IFN-gamma promotes Fas ligand- and perforin-mediated liver cell destruction by cytotoxic CD8 T cells

To study liver cell damage by CTL, CD8 T cells from P14 TCR transgenic (tg) mice specific for the gp33 epitope of lymphocytic choriomeningitis virus with either deficiency in IFN-gamma (P14.IFN-gamma(null)), functional Fas ligand (P14.gld), or perforin (P14.PKO) were transferred into H8 tg mice ubiquitously expressing gp33 Ag. Treatment of H8 recipient mice with agonistic anti-CD40 Abs induced vigorous expansion of the transferred P14 T cells and led to liver cell destruction determined by increase of glutamate dehydrogenase serum levels and induction of caspase-3 in hepatocytes. Liver injury was mediated by the Fas/Fas ligand (FasL) pathway and by perforin, because P14.gld and P14.PKO T cells failed to induce increased glutamate dehydrogenase levels despite strong in vivo proliferation. In addition, H8 tg mice lacking Fas were resistant to the pathogenic effect of P14 T cells. Besides FasL and perforin, IFN-gamma was also required for liver cell damage, because P14.IFN-gamma(null) T cells adoptively transferred into H8 mice failed to induce disease. Moreover, Fas expression on hepatocytes from H8 recipient mice was increased after transfer of wild-type compared with P14.IFN-gamma(null) T cells, and wild-type P14 T cells expressed higher levels of FasL than P14 T cells lacking IFN-gamma. Thus, our data suggest that IFN-gamma released by activated CD8 T cells upon Ag contact facilitates liver cell destruction.     

CD40 ligation in vivo induces bystander proliferation of memory phenotype CD8 T cells

Injection of agonistic anti-CD40 Abs into mice has been shown to amplify weak CD8 T cell responses to poorly immunogenic compounds and to convert T cell tolerance to T cell priming. In this study we demonstrate that anti-CD40 treatment of C57BL/6 mice, without Ag delivery, led to a marked increase in the number of memory phenotype CD4 and CD8 T cells. Adoptive transfer experiments using CD40-deficient hosts further revealed that the proliferative response of memory T cells, induced by systemic CD40 signaling, was dependent on CD40 expression of host APCs. CD40 ligation in vivo induced vigorous cell division of both memory phenotype and bona fide virus-specific memory CD8 T cells in a partially IL-15-dependent manner. However, only memory phenotype, but not Ag-experienced memory CD8 T cells increased in cell number after anti-CD40 treatment in vivo. Taken together our data show that activation of APC via CD40 induces a marked bystander proliferation of memory phenotype T cells. In addition, we demonstrate that bona fide Ag-experienced memory CD8 T cells respond differently to anti-CD40-induced signals than memory phenotype CD8 T cells.     

Cutting edge: in vitro-generated tolerogenic APC induce CD8+ T regulatory cells that can suppress ongoing experimental autoimmune encephalomyelitis

APC exposed to TGFbeta2 and Ag (tolerogenic APC) promote peripheral Ag-specific tolerance via the induction of CD8(+) T regulatory cells capable of suppressing Th1 and Th2 immunity. We postulated that tolerogenic APC might reinstate tolerance toward self-neuronal Ags and ameliorate ongoing experimental autoimmune encephalomyelitis (EAE). Seven days after immunization with myelin basic protein (MBP), mice received MBP-specific tolerogenic APC, and EAE was evaluated clinically. To test for the presence and the phenotype of T regulatory cells, CD4 and/or CD8 T cells from tolerogenic APC-treated mice were transferred to naive mice before their immunization with MBP. The MBP-specific tolerogenic APC decreased both the severity and incidence of ongoing EAE. Tolerance to self-neuronal Ags was induced in naive recipient mice via adoptive transfer of CD8(+), but not CD4(+) T cells. Rational use of in vitro-generated tolerogenic APC may lead to novel therapy for autoimmune disease.     

Lack of effector cell function and altered tetramer binding of tumor-infiltrating lymphocytes

Tumor-specific CD8 T cell responses to MCA102 fibrosarcoma cells expressing the cytotoxic T cell epitope gp33 from lymphocytic choriomeningitis virus were studied. MCA102(gp33) tumors grew progressively in C57BL/6 mice, despite induction of peripheral gp33-tetramer(+) T cells that were capable of mediating antiviral protection, specific cell rejection, and concomitant tumor immunity. MCA102(gp33) tumors were infiltrated with a high number ( approximately 20%) of CD11b(+)CD11c(-) macrophage-phenotype cells that were able to cross-present the gp33 epitope to T cells. Tumor-infiltrating CD8 T cells exhibited a highly activated phenotype but lacked effector cell function. Strikingly, a significant portion of tumor-infiltrating lymphocytes expressed TCRs specific for gp33 but bound MHC tetramers only after cell purification and a 24-h resting period in vitro. The phenomenon of "tetramer-negative T cells" was not restricted to tumor-infiltrating lymphocytes from MCA102(gp33) tumors, but was also observed when Ag-specific T cells derived from an environment with high Ag load were analyzed ex vivo. Thus, using a novel tumor model, allowing us to trace tumor-specific T cells at the single cell level in vivo, we demonstrate that the tumor microenvironment is able to alter the functional activity of T cells infiltrating the tumor mass.     

In vivo ligation of CD40 enhances priming against the endogenous tumor antigen and promotes CD8+ T cell effector function in SV40 T antigen transgenic mice

The ability to initiate and sustain CD8(+) T cell responses to tumors in vivo is hindered by the development of peripheral T cell tolerance against tumor-associated Ags. Approaches that counter the onset of T cell tolerance may preserve a pool of potentially tumor-reactive CD8(+) T cells. Administration of agonist Ab to the CD40 molecule, expressed on APCs, can enhance immunization approaches targeting T lymphocytes in an otherwise tolerance-prone environment. In this report, the effects of anti-CD40 administration on priming of naive CD8(+) T cells against an endogenous tumor Ag were investigated. Line 501 mice express the SV40 large T Ag oncoprotein as a transgene from the alpha-amylase promoter, resulting in the development of peripheral CD8(+) T cell tolerance to the H-2-D(b)-restricted immunodominant epitope I of T Ag by 6 mo of age, before the appearance of osteosarcomas. We demonstrate that naive epitope I-specific TCR transgenic (TCR-I) T cells undergo peripheral tolerance following adoptive transfer into 6-mo-old 501 mice. In contrast, administration of agonistic anti-CD40 Ab led to increased expansion of TCR-I T cells in 501 mice, the acquisition of effector function by TCR-I T cells and the establishment of T cell memory. Importantly, this enhanced priming effect of anti-CD40 administration did not require immunization and was effective even if administered after naive TCR-I T cells had encountered the endogenous T Ag. Thus, anti-CD40 administration can block the onset of peripheral tolerance and enhance the recruitment of functionally competent effector T cells toward an endogenous tumor Ag.     

CD28, IL-2-independent costimulatory pathways for CD8 T lymphocyte activation.

We investigate, here, the mechanism of the costimulatory signals for CD8 T cell activation and confirm that costimulation signals via CD28 do not appear to be required to initiate proliferation, but provide survival signals for CD8 T cells activated by TCR ligation. We show also that IL-6 and TNF-alpha can provide alternative costimulatory survival signals. IL-6 and TNF-alpha costimulate naive CD8 T cells cultured on plate-bound anti-CD3 in the absence of CD28 ligation. They act directly on sorted CD8-positive T cells. They also costimulate naive CD8 T cells from Rag-2-deficient mice, bearing transgenic TCRs for HY, which lack memory cells, a potential source of IL-2 secretion upon activation. IL-6 and TNF-alpha provide costimulation to naive CD8 T cells from CD28, IL-2, or IL-2Ralpha-deficient mice, and thus function in the absence of the B7-CD28 and IL-2 costimulatory pathways. The CD8 T cell generated via the anti-CD3 plus IL-6 and TNF-alpha pathway have effector function in that they express strong cytolytic activity on Ag-specific targets. They secrete only very small amounts of any of the cytokines tested upon restimulation with peptide-loaded APC. The ability of the naive CD8 T cells to respond to TCR ligation and costimulatory signals from IL-6 and TNF-alpha provides a novel pathway that can substitute for signals from CD4 helper cells or professional APC. This may be significant in the response to viral Ags, which can be potentially expressed on the surface of any class I MHC-expressing cell.     

Role of CD4 T cell help and costimulation in CD8 T cell responses during Listeria monocytogenes infection

CD4 T cells are known to assist the CD8 T cell response by activating APC via CD40-CD40 ligand (L) interactions. However, recent data have shown that bacterial products can directly activate APC through Toll-like receptors, resulting in up-regulation of costimulatory molecules necessary for the efficient priming of naive T cells. It remains unclear what role CD4 T cell help and various costimulation pathways play in the development of CD8 T cell responses during bacterial infection. In this study, we examined these questions using an intracellular bacterium, Listeria monocytogenes, as a model of infection. In CD4 T cell-depleted, CD4(-/-), and MHC class II(-/-) mice, L. monocytogenes infection induced CD8 T cell activation and primed epitope-specific CD8 T cells to levels commensurate with those in normal C57BL/6 mice. Furthermore, these epitope-specific CD8 T cells established long-term memory in CD4(-/-) mice that was capable of mounting a protective recall response. In vitro analysis showed that L. monocytogenes directly stimulated the activation and maturation of murine dendritic cells. The CD8 T cell response to L. monocytogenes was normal in CD40L(-/-) mice but defective in CD28(-/-) and CD137L(-/-) mice. These data show that in situations where infectious agents or immunogens can directly activate APC, CD8 T cell responses are less dependent on CD4 T cell help via the CD40-CD40L pathway but involve costimulation through CD137-CD137L and B7-CD28 interactions.     

CD40 engagement enhances antigen-presenting langerhans cell priming of IFN-gamma-producing CD4+ and CD8+ T cells independently of IL-12

The delivery of CD40 signaling to APCs during T cell priming enhances many T cell-mediated immune responses. Although CD40 signaling up-regulates APC production of IL-12, the impact of this increased production on T cell priming is unclear. In this study an IL-12-independent T cell-mediated immune response, contact hypersensitivity (CHS), was used to further investigate the effect of CD40 ligation on the phenotypic development of Ag-specific CD4(+) and CD8(+) T cells. Normally, sensitization for CHS responses induces hapten-specific CD4(+) T cells producing type 2 cytokines and CD8(+) T cells producing IFN-gamma. Treatment of mice with agonist anti-CD40 mAb during sensitization with the hapten 2,4-dinitrofluorobenzene resulted in CHS responses of increased magnitude and duration. These augmented responses in anti-CD40 Ab-treated mice correlated with increased numbers of hapten-specific CD4(+) and CD8(+) T cells producing IFN-gamma in the skin draining lymph nodes. Identical results were observed using IL-12(-/-) mice, indicating that CD40 ligation promotes CHS responses and development of IFN-gamma-producing CD4(+) and CD8(+) T cells in the absence of IL-12. Engagement of CD40 on hapten-presenting Langerhans cells (hpLC) up-regulated the expression of both class I and class II MHC and promoted hpLC migration into the T cell priming site. These results indicate that hpLC stimulated by CD40 ligation use a mechanism distinct from increased IL-12 production to promote Ag-specific T cell development to IFN-gamma-producing cells.     

The development of functional CD8 T cell memory after Listeria monocytogenes infection is not dependent on CD40

The immunologic requirements for generating long-lived protective CD8 T cell memory remain unclear. Memory CD8 populations generated in the absence of CD4 Th cells reportedly have functional defects, and at least a subset of CD8 T cells transiently express CD40 after activation, suggesting that direct CD4-CD8 T cell interactions through CD40 may influence the magnitude and functional quality of memory CD8 populations. To ascertain the role of CD40 in such direct T cell interactions, we investigated CD8 T cell responses in CD40-/- mice after infection with Listeria monocytogenes, an intracellular bacterium that induces APC activation and thus priming of CD8 T cells independently of CD4 Th cell help through CD40. In this study we show that memory CD8 T cells generated in CD40-deficient mice show in vivo cytotoxicity and cytokine production equivalent to CD8 memory T cells from wild-type mice. Upon secondary Listeria infection, CD40-/- memory CD8 T cells expand to greater numbers than seen in wild-type mice. These results indicate that CD40 ligation on CD8 T cells, although reportedly a part of CD8 T cell memory development in an H-Y-directed response, is not needed for the development of functional memory CD8 T cell populations after Listeria infection.     

CD40 activation in vivo overcomes peptide-induced peripheral cytotoxic T-lymphocyte tolerance and augments anti-tumor vaccine efficacy.

The outcome of antigen recognition by naive CD8+ cytotoxic T lymphocytes (CTLs) in the periphery is orchestrated by CD4+ T-helper cells, and can either lead to priming or tolerization. The presence of T-helper cells favors the induction of CTL immunity, whereas the absence of T-helper cells can result in CTL tolerance. The action of T helper cells in CTL priming is mediated by CD40-CD40 ligand interactions. We demonstrate here that triggering of CD40 in vivo can considerably enhance the efficacy of peptide-based anti-tumor vaccines. The combination of a tolerogenic peptide vaccine containing a minimal essential CTL epitope with an activating antibody against CD40 converts tolerization into strong CTL priming. Moreover, CD40 ligation can provide an already protective tumor-specific peptide vaccine with the capacity to induce therapeutic CTL immunity in tumor-bearing mice. These findings indicate that the CD40-CD40 ligand pair can act as a 'switch', determining whether naive peripheral CTLs are primed or tolerized, and support the clinical use of CD40-stimulating agents as components of anti-cancer vaccines.     

OX40 costimulation synergizes with GM-CSF whole-cell vaccination to overcome established CD8+ T cell tolerance to an endogenous tumor antigen

T cell costimulation via OX40 is known to increase CD4+ T cell expansion and effector function and enhances the development of T cell memory. OX40 costimulation can also prevent, and even reverse, CD4+ T cell anergy. However, the role of OX40 in CD8+ T cell function is less well defined, particularly in the setting of immune tolerance. To determine the effects of OX40 costimulation on the induction of the host CD8+ T cell repertoire to an endogenous tumor Ag, we examined the fate of CD8+ T cells specific for the immunodominant rat HER-2/neu epitope, RNEU420-429, in FVB MMTV-neu (neu-N) mice, which express rat HER-2/neu protein in a predominantly mammary-restricted fashion. We show that the RNEU420-429-specific T cell repertoire in neu-N mice expands transiently after vaccination with a neu-targeted GM-CSF-secreting whole-cell vaccine, but quickly declines to an undetectable level. However, OX40 costimulation, when combined with GM-CSF-secreting tumor-targeted vaccination, can break established CD8+ T cell tolerance in vivo by enhancing the expansion, and prolonging the survival and effector function of CD8+ T cells specific for RNEU420-429. Moreover, we demonstrate that OX40 expression is up-regulated on both CD4+ and CD8+ T cells shortly after administration of a GM-CSF expressing vaccine. These studies highlight the increased efficacy of OX40 costimulation when combined with a GM-CSF-secreting vaccine, and define a new role for OX40 costimulation of CD8+ T cells in overcoming tolerance and boosting antitumor immunity.     

CD4 T cell-mediated alloresistance to fully MHC-mismatched allogeneic bone marrow engraftment is dependent on CD40-CD40 ligand interactions, and lasting T cell tolerance is induced by bone marrow transplantation with initial blockade of this pathway

Costimulatory blockade can be used to promote allogeneic marrow engraftment and tolerance induction, but on its own is not 100% reliable. We sought to determine whether one or the other of the CD4 or CD8 T cell subsets of the recipient was primarily responsible for resistance to allogeneic marrow engraftment in mice receiving costimulatory blockade, and to use this information to develop a more reliable, minimal conditioning regimen for induction of mixed chimerism and transplantation tolerance. We demonstrate that a single anti-CD40 ligand mAb treatment is sufficient to completely overcome CD4 cell-mediated resistance to allogeneic marrow engraftment and rapidly induce CD4 cell tolerance, but does not reliably overcome CD8 CTL-mediated alloresistance. The data suggest that costimulation, which activates alloreactive CTL, is insufficient to activate alloreactive CD4 cells when the CD40 pathway is blocked. The addition of host CD8 T cell depletion to anti-CD40 ligand treatment reliably allows the induction of mixed chimerism and donor-specific skin graft tolerance in 3 Gy-irradiated mice receiving fully MHC-mismatched bone marrow grafts. Thus, despite the existence of multiple costimulatory pathways and pathways of APC activation, our studies demonstrate an absolute dependence on CD40-mediated events for CD4 cell-mediated rejection of allogeneic marrow. Exposure to donor bone marrow allows rapid tolerization of alloreactive CD4 cells when the CD40 pathway is blocked, leading to permanent marrow engraftment and intrathymic tolerization of T cells that develop subsequently.     

Breaking of CD8+ T cell tolerance through in vivo ligation of CD40 results in inhibition of chronic graft-versus-host disease and complete donor cell engraftment

In the DBA/2 --> unirradiated (C57BL/6 x DBA/2)F(1) model of chronic graft-vs-host disease (cGVHD), donor CD4(+) T cells play a critical role in breaking host B cell tolerance, while donor CD8(+) T cells are rapidly removed and the remaining cells fall into anergy. Previously we have demonstrated that in vivo ligation of GITR (glucocorticoid-induced TNF receptor-related gene) can activate donor CD8(+) T cells, subsequently converting the disease pattern from cGVHD to an acute form. In this study, we investigated the effect of an agonistic mAb against CD40 on cGVHD. Treatment of anti-CD40 mAb inhibited the production of anti-DNA IgG1 autoantibody and the development of glomerulonephritis. The inhibition of cGVHD occurred because anti-CD40 mAb prevented donor CD8(+) T cell anergy such that subsequently activated donor CD8(+) T cells deleted host CD4(+) T cells and host B cells involved in autoantibody production. Additionally, functionally activated donor CD8(+) T cells induced full engraftment of donor hematopoietic cells and exhibited an increased graft-vs-leukemia effect. However, induction of acute GVHD by donor CD8(+) T cells seemed to be not so apparent. Further CTL analysis indicated that there were lower levels of donor CTL activity against host cells in mice that received anti-CD40 mAb, compared with mice that received anti-GITR mAb. Taken together, our results suggest that a different intensity of donor CTL activity is required for removal of host hematopoietic cells, including leukemia vs induction of acute GVHD.     

Effects of an epitope-specific CD8+ T-cell response on murine coronavirus central nervous system disease: protection from virus replication and antigen spread and selection of epitope escape mutants

Both CD4(+) and CD8(+) T cells are required for clearance of the murine coronavirus mouse hepatitis virus (MHV) during acute infection. We investigated the effects of an epitope-specific CD8(+) T-cell response on acute infection of MHV, strain A59, in the murine CNS. Mice with CD8(+) T cells specific for gp33-41 (an H-2D(b)-restricted CD8(+) T-cell epitope derived from lymphocytic choriomeningitis glycoprotein) were infected with a recombinant MHV-A59, also expressing gp33-41, as a fusion protein with enhanced green fluorescent protein (EGFP). By 5 days postinfection, these mice showed significantly (approximately 20-fold) lower titers of infectious virus in the brain compared to control mice. Furthermore mice with gp33-41-specific CD8(+) cells exhibited much reduced levels of viral antigen in the brain as measured by immunohistochemistry using an antibody directed against viral nucleocapsid. More than 90% of the viruses recovered from brain lysates of such protected mice, at 5 days postinfection, had lost the ability to express EGFP and had deletions in their genomes encompassing EGFP and gp33-41. In addition, genomes of viruses from about half the plaques that retained the EGFP gene had mutations within the gp33-41 epitope. On the other hand, gp33-41-specific cells failed to protect perforin-deficient mice from infection by the recombinant MHV expressing gp33, indicating that perforin-mediated mechanisms were needed. Virus recovered from perforin-deficient mice did not exhibit loss of EGFP expression and the gp33-41 epitope. These observations suggest that the cytotoxic T-cell response to gp33-41 exerts a strong immune pressure that quickly selects epitope escape mutants to gp33-41.     

Inefficient cross-presentation limits the CD8+ T cell response to a subdominant tumor antigen epitope

CD8(+) T lymphocytes (T(CD8)) responding to subdominant epitopes provide alternate targets for the immunotherapy of cancer, particularly when self-tolerance limits the response to immunodominant epitopes. However, the mechanisms that promote T(CD8) subdominance to tumor Ags remain obscure. We investigated the basis for the lack of priming against a subdominant tumor epitope following immunization of C57BL/6 (B6) mice with SV40 large tumor Ag (T Ag)-transformed cells. Immunization of B6 mice with wild-type T Ag-transformed cells primes T(CD8) specific for three immunodominant T Ag epitopes (epitopes I, II/III, and IV) but fails to induce T(CD8) specific for the subdominant T Ag epitope V. Using adoptively transferred T(CD8) from epitope V-specific TCR transgenic mice and immunization with T Ag-transformed cells, we demonstrate that the subdominant epitope V is weakly cross-presented relative to immunodominant epitopes derived from the same protein Ag. Priming of naive epitope V-specific TCR transgenic T(CD8) in B6 mice required cross-presentation by host APC. However, robust expansion of these T(CD8) required additional direct presentation of the subdominant epitope by T Ag-transformed cells and was only significant following immunization with T Ag-expressing cells lacking the immunodominant epitopes. These results indicate that limited cross-presentation coupled with competition by immunodominant epitope-specific T(CD8) contributes to the subdominant nature of a tumor-specific epitope. This finding has implications for vaccination strategies targeting T(CD8) responses to cancer.     

Critical role of costimulation in the activation of naive antigen-specific TCR transgenic CD8+ T cells in vitro.

The influence of costimulation on the activation of naive CD8+ T cells and thymocytes was studied in vitro using H-Y-specific TCR-transgenic mice and H-Y antigenic peptide. Using a variety of physiological APC types, the activation of naive CD8+ T cells depended strictly on costimulation, which could not be substituted by high epitope density. T cell activation is known to be regulated by the interactions between CD86/CD80 and CD28/CD152, although it remains unclear whether the B7 isoforms have distinct roles. Addition of soluble anti-CD86 Ab led to profound inhibition of T cell reactivity, further confirming the importance of costimulation in naive CD8+ T cell activation. Finally, TCR engagement in the absence of costimulation had no effect on the subsequent reactivity of peripheral naive transgenic CD8+ T cells, but induced nonresponsiveness in mature CD8+ transgenic thymocytes. Collectively, these results demonstrate the importance of costimulation for naive CD8+ T cell activation, suggest that CD80 and CD86 can mediate opposing effects, possibly due to differential interaction with CD152 and CD28, and indicate differences in the sensitivity of immature vs mature CD8+ T cells to the induction of nonresponsiveness following costimulation-deficient Ag presentation.     

Soluble antigen and CD40 triggering are sufficient to induce primary and memory cytotoxic T cells

The signals directing induction of tolerance rather than immunity are largely unknown. The CD8 T cell response to soluble Ags generally results in deletional tolerance following transient, costimulation-dependent activation. We demonstrated that CD40 signaling reversed the outcome of this response. Adoptive transfer of OVA-specific CD8 T cells followed by soluble OVA immunization resulted in induction of lytic activity and optimal clonal expansion only when CD40 was triggered via an agonistic mAb. Activation of CD8 T cells by CD40 signaling was indirect, because CD40 expression by host cells was required. CD40 signaling along with soluble Ag immunization also induced expansion of secondary lymphoid and intestinal mucosal endogenous OVA-specific CD8 T cells as detected by MHC tetramer reactivity. When CD40 activation was included, long-lived secondary lymphoid and mucosal memory CD8 cells were generated from adoptively transferred and endogenous CD8 T cells. Mucosal and peripheral CD8 memory cells exhibited constitutive Ag-specific lytic activity, with mucosal memory cells being 10-fold more lytic than splenic or lymph node memory cells. These results demonstrated that CD40 signaling during a response to a poorly immunogenic soluble Ag was necessary and sufficient for CTL and memory T cell induction.     

Break of T cell ignorance to a viral antigen in the liver induces hepatitis

To study peripheral tolerance of CD8 T cells to a classically MHC-restricted peptide Ag expressed in hepatocytes, ALB1 transgenic (tg) mice expressing the CTL epitope GP33 of the lymphocytic choriomeningitis virus glycoprotein under control of the mouse albumin promoter were generated. ALB1 mice exclusively expressed the GP33 transgene in the liver and, at a 100- to 1000-fold lower level, in the thymus. TCR-tg mice specific for the GP33 epitope were used to directly follow GP33-specific T cells in vivo. These experiments revealed that 1) thymic expression of the GP33 transgene led to incomplete central deletion of TCR-tg cells; and 2) peripheral TCR-tg cells in ALB1 mice ignored the GP33 transgene expressed in hepatocytes. Ignorance of adoptively transferred TCR-tg cells in ALB1 mice was broken by infection with lymphocytic choriomeningitis virus, leading to induction of hepatitis in ALB1, but not in control, mice. Taken together, we have established a novel model of virus-induced CD8 T cell-mediated autoimmune hepatitis in mice and demonstrate that naive CD8 T cells may ignore Ags expressed in the liver.     

Impaired CD4 T cell activation due to reliance upon B cell-mediated costimulation in nonobese diabetic (NOD) mice

Diabetes in nonobese diabetic (NOD) mice results from the activation of I-A(g7)-restricted, islet-reactive T cells. This study delineates several characteristics of NOD CD4 T cell activation, which, independent of I-A(g7), are likely to promote a dysregulated state of peripheral T cell tolerance. NOD CD4 T cell activation was found to be resistant to antigenic stimulation via the TCR complex, using the progression of cell division as a measure. The extent of NOD CD4 T cell division was highly sensitive to changes in Ag ligand density. Moreover, even upon maximal TCR complex-mediated stimulation, NOD CD4 T cell division prematurely terminated. Maximally stimulated NOD CD4 T cells failed to achieve the threshold number of division cycles required for optimal susceptibility to activation-induced death, a critical mechanism for the regulation of peripheral T cell tolerance. Importantly, these aberrant activation characteristics were not T cell-intrinsic but resulted from reliance on B cell costimulatory function in NOD mice. Costimulation delivered by nonautoimmune strain APCs normalized NOD CD4 T cell division and the extent of activation-induced death. Thus, by disrupting the progression of CD4 T cell division, polarization of APC costimulatory function to the B cell compartment could allow the persistence and activation of diabetogenic cells in NOD mice.     

Defective CD8+ T cell peripheral tolerance in nonobese diabetic mice.

Nonobese diabetic (NOD) mice develop spontaneous autoimmune diabetes that involves participation of both CD4+ and CD8+ T cells. Previous studies have demonstrated spontaneous reactivity to self-Ags within the CD4+ T cell compartment in this strain. Whether CD8+ T cells in NOD mice achieve and maintain tolerance to self-Ags has not previously been evaluated. To investigate this issue, we have assessed the extent of tolerance to a model pancreatic Ag, the hemagglutinin (HA) molecule of influenza virus, that is transgenically expressed by pancreatic islet beta cells in InsHA mice. Previous studies have demonstrated that BALB/c and B10.D2 mice that express this transgene exhibit tolerance of HA and retain only low-avidity CD8+ T cells specific for the dominant peptide epitope of HA. In this study, we present data that demonstrate a deficiency in peripheral tolerance within the CD8+ T cell repertoire of NOD-InsHA mice. CD8+ T cells can be obtained from NOD-InsHA mice that exhibit high avidity for HA, as measured by tetramer (K(d)HA) binding and dose titration analysis. Significantly, these autoreactive CD8+ T cells can cause diabetes very rapidly upon adoptive transfer into NOD-InsHA recipient mice. The data presented demonstrate a retention in the repertoire of CD8+ T cells with high avidity for islet Ags that could contribute to autoimmune diabetes in NOD mice.