A central role of CD40 ligand in the regulation of CD4+ T-cell responses.
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Animals↗
Biomedical subjects
R A Flavell
Publications and source records attributed to R A Flavell.
Disruption of CD40-CD40 ligand interactions results in an enhanced susceptibility to Leishmania amazonensis infection.
To study the role of CD40 ligand (CD40L) in the host immune responses against intracellular pathogens, we infected CD40L knockout (CD40L-/-) mice with Leishmania amazonensis. Although wild-type mice were susceptible to infection and developed progressive ulcerative lesions, tissue parasite burdens in CD40L-/- mice were significantly higher. This heightened susceptibility to infection was associated with an impaired T cell and macrophage activation and altered inflammatory response, as reflected by low levels of IFN gamma, lymphotoxin-tumor necrosis factor (LT-TNF), and nitric oxide (NO) production. Furthermore, CD40L-/- mice failed to generate a protective immune response after immunization. These results indicate an essential role of cognate CD40-CD40L interactions in the generation of cellular immune responses against an intracellular parasite.
Animals↗
Mice lacking the MHC class II transactivator (CIITA) show tissue-specific impairment of MHC class II expression.
CIITA activates the expression of multiple genes involved in antigen presentation and it is believed to be required for both constitutive and IFN gamma-inducible expression of these genes. To understand the role of CIITA in vivo, we have used gene targeting to generate mice that lack CIITA. CIITA-deficient (-/-) mice do not express conventional MHC class II molecules on the surface of splenic B cells and dendritic cells. In addition, macrophages resident in the peritoneal cavity do not express MHC class II molecules upon IFN gamma stimulation nor do somatic tissues of mice injected with IFN gamma, in contrast with wild-type mice. The levels of Ii and H-2M gene transcripts are substantially decreased but absent in CIITA (-/-) mice. The transcription of nonconventional MHC class II genes is, however not affected by CIITA deficiency. A subset of thymic epithelial cells express MHC class II molecules. Nonetheless, very few mature CD4 T cells are present in the periphery of CIITA (-/-) mice despite MHC class II expression in the thymus. Consequently, CIITA(-/-) mice are impaired in T-dependent antigen responses and MHC class II-mediated allogeneic responses.
Animals↗
Disruption of cytokine signaling in lymphoid development: unique contributions of the common cytokine gamma chain and the Jak3 kinase.
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Animals↗
The role of CD40 ligand in costimulation and T-cell activation.
It is clear by now that cell-to-cell interactions involving a variety of signals are required for effective immune response. The data reviewed here suggest that CD40-CD40L interactions are critical for development of CD4 T-cell-dependent effector functions. Lack of this important interaction results in greatly reduced activation of CD4 T cells, while successful interaction of these molecules results in full activation of these T cells. Consequently, the absence of CD40-CD40L interactions leads to impairment of T-cell effector such as help for B-cell differentiation and class switch, activation of monocytes and macrophages to produce cytokines and to kill intracellular pathogens, and activation of autoreactive T cells to mount an autoimmune response. The effector functions of T cells controlled by CD40-CD40L interactions in a successful immune response are given in Table I. Data presented so far suggest that CD40-CD40L interactions play a role in early signalling events, where interactions of this kind are required to induce expression of costimulatory molecules on APC. One possible sequence of events in that APC, like DC, take up antigens at the site of injury or infection and migrate to lymph nodes, where they present antigens complexed with MHC class II molecules to naive T cells. This results in expression of CD40L on T cells. Coupling of this newly expressed CD40L on T cells with CD40 on APC results in expression of the costimulatory activity of the APC. At this time the costimulatory signal provided by the APC is received by the T cells via CD28/CTLA-4, which drives the cell to enter into cell cycle and complete T cell activation. T cells thereby activated can now enter into secondary cognate CD40-CD40L-dependent effector recognition with B cells to switch Ig class, macrophages to produce cytokines and new DC carrying the same antigen to up-regulate costimulatory activity. A tight regulation of expression of CD40L on T cells and costimulatory activity on APC would prevent activation of unwanted bystander T cells. The coupling of activation of the APC primed with the cognate antigen to the activation of the T-cell specific for that antigen in this model provides an additional regulatory step in the initiation of the immune response. This also suggests that a limited number of T cells/APC will be activated, both of which will be specific in nature. This additional step may be important for safeguarding against an autoimmune response. In addition, the fact that CD40L uniquely seems to play this role suggests that selective immunotherapies to treat autoimmune disease and prevent graft rejection can be targeted on this molecule. On the other hand, CD40-directed approaches to up-regulate costimulatory activity on APC could be developed to fight tumor growth, contain infections and treat immunodeficiencies.
Abatacept↗
Use of recombinant antigens of Borrelia burgdorferi in serologic tests for diagnosis of lyme borreliosis.
Recombinant antigens of outer surface proteins (Osps) OspA, OspB, OspC, OspE, and OspF of Borrelia burgdorferi sensu stricto and of p41-G, an antigenic region of flagellin of this spirochete, were tested with human sera in class-specific and polyvalent enzyme-linked immunosorbent assays (ELISAs). In analyses for immunoglobulin M (IgM) antibodies, 18 (85.7%) of 21 serum samples from persons who had been diagnosed as having Lyme borreliosis on the basis of the presence of erythema migrans reacted positively in ELISAs with one or more Osp antigens or the p41-G antigen. Eleven serum samples contained antibodies to OspC antigen, and of these, six also reacted to the p41-G antigen and to one or more of the other recombinant antigens. The remaining five serum samples reacted solely to OspC (n = 4) or to OspC plus OspA and OspE without reactivity to p41-G (n = 1). In analyses for IgG antibodies, seropositivity was comparable to that of IgM analyses and was marked by predominant reactivity to p41-G, OspC, and OspF. Similarly, all 21 serum samples were positive in polyvalent and class-specific ELISAs with whole-cell B. burgdorferi. Minor cross-reactivity was noted when sera from persons who had syphilis, periodontitis or other oral infections, or rheumatoid arthritis were tested with OspC, OspE, OspF, and p41-G. With relatively high degrees of specificity, ELISAs with recombinant antigens, particularly OspC and p41-G, can help to confirm B. burgdorferi infections.
Antibodies, Bacterial↗
Novel Borrelia burgdorferi isolates from Ixodes scapularis and Ixodes dentatus ticks feeding on humans.
Seven cultures of Borrelia burgdorferi differing from strains B31 and ZS7 were identified from among 99 isolates from Ixodes scapularis ticks and from white-footed mice (Peromyscus leucopus) and 1 isolate from an Ixodes dentatus tick. Five of the six novel isolates from I. scapularis and the isolate from I. dentatus were from ticks feeding on humans. The six isolates from I. scapularis lacked OspA and OspB, four possessed an OspD band, and two reacted with an anti-OspC monoclonal antibody. Restriction fragment length polymorphisms of HindIII-digested DNAs from six OspA-negative isolates did not hybridize with radiolabeled ospA or LA88 DNA, and only isolate 46047 hybridized with the pG gene. Fragments similar to those recorded for the standard B. burgdorferi sensu stricto strains B31 and ZS7 were obtained with the fla and the HSP70 genes. Pulsed-field gel electrophoresis patterns of DNA digested with MluI included the specific B. burgdorferi sensu stricto band at 135 kbp for the five OspA-negative isolates from I. scapularis ticks. The six novel isolates apparently lack the 55-kbp plasmid encoding OspA. The pG-containing plasmid may be missing from all but isolate 46047. The isolate from the I. dentatus tick was similar to previous isolates from I. dentatus ticks feeding on rabbits. None of the isolates could be recovered from inoculated C3H/HeNCrlBR or white-footed mice. All isolates reacted with sera from humans with early or late Lyme disease. Our studies demonstrate that these borreliae occur in ticks feeding on humans, and therefore, at least some humans in the northeastern United States are likely being exposed to borreliae other than the classic B31-type strains that have thus far been isolated from humans.
Animals↗
CD40 ligand-deficient mice generate a normal primary cytotoxic T-lymphocyte response but a defective humoral response to a viral infection.
CD40 ligand is expressed on activated T cells and interacts with CD40 on B cells and monocytes. It is not known what role CD40 ligand plays in the generation of immune responses to viral infection. To address this issue, we examined virus-specific T- and B-cell responses in CD40 ligand-deficient (CD40L-/-) mice following infection with lymphocytic choriomeningitis virus (LCMV). We found that primary anti-LCMV specific antibody responses were severely impaired in CD40L-/- mice, with the defect being most striking for antibody of the immunoglobulin G1 (IgG1) isotype. Interestingly, low levels of LCMV-specific antibodies of the IgG2a, IgG2b, and IgG3 isotypes were made in the CD40L-/- mice, showing that IgG1 responses are totally dependent on CD40L but that at least some IgG2a, IgG2b, and IgG3 responses can be CD40L independent. However, unlike CD40L+/+ mice, CD40L-/- mice were unable to sustain virus-specific antibody responses and showed a gradual decline in serum antibody levels over time. The CD40L-/- mice were also deficient in the generation of memory B cells. In contrast to the severely impaired humoral responses, CD40L-/- mice generated potent virus-specific CD8+ cytotoxic T-lymphocyte responses after LCMV infection and were able to clear the virus. These results show that CD40L does not play a role in generating primary virus-specific CD8+ cytotoxic T-lymphocyte responses but does affect the primary antibody response and the generation of memory B cells.
Acute Disease↗
Transient subversion of CD40 ligand function diminishes immune responses to adenovirus vectors in mouse liver and lung tissues.
First-generation adenovirus vectors will have limited application in gene therapy for chronic diseases because of destructive host immune responses. Important immune effectors include CD8+ T cells, which mediate target cell destruction and ablate transgene expression, and B cells, which produce neutralizing antibodies that block effective readministration of vector. Previous studies indicated that activation of CD4+ T cells by virus capsid proteins is necessary for full realization of effector function of CD8+ T cells and B cells. In this paper, we present a strategy for preventing CD4+ T-cell activation by an adenovirus vector delivered to mouse liver and lung tissues which is based on interfering with T-cell priming via CD40 ligand-CD40 interactions. Adenovirus transgene expression was stabilized in mice genetically deficient in CD40 ligand (CD40L), and neutralizing antibody to adenovirus did not develop, allowing efficient readministration of vector. A transient blockade of T-cell activation with an antibody to CD40L infused into the animal at the time of adenovirus vector-mediated gene transfer led to stabilization of transgene expression and diminished production of neutralizing antibody, allowing readministration of vector. In vitro T-cell assays suggested that a block in the primary activation of CD4+ T cells was responsible for the lack of B-cell- and cytotoxic-T-cell-dependent responses. This suggests a strategy for improving the potential of adenovirus vectors based on administration of an antibody to CD40L at the time of vector administration.
Actins↗
Regulation of AP-1 and NFAT transcription factors during thymic selection of T cells.
The ability of thymocytes to express cytokine genes changes during the different stages of thymic development. Although CD4- CD8- thymocytes are able to produce a wide spectrum of cytokines in response to a T-cell receptor (TcR)-independent stimulus, as they approach the double-positive (DP) CD4+ CD8+ stage, they lose the ability to produce cytokine. After the DP stage, thymocytes become single-positive CD4+ or CD8+ thymocytes which reacquire the ability to secrete cytokines. In an attempt to understand the molecular basis of this specific regulatin, we use AP-1-luciferase and newly generated NFAT-luciferase transgenic mice to analyze the transcriptional and DNA-binding activities of these two transcription factors that are involved in the regulation of cytokine gene expression. Here, we show that both AP-1 and NFAT transcriptional activities are not inducible in the majority of DP cells but that during the differentiation of DP cells to the mature single-positive stage, thymocytes regain this inducibility. Subpopulation analysis demonstrates that this inducibility is reacquired at the DP stage before the down-modulation of one of the coreceptors. Indeed AP-1 inducibility, just like the ability to express the interleukin-2 gene, is reacquired during the differentiation of DP TcRlow CD69low heat-stable antigen (HSA)high thymocytes to DP TcRhigh CD69high HSAhigh cells, which is considered to be the consequence of the first signal that initiates positive selection. We therefore propose that the inability of DP thymocytes to induce AP-1 and NFAT activities is one of the causes for the lack of cytokine gene expression at this stage and that this inducibility is reacquired at the latest stage of DP differentiation as a consequence of positive selection. This could be a mechanism to prevent the activation of DP thymocytes before selection has taken place.
Animals↗
Transgenic modeling of cytokines in the investigation of pulmonary disease.
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Animals↗
Breaking immunologic ignorance to an antigenic peptide of simian virus 40 large T antigen.
Expression of antigenic proteins in the periphery may result in immunologic tolerance, immunologic ignorance, or autoimmunity. It is not clear why some Ags induce tolerance, whereas others activate Ag-reactive T cells. The physical nature of the Ag, the developmental timing of Ag expression, the priming of reactive lymphocytes, and the level of Ag expression are possible factors determining the immunologic response to extra-thymic Ags. Expression of the whole SV40 large T Ag (SV40-T) induces transformation of T antigen-expressing cells in vivo, and this phenomenon has been postulated to be the triggering event that leads to autoimmunity in some transgenic mouse models. Here we present a model in which a nononcogenic, yet antigenic, fragment of the SV40-T (SV40-Tfrag) is expressed specifically in pancreatic islet beta-cells. In contrast to whole SV40-T transgenic mice, SV40-Tfrag mice that are also transgenic for a TCR specific for the SV40-T are ignorant of Ag in vivo. They do not respond in vivo to the tissue-specific SV40-Tfrag Ag even after priming, but are fully responsive in vitro. This immunologic ignorance cannot be broken after activated SV40-T reactive T cells are transferred into sublethally irradiated mice expressing the islet-specific SV40-Tfrag. However, similar adoptive transfer experiments in mice co-expressing B7-1 and SV40-Tfrag on islet cells specifically lead to Ag recognition and diabetes. This demonstrates that in some circumstances the presence of (primed) reactive T cells is not sufficient to break tolerance; rather, costimulation is additionally required to elicit an autoimmune response. This also suggests that SV40-T-induced cellular transformation is important for the autoimmune response directed against SV40-T in other tissue-specific transgenic models.
Animals↗
An ospA frame shift, identified from DNA in Lyme arthritis synovial fluid, results in an outer surface protein A that does not bind protective antibodies.
Passive immunization with murine or human Abs to outer surface protein A (OspA) can protect mice against Borrelia burgdorferi, but OspA Abs elicited during natural infection in mice or humans are unable to clear the spirochete from the infected host. To examine Ab binding by OspA during the course of human infection, we amplified the operon encoding full-length ospA and ospB from synovial fluids of a patient with chronic Lyme arthritis, the first such recoveries from human material, at four separate time points over 4.5 mo, and expressed OspA in Escherichia coli. OspA mAbs that passively protected mice from infection did not bind one of the expressed OspAs, because of a deletion in ospA that resulted in a frame shift and premature stop codon near the carboxyl terminus. However, expressed OspA from a later synovial fluid sample did not contain this deletion. Thus, although altered forms of OspA, which potentially can influence host immune effectiveness, do occur in the human host, they cannot be the only factors responsible for microbial persistence.
Adolescent↗
Impairment of antigen-specific T-cell priming in mice lacking CD40 ligand.
Lack of functional expression of CD40 ligand (CD40L) on T cells results in hyper-IgM syndrome (HIGM1), a human immunodeficiency associated with a severely impaired humoral immune response that is consistent with defects in B-cell responses. Patients also succumb to recurrent opportunistic infections such as Pneumocystis carinii and Cryptosporidial diarrhoea, suggesting that T-cell functions are also compromised in these individuals, but so far this has not been explained. We have previously shown that mice deficient for CD40L, like HIGM1 patients, show grossly abnormal humoral responses. Here we report that CD40L-deficient mice are defective in antigen-specific T-cell responses. Adoptively transferred antigen-specific CD4+ T cells lacking CD40L failed to expand upon antigen challenge of the recipients, showing that expression of CD40L on T cells is required for in vivo priming of CD4+ T cells and therefore for the initiation of specific T-cell immune responses.
Animals↗
B7 costimulation is necessary for the activation of the lytic function in cytotoxic T lymphocyte precursors.
B7-CD28 costimulation is essential for the activation of CD4+ T helper cells, mainly by regulating IL-2 and other cytokine production. The requirement for this costimulatory pathway in the activation of CD8+ T cells, however, is still poorly understood. Here we analyzed the role of B7-CD28 costimulation in the differentiation of Ag-specific CTL precursor. We found that the activation of not only IL-2 production but also cytotoxic function in CD8 T cells requires B7 costimulation. The costimulatory signal, which cannot be replaced by exogenous IL-2, is directly implicated in the activation of the lytic machinery in CD8 T cells. Moreover, B7-CD28 costimulation appears to play a critical role in the accumulation of mRNA encoding at least one of the granzymes required for cytolytic function, granzyme B or CTLA-1. The production of IFN-gamma by CD8 T cells, however, does not appear to require costimulation.
Abatacept↗
Rapid induction of a novel costimulatory activity on B cells by CD40 ligand.
BACKGROUND: T cells and B cells communicate by direct cell--cell interaction that is crucial to the functioning of the immune system. It is well established that the interaction between B-cell-expressed CD40 and T-cell-expressed CD40 ligand (CD40L) is critical for T-cell-dependent antibody responses, but the role of this interaction in T-cell responses is less clear. In this study, we have used mice with targeted mutations in the genes encoding CD40L or CD28 to investigate how the CD40-CD40L interaction induces on B cells a costimulatory activity that acts in addition to antigen to trigger T-cell growth. RESULTS: We show that T cells from Cd40L-deficient mice induce a substantially reduced costimulatory activity on B cells compared to wild-type T cells, particularly at early time points. Surprisingly, T cells, from CD40L-deficient mice induce similar levels of B7-1 and B7-2 as do wild-type T cells. We further show that the CD40L-mediated induction of costimulatory activity precedes the induction of B7-1, B7-2 and the heat-stable antigen (HSA). CD4 T cells isolated from the CD28-deficient mice can receive costimulatory activity from CD40L-induced B cells, demonstrating that the induced molecules can costimulate T cells by a CD28-independent mechanism. We have generated a novel monoclonal antibody that inhibits the CD40L-induced costimulatory activity. Expression of the epitope detected by this monoclonal antibody correlates with the induction of the costimulatory activity, and the molecule recognized by the monoclonal antibody is a single chain of around 85 kDa, distinct from B7-1, B7-2, ICAM-1, ICAM-2, ICAM-3, HSA CD5, integrin and 4-1BB ligand. CONCLUSIONS: Our results demonstrate that CD40L is both necessary and sufficient for rapid, T-cell-mediated induction of costimulatory activity on B cells. This costimulatory activity is distinct from B7-1 and B7-2, and is independent of CD28.
Animals↗
An essential role for gp39, the ligand for CD40, in thymic selection.
The interactions between CD40 on B cells and its ligand gp39 on activated T helper cells are known to be essential for the development of thymus-dependent humoral immunity. However, CD40 is also functionally expressed on thymic epithelial cells and dendritic cells, suggesting that gp39-CD40 interactions may also play a role in thymic education, the process by which self-reactive cells are deleted from the T cell repertoire. Six systems of negative selection were studied for their reliance on gp39-CD40 interactions to mediate negative selection. In all cases, when the antigen/superantigen was endogenously expressed (in contrast to exogenously administered), negative selection was blocked by loss of gp39 function. Specifically, blockade of gp39-CD40 interactions prevented the deletion of thymocytes expressing V beta 3, V beta 11, and V beta 12, specificities normally deleted in BALB/c mice because of the endogenous expression of minor lymphocyte-stimulating determinants. Independent verification of a role of gp39 in negative selection was provided by studies in gp39-deficient mice where alterations in T cell receptor (TCR) V beta expression were also observed. Studies were also performed in the AND TCR transgenic (Tg) mice, which bear the V alpha 11, V beta 3 TCR and recognize both pigeon cytochrome c (PCC)/IEk and H-2As. Neonatal administration of anti-gp39 to AND TCR Tg mice that endogenously express H-2As or endogenously produce PCC prevented the deletion of TCR Tg T cells. In contrast, deletion mediated by high-dose PCC peptide antigen (administered exogenously) in AND TCR mice was unaltered by administration of anti-gp39. In addition, deletion by Staphylococcus enterotoxin B in conventional mice was also unaffected by anti-gp39 administration. gp39 expression was induced on thymocytes by mitogens or by antigen on TCR Tg thymocytes. Immunohistochemical analysis of B7-2 expression in the thymus indicated that, in the absence of gp39, B7-2 expression was substantially reduced. Taken together, these data suggest that gp39 may influence negative selection through the regulation of costimulatory molecule expression. Moreover, the data support the hypothesis that, for negative selection to some endogenously produced antigens, negative selection may be dependent on TCR engagement and costimulation.
Amino Acid Sequence↗
T-cell activation. Two for T.
T-cell activation requires costimulation in addition to the antigen-specific signal. Recent results suggest that distinct costimulatory molecules can specifically activate different types of T-cell response.
Animals↗