (e.g. fish, shellfish, nuts) are lifelong, without any disease-transforming therapies and unclear in their underlying immunology. Clinical manifestations of food allergy are largely mediated by IgE. Although persistent IgE titres have conventionally been attributed to long-lived IgE+ plasma cells (PCs), this has not been directly and comprehensively tested.
Objective
To evaluate mechanisms underlying persistent IgE and allergic responses to food allergens.
Methods
We used a model of peanut allergy and anaphylaxis, various knockout mice, adoptive transfer experiments and in vitro assays, to identify mechanisms underlying persistent IgE humoral immunity over almost the entire life-span of the mouse (18-20 months).
Results
Contrary to conventional paradigms, our data show that clinically relevant lifelong IgE titres are not sustained by long-lived IgE+ PCs. Instead, lifelong reactivity is conferred by allergen-specific long-lived memory B cells that replenish the IgE+ PC compartment. B cell re-activation requires allergen re-exposure and IL-4 production by CD4 T cells. We define the half-lives of antigen-specific: germinal centers (23.3 days); IgE+ and IgG1+ PCs (60 days and 234.4 days respectively) and clinically-relevant cell-bound IgE (67.3 days).
Conclusions
These findings can explain lifelong food allergies observed in humans as the consequence of allergen exposures that recurrently activate memory B cells and identify these as a therapeutic target with disease-transforming potential.
International Congress of Immunology (ICI), Melbourne, AUSTRALIA, AUG 21-26, 2016
Prophylactic vaccinations have been one of the greatest advances in modern medicine, both eradicating disease and reducing mortality. The translation of this advance into cancer therapy has been challenging and dates back to the turn of the twentieth century (Currie, Br J Cancer 26: 141–153, 1972). Cancer cells, derived from an aberrant clone, bear predominantly self-antigens and thus avoid alerting the immune system. In addition, the tumor microenvironment can be severely immunosuppressive; adding an extra layer of protection against the host immune response. Tumor cells can actively suppress immune responses through the downregulation of antigen presentation and the production of membrane-bound and secreted immuneregulatory molecules (Upadhyay et al, Cancers (Basel), 7: 736-62, 2015). To overcome such obstacles, a successful cancer vaccine must be able to induce a powerful immune response against tumor-associated antigens (TAAs) while avoiding normal host cells. This strategy has proven difficult because TAAs are highly variable in their immunogenicity and undergo immune editing to escape recognition. In addition, they can differ between tumor types and more importantly between individuals (Escors, New J Sci, 2014: 25, 2014). The presence of antigen-presenting cells (APCs) is generally low in the tumor microenvironment. Some efficacy in the treatment of cancer has been demonstrated by the use of autologous dendritic cells (DCs) pulsed with tumor cell lysates containing a whole array of antigens as well as single TAAs (Reichardt et al, Blood Rev: 18, 235-43, 2004). DC can be differentiated and expanded from peripheral blood ex vivo, and a resected tumor mass can be used to subsequently load the DC with TAAs. These strategies, while successful in developing a patient-specific vaccine, are labor and time intensive limiting the ability to experiment with numerous iterations to optimize the approach.
The immune system of higher vertebrates is capable of protecting the individual against a vast array of invading organisms and can be broadly divided into innate and adaptive components. Innate immunity protects the host via an assortment of mechanisms that include epithelial barriers, antimicrobial peptides, the complement cascade, and genetically encoded pattern recognition receptors specific for certain commonly encountered bacterial and viral antigens1-5. Adaptive immunity supplements and complements the innate immune response, and through humoral and cellular components has the capacity to recognize and ultimately destroy essentially any pathogen that has not been previously encountered by the host.
Transplantation between genetically disparate individuals of the same species evokes numerous changes in the host and the graft. In general, the outcome of a transplanted organ is determined by two closely interrelated components. The first component is the biology of the foreign graft (i.e., type of organ and its macro-and microenvironments, the anatomic location, the type of vascular connection, the circumstance surrounding organ harvest and transplant, and the genetic differences between the donor and the recipient). The second component is the nature and intensity of immune responses that develop in the recipient after encountering that specific foreign tissue. The immunity of allograft rejection involves a number of powerful, complex, interrelated, and well-orchestrated immune mechanisms and is only partially understood. This destructive immune response, if left unchecked, can and almost universally does, lead to complete destruction of the transplanted organ over time. This chapter offers an overview of the features and mechanisms of the immune system that work in concert to interact with and reject the organ allograft. Subsequent chapters will describe the role of distinct components that affect the immunology characteristic of the rejection response.
Antigen-specific immune tolerance promises to provide safe and effective therapies to prevent type 1 diabetes (T1D). Antigen-specific therapy requires two components: well-defined, clinically relevant autoantigens; and safe approaches to inducing tolerance in T cells specific for these antigens. Proinsulin is a critical autoantigen in both NOD mice, based on knockout mouse studies and induction of immune tolerance to proinsulin preventing disease whereas most antigens cannot, and also in human T1D based on proinsulin-specific T cells being found in the islets of affected individuals and the early appearance of insulin autoantibodies. Effective antigen-specific therapies that prevent T1D in humans have not yet been developed although doubt remains about the best molecular form of the antigen, the dose and the route of administration. Preclinical studies suggest that antigen specific therapy is most useful when administered before onset of autoimmunity but this time-window has not been tested in humans until the recent "pre-point" study. There may be a 'window of opportunity' during the neonatal period when 'vaccine' like administration of proinsulin for a short period may be sufficient to prevent diabetes. After the onset of autoimmunity, naive antigen-specific T cells have differentiated into antigen-experienced memory cells and the immune responses have spread to multiple antigens. Induction of tolerance at this stage becomes more difficult although recent studies have suggested generation of antigen-specific TR1 cells can inhibit memory T cells. Preclinical studies are required to identify additional 'help' that is required to induce tolerance to memory T cells and develop protocols for effective therapy in individuals with established autoimmunity.
Immunological memory may not represent a special characteristic of lymphocytes but simply re£ect low-level responses driven by antigen that is re-encountered or persists within the host. T-cell memory is important to control persistent infections within the individual host and cannot be transmitted to o¡spring because of MHC polymorphism and MHC-restricted T-cell recognition. In contrast, antibody memory is transmissible from mother to o¡spring and may function essentially to protect o¡spring during the phase of physiological immuno-incompetence before, at and shortly after birth. This physiological immuno-incompetence is a result of MHC polymorphism and the dangers of the graft-versus-host and host-versus-graft reaction between mother and embryo, which necessitate immunosuppression of the mother and immuno-incompetence of the o¡spring. One may argue therefore that immunological memory of transmissible immunological experience is the basis on which MHC-restricted T-cell recognition could develop or coevolve.
In this report, we have addressed two questions concerning immunological memory: the way in which naive and memory T cells recirculate through the body, and the intrinsic rate of division within the naive and memory populations. We identified naive and memory T cells in sheep by their cell surface phenotype and their ability to respond to recall antigen. Memory T cells were CD2hi, CD58hi, CD44hi, CD11ahi, and CD45R-, as pertains in man. T cells that crossed from blood to the tissues of the hind leg and accumulated in the popliteal afferent lymph were all of memory phenotype. Conversely, T cells in efferent lymph, 90% of which entered the lymph node (LN) via high endothelial venules (HEV), were mostly of the naive phenotype (CD2lo, CD58lo, CD44lo, CD11alo, and CD45R+). The marked enrichment of these two phenotypes in different recirculatory compartments indicated that memory T cells selectively traffic from blood to peripheral tissues to LN (via afferent lymph), whereas naive T cells selectively traffic from blood to LN (via HEV). We argue that the differential use of these two recirculation pathways probably optimizes lymphocyte interactions with antigen. The nonrandom distribution of T cell subsets in various recirculatory compartments may be related to the relative proportion of memory cells in each subset. In particular, gamma/delta T cells in blood were almost exclusively of memory phenotype, and accumulated preferentially in afferent, but not in efferent, lymph. Finally, using the bromo-deoxyuridine labeling technique, we found that at least a sizeable proportion of memory T cells, whether in blood or afferent lymph, were a dividing population of cells, whereas naive T cells were a nondividing population. This result supports an alternative model of lymphocyte memory that assumes that maintenance of memory requires persistent antigenic stimulation.
Binding of L-selectin expressed on lymphocytes to carbohydrate ligand(s) on lymph node high endothelial venules is thought to initiate lymphocyte extravasation from blood to lymph during recirculation and localization to sites of antigen (Ag) exposure. Previous studies have shown that treatment of lymphocytes with antibody to L-selectin (MEL-14) ablates trafficking to peripheral lymph nodes (PLN). In mice, naive but not memory CD4 cells express L-selectin. To examine the role of L-selectin in helper T cell migration, we studied the effects of in vivo administration of MEL-14 on CD4 cell responses. Systemic exposure of mice to MEL-14 depleted CD4 cells expressing a naive phenotype (CD45RBhi, CD44lo) from PLN but not from spleen. The majority of residual lymph node CD4 cells exhibited the reciprocal, memory phenotype (CD45RBlo, CD44hi). MEL-14 treatment prevented priming of naive CD4 cells for proliferation and cytokine production (IL-2 and IL-4) to keyhole limpet hemocyanin in PLN draining the site of Ag injection, but not in the spleen. The results suggest that naive cells were not depleted, but rather diverted to other sites where priming occurred. The data demonstrate that L-selectin mediates extravasation of naive CD4 cells into PLN and that its function cannot be replaced by other homing receptors.
Memory or antigen-experienced CD4 T cells differ from naive CD4 T cells both phenotypically by cell surface marker expression, and functionally by their dissimilar pattern of cytokine secretion and activation requirements through their T cell receptor (TCR). We show here that activation of memory CD4 T cells (CD45RBlo subset), but not naive CD4 T cells (CD45RBhi subset), is inhibited by MHC class II molecules on antigen-presenting cells and by CD4 ligation. We propose that the selective negative signal in memory cells is a direct result of the differences in signaling via CD4 and CD3, exemplified in the disparate pattern of tyrosine-phosphorylated proteins visible after activation of the two subsets. In vivo, this inhibitory signal may serve to prevent irrelevant interactions between memory CD4 T cells and bystander MHC class II+ cells, and may also be responsible for the defective functioning of memory CD4 T cells in AIDS.
A host's ability to resist certain pathogens such as Leishmania major can depend upon the phenotype of T helper (Th) subset that develops. Different murine genetic backgrounds are known to significantly alter the direction of Th subset development, although the cellular basis of this influence is poorly understood. To examine the basis of this effect we used an in vitro alpha/beta-T cell receptor (TCR) transgenic system for analysis of Th phenotype development. To control for TCR usage, we derived the DO11.10 alpha/beta-TCR transgene in several genetic backgrounds. Our findings suggest that the effects of genetic background on Th phenotype development reside within the T cell, and not the antigen-presenting cell compartment. Transgenic T cells from both the B10.D2 and BALB/c backgrounds showed development toward either the Th1 or Th2 phenotype under the strong directing influence of interleukin (IL) 12 and IL4, respectively. However, when T cells were activated in vitro under neutral conditions in which exogenous cytokines were not added, B10.D2-derived T cells acquired a significantly stronger Th1 phenotype than T cells from the BALB/c background, correspondent with in vivo Th responses to Leishmania in these strains. Importantly, these cytokine differences resulted in distinct functional properties, because B10.D2- but not BALB/c-derived T cells could induce macrophage production of nitric oxide, an important antimicrobial factor. Thus, the genetically determined default Th phenotype development observed in vitro may correspond to in vivo Th subset responses for pathogens such as Leishmania which do not initiate strong Th phenotype-directing signals.
Self-reactive B cells from tolerant double-transgenic (Dbl-Tg) mice coexpressing hen egg lysozyme (HEL) and rearranged anti-HEL immunoglobulin genes have a relatively short life span when compared to normal B cells, irrespective of whether they are exposed to antigen in multivalent membrane-bound form (mHEL-Dbl-Tg mice) or soluble form (sHEL-Dbl-Tg mice). The factors responsible for determining the fate of these B cells after encounter with self-antigen were investigated using a cell-tracking technique in which anti-HEL Ig-Tg spleen cells were labeled with the intracellular dye 5-carboxyfluorescein diacetate-succinimidyl ester (CFSE) and injected either into non-Tg recipients or a variety of HEL-Tg hosts. In non-Tg recipients, HEL-binding B cells persisted in the circulation and could be detected in the follicles of the spleen for at least 5 d. On transfer into either mHEL-Tg or sHEL-Tg hosts, they underwent activation and then rapidly disappeared from the blood and spleen over the next 3 d, consistent with the short life span reported previously. Immunohistology of spleens from sHEL-Tg recipients indicated that the transferred B cells had migrated to the outer margins of the periarteriolar lymphoid sheath (PALS), where they were detectable for 24 h before being lost. The positioning of B cells in the outer PALS depended on a critical threshold of Ig receptor binding corresponding to a serum HEL concentration between 0.5 and 15 ng/ml, but was not restricted to endogenously expressed HEL in that the same migratory pattern was observed after transfer into non-Tg recipients given exogenous (foreign) HEL. Moreover, bone marrow-derived immature Ig-Tg B cells homed to the outer PALS of sHEL-Tg mice and then disappeared at the same rate as mature B cells, indicating that the stage of maturation did not influence the fate of self-reactive B cells in a tolerant environment. On the other hand, HEL-binding B cells transferred into sHEL-Dbl-Tg recipients persisted over the 3-d period of study, apparently due to insufficient availability of antigen, as indicated by the fact that the degree of Ig receptor downregulation on the transferred B cells was much less than in sHEL-Tg recipients. If T cell help was provided to Ig-Tg B cells at the time of transfer into sHEL-Tg recipients in the form of preactivated CD4+ T cells specific for major histocompatibility complex-peptide complexes on the B cell surface, HEL-binding B cells migrated through the outer PALS of the spleen to the follicle, where they formed germinal centers, or to adjacent red pulp, where they formed proliferative foci and secreted significant amounts of anti-HEL antibody. Taken together, these results indicated that the outcome of the interaction between self-antigen and B cells is largely determined by a combination of the degree of receptor engagement and availability of T cell help.
The cellular basis of T cell memory is a controversial issue and progress has been hampered by the inability to induce and to trace long-term memory T cells specific for a defined antigen in vivo. By using the murine model of lymphocytic choriomeningitis virus (LCMV) infection and an adoptive transfer system with CD8+ T cells from transgenic mice expressing an LCMV-specific T cell receptor, a population of authentic memory T cells specific for LCMV was generated and analyzed in vivo. The transgenic T cells that have expanded (1,000-fold) and then decreased (10-fold) in LCMV-infected C57BL/6 recipient mice exhibited the following characteristics: they were (a) of larger average cell size than their naive counterparts but smaller than day 8 effector cells; (b) heterogeneous with respect to expression of cell surface "memory" markers; and (c) directly cytolytic when isolated from recipient spleens. The time-dependent proliferative activity of these LCMV-specific memory T cells was analyzed in the recipients by bromodeoxyuridine labeling experiments in vivo. The experiments revealed that LCMV-specific CD8+ memory T cells can persist in LCMV-immune mice for extended periods of time (>2 mo) in the absence of cell division; the memory population as a whole survived beyond 11 mo.
While interactions between CD28 and members of the B7 family costimulate and enhance T cell responses, recent evidence indicates that the CD28 homologue CTLA-4 plays a downregulatory role. The mechanism by which this occurs is not clear, but it has been suggested that CTLA-4 terminates ongoing responses of activated T cells, perhaps by induction of apoptosis. Here we demonstrate that CTLA-4 engagement by antibody cross-linking or binding to B7 inhibits proliferation and accumulation of the primary T cell growth factor, IL-2, by cells stimulated with anti-CD3 and anti-CD28. This inhibition is not a result of enhanced cell death. Rather it appears to result from restriction of transition from the G1 to the S phase of the cell cycle. Our observation that upregulation of both the IL-2R alpha chain and the CD69 activation antigen are inhibited by CTLA-4 engagement supplies further evidence that CTLA-4 restricts the progression of T cells to an activated state. Together this data demonstrates that CTLA-4 can regulate T cell activation in the absence of induction of apoptotic cell death.
The induction of T cell anergy in vivo is thought to result from antigen recognition in the absence of co-stimulation and inflammation, and is associated with a block in T cell proliferation and Th1 differentiation. Here we have examined the role of interleukin (IL)-12, a potent inducer of Th1 responses, in regulating this process. T cell tolerance was induced by the administration of protein antigen without adjuvant in normal mice, and in recipients of adoptively transferred T cells from T cell receptor transgenic mice. The administration of IL-12 at the time of tolerance induction stimulates Th1 differentiation, but does not promote antigen-specific T cell proliferation. Conversely, inhibiting CTLA-4 engagement during anergy induction reverses the block in T cell proliferation, but does not promote full Th1 differentiation. T cells exposed to tolerogenic antigen in the presence of both IL-12 and anti-CTLA-4 antibody are not anergized, and behave identically to T cells which have encountered immunogenic antigen. These results suggest that two processes contribute to the induction of anergy in vivo; CTLA-4 engagement, which leads to a block in the ability of T cells to proliferate to antigen, and the absence of a prototypic inflammatory cytokine, IL-12, which prevents the differentiation of T cells into Th1 effector cells. The combination of IL-12 and anti-CTLA-4 antibody is sufficient to convert a normally tolerogenic stimulus to an immunogenic one.
A small population of CD4+ OVA-specific TCR transgenic T cells was tracked following the induction of peripheral tolerance by soluble Ag to address whether functionally unresponsive, or anergic T cells, persist in vivo for extended periods of time. Although injection of OVA peptide in the absence of adjuvant caused a transient expansion and deletion of the Ag-specific T cells, a population that showed signs of prior activation persisted in the lymphoid tissues for several months. These surviving OVA-specific T cells had long-lasting, but reversible defects in their ability to proliferate in lymph nodes and secrete IL-2 and TNF-alpha in vivo following an antigenic challenge. These defects were not associated with the production of Th2-type cytokines or the capacity to suppress the clonal expansion of a bystander population of T cells present in the same lymph nodes. Therefore, our results provide direct evidence that a long-lived population of functionally impaired Ag-specific CD4+ T cells is generated in vivo after exposure to soluble Ag.
Early events in the humoral immune response were visualized in lymph nodes by simultaneous tracking of antigen-specific CD4
T and B cells after immunization. The T cells were initially activated in the T cell areas when the B cells were still randomly
dispersed in the B cell–rich follicles. Both populations then migrated to the edges of the follicles and interacted there,
resulting in CD154-dependent B cell proliferation and germinal center formation. These results provide visual documentation
of cognate T-B cell interactions and localize them to the follicular border.
CD8+ T cells are present at a higher frequency following a primary response, and these memory cells exhibit qualitative differences from naive cells. The importance of these differences vs increased precursor frequency in making a memory response more rapid and efficient has been unclear. Adoptive transfer of 2C TCR transgenic CD8+ T cells into normal recipients, followed by i.p. challenge with allogeneic P815 tumor, results in a long-lived memory population that includes both endogenous host CD8+ T cells and 2C cells. The 2C cells can be identified, using 1B2 mAb specific for the TCR, and thus used as an indicator of the properties of the memory cells. The memory cells have a heterogeneous surface phenotype, and their distribution in lymphoid organs, blood, and peripheral sites is distinct from that of naive cells. Upon rechallenge with Ag, memory cells access the peritoneal cavity much more rapidly than do naive cells (12 h vs 5 days). This appears to result from a requirement for naive cells to interact with Ag before they can efficiently migrate to inflammatory sites, while this is not required for memory cells. In addition, memory cells exhibit some cytolytic activity before rechallenge with Ag, and potent cytolytic activity is present in the peritoneal cavity within 12 h of rechallenge. Comparison of primary and memory responses in mice having similar frequencies of Ag-specific precursors demonstrated that the more rapid migration and the immediate effector function of at least some memory cells contribute very substantially to making a memory response at a peripheral site more rapid and efficient.
CTLA-4, a negative regulator of T cell function, was found to associate with the T cell receptor (TCR) complex ζ chain in
primary T cells. The association of TCRζ with CTLA-4, reconstituted in 293 transfectants, was enhanced by p56lck-induced tyrosine phosphorylation. Coexpression of the CTLA-4–associated tyrosine phosphatase, SHP-2, resulted in dephosphorylation
of TCRζ bound to CTLA-4 and abolished the p56lck-inducible TCRζ–CTLA-4 interaction. Thus, CTLA-4 inhibits TCR signal transduction by binding to TCRζ and inhibiting tyrosine
phosphorylation after T cell activation. These findings have broad implications for the negative regulation of T cell function
and T cell tolerance.
To determine how murine memory and naive T cells differ, we generated large numbers of long-lived memory CD8+ T cells and compared them to naive cells expressing the same antigen-specific receptor (T cell receptor; TCR). Although both
populations expressed similar levels of TCR and CD8, on antigen stimulation in vitro memory T cells down-regulated their TCR faster and more extensively and secreted IFN-γ and IL-2 faster than naive T cells.
Memory cells were also larger, and when freshly isolated from mice they contained perforin and killed target cells without
having to be restimulated. They further differed from naive cells in requiring IL-15 for proliferation and in having a greater
tendency to undergo apoptosis in vitro. On antigen stimulation in vivo, however, they proliferated more rapidly than naive cells. These findings suggest that, unlike naive T cells, CD8 memory
T cells are intrinsically programmed to rapidly express their effector functions in vivo without having to undergo clonal expansion and differentiation.
In this report, we have addressed two questions concerning immunological memory: the way in which naive and memory T cells recirculate through the body, and the intrinsic rate of division within the naive and memory populations. We identified naive and memory T cells in sheep by their cell surface phenotype and their ability to respond to recall antigen. Memory T cells were CD2hi, CD58hi, CD44hi, CD11ahi, and CD45R-, as pertains in man. T cells that crossed from blood to the tissues of the hind leg and accumulated in the popliteal afferent lymph were all of memory phenotype. Conversely, T cells in efferent lymph, 90% of which entered the lymph node (LN) via high endothelial venules (HEV), were mostly of the naive phenotype (CD2lo, CD58lo, CD44lo, CD11alo, and CD45R+). The marked enrichment of these two phenotypes in different recirculatory compartments indicated that memory T cells selectively traffic from blood to peripheral tissues to LN (via afferent lymph), whereas naive T cells selectively traffic from blood to LN (via HEV). We argue that the differential use of these two recirculation pathways probably optimizes lymphocyte interactions with antigen. The nonrandom distribution of T cell subsets in various recirculatory compartments may be related to the relative proportion of memory cells in each subset. In particular, gamma/delta T cells in blood were almost exclusively of memory phenotype, and accumulated preferentially in afferent, but not in efferent, lymph. Finally, using the bromo-deoxyuridine labeling technique, we found that at least a sizeable proportion of memory T cells, whether in blood or afferent lymph, were a dividing population of cells, whereas naive T cells were a nondividing population. This result supports an alternative model of lymphocyte memory that assumes that maintenance of memory requires persistent antigenic stimulation.
A host's ability to resist certain pathogens such as Leishmania major can depend upon the phenotype of T helper (Th) subset that develops. Different murine genetic backgrounds are known to significantly alter the direction of Th subset development, although the cellular basis of this influence is poorly understood. To examine the basis of this effect we used an in vitro alpha/beta-T cell receptor (TCR) transgenic system for analysis of Th phenotype development. To control for TCR usage, we derived the DO11.10 alpha/beta-TCR transgene in several genetic backgrounds. Our findings suggest that the effects of genetic background on Th phenotype development reside within the T cell, and not the antigen-presenting cell compartment. Transgenic T cells from both the B10.D2 and BALB/c backgrounds showed development toward either the Th1 or Th2 phenotype under the strong directing influence of interleukin (IL) 12 and IL4, respectively. However, when T cells were activated in vitro under neutral conditions in which exogenous cytokines were not added, B10.D2-derived T cells acquired a significantly stronger Th1 phenotype than T cells from the BALB/c background, correspondent with in vivo Th responses to Leishmania in these strains. Importantly, these cytokine differences resulted in distinct functional properties, because B10.D2- but not BALB/c-derived T cells could induce macrophage production of nitric oxide, an important antimicrobial factor. Thus, the genetically determined default Th phenotype development observed in vitro may correspond to in vivo Th subset responses for pathogens such as Leishmania which do not initiate strong Th phenotype-directing signals.
Binding of L-selectin expressed on lymphocytes to carbohydrate ligand(s) on lymph node high endothelial venules is thought to initiate lymphocyte extravasation from blood to lymph during recirculation and localization to sites of antigen (Ag) exposure. Previous studies have shown that treatment of lymphocytes with antibody to L-selectin (MEL-14) ablates trafficking to peripheral lymph nodes (PLN). In mice, naive but not memory CD4 cells express L-selectin. To examine the role of L-selectin in helper T cell migration, we studied the effects of in vivo administration of MEL-14 on CD4 cell responses. Systemic exposure of mice to MEL-14 depleted CD4 cells expressing a naive phenotype (CD45RBhi, CD44lo) from PLN but not from spleen. The majority of residual lymph node CD4 cells exhibited the reciprocal, memory phenotype (CD45RBlo, CD44hi). MEL-14 treatment prevented priming of naive CD4 cells for proliferation and cytokine production (IL-2 and IL-4) to keyhole limpet hemocyanin in PLN draining the site of Ag injection, but not in the spleen. The results suggest that naive cells were not depleted, but rather diverted to other sites where priming occurred. The data demonstrate that L-selectin mediates extravasation of naive CD4 cells into PLN and that its function cannot be replaced by other homing receptors.
The dynamic and reciprocal communication between T helper (Th) cells and B cells appears to rely on the provision of multiple signals. The first is antigen specific and is mediated by the interaction between the T-cell receptor (TCR) and antigen bound to the major histocompatibility complex (MHC). The subsequent signals are provided by the binding of accessory molecules such as CD28 and CD40 to their respective ligands. Here, Fiona Durie and colleagues discuss the co-stimulatory role of the interaction between CD40 on B cells and CD40 ligand (CD40L, gp39) on T cells, and review evidence that suggests blocking this interaction may induce T-cell tolerance.
We have characterized the surface phenotype and function of long-lived, Ag-specific memory CD4+ T cells generated in vivo by immunization with keyhole limpet hemocyanin (KLH). CD4+ T cells from the spleens of mice primed more than 2 mo previously with KLH, produced high levels of IL-2 and IL-3, and low levels of IL-4 and IFN-gamma in response to in vitro restimulation with specific Ag. The KLH-primed T cells mediated carrier-specific helper activity for the antibody production by NIP-primed B cells in secondary in vitro responses to NIP-KLH. Subsets of CD4+ T cells from KLH-primed mice were isolated on the basis of surface CD45RB (23G2) by magnetic separation and were examined for functional capacity in several assays of Ag-specific recall. Virtually all of the secretion of IL-2, IL-3, IL-4, and IFN-gamma in response to restimulation with Ag in vitro was associated with, and considerably enriched in, the CD45RB- subset of CD4+ T cells. Similarly, carrier-specific helper function and Ag-specific proliferation in vitro were also confined to the CD45RB-, CD4+ subset of T cells, confirming the previous association of this surface phenotype with memory Th cell activity. We also examined expression of the lymphocyte homing receptor, MEL-14 (gp90MEL), which is required for lymphocyte extravasation to peripheral lymph nodes and is present in high levels on naive T cells. MEL-14 positive and negative subsets of CD4+ T cells from long term KLH-primed mice were evaluated for Ag-specific memory function in terms of lymphokine production, Ag-induced proliferation, and helper activity. Each of these functions was associated exclusively with the MEL-14- subset of CD4+ T cells, which exhibited responses comparable to the CD45RB- subset. These data indicate that memory Th cell function in the spleen is contained within the MEL-14-, CD45RB- subset of CD4+ T cells and suggest that memory helper cells may have different patterns of recirculation from naive T cells.
The CD45 Ag family is comprised of at least five isoforms generated by the alternative splicing of three exons encoded by a single leukocyte common Ag gene. These isoforms, differentially expressed on subsets of T cells, are widely believed to delineate maturational status. Thus, cells expressing the high molecular mass CD45RA+ isoforms (220 and 205 kDa) are believed to represent naive cells, whereas those lacking CD45RA and expressing high density CD45RO (180 kDa) represent memory cells. This is based on findings that after activation, CD45RA+ cells "convert" to the CD45RA- phenotype, losing CD45RA and gaining CD45RO. This conversion is believed to be both unidirectional and irreversible. We now report a strategy for the development of polyclonal CD45RA+ cell lines derived from peripheral blood CD4+CD45RA+ cells. However, in such cell lines, CD45RA expression was not constant, but varied cyclically with restimulation. After restimulation, CD45RA expression decreased significantly, nadiring on days 5 to 7. However, CD45RA was fully re-expressed by days 14 to 16. These cells coexpressed high density CD45RO, which did not vary through the stimulatory cycle. Cells sorted CD45RA- on day 7 of the cycle rapidly reexpress CD45RA. Metabolic labeling revealed that the 220 kDa CD45RA isoform is no longer synthesized. Synthesis of the 205 kDa (CD45RA) isoform decreased significantly early after stimulation, but then increased back to baseline after day 12. The 180-kDa (CD45RO) isoform followed an opposite pattern. Our results document that alternative splicing of the different CD45 isoforms is highly regulated after activation and that conversion from CD45RA+ to CD45RA- is neither unidirectional, nor irreversible.
An engagement of CD40 with CD40 ligand (CD40L) expressed on activated T cells is known to provide an essential costimulatory signal to B cells in vitro. To investigate the role of CD40 in in vivo immune responses, CD40-deficient mice were generated by gene targeting. The significant reduction of CD23 expression on mature B cells and relatively decreased number of IgM bright and IgD dull B cells were observed in the mutant mice. The mutant mice mounted IgM responses but no IgG, IgA, and IgE responses to thymus-dependent (TD) antigens. However, IgG as well as IgM responses to thymus-independent (TI) antigens were normal. Furthermore, the germinal center formation was defective in the mutant mice. These results suggest that CD40 is essential for T cell-dependent immunoglobulin class switching and germinal center formation, but not for in vivo T cell-dependent IgM responses and T cell-independent antibody responses.
Whether the L-selectin-low (L-sel-lo) phenotype of acutely stimulated CD8+ T cells is a permanent characteristic of long-term memory CTL precursors (p) is addressed for mice primed with an influenza A virus or the murine parainfluenza type 1 virus, Sendai virus. In both cases, many of the splenic CD8+ CTLp gradually lose the predominantly L-sel-lo profile associated with recently generated CTLp populations. The influenza-specific CTLp also tend to revert from the activated alpha 4-integrin-high to the resting alpha 4-integrin-low form. The kinetics of the switch back to the "naive" L-sel-hi phenotype differs for the influenza and Sendai virus models, perhaps reflecting events occurring during the acute phases of these responses. The return to being L-sel-hi is not due to irreversible lymphocyte senescence, because restimulation of this set with the inducing virus in vitro causes most of the cells to become L-sel-lo. Also, despite the time-related drift of these particular memory CTLp to the L-sel-hi state, the size of the total pool of L-sel-lo CD8+ T cells increases with age.
Evidence from tissue culture experiments and in vivo studies indicates that certain forms of antigen presentation render T cells unresponsive to subsequent antigenic stimulation. Great progress has been made recently in understanding the induction, maintenance, and in vivo relevance of this unresponsive state, called clonal anergy.
In vitro generated CD4 cell lines and effectors often produce either Th1 or Th2 cytokines, but stimulation of resting CD4 cells from animals leads to production mostly of IL-2. To determine whether polarization of CD4 effector cells results in development of polarized memory, I generated antigen-specific Th1 and Th2 effectors in vitro and transferred them to adoptive hosts. The effectors gave rise to long-lived populations of CD4 T cells with the phenotype of resting memory cells. Recovered cells responded vigorously to specific antigen, producing a pattern of cytokines closely related to that of the transferred effectors. Thus, encounter with a peptide antigen and directive cytokines at the initiation of culture can lead to generation of both effector and long-lived memory cell populations that produce restricted patterns of cytokines.
An adoptive transfer system was used to monitor physically the behavior of a trace population of TCR transgenic T cells in vivo. After subcutaneous injection of antigen in adjuvant, the antigen-specific cells accumulated first in the paracortical region of the draining lymph nodes, proliferated there for several days, and then moved into lymph node follicles, where they accounted for most of the T cells. They then disappeared slowly from the draining nodes, and the remaining cells were hypersensitive to antigenic stimulation in vitro. In contrast, when the antigen was introduced into the blood, the antigen-specific cells rapidly accumulated in the paracortical regions of all lymph nodes, proliferated there for a short time, but never entered follicles. Most of the cells then rapidly disappeared, leaving behind a population that was hyporesponsive to antigenic stimulation. These results provide a physical basis for the classical finding that antigen-specific memory and tolerance can be influenced by the form of antigen administration.
Secondary responses to Ag in vivo are characterized by more rapid kinetics and greatly enhanced magnitude compared with primary responses. For CD4+ T cells, this is in part due to a greater frequency of Ag-specific memory cells, and may also reflect differences in responsiveness of memory vs naive cells to stimulation. To compare activation requirements and the role of accessory cells, naive and memory cells were stimulated with immobilized anti-CD3 in the presence or absence of APC. With anti-CD3 alone, naive cells proliferated slightly but produced no detectable IL-2, whereas memory cells proliferated well with significant IL-2 production. Increasing numbers of T-depleted APC greatly enhanced responses of naive cells to levels equivalent to those of memory cells, whereas for memory cells only IL-2 production increased slightly. The response of naive cells was equivalent in magnitude and kinetics to that of memory cells when low density APC, enriched in dendritic cells and depleted of resting B cells, were used with anti-CD3. To directly compare naive and memory responses in an Ag-specific model, we examined CD4+ cells specific for a peptide of pigeon cytochrome c fragment isolated from TCR-alpha beta transgenic mice. Naive cells were compared with 4-day activated blasts (effectors) and memory cells generated by adoptive transfer of effectors to adult thymectomized bone marrow reconstituted mice, in which the cells return to a resting state but still respond to recall Ag. Naive cells responded to Ag on dendritic cells and activated B cells but not on resting B cells or macrophages. In contrast, both memory cells and effectors were stimulated by all APCs, including resting B cells and macrophage to a limited extent. The ability of memory cells to respond to all APC types was confirmed using Ag-specific cells generated by in vivo priming with keyhole limpet hemocyanin. These results suggest that memory cells are considerably less dependent on accessory cell costimulation than naive cells, but that naive cells can respond equivalently in both magnitude and kinetics if Ag is presented on costimulatory APCs such as dendritic cells. In addition, these studies suggest that the enhanced secondary T cell response is due to a combination of the increased frequency of Ag-specific cells and their ability to react to Ag presented on a wider range of APC types, rather than an inherent capacity of memory T cells to respond better and faster.
An increased understanding of the types of T-cell subsets that exist in vivo, their relationships to one another, and how to identify and isolate them or effect their generation, has led to a comprehensive view of the antigen-presenting cells (APCs) which may be active and regulatory during the course of an immune response. Recent studies show that naive T cells only respond efficiently to dendritic cells and activated B cells whereas memory and effector cells respond to all APC types to some extent, including resting B cells. High level co-stimulatory molecule expression largely explains why APCs such as dendritic cells are far more effective stimulators than resting B cells. The available data, therefore, suggest that the requirement for co-stimulation, and hence capacity to respond to various APCs, is largely a function of the differentiation state of the T cell, and that previous encounter with antigen fundamentally increases the ability of T cells to subsequently respond to antigen rechallenge.
CD4 T cells bearing high (240-190 kDa) and low (180 kDa) molecular mass isoforms of the leukocyte common antigen CD45 define functionally distinct subsets which have been equated with naive and memory T cells. In the rat, CD4 T cells expressing a high molecular mass isoform [identified by monoclonal antibody MRC-OX22 (anti-CD45RC)] exchange this for the 180 kDa molecule (CD45RC-) when stimulated by antigen. Here we show, by transferring mature allotype-marked CD45RC- CD4 T cells (depleted of immature Thy-1+ CD45RC- recent thymic emigrants) into normal euthymic recipients, that many T cells re-express the high molecular mass isoform in less than 6 h. By 24 h, 30-60% of CD45RC- CD4 T cells became CD45RC+; within a week the entire cohort appeared to exchange the low for the high molecular mass isoform. Isoform exchange was dynamic and many CD4 T cells returned once again to the CD45RC- state. CD45RC- CD4 T cells declined in number more rapidly than the CD45RC+ subset after transfer. The results suggest that CD45R isoforms distinguish between resting T cells (CD45RC+) and those which have encountered antigen in the recent past. CD45R isoforms would appear to be unsuitable markers of naive and memory T cells.
The immune system can remember, sometimes for a lifetime, the identity of a pathogen. Understanding how this is accomplished has fascinated immunologists and microbiologists for many years, but there is still considerable debate regarding the mechanisms by which long-term immunity is maintained. Some of the controversy stems from a failure to distinguish between effector and memory cells and to define their roles in conferring protection against disease. Here the current understanding of the cellular basis of immune memory is reviewed and the relative contributions made to protective immunity by memory and effector T and B cells are examined.
The appropriate recirculation and migration of naive, effector and memory T cells into inflamed tissue are precisely controlled by adhesive interactions with vascular endothelium. Analyses of CD4 lymphocytes have indicated that naive and antigen-experienced cells exhibit distinctive patterns of homing and recirculation, and that subsets of cells preferentially localize in different anatomical locations as a consequence of previous antigen exposure and differences in adhesion receptor usage.
The concept that ‘naive’ T cells (CD4+CD45RAhi) selectively migrate into lymphoid organs and ‘memory’ T cells (CD4+CD45R0hi) migrate into nonlymphoid organs has been enthusiastically taken up by the scientific community. However, even today, this premise is based mainly on indirect evidence obtained in one species. Here, Jürgen Westermann and Reinhard Pabst argue that, in the light of recent data, the generalization of this concept was too early.
Humoral immune responses were characterized in mouse strains lacking either or both B7 molecules. Mice deficient in both B7-1 and B7-2 failed to generate antigen-specific IgG1 and IgG2a responses and lacked germinal centers when immunized by a number of routes and even in the presence of complete Freund's adjuvant. These results demonstrate that B7-mediated signaling plays a critical role in germinal center formation and immunoglobulin class switching in vivo. Mice lacking only B7-1 or B7-2 mounted high-titer antigen-specific IgG responses when immunized in complete Freund's adjuvant, indicating that B7-1 and B7-2 can have overlapping, compensatory functions for IgG responses. When immunized intravenously without adjuvant, B7-2-deficient mice failed to switch antibody isotypes or form germinal centers, whereas B7-1-deficient mice gave antibody responses comparable with wild-type mice. Thus, B7-2 has an important role in initiating antibody responses in the absence of adjuvant, but the induction of B7-1 by adjuvant in B7-2-deficient mice can compensate for the absence of B7-2.
Studies of T cell anergy in vitro have led to the widely accepted view that anergy is induced by T cell antigen recognition without costimulation. We show that the induction of T cell anergy in vivo is due to an abortive T cell response that requires recognition of B7 molecules, since blocking B7 maintains T cells in an unactivated but functionally competent state. Furthermore, the induction of anergy is prevented by blocking CTLA-4, the inhibitory T cell receptor for B7 molecules. Thus, in vivo T cell anergy may be induced not because of a lack of costimulation, but as a result of specific recognition of B7 molecules by CTLA-4. In contrast, blocking CD28 on T cells prevents priming but not the induction of tolerance. Therefore, the outcome of antigen recognition by T cells is determined by the interaction of CD28 or CTLA-4 on the T cells with B7 molecules.
Adoptive transfer of TCR-transgenic T cells uniformly expressing an identifiable TCR of known peptide/MHC specificity can be used to monitor the in vivo behavior of antigen-specific T cells. We have used this system to show that naive T cells are initially activated within the T-cell zones of secondary lymphoid tissue to proliferate in a B7-dependent manner. If adjuvants or inflammatory cytokines are present during this period, enhanced numbers of T cells accumulate, migrate into B-cell-rich follicles, and acquire the capacity to produce IFN-gamma and help B cells produce IgG2a. If inflammation is absent, most of the initially activated antigen-specific T cells disappear without entering the follicles, and the survivors are poor producers of IL-2 and IFN-gamma. Our results indicate that inflammatory mediators play a key role in regulating the anatomic location, clonal expansion, survival and lymphokine production potential of antigen-stimulated T cells in vivo.
Most of the antigen-specific T and B cells participating in the primary immune response are rapidly eliminated, but some of the cells survive and become long-lived memory cells. There have been a number of recent developments on the features and functions of memory cells.
Naive and memory CD4 T cells differ in cell surface phenotype, function, activation requirements, and modes of regulation. To investigate the molecular bases for the dichotomies between naive and memory CD4 T cells and to understand how the T cell receptor (TCR) directs diverse functional outcomes, we investigated proximal signaling events triggered through the TCR/CD3 complex in naive and memory CD4 T cell subsets isolated on the basis of CD45 isoform expression. Naive CD4 T cells signal through TCR/CD3 similar to unseparated CD4 T cells, producing multiple tyrosine-phosphorylated protein species overall and phosphorylating the T cell-specific ZAP-70 tyrosine kinase which is recruited to the CD3zeta subunit of the TCR. Memory CD4 T cells, however, exhibit a unique pattern of signaling through TCR/CD3. Following stimulation through TCR/CD3, memory CD4 T cells produce fewer species of tyrosine-phosphorylated substrates and fail to phosphorylate ZAP-70, yet unphosphorylated ZAP-70 can associate with the TCR/CD3 complex. Moreover, a 26/28-kDa phosphorylated doublet is associated with CD3zeta in resting and activated memory but not in naive CD4 T cells. Despite these differences in the phosphorylation of ZAP-70 and CD3-associated proteins, the ZAP-70-related kinase, p72syk, exhibits similar phosphorylation in naive and memory T cell subsets, suggesting that this kinase could function in place of ZAP-70 in memory CD4 T cells. These results indicate that proximal signals are differentially coupled to the TCR in naive versus memory CD4 T cells, potentially leading to distinct downstream signaling events and ultimately to the diverse functions elicited by these two CD4 T cell subsets.
This study addresses the question of whether human peripheral CD4+ CD45RA+ T cells possess antigen-specific immune memory. CD4+ CD45RA+ T cells were isolated by a combination of positive and negative selection. Putative CD4+ CD45RA+ cells expressed CD45RA (98.9%) and contained < 0.1% CD4+ CD45RO+ and < 0.5% CD4+ CD45RA+ CD45RO+ cells. Putative CD45RO+ cells expressed CD45RO (90%) and contained 9% CD45RA+ CD45RO+ and < 0.1% CD4+ CD45RA+ cells. The responder frequency of Dermatophagoides pteronyssinus-stimulated CD4+ CD45RA+ and CD4+ CD45RO+ T cells was determined in two atopic donors and found to be 1:11,314 and 1:8031 for CD4+ CD45RA+ and 1:1463 and 1:1408 for CD4+ CD45RO+ T cells. The responder frequencies of CD4+ CD45RA+ and CD4+ CD45RO+ T cells from two non-atopic, but exposed, donors were 1:78031 and 1:176,903 for CD4+ CD45RA+ and 1:9136 and 1:13,136 for CD4+ CD45RO+ T cells. T cells specific for D. pteronyssinus were cloned at limiting dilution following 10 days of bulk culture with D. pteronyssinus antigen. Sixty-eight clones were obtained from CD4+ CD45RO+ and 24 from CD4+ CD45RA+ T cells. All clones were CD3+ CD4+ CD45RO+ and proliferated in response to D. pteronyssinus antigens. Of 40 clones tested, none responded to Tubercule bacillus purified protein derivative (PPD). No difference was seen in the pattern of interleukin-4 (IL-4) or interferon-gamma (IFN-gamma) producing clones derived from CD4+ CD45RA+ and CD4+ CD45RO+ precursors, although freshly isolated and polyclonally activated CD4+ CD45RA+ T cells produced 20-30-fold lower levels of IL-4 and IFN-gamma than their CD4+ CD45RO+ counterparts. Sixty per cent of the clones used the same pool of V beta genes. These data support the hypothesis that immune memory resides in CD4+ CD45RA+ as well as CD4+ CD45RO+ T cells during the chronic immune response to inhaled antigen.
Naive and memory CD4 T cells are frequently defined by exon-specific monoclonal antibodies (mAb) which stain (or not) high- or low-molecular-weight (MW) isoforms of the leucocyte common antigen CD45. The link between isoform and the naive/memory designation is complicated by the fact that CD4 T cells with a 'memory' phenotype (CD45RA-, RB-, RC-, or CD45RO+) may revert ('revertants') and re-express the high mw isoform (CD45RA+, RB+, RC+). Isoform expression also changes during normal T-cell development. Furthermore, the picture may be incomplete since an exon-specific mAb will not detect all possible isoforms on a cell. We have used molecular techniques to determine whether revertant CD4 memory T cells were different from naive T cells with respect to CD45R isoform expression. Using the anti-CD45RC mAb OX22 to purify rat lymphocyte subsets, CD45R isoform expression was examined at the mRNA level in CD4 T cells at different stages of development and compared with that of B cells and unseparated lymphocytes. B cells contained abundant message for the highest MW 3-exon isoform ABC, the 2-exon isoforms AB and BC, and the null isoform O. Both immature CD45RC- (i.e. CD4+8- 'single positive' thymocytes, and peripheral Thy-1+ recent thymic emigrants) and mature CD45RC- 'antigen-experienced' CD4 T cells had message for single-exons B, possibly C and for the O exon. In contrast, CD45RC+ CD4 T cells contained mRNA coding for ABC (low level), AB, BC, B, C (low level) and O (low level). Importantly, there was no difference between CD45RC+ T cells that had not seen antigen ('truly native') and CD45RC+ antigen-experienced revertant memory T cells. This observation has implications for understanding long-term immunological memory.
Lymphocyte differentiation is connected with profound alterations in the migratory pattern of lymphocytes. Whereas naive cells predominantly recirculate through lymphoid tissues, activated lymphocytes acquire an increased preference for immigration into non-lymphoid tissues and a reduced capacity for recirculation via high endothelial venules (HEV). A variety of data had indicated that memory-related subpopulations of cells in man and sheep, classified by the low expression of the CD45RA isotype, also lack the capacity to recirculate via HEV. However, recent data in the rat called these results into question. We therefore analyzed the migration properties of murine CD4+ T cell subpopulations defined by several markers used to distinguish memory from naive CD4+ cells in mice, namely CD45RB, L-selectin and CD44. Our data clearly show that the majority of putative memory cells expressing either low levels of CD45RB, low levels of L-selectin or high levels of CD44 display a strongly reduced capacity for direct entry into lymphoid tissues, including the spleen, from the blood stream. The accumulation in peripheral lymph nodes is further reduced by treatment with anti-L-selectin antibody, which blocks their entry via HEV. This indicates that memory CD4+ T cells are not excluded from crossing lymph node HEV, and that the numbers of cells entering the node via this route exceed the numbers entering via the afferent lymph, at least in the absence of local inflammation. Concomitantly, a strongly enhanced localization of cells of the memory phenotype is observed in lung and liver as compared with naive cells. Trafficking to specific sites such as skin or gut mucosa is not a prominent feature of the total population of memory cells. The trafficking to lung and liver and an increased ability to bind to dendritic cells, demonstrable in in vitro adhesion assays, suggest a more sessile phenotype of most memory cells. With respect to these properties, memory cells have a surprizing similarity to fully activated lymphocytes.
It is thought that naive T cells predominantly enter lymphoid organs such as lymph nodes (LN) and Peyer's patches (PP) via high endothelial venules (HEV), whereas memory T cells migrate mainly into non-lymphoid organs. However, direct evidence for the existence of these distinct migration pathways in vivo is incomplete, and nothing is known about their migration through the different compartments of lymphoid organs. Such knowledge would be of considerable interest for understanding T cell memory in vivo. In the present study we separated naive and memory CD4+ T cells from the rat thoracic duct according to the expression of the high and low molecular weight isoforms of CD45R, respectively. At various time points after injection into congenic animals, these cells were identified by quantitative immunohistology in HEV, and T and B cell areas of different LN and PP. Three major findings emerged. First, both naive and memory CD4+ T cells enter lymphoid organs via the HEV in comparable numbers. Second, naive and memory CD4+ T cells migrate into the B cell area, although in small numbers and continuously enter established germinal centers (GC) with a bias for memory CD4+ T cells. Third, memory CD4+ T cells migrate faster through the T cell area of lymphoid organs than naive CD4+ T cells. Thus, our study shows that memory CD4+ T cells are not excluded from the HEV route. In addition, "memory" might depend in part on the ability of T cells to specifically enter the B cell area and GC and to screen large quantities of lymphoid tissues in a short time.
The cellular basis of immunological memory has been reassessed in the light of recent evidence identifying two types of CD4+ memory T cell. Here, Eric Bell and colleagues argue that two crucial components of memory—rapid recall and long life—belong not to one but to two distinct subsets of CD4+ memory T cells: the CD45Rlo and CD45Rhi subsets, respectively. Importantly, only the CD45Rlo subset depends on persisting antigen.
Immunological memory can be defined as the faster and stronger response of an animal that follows reexposure to the same antigen. By this definition, it is an operational property of the whole animal or the immune system. Memory cells express a different pattern of cell surface markers, and they respond in several ways that are functionally different from those of naive cells. Murine memory cells are CD44 high and low in the expression of activation markers such as CD25 (IL-2R), whereas human memory cells are CD45RA-, CD45RO+. In contrast to naive cells, memory cells secrete a full range of T cell cytokines and can be polarized to secrete particular restricted patterns of secretion for both CD4 and CD8 T cells. The requirements for the activation of memory cells for proliferation and cytokine production are not quite as strict as those of naive cells, but costimulation in the broad sense is required for optimum responses and for responses to suboptimum antigen concentrations. It would appear that memory cells can persist in the absence of antigenic stimulation and persist as nondividing cells. Reencounter with the same antigen can expand the population to a new, stable, higher level and generate a separate population of CD44 high effectors that may be required for protection, while competition from other antigens can drive it down to a lower stable level. It is unclear how or where memory cells arise, but once generated they have different pathways of recirculation and homing.
Proliferation of memory-phenotype (CD44hi) CD8+ cells induced by infectious agents can be mimicked by injection of type I interferon (IFN I) and by IFN I-inducing agents such as lipopolysaccharide and Poly I:C; such proliferation does not affect naive T cells and appears to be TCR independent. Since IFN I inhibits proliferation in vitro, IFN I-induced proliferation of CD8+ cells in vivo presumably occurs indirectly through production of secondary cytokines, e.g., interleukin-2 (IL-2) or IL-15. We show here that, unlike IL-2, IL-15 closely mimics the effects of IFN I in causing strong and selective stimulation of memory-phenotype CD44hi CD8+ (but not CD4+) cells in vivo; similar specificity applies to purified T cells in vitro and correlates with much higher expression of IL-2Rbeta on CD8+ cells than on CD4+ cells.
Antigen encounter not only induces a change in surface expression of CD45RC isoforms in the rat from a high (CD45RC+) to a low molecular weight molecule (CD45RC-), but also stimulates changes in expression of adhesion molecules that regulate CD4 T-cell migration. T cells with an activated or 'memory' phenotype (CD45RC-) are thought to enter lymph nodes almost exclusively via afferent lymphatics whereas T cells in a resting state (CD45RC+) migrate across high endothelial venules (HEV). The present study monitored the rapid recirculation from blood to lymph of allotype-marked CD45RC T-cell subsets. Surprisingly, we found that CD45RC- CD4 T cells entered the thoracic duct slightly faster and reached peak numbers 3 hr earlier (18 hr) than did the CD45RC+ subset. To determine whether the entrance of CD45RC+ and RC- subsets was restricted to HEV and afferent lymphatics, respectively, recirculation of CD4 T cells was monitored in mesenteric lymphadenectomized (MLNx) rats (on healing the intestinal afferent lymphatics are joined directly to the thoracic duct), or in recipients that had had the mesenteric lymph node (MLN) acutely (2-3 hr) deafferentized (entry would be restricted to HEV). In these studies CD45RC- CD4 T cells entered the MLN across HEV on an equal basis with T cells expressing a CD45RC+ phenotype. Contrary to currently held dogma the results showed that, in vivo, CD4 T cells with a memory phenotype freely enter lymph nodes (LN) across HEV as well as via afferent lymphatics.
Helper T (Th) cell differentiation is highly regulated by cytokines but initiated by mitogens. By examining gene expression in discrete generations of dividing cells, we have delineated the relationship between proliferation and differentiation. Initial expression of IL-2 is cell cycle-independent, whereas effector cytokine expression is cell cycle-dependent. IFNgamma expression increases in frequency with successive cell cycles, while IL-4 expression requires three cell divisions. Cell cycle progression and cytokine signaling act in concert to relieve epigenetic repression and can be supplanted by agents that hyperacetylate histones and demethylate DNA. Terminally differentiated cells exhibit stable epigenetic modification and cell cycle-independent gene expression. These data reveal a novel mechanism governing Th cell fate that initially integrates proliferative and differentiative signals and subsequently maintains stability of the differentiated state.
Phenotypically and functionally, the early steps of T cell differentiation are not well characterized. In addition, the effector T cell stage shares several phenotypic characteristics with memory T cells, which has made the analysis of T cell memory difficult. In this study, we have investigated in vitro and in vivo the differentiation of naive CTL into effector and memory CTL as a function of cell division using lymphocytic choriomeningitis virus-specific TCR-transgenic spleen cells labeled with the vital dye carboxyfluorescein diacetate, succinimidyl ester. The following major points emerged. 1) During the first nine cell divisions, the investigated cell surface markers were strongly modulated. 2) The TCR was stepwise down-regulated during viral infection. 3) Cytotoxic effector function was acquired within one cell division and was retained during the next four to five divisions. 4) In vitro, CTL reached a CD44highCD62L+ memory phenotype after 6-10 cell divisions and required restimulation to exert effector function. 5) Lymphocytic choriomeningitis virus memory mice contained two distinct memory populations: a CD44highCD62L- population, predominately located in the spleen and exerting rapid effector function, and a CD44highCD62L+ population found in the spleen and the lymph nodes, which had lost immediate effector function. This finding suggests that two types of memory CTL exist. The correlation between CD62L expression, effector function, and Ag persistence is discussed.
The phenotypic and functional characteristics of Ag-specific memory CD4+ lymphocytes are poorly defined. To examine the properties and cytokine responsiveness of these cells, we have developed an adoptive transfer system using in vitro-activated T cells expressing the DO.11 transgenic TCR specific for OVA(323-339)+ I-Ad. In vitro-activated DO.11 CD4+ cells exhibit comparable survival patterns at 1, 6, and 10 wk after adoptive transfer, indicating that a stable population of memory cells has been generated. In the absence of Ag, previously activated T cells survive longer than their naive counterparts in vivo, rapidly revert to a partially naive phenotype, and maintain their effector cytokine profile. The DO.11 CD4+ memory cells are capable of proliferating in response to IL-2 and IL-4, while naive DO.11 CD4+ cells exhibit no such proliferative responses.
In the present study, naive T cells were compared with in vivo generated effector and memory T cells expressing the same TCR specific for lymphocytic choriomeningitis virus. Upon restimulation in vitro, the same minimal concentrations of the full agonist peptide p33 and also of weak and partial agonist peptides were required for proliferation of naive, effector and memory T cells, indicating no difference in threshold of activation. However, activation kinetics were distinct. While effector cytotoxic T cells exhibited immediate ex vivo lytic effector function, naive and memory T cells required 12 h and more exposure to antigen to develop lytic activity. However, both effector and memory T cells contained IFN-gamma mRNA in vivo and required less than 3 h for secretion of cytokines upon restimulation in vitro. In contrast, naive T cells did not contain IFN-gamma mRNA and required more than 12 h for cytokine secretion. Our results show that memory T cells exhibit a unique phenotype in that they produce cytokines and commit to proliferation as rapidly as effector cells, whereas they resemble naive T cells in the time requirement for development of cytolytic function.
We have studied the kinetics of the antigen induced response of naive and memory CD8 T cells expressing a transgenic T cell receptor (TCR) specific for the glycoprotein peptide amino acid 33-41 (GP33) of the lymphocytic choriomeningitis virus (LCMV). Memory T cells were generated in vivo by adoptive transfer of LCMV TCR transgenic T cells into normal recipient mice, followed by LCMV infection. The results demonstrated that the cell cycle progression and kinetics of TCR down-modulation, CD25 and CD69 up-regulation were identical in naive and memory T cells after antigen recognition. Moreover, the two T cell populations did not differ in respect of activation thresholds and in their proliferative capacities neither in vitro nor in vivo. However, memory CD8 T cells could be more rapidly induced to become cytolytic and to secrete high levels of interleukin-2 and interferon-gamma than naive T cells. LCMV GP33-specific CD8 memory T cells were only slightly more efficient in reducing LCMV titers in the spleen but were far more effective than naive LCMV GP33-specific T cells in controlling subcutaneous tumor growth of B16.F10 melanoma cells which expressed the LCMV GP33 epitope as tumor-associated antigen. Thus, in our experiments the main difference between CD8 memory T cells and naive cells is the ability of the former to rapidly acquire effector cell functions.
- F Boriello
- M P Sethna
- S D Boyd
- A N Schweitzer
- E A Tivol
- D Jacoby
- T B Strom
- E M Simpson
- G J Freeman
- A H Sharpe
Boriello, F., M. P. Sethna, S. D. Boyd, A. N. Schweitzer, E. A. Tivol, D. Jacoby,
T. B. Strom, E. M. Simpson, G. J. Freeman, and A. H. Sharpe. 1997. B7-1 and
B7-2 have overlapping, critical roles in immunoglobulin class switching and
germinal center formation. Immunity 6:303.