Tolerance Of CD8 Research Articles

LPS에 의해 자극된 B 림프구에 의한 Th1 림프구 분화 억제

The lymphocyte component of the immune system is divided into B lymphocytes and T lymphocytes. B lymphocytes produce antibodies (humoral immunity) via maturation into plasma cells, and T lym-phocytes kill other cells or organisms (cellular immunity). A traditional immunological paradigm is that B lymphocyte and T lymphocyte interactions are a one-way phenomenon, with T lymphocytes helping to induce the terminal differentiation of B lymphocytes into immunoglobulin class-switched plasma cells. A deficiency of T lymphocytes was reported to result in defective B lymphocyte function. However, evidence for a reciprocal interaction between B and T lymphocytes is emerging, with B lym-phocytes influencing the differentiation and effector function of T lymphocytes. For example, B lym-phocytes have been shown to induce direct tolerance of antigen-specific CD8

Suppression of Dendritic Cell-Derived IL-12 by Endogenous Glucocorticoids Is Protective in LPS-Induced Sepsis.

Sepsis, an exaggerated systemic inflammatory response, remains a major medical challenge. Both hyperinflammation and immunosuppression are implicated as causes of morbidity and mortality. Dendritic cell (DC) loss has been observed in septic patients and in experimental sepsis models, but the role of DCs in sepsis, and the mechanisms and significance of DC loss, are poorly understood. Here, we report that mice with selective deletion of the glucocorticoid receptor (GR) in DCs (GRCD11c-cre) were highly susceptible to LPS-induced septic shock, evidenced by elevated inflammatory cytokine production, hypothermia, and mortality. Neutralizing anti-IL-12 antibodies prevented hypothermia and death, demonstrating that endogenous GC-mediated suppression of IL-12 is protective. In LPS-challenged GRCD11c-cre mice, CD8+ DCs were identified as the major source of prolonged IL-12 production, which correlated with elevations of NK cell-derived IFN-γ. In addition, the loss of GR in CD11c+ cells rescued LPS-induced loss of CD8+ DCs but not other DC subsets. Unlike wild-type animals, exposure of GRCD11c-cre mice to low-dose LPS did not induce CD8+ DC loss or tolerance to subsequent challenge with high dose, but neutralization of IL-12 restored the ability of low-dose LPS to tolerize. Therefore, endogenous glucocorticoids blunt LPS-induced inflammation and promote tolerance by suppressing DC IL-12 production.

IL-2 complex rescues CD8+ T cell tolerance for cancer immunotherapy (VAC7P.1041)

Abstract IL-2 is a well-characterized T cell growth factor that promotes proliferation and effector function of CD8+ T cells. For this reason, IL-2 has been used to boost immune responses in patients with melanoma. However, IL-2 treatment requires large potentially toxic doses of IL-2, and often only limited therapeutic efficacy is acheived. Efforts to enhance the efficacy and safety of IL-2 have recently focused on the use of cytokine/antibody complexes. These complexes demonstrate greater bioactivity in vivo relative to IL-2 alone, making them an attractive approach for improved immunotherapy. One of the major impediments to anti-tumor immunity is CD8+ T cell tolerance, where tumor-reactive T cells are either eliminated by apoptosis or rendered dysfunctional/anergic. The influence of IL-2 complex (IL-2c) on tolerant CD8+ T cells has not been explored. We utilized a mouse model of T cell tolerance to examine how IL-2c treatment impacts tolerant tumor-reactive T cells during immunotherapy for leukemia. Specifically, IL-2c was administered daily starting 3 days after T cell transfer into tumor-bearing mice. The function and persistence of transferred tumor-reactive T cells was assessed 5 days later. Treatment with IL-2c prevented apoptotic deletion of transferred CD8+ T cells and improved effector function. These results suggest that IL-2c treatment is sufficient to overcome peripheral T cell tolerance, and support further exploration of IL-2c as an immunotherapeutic for cancer.

Micro-RNA-155-mediated control of heme oxygenase 1 (HO-1) is required for restoring adaptively tolerant CD4+ T-cell function in rodents.

T cells chronically stimulated by a persistent antigen often become dysfunctional and lose effector functions and proliferative capacity. To identify the importance of micro-RNA-155 (miR-155) in this phenomenon, we analyzed mouse miR-155-deficient CD4(+) T cells in a model where the chronic exposure to a systemic antigen led to T-cell functional unresponsiveness. We found that miR-155 was required for restoring function of T cells after programmed death receptor 1 blockade. Heme oxygenase 1 (HO-1) was identified as a specific target of miR-155 and inhibition of HO-1 activity restored the expansion and tissue migration capacity of miR-155(-/-) CD4(+) T cells. Moreover, miR-155-mediated control of HO-1 expression in CD4(+) T cells was shown to sustain in vivo antigen-specific expansion and IL-2 production. Thus, our data identify HO-1 regulation as a mechanism by which miR-155 promotes T-cell-driven inflammation.

Cyclic dinucleotides (CDNs) reversed T cells tolerance in anti-tumor immunotherapy

Meeting abstracts One important barrier to cancer immunotherapy. We used cyclic dinucleotides (CDNs) and Listeria vaccine and LPS treated HA-specific CD8 tolerance B10. D2 mice. Our results demonstrated T cells IFN-gamma increased in CDNs and Listeria vaccine plus CDNs treatment groups. But CDNs

JQ 1, a Selective Bromodomain Inhibitor, Decreased the Expression of the Tolerogenic Molecule PDL1 in Antigen-Presenting Cells (APCs) and Restores the Responsiveness of Anergic CD4+ T Cells

Bromodomain and extraterminal (BET) is a protein domain that recognizes acetylated lysine residues such as those on the N-terminal tails of histones. This recognition is often a prerequisite for protein-histone association, chromatin remodeling and gene transcription. The role of BET proteins in regulating the response of inflammatory cytokine genes through translation of histone marks is poorly understood.Given that the inflammatory status of the APC is critical in determining T-cell activation versus T-cell tolerance and that epigenetic modifications of specific genes in the APC play a key role in this process, we recently determined the functional consequences of inhibiting BET in APCs.First, we evaluated the effects of JQ 1, a selective small-molecule BET bromodomain inhibitor on APC’s function and its regulation of antigen-specific CD4+ T-cells response. In vitro treatment of peritoneal elicited macrophages (PEM) or bone marrow derived dendritic cells (DCs) with increasing concentrations of JQ 1 resulted in decreased expression and protein production of the anti-inflammatory cytokine IL-10 and IL-6 in response to LPS stimulation. At the concentration used, JQ 1 did not affect the viability of treated APCs. Second, analysis of the expression of MHC class molecules and co-stimulatory molecules revealed a decreased expression of the tolerogenic PDL1 molecule in JQ 1- treated APCs as compared to untreated APCs. Third, we evaluated the ability of JQ 1 treated APCs to present cognate antigen to naïve or tolerant antigen-specific CD4+ T-cells. We found that treatment of either PEM or DC with JQ 1 enhanced their antigen-presenting capabilities leading to effective priming of naïve CD4+ T-cells confirmed by their increased production of IL-2 and IFN-gamma in response to cognate antigen. More importantly, JQ 1- treated APCs were able to restore the responsiveness of tolerant CD4+ T-cells isolated from lymphoma bearing hosts.Taken together, we have found that APCs treated with the Bromodomain specific inhibitor JQ 1 are more inflammatory, display lower expression of the immunosuppressive molecule PDL1 and more importantly, are capable of restoring the responsiveness of tolerant T-cells. Our studies therefore have unveiled a previously unknown immunological effect of BET inhibitors and have broadened their clinical scope as promising adjuvants in cancer immunotherapy. DisclosuresNo relevant conflicts of interest to declare.

Neuropilin-1 expression is induced on tolerant self-reactive CD8+ T cells but is dispensable for the tolerant phenotype.

Establishing peripheral CD8+ T cell tolerance is vital to avoid immune mediated destruction of healthy self-tissues. However, it also poses a major impediment to tumor immunity since tumors are derived from self-tissue and often induce T cell tolerance and dysfunction. Thus, understanding the mechanisms that regulate T cell tolerance versus immunity has important implications for human health. Signals received from the tissue environment largely dictate whether responding T cells become activated or tolerant. For example, induced expression and subsequent ligation of negative regulatory receptors on the surface of self-reactive CD8+ T cells are integral in the induction of tolerance. We utilized a murine model of T cell tolerance to more completely define the molecules involved in this process. We discovered that, in addition to other known regulatory receptors, tolerant self-reactive CD8+ T cells distinctly expressed the surface receptor neuropilin-1 (Nrp1). Nrp1 was highly induced in response to self-antigen, but only modestly when the same antigen was encountered under immune conditions, suggesting a possible mechanistic link to T cell tolerance. We also observed a similar Nrp1 expression profile on human tumor infiltrating CD4+ and CD8+ T cells. Despite high expression on tolerant CD8+ T cells, our studies revealed that Nrp1 had no detectable role in the tolerant phenotype. Specifically, Nrp1-deficient T cells displayed the same functional defects as wild-type self-reactive T cells, lacking in vivo cytolytic potential, IFNγ production, and antitumor responses. While reporting mostly negative data, our findings have therapeutic implications, as Nrp1 is now being targeted for human cancer therapy in clinical trials, but the precise molecular pathways and immune cells being engaged during treatment remain incompletely defined.

γδT cells drive myeloid-derived suppressor cell-mediated CD8+ T cell exhaustion in hepatitis B virus-induced immunotolerance.

The mechanisms of liver hepatitis B virus (HBV)-induced systemic immune tolerance are still elusive, and the role of γδT cells has not yet been described. We examined the function of γδT cells in HBV-carrier mice--immunocompetent mice with plasmid-mediated persistent HBV expression in the liver. In this study, we found that γδT cell deficiency led to a break in HBV-induced tolerance and subsequent recovery of hepatic HBV-specific CD8(+) T cells. Of interest, IL-17(-/-) mice phenocopied TCRδ(-/-) mice in terms of losing HBV persistence, and adoptive transfer of γδT cells restored HBV-persistent expression in TCRδ(-/-) mice. We further observed that hepatic CD11b(+)Gr1(+) myeloid-derived suppressor cells (MDSCs) play a major role in this mechanism, as they were significantly reduced in both HBV-carrier TCRδ(-/-) and IL-17(-/-) mice. MDSC numbers also recovered after adoptive transfer of γδT cells, particularly Vγ4(+) T cells. Furthermore, anti-Gr1-mediated MDSC depletion in HBV-carrier mice accelerated HBV elimination from the host, whereas MDSCs transferred to γδT cell-deficient mice restored HBV-induced tolerance. Accordingly, inhibition of MDSCs by the arginase-1 inhibitor norNOHA enhanced the number of HBV-specific CD8(+) T cells and promoted HBV clearance. We also observed enhanced CD8(+) T cell number with a notable decline of MDSCs in TCRδ(-/-) mice compared with wild-type mice during the recombinant adeno-associated virus/HBV1.3 virus infection. Importantly, HBV-carrier TCRδ(-/-) mice not only exhibited increased anti-HBV CD8(+) T cells but also markedly reduced MDSCs. Overall, the current study reveals that γδT cells play a previously unrecognized regulatory role in liver tolerance by mobilizing MDSC infiltration to the liver, leading to MDSC-mediated CD8(+) T cell exhaustion.

Neonatal Tolerance: Donor B Cells Home to Host Lymphoid Organs, Interact With Apoptotic Cells, Produce IL-10 and Prolong Heart Allograft Survival.

Introduction: Newborn mice injected with a mixture of semi-allogeneic (F1) bone marrow (BMC) and spleen cells (SC) accept donor-type transplants as adults. B cells are abundant in both BMC and SC and have been reported to tolerize CD8+ T cells. To determine what role, if any, B cells in the tolerizing inoculum play in this classic tolerance model, we studied the effect of purified B cells injected into newborn mice. Methods: F1-B cells and allogeneic-B cells (allo-B) were purified from spleen using StemCell Technologies EasySep® CD19 positive selection kits. C3H/He (C3H; H-2k) neonates were injected within 24 hours of birth with 106 purified B cells, GFP-expressing (C57BL/6 [B6; H-2b] or C3H X B6 [H-2k/b]) or CFSE-labelled (BALB/c [BALB; H-2d] or C3H X BALB [H-2k/d]). Injected cells were tracked by whole body/organ imaging; their interactions and fates were studied with high-resolution optical-sectioning microscopy. Impact on heart allograft survival was also determined. Results: By whole organ imaging (day 3), purified F1-B cells were detected in primary and secondary lymphoid organs (spleen, lymph node and bone). At day 6, injected B cells in spleen were positioned in follicular regions. F1- and allo-B cells both interacted with host CD8+ T cells; F1-B cells remained intact while allo-B cells fragmented and were taken up by host DC. Survival of F1-B cells suggested they induced tolerance in host CD8+ T cells. Consistent with that, F1-B cells also interacted with host apoptotic cells in spleen. Apoptotic cells have been reported to induce B cells to produce IL-10 and in turn convert T cells into Tr1-type regulatory cells. IL-10-producing F1-B cells were observed interacting with host T cells in day 3 spleens. To determine if other mechanisms played a role in prolonging F1-B cell survival we examined host Foxp3 regulatory and apoptotic T cells; in both cases events were few. Purified F1-B cells prolonged survival of donor-type hearts to 100 days in 6 of 7 transplanted mice. Beat scores were low, however, and one day-105 beating heart showed infiltrating host macrophages and T cells including CD8+ T cells. Conclusion: Neonatally-injected purified F1-B cells actively regulate the neonatal immune system and prolong survival of donor-type heart grafts transplanted as adults. By themselves, however, F1-B cells do not induce a robust tolerance suggesting additional F1 cell types are required for the process.

PD-1 and CTLA-4 signals are not important for the maintenance of tolerant self-reactive CD8 T cells in the small intestine (MUC8P.808)

Abstract Tolerance is an important mechanism by which the immune system distinguishes between self and non-self. Peripheral T cell tolerance is critical for the control of self-reactive cells that escape negative selection. Anergy is one mechanism that controls self-reactive cells in the periphery and prevents autoimmunity. Using a system that allows for the study self-specific CD8 T cell responses to a known intestinal auto-antigen, we determined that mucosal antigen signaling can lead to a long-term, stable population of cells in the intestine that have a classic anergic phenotype. These cells do not produce effector cytokines or induce intestinal damage. Inhibitory molecules, like PD-1 and CTLA-4, are believed to be important for both the induction and maintenance of T cell tolerance. We show that the induction phase of CD8 T cell tolerance to self was highly sensitive to control through both PD-1 and CTLA-4. However, once tolerance was established, the maintenance of tolerant CD8 T cells was independent of PD-1 or CTLA-4 signaling. Overall, this indicates that distinct mechanisms are responsible for the induction and maintenance of tolerant T cells. In addition, there may be many tolerant T cells resistant to immunotherapeutics targeting these pathways. Determining which subsets of self-reactive cells are susceptible to modulation through these pathways is important for understanding basic mechanisms tolerance and autoimmunity and for improving cancer therapeutics.

Transcriptional Repression of the Transforming Growth Factor β (TGF-β) Pseudoreceptor BMP and Activin Membrane-bound Inhibitor (BAMBI) by Nuclear Factor κB (NF-κB) p50 Enhances TGF-β Signaling in Hepatic Stellate Cells

TLR4 signaling induces down-regulation of the bone morphogenic protein (BMP) and activin membrane-bound inhibitor (BAMBI), which enhances TGF-β signaling during hepatic stellate cell (HSC) activation. We investigated the mechanism by which TLR4 signaling down-regulates BAMBI expression in HSCs and found that TLR4- and TNF-α-mediated BAMBI down-regulation is dependent on regulation of BAMBI promoter activity through the interaction with NF-κBp50 and HDAC1 in HSCs. Bambi was predominantly expressed in HSCs, at high levels in quiescent HSCs but at low levels in in vivo-activated and LPS-stimulated HSCs. In human HSCs, BAMBI expression was down-regulated in response to LPS and TNF-α. A BAMBI reporter assay demonstrated that the regulatory element to repress BAMBI transcription is located between 3384 and 1560 bp upstream from the transcription start site. LPS stimulation down-regulated BAMBI expression in cells with NF-κBp65 knockdown. However, it failed to down-regulate BAMBI in cells with inactivation of NF-κB or NF-κBp50 silencing, indicating that NF-κBp50 is a factor for BAMBI down-regulation. ChIP analysis revealed that LPS and TNF-α induced binding of the NF-κBp50/p50 homodimer to the BAMBI promoter region. We also found that HDAC1 is bound to this region as part of the NF-κBp50-HDAC1 complex, repressing transcriptional activity of the BAMBI promoter. Finally, we confirmed that LPS does not repress BAMBI reporter activity using a BAMBI reporter construct with a mutation at 3166 bp upstream of the coding region. In summary, our study demonstrates that LPS- and TNF-α-induced NF-κBp50-HDAC1 interaction represses BAMBI transcriptional activity, which contributes to TLR4-mediated enhancement of TGF-β signaling in HSCs during liver fibrosis.

Allospecific Rejection of MHC Class I-Deficient Bone Marrow by CD8 T Cells

Avoidance of long-term immunosuppression is a desired goal in organ transplantation. Mixed chimerism offers a promising approach to tolerance induction, and we have aimed to develop low-toxicity, nonimmunodepleting approaches to achieve this outcome. In a mouse model achieving fully MHC-mismatched allogeneic bone marrow engraftment with minimal conditioning (3 Gy total body irradiation followed by anti-CD154 and T cell–depleted allogeneic bone marrow cells), CD4 T cells in the recipient are required to promote tolerance of preexisting alloreactive recipient CD8 T cells and thereby permit chimerism induction. We now demonstrate that mice devoid of CD4 T cells and NK cells reject MHC Class I-deficient and Class I/Class II-deficient marrow in a CD8 T cell–dependent manner. This rejection is specific for donor alloantigens, since recipient hematopoiesis is not affected by donor marrow rejection and MHC Class I-deficient bone marrow that is syngeneic to the recipient is not rejected. Recipient CD8 T cells are activated and develop cytotoxicity against MHC Class I-deficient donor cells in association with rejection. These data implicate a novel CD8 T cell–dependent bone marrow rejection pathway, wherein recipient CD8 T cells indirectly activated by donor alloantigens promote direct killing, in a T cell receptor–independent manner, of Class I-deficient donor cells.

TLR1/2 Ligand–Stimulated Mouse Liver Endothelial Cells Secrete IL-12 and Trigger CD8+ T Cell Immunity In Vitro

Liver sinusoidal endothelial cells (LSECs) are unique organ-resident APCs capable of Ag cross-presentation and subsequent tolerization of naive CD8(+) T cells. Under certain conditions, LSECs can switch from a tolerogenic to an immunogenic state and promote the development of T cell immunity. However, little is known about the mechanisms of LSECs to induce T cell immunity. In this study, we investigated whether functional maturation of LSECs can be achieved by TLR ligand stimulation and elucidated the mechanisms involved in LSEC-induced T cell immunity. We demonstrate that pretreatment of LSECs with palmitoyl-3-cysteine-serine-lysine-4 (P3C; TLR1/2 ligand) but not poly(I:C) (TLR3 ligand) or LPS (TLR4 ligand) reverted their suppressive properties to induce T cell immunity. Importantly, P3C stimulation caused functional maturation of Ag-presenting LSECs and enabled them to activate virus-specific CD8(+) T cells. The LSEC-mediated CD8(+) T cell immunity was initiated by soluble mediators, one of which was IL-12 secreted at a low but sustained level after P3C stimulation. P3C stimulation did not induce programmed death ligand 1 expression on LSECs, thereby favoring T cell proliferation and activation instead of suppression. Our data suggest that LSECs undergo maturation exclusively in response to TLR1/2 ligand stimulation and that the immunological status of LSECs was dependent upon the balance between programmed death ligand 1 and IL-12 expression. These results have implications for our understanding of liver-specific tolerance and autoimmunity and for the development of strategies to overcome T cell tolerance in situations such as chronic viral liver infections or liver cancer.

Cytokines affecting CD4+ T regulatory cells in transplant tolerance. Interleukin-4 does not maintain alloantigen specific CD4+CD25+ Treg

IL-4 is thought to promote induction of transplantation tolerance and alloantigen-specific CD4+CD25+ T regulatory cells (Treg). This study examined the effect of IL-4 on the induction and maintenance of the CD4+ T regulatory cells (Treg) that mediate transplantation tolerance. Tolerance was induced in DA rats with PVG heterotopic cardiac allografts by a short course of cyclosporine. Naïve and tolerant lymphocytes, including the CD4+ and CD4+CD25+ T cell subsets, were assayed in mixed lymphocyte cultures with or without recombinant (r)IL-4 or other cytokines. The proliferation, cell surface and cytokine phenotype of these cells was examined, as was their capacity to adoptively transfer tolerance. rIL-4 enhanced the proliferation of naïve and tolerant lymphoid cells, including CD4+ and CD4+CD25+ T cells, but this was not alloantigen specific. Naïve or tolerant CD4+ T cells cultured with rIL-4 and donor PVG antigen effected rapid graft rejection, even though before culture tolerant CD4+ T cells transferred antigen-specific tolerance. These rIL-4 cultured CD4+ T cells had a phenotype consistent with activated CD4+CD25+FoxP3− Th2 cells. While naïve natural CD4+CD25+ T cells (nTreg) cultured with alloantigen and rIL-4 had enhanced proliferation and capacity to suppress rejection in vivo, the culture of tolerant CD4+CD25+ T cells with alloantigen and rIL-4 could not sustain their proliferation against specific donor, nor their capacity to transfer tolerance to specific donor allograft. Thus, IL-4 promotes both regulatory and effector T cells early in the immune response, but once alloimmune tolerance is established, IL-4 promoted the activation of effector cells to mediate rejection and did not support alloantigen-specific Treg that could transfer specific tolerance.

CRYAB modulates the activation of CD4+ T cells from relapsing–remitting multiple sclerosis patients

Background: Suppression of activation of pathogenic CD4+ T cells is a potential therapeutic intervention in multiple sclerosis (MS). We previously showed that a small heat shock protein, CRYAB, reduced T cell proliferation, pro-inflammatory cytokine production and clinical signs of experimental allergic encephalomyelitis, a model of MS. Objective: We assessed whether the ability of CRYAB to reduce the activation of T cells translated to the human disease. Methods: CD4+ T cells from healthy controls and volunteers with MS were activated in vitro in the presence or absence of a CRYAB peptide (residues 73–92). Parameters of activation (proliferation rate, cytokine secretion) and tolerance (anergy, activation-induced cell death, microRNAs) were evaluated. Results: The secretion of pro-inflammatory cytokines by CD4+ T cells was decreased in the presence of CRYAB in a subset of relapsing–remitting multiple sclerosis (RRMS) participants with mild disease severity while no changes were observed in healthy controls. Further, there was a correlation for higher levels of miR181a microRNA, a marker upregulated in tolerant CD8+ T cells, in CD4+ T cells of MS patients that displayed suppressed cytokine production (responders). Conclusion: CRYAB may be capable of suppressing the activation of CD4+ T cells from a subset of RRMS patients who appear to have less disability but similar age and disease duration.

Lymphatic endothelial cells induce anergy of β-galactosidase specific CD4 T cells by providing antigen to CD11c cells (P1147)

Abstract Lymphatic endothelial cells (LEC) express a variety of peripheral tissue antigens (Ags) and induce deletional tolerance of tyrosinase-specific CD8 T cells. LEC express MHC II molecules, suggesting they may tolerize CD4 T cells. We expressed β-galactosidase (β-gal) selectively in LEC using Lyve-1cre x β-galflox mice to investigate the ability of LEC to tolerize β-gal specific CD4 and CD8 T cells (Bg2 and Bg1, respectively). Using in vitro co-cultures and MHC I-/- bone marrow chimeras, we demonstrated that Bg1 CD8 T cells undergo deletional tolerance after recognizing Ag presented by MHC I molecules on LEC. Peptide-pulsed LEC induce Bg2 CD4 T cell proliferation in vitro, indicating LEC express functional MHC II. However, Bg2 cells proliferate when co-cultured with CD11c cells but not with LEC from β-gal+ mice. When Ag presentation is restricted to LEC in MHC II-/- bone marrow chimeras, Bg2 cells do not proliferate or upregulate CD69, CD25 or CD44. However, Bg2 cells do proliferate when transferred into chimeras with MHC II+ β-galneg bone marrow, demonstrating that hematopoietic cells acquire and present Ag from LEC. Bg2 cells activated in vivo by hematopoietic presentation of LEC-derived Ag do not proliferate in response to subsequent rechallenge, demonstrating that they have become anergic. Thus, LEC show a diminished ability to directly present MHC II Ags, but play an important role in CD4 tolerance by serving as a reservoir of Ag used by hematopoietic cells to induce anergy.

PD-1-mediated deletion induces tolerance of CD8 T cell response specific for H60 under helpless condition (P2225)

Abstract H60 is a dominant minor H antigen, for which specific CD8 T cell response dominates over those for other minor H antigens in B6 anti-BALB.B response. The specific CD8 T cell response for H60 is CD4 help-dependent, as burst expansion of the specific cells during the primary and memory responses occurs only when CD4 T cell activation accompanies. When the CD4 help is not provided, primary expansion was limited and, then, recall expansion of the specific CD8 T cells is impaired even when the CD4 help is provided during the memory response. This tolerance phenomenon was ascribed to intrinsic property of H60-specific CD8 T cells activated under the helpless condition. The H60-specific CD8 T cells activated under helpless condition could develop into functionally active effector cells, migrating to inflamed sites. However, they were more prone to cell death with up-regulation of PD-1 on the H60-specific CD8 T cells activated under helpless condition. Recall expansion of H60-specific CD8 T was restored in the PD-1 deficient mice that had been primed under helpless condition, suggesting the significance of negative regulation by PD-1 in the tolerance induction of H60-specific CD8 T cell response after the helpless priming. These results will provide useful information for induction of tolerance after allogeneic transplantation.

CD8+ T Cell Exhaustion During Persistent Viral Infection is Regulated Independently of the Virus-Specific T Cell Receptor

During chronic viral infections, responses by virus-specific CD8+ T cells become marginalized by the acquisition of functional defects and reduced cell numbers in a process defined as T cell exhaustion. Similarly, T cell tolerance to self-antigen is also characterized by impaired effector function and eventual deletion of self-reactive T cells. Induction of both tolerance and exhaustion involve many shared inhibitory mechanisms, thus similar therapeutic approaches have proven effective in these distinct environments. We previously demonstrated that tolerant self-reactive CD8+ T cells expressing dual-T cell receptors (i.e., dual-TCR) could be rescued by immunization through a second TCR specific for a foreign antigen. These data revealed that T cell tolerance was regulated at the level of the self-reactive TCR. Here, dual-TCR CD8+ T cells were used to examine if exhaustion during persistent viral infection could be rescued by an analogous strategy of immunization through a second TCR not involved in recognition of virus. In direct contrast to the rescue achievable in tolerant CD8+ T cells, exhausted T cells were equally impaired through both TCR. These findings suggest that exhaustion is maintained by defects downstream of the virus-specific TCR, and establish that exhaustion and tolerance are distinctly regulated states of T cell dysfunction.

Liver-Primed Memory T Cells Generated under Noninflammatory Conditions Provide Anti-infectious Immunity

Development of CD8(+) T cell (CTL) immunity or tolerance is linked to the conditions during T cell priming. Dendritic cells (DCs) matured during inflammation generate effector/memory T cells, whereas immature DCs cause T cell deletion/anergy. We identify a third outcome of T cell priming in absence of inflammation enabled by cross-presenting liver sinusoidal endothelial cells. Such priming generated memory T cells that were spared from deletion by immature DCs. Similar to central memory T cells, liver-primed T cells differentiated into effector CTLs upon antigen re-encounter on matured DCs even after prolonged absence of antigen. Their reactivation required combinatorial signaling through the TCR, CD28, and IL-12R and controlled bacterial and viral infections. Gene expression profiling identified liver-primed T cells as a distinct Neuropilin-1(+) memory population. Generation of liver-primed memory T cells may prevent pathogens that avoid DC maturation by innate immune escape from also escaping adaptive immunity through attrition of the T cell repertoire.

CD4 Cells Engineered to Express an MHC Class I Restricted TCR Can Rescue CD8 Cells Tolerized to Tumour-Associated Antigens

AbstractAbstract 952 Background:The efficacy of T cell therapies for cancer may be limited when targeting tumour-associated antigens (TAA) which are also self-antigens. Ongoing exposure to TAA on normal cells may lead to tolerance via anergy or exhaustion of antigen-specific T cells. Methods:We have designed a model of tolerance to TAA in which T cell receptor (TCR)-transduced CD8 T cells recognise pMDM2, a TAA that is also a ubiquitous self-antigen. CD8+ T cells were transduced with pMDM2-specific TCR (MDM-CD8) and transferred to sub-lethally irradiated B6 mice that express pMDM2 in the context of MHC Class I (H2-Kb). MDM-CD8 cells are detectable 4 weeks after transfer but show defective in vivo killing of target cells pulsed with MDM2 peptide. We have used this model to determine the mechanism of tolerance and to evaluate whether tolerant CD8+ T cells can be rescued by CD4 help. Results:To determine whether tolerance of MDM-CD8 cells was dependent upon recognition of cognate antigen, we transferred MDM-CD8 cells into mice of a different MHC background (BALB/c) which lack H2-Kb required for presentation of the TCR-recognised MDM2 peptide. When BALB/c MDM-CD8 cells were transferred to BALBc hosts their functions were preserved and they retained efficient antigen-specific cytolysis.To determine whether tolerance could be modified by provision of CD4+ T cell help, we co-transferred MDM-CD8 with transgenic OT-II CD4+ cells. OT-II cells were primed with dendritic cells (DCs) loaded with cognate pOVA323-339 or irrelevant peptide. When activated through their TCR, OT-II cells increased both the frequency of MDM2-specific CD8 cells and their cytotoxic functions, indicating that CD4 help can overcome CD8 tolerance to TAA.Ineffective antigen presentation to CD4 cells and lack of known MHC class II-restricted TAA are major limitations to providing CD4 help in T cell therapy for cancer. We therefore tested whether transfer of the MHC Class I-restricted MDM2 TCR into CD4 cells could provide help upon transfer to antigen-expressing hosts. Co-transfer of MDM2-TCR-transduced CD4 cells with CD8 cells improved antigen-specific killing of target cells when compared to single transfer of either TCR-transduced CD8 or CD4 cells. Conclusion:CD4 cells rendered capable of responding to an MHC class I restricted TAA by TCR transfer can rescue tolerance developing in a CD8 population with the same specificity. This is potentially a novel way to circumvent defective immune responses arising in adoptively transferred effector cells due to prolonged exposure to cognate antigen on normal host cells. Disclosures:Stauss:Cell Medica: Scientific Advisor Other.