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Cancer immunotherapy has seen significant success in the last decade for cancer management by enhancing endogenous cancer immunity. However, immunotherapies developed thus far have seen limited success in the majority of high-grade serous carcinoma (HGSC) ovarian cancer patients. This is largely due to the highly immunosuppressive tumour microenvironment of HGSC and late-stage identification. Thus, novel treatment interventions are needed to overcome this immunosuppression and complement existing immunotherapies. Here, we have identified through analysis of > 600 human HGSC tumours a critical role for Let-7i in modulating the tumoural immune network. Tumoural expression of Let-7i had high positive correlation with anti-cancer immune signatures in HGSC patients. Confirming this role, enforced Let-7i expression in murine HGSC tumours resulted in a significant decrease in tumour burden with a significant increase in tumour T cell numbers in tumours. In concert with the improved tumoural immunity, Let-7i treatment also significantly increased CD86 expression in antigen presenting cells (APCs) in the draining lymph nodes, indicating enhanced APC activity. Collectively, our findings highlight an important role of Let-7i in anti-tumour immunity and its potential use for inducing an anti-tumour effect in HGSC.
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MicroRNAs , Neoplasias Ovarianas , Animais , Feminino , Humanos , Camundongos , MicroRNAs/genética , Neoplasias Ovarianas/patologia , Linfócitos T/metabolismo , Microambiente TumoralRESUMO
Immunotherapies have emerged as promising strategies for cancer treatment. However, existing immunotherapies have poor activity in high-grade serous ovarian cancer (HGSC) due to the immunosuppressive tumor microenvironment and the associated low tumoral CD8+ T cell (CTL) infiltration. Through multiple lines of evidence, including integrative analyses of human HGSC tumors, we have identified miR-146a as a master regulator of CTL infiltration in HGSC. Tumoral miR-146a expression is positively correlated with anti-cancer immune signatures in human HGSC tumors, and delivery of miR-146a to tumors resulted in significant reduction in tumor growth in both ID8-p53-/- and IG10 murine HGSC models. Increasing miR-146a expression in tumors improved anti-tumor immune responses by decreasing immune suppressive neutrophils and increasing CTL infiltration. Mechanistically, miR-146a targets IL-1 receptor-associated kinase 1 and tumor necrosis factor receptor-associated factor 6 adaptor molecules of the transcription factor nuclear factor κB signaling pathway in ID8-p53-/- cells and decreases production of the downstream neutrophil chemoattractant, C-X-C motif chemokine ligand 1. In addition to HGSC, tumoral miR-146a expression also correlates strongly with CTL infiltration in other cancer types including thyroid, prostate, breast, and adrenocortical cancers. Altogether, our findings highlight the ability of miR-146a to overcome immune suppression and improve CTL infiltration in tumors.
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Type 1 diabetes (T1D) is caused by aberrant activation of autoreactive T cells specific for the islet beta cells. How islet-specific T cells evade tolerance to become effector T cells is unknown, but it is believed that an altered gut microbiota plays a role. Possible mechanisms include bystander activation of autoreactive T cells in the gut or "molecular mimicry" from cross-reactivity between gut microbiota-derived peptides and islet-derived epitopes. To investigate these mechanisms, we use two islet-specific CD8+ T cell clones and the non-obese diabetic mouse model of type 1 diabetes. Both insulin-specific G9C8 cells and IGRP-specific 8.3 cells underwent early activation and proliferation in the pancreatic draining lymph nodes but not in the Peyer's patches or mesenteric lymph nodes. Mutation of the endogenous epitope for G9C8 cells abolished their CD69 upregulation and proliferation, ruling out G9C8 cell activation by a gut microbiota derived peptide and molecular mimicry. However, previously activated islet-specific effector memory cells but not naïve cells migrated into the Peyer's patches where they increased their cytotoxic function. Oral delivery of butyrate, a microbiota derived anti-inflammatory metabolite, reduced IGRP-specific cytotoxic function. Thus, while initial activation of islet-specific CD8+ T cells occurred in the pancreatic lymph nodes, activated cells trafficked through the gut lymphoid tissues where they gained additional effector function via non-specific bystander activation influenced by the gut microbiota.
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Diabetes Mellitus Tipo 1 , Ilhotas Pancreáticas , Camundongos , Animais , Linfócitos T CD8-Positivos , Diabetes Mellitus Tipo 1/genética , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/patologia , Peptídeos/metabolismo , Linfonodos , Epitopos/metabolismoRESUMO
Type 1 diabetes is an autoimmune disease with no cure, where clinical translation of promising therapeutics has been hampered by the reproducibility crisis. Here, short-term administration of an antagonist to the receptor for advanced glycation end products (sRAGE) protected against murine diabetes at two independent research centers. Treatment with sRAGE increased regulatory T cells (Tregs) within the islets, pancreatic lymph nodes, and spleen, increasing islet insulin expression and function. Diabetes protection was abrogated by Treg depletion and shown to be dependent on antagonizing RAGE with use of knockout mice. Human Tregs treated with a RAGE ligand downregulated genes for suppression, migration, and Treg homeostasis (FOXP3, IL7R, TIGIT, JAK1, STAT3, STAT5b, CCR4). Loss of suppressive function was reversed by sRAGE, where Tregs increased proliferation and suppressed conventional T-cell division, confirming that sRAGE expands functional human Tregs. These results highlight sRAGE as an attractive treatment to prevent diabetes, showing efficacy and reproducibility at multiple research centers and in human T cells.
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Doenças Autoimunes , Diabetes Mellitus Tipo 1 , Animais , Humanos , Insulina/uso terapêutico , Camundongos , Receptor para Produtos Finais de Glicação Avançada/genética , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Reprodutibilidade dos Testes , Linfócitos T ReguladoresRESUMO
The autoimmune disease type 1 diabetes is predominantly mediated by CD8+ cytotoxic T-cell destruction of islet beta cells, of which islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP)206-214 is a dominant target antigen specificity. Previously, we found that a liposome-based antigen-specific immunotherapy encapsulating the CD4+ T-cell islet epitope 2.5mim together with the nuclear factor-κB inhibitor calcitriol induced regulatory T cells and protected from diabetes in NOD mice. Here we investigated whether the same system delivering IGRP206-214 could induce antigen-specific CD8+ T-cell-targeted immune regulation and delay diabetes. Subcutaneous administration of IGRP206-214 /calcitriol liposomes transiently activated and expanded IGRP-specific T-cell receptor transgenic 8.3 CD8+ T cells. Liposomal co-delivery of calcitriol was required to optimally suppress endogenous IGRP-specific CD8+ T-cell interferon-γ production and cytotoxicity. Concordantly, a short course of IGRP206-214 /calcitriol liposomes delayed diabetes progression and reduced insulitis. However, when IGRP206-214 /calcitriol liposomes were delivered together with 2.5mim /calcitriol liposomes, disease protection was not observed and the regulatory effect of 2.5mim /calcitriol liposomes was abrogated. Thus, tolerogenic liposomes that target either a dominant CD8+ or a CD4+ T-cell islet epitope can delay diabetes progression but combining multiple epitopes does not enhance protection.
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Diabetes Mellitus Tipo 1 , Animais , Linfócitos T CD8-Positivos , Epitopos de Linfócito T , Glucose-6-Fosfatase/metabolismo , Tolerância Imunológica , Lipossomos/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Linfócitos T ReguladoresRESUMO
Mechanisms by which advanced glycation end products (AGEs) contribute to type 1 diabetes (T1D) pathogenesis are poorly understood. Since life-long pharmacotherapy with alagebrium chloride (ALT) slows progression to experimental T1D, we hypothesized that acute ALT therapy delivered prediabetes, may be effective. However, in female, non-obese diabetic (NODShiLt) mice, ALT administered prediabetes (day 50-100) did not protect against experimental T1D. ALT did not decrease circulating AGEs or their precursors. Despite this, pancreatic ß-cell function was improved, and insulitis and pancreatic CD45.1+ cell infiltration was reduced. Lymphoid tissues were unaffected. ALT pre-treatment, prior to transfer of primed GC98 CD8+ T cell receptor transgenic T cells, reduced blood glucose concentrations and delayed diabetes, suggesting islet effects rather than immune modulation by ALT. Indeed, ALT did not reduce interferon-γ production by leukocytes from ovalbumin-pre-immunised NODShiLt mice and NODscid recipients given diabetogenic ALT treated NOD splenocytes were not protected against T1D. To elucidate ß-cell effects, NOD-derived MIN6N8 ß-cell major histocompatibility complex (MHC) Class Ia surface antigens were examined using immunopeptidomics. Overall, no major changes in the immunopeptidome were observed during the various treatments with all peptides exhibiting allele specific consensus binding motifs. As expected, longer MHC Class Ia peptides were captured bound to H-2Db than H-2Kb under all conditions. Moreover, more 10-12 mer peptides were isolated from H-2Db after AGE modified bovine serum albumin (AGE-BSA) treatment, compared with bovine serum albumin (BSA) or AGE-BSA+ALT treatment. Proteomics of MIN6N8 cells showed enrichment of processes associated with catabolism, the immune system, cell cycling and presynaptic endocytosis with AGE-BSA compared with BSA treatments. These data show that short-term ALT intervention, given prediabetes, does not arrest experimental T1D but transiently impacts ß-cell function.
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Type-2 immunity elicits tissue repair and homeostasis, however dysregulated type-2 responses cause aberrant tissue remodelling, as observed in asthma. Severe respiratory viral infections in infancy predispose to later asthma, however, the processes that mediate tissue damage-induced type-2 inflammation and the origins of airway remodelling remain ill-defined. Here, using a preclinical mouse model of viral bronchiolitis, we find that increased epithelial and mesenchymal high-mobility group box 1 (HMGB1) expression is associated with increased numbers of IL-13-producing type-2 innate lymphoid cell (ILC2s) and the expansion of the airway smooth muscle (ASM) layer. Anti-HMGB1 ablated lung ILC2 numbers and ASM growth in vivo, and inhibited ILC2-mediated ASM cell proliferation in a co-culture model. Furthermore, we identified that HMGB1/RAGE (receptor for advanced glycation endproducts) signalling mediates an ILC2-intrinsic IL-13 auto-amplification loop. In summary, therapeutic targeting of the HMGB1/RAGE signalling axis may act as a novel asthma preventative by dampening ILC2-mediated type-2 inflammation and associated ASM remodelling.
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Remodelação das Vias Aéreas/imunologia , Proteína HMGB1/imunologia , Inflamação/imunologia , Linfócitos/imunologia , Músculo Liso/imunologia , Animais , Camundongos , Músculo Liso/patologia , Receptor para Produtos Finais de Glicação Avançada/imunologiaRESUMO
A series of layered peripheral checkpoints maintain self-reactive B cells in an unresponsive state. Autoantibody production occurs when these checkpoints are breached; however, when and how this occurs is largely unknown. In particular, how self-reactive B cells are restrained during bystander inflammation in otherwise healthy individuals is poorly understood. A weakness has been the unavailability of methods capable of dissecting physiologically relevant B cell responses without the use of an engineered BCR. Resolving this will provide insights that decipher how this process goes awry during autoimmunity or could be exploited for therapy. In this study, we use a strong adjuvant to provide bystander innate and adaptive signals that promote B cell responsiveness in conjunction with newly developed B cell detection tools to study in detail the ways that peripheral tolerance mechanisms limit the expansion and function of self-reactive B cells activated under these conditions. We show that although self-reactive B cells are recruited into the germinal center, their development does not proceed, possibly because of rapid counterselection. Consequently, differentiation of plasma cells is blunted, and Ab responses are transient and devoid of affinity maturation. We propose this approach, and these tools can be more widely applied to track Ag-specific B cell responses to more disease-relevant Ags, without the need for BCR transgenic mice, in settings where tolerance pathways are compromised or have been genetically manipulated to drive stronger insights into the biology underlying B cell-mediated autoimmunity.
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Formação de Anticorpos/imunologia , Linfócitos B/imunologia , Tolerância Imunológica/imunologia , Tolerância Periférica/imunologia , Receptores de Antígenos de Linfócitos B/imunologia , Animais , Autoanticorpos/imunologia , Autoantígenos/imunologia , Autoimunidade/imunologia , Diferenciação Celular/imunologia , Feminino , Centro Germinativo/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Plasmócitos/imunologiaRESUMO
BACKGROUND AIMS: Pathological activation and collaboration of T and B cells underlies pathogenic autoantibody responses. Existing treatments for autoimmune disease cause non-specific immunosuppression, and induction of antigen-specific tolerance remains an elusive goal. Many immunotherapies aim to manipulate the T-cell component of T-B interplay, but few directly target B cells. One possible means to specifically target B cells is the transfer of gene-engineered BM that, once engrafted, gives rise to widespread specific and tolerogenic antigen expression within the hematopoietic system. METHODS: Gene-engineered bone marrow encoding ubiquitous ovalbumin expression was transferred after low-dose (300-cGy) immune-preserving irradiation. B-cell responsiveness was monitored by analyzing ovalbumin-specific antibody production after immunization with ovalbumin/complete Freund's adjuvant. Ovalbumin-specific B cells and their response to immunization were analyzed using multi-tetramer staining. When antigen-encoding bone marrow was transferred under immune-preserving conditions, cognate antigen-specific B cells were purged from the recipient's preexisting B-cell repertoire and the repertoire that arose after bone marrow transfer. RESULTS: OVA-specific B-cell deletion was apparent within the established host B-cell repertoire as well as that developing after gene-engineered bone marrow transfer. OVA-specific antibody production was substantially inhibited by transfer of OVA-encoding BM and activation of OVA-specific B cells, germinal center formation and subsequent OVA-specific plasmablast differentiation were all inhibited. Low levels of gene-engineered bone marrow chimerism were sufficient to limit antigen-specific antibody production. RESULTS: These data show that antigen-specific B cells within an established B-cell repertoire are susceptible to de novo tolerance induction, and this can be achieved by transfer of gene-engineered bone marrow. This adds further dimensions to the utility of antigen-encoding bone marrow transfer as an immunotherapeutic tool.
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Formação de Anticorpos , Antígenos/metabolismo , Antígeno de Maturação de Linfócitos B/metabolismo , Linfócitos B/imunologia , Transplante de Medula Óssea , Medula Óssea/imunologia , Depleção Linfocítica , Animais , Diferenciação Celular , Centro Germinativo/citologia , Centro Germinativo/metabolismo , Tolerância Imunológica/imunologia , Camundongos Endogâmicos C57BL , Ovalbumina/biossíntese , Ovalbumina/imunologia , Linfócitos T/imunologiaRESUMO
The type-2 inflammatory response that promotes asthma pathophysiology occurs in the absence of sufficient immunoregulation. Impaired regulatory T cell (Treg) function also predisposes to severe viral bronchiolitis in infancy, a major risk factor for asthma. Hence, we hypothesized that long-lived, aberrantly programmed Tregs causally link viral bronchiolitis with later asthma. Here we found that transient plasmacytoid dendritic cell (pDC) depletion during viral infection in early-life, which causes the expansion of aberrant Tregs, predisposes to allergen-induced or virus-induced asthma in later-life, and is associated with altered airway epithelial cell (AEC) responses and the expansion of impaired, long-lived Tregs. Critically, the adoptive transfer of aberrant Tregs (unlike healthy Tregs) to asthma-susceptible mice failed to prevent the development of viral-induced or allergen-induced asthma. Lack of protection was associated with increased airway epithelial cytoplasmic-HMGB1 (high-mobility group box 1), a pro-type-2 inflammatory alarmin, and granulocytic inflammation. Aberrant Tregs expressed lower levels of CD39, an ectonucleotidase that hydrolyzes extracellular ATP, a known inducer of alarmin release. Using cultured mouse AECs, we identify that healthy Tregs suppress allergen-induced HMGB1 translocation whereas this ability is markedly impaired in aberrant Tregs. Thus, defective Treg programming in infancy has durable consequences that underlie the association between bronchiolitis and subsequent asthma.
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Asma/etiologia , Asma/metabolismo , Bronquiolite/etiologia , Bronquiolite/metabolismo , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Alérgenos/imunologia , Animais , Asma/patologia , Biomarcadores , Bronquiolite/patologia , Citocinas/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Suscetibilidade a Doenças , Proteína HMGB1/metabolismo , Imunização , Camundongos , Transporte Proteico , Índice de Gravidade de Doença , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismoRESUMO
Actinic Keratosis (AK), Intraepidermal Carcinoma (IEC), and Squamous Cell Carcinoma (SCC) are generally considered to be advancing stages of the same disease spectrum. However, while AK often regress spontaneously, and IEC often regress in response to immune-activating treatments, SCC typically do not regress. Therefore, it is vital to define whether fundamental immunological changes occur during progression to SCC. Here we show that proinflammatory cytokine expression, chemokine expression, and immune cell infiltration density change during progression to SCC. Our findings suggest a switch from predominantly proinflammatory cytokine production to chemokine production is a key feature of progression from precancer to cancer. Together, these observations propose a model that can underpin current research and open new avenues of exploration into the clinical significance of these profiles with respect to immunotherapeutic or other treatment outcomes.
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Carcinoma de Células Escamosas/patologia , Quimiocinas/análise , Citocinas/análise , Neoplasias Cutâneas/patologia , Idoso , Idoso de 80 Anos ou mais , Carcinoma in Situ/patologia , Progressão da Doença , Feminino , Humanos , Ceratose Actínica/patologia , Masculino , Pessoa de Meia-Idade , Lesões Pré-Cancerosas/patologia , Pele/patologiaRESUMO
Current HLA-typing methods are typically designed to provide exquisitely-detailed identification of multiple HLA-alleles to satisfy the requirements for organ and bone marrow transplantation or genetic studies. Many human immunological studies, on the other hand, focus around only a small number of HLA alleles that are abundant or of relevance to specific diseases. Consequently, for such studies, many HLA typing approaches are not cost-effective and are potentially complicated, slow and not easily performed in-house. Work-flow would be streamlined by a simple, inexpensive and rapid typing method able to be performed in-house. We outline a straightforward approach that provides appropriate data for much immunological research. In a predominantly Caucasian population, flow cytometry using anti-HLA-A2, -B8 and -B7 antibodies consistently and accurately screened for samples carrying the highly-abundant HLA class I alleles HLA-A*02:01, -B*08:01 and -B*07:02 that form the focus of immunological studies. Next, we describe a straightforward and simple strategy for design and use of allele-specific PCR primers to identify, at high-resolution, alleles of interest. When combined with a simple gDNA extraction technique this provides reliable, simple and inexpensive in-house HLA typing demonstrated here for highly-abundant HLA class I alleles.
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Alelos , Primers do DNA/genética , Antígeno HLA-A2/genética , Antígeno HLA-B7/genética , Antígeno HLA-B8/genética , Teste de Histocompatibilidade , Reação em Cadeia da Polimerase , HumanosRESUMO
Patients receiving immunosuppressive drugs to prevent organ transplant rejection exhibit a greatly increased risk of developing cutaneous squamous cell carcinoma (SCC). However, not all immunosuppressive drugs confer the same risk. Randomised, controlled trials demonstrate that switching renal transplant recipients receiving calcineurin inhibitor-based therapies to mammalian target of rapamycin (mTOR) inhibitors results in a reduced incidence of de novo SSC formation, and can even result in the regression of pre-existing premalignant lesions. However, the contribution played by residual immune function in this setting is unclear. We examined the hypotheses that mTOR inhibitors promote the enhanced differentiation and function of CD8+ memory T cells in the skin. Here, we demonstrate that the long-term oral administration of rapamycin to achieve clinically-relevant whole blood drug target thresholds, creates a "low rapamycin dose" environment in the skin. While both rapamycin and the calcineurin inhibitor tacrolimus elongated the survival of OVA-expressing skin grafts, and inhibited short-term antigen-specific CD8+ T cell responses, rapamycin but not tacrolimus permitted the statistically significant infiltration of CD8+ effector memory T cells into UV-induced SCC lesions. Furthermore, rapamycin uniquely enhanced the number and function of CD8+ effector and central memory T cells in a model of long-term contact hypersensitivity provided that rapamycin was present during the antigen sensitization phase. Thus, our findings suggest that patients switched to mTOR inhibitor regimens likely experience enhanced CD8+ memory T cell function to new antigen-challenges in their skin, which could contribute to their lower risk of de novo SSC formation and regression of pre-existing premalignant lesions.
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Natural killer T (NKT) cells are prominent innate-like lymphocytes in the liver with critical roles in immune responses during infection, cancer, and autoimmunity. Interferon gamma (IFN-γ) and IL-4 are key cytokines rapidly produced by NKT cells upon recognition of glycolipid antigens presented by antigen-presenting cells (APCs). It has previously been reported that the transcriptional coactivator ß-catenin regulates NKT cell differentiation and functionally biases NKT cell responses toward IL-4, at the expense of IFN-γ production. ß-Catenin is not only a central effector of Wnt signaling but also contributes to other signaling networks. It is currently unknown whether Wnt ligands regulate NKT cell functions. We thus investigated how Wnt ligands and ß-catenin activity shape liver NKT cell functions in vivo in response to the glycolipid antigen, α-galactosylceramide (α-GalCer) using a mouse model. Pharmacologic targeting of ß-catenin activity with ICG001, as well as myeloid-specific genetic ablation of Wntless (Wls), to specifically target Wnt protein release by APCs, enhanced early IFN-γ responses. By contrast, within several hours of α-GalCer challenge, myeloid-specific Wls deficiency, as well as pharmacologic targeting of Wnt release using the small molecule inhibitor IWP-2 impaired α-GalCer-induced IFN-γ responses, independent of ß-catenin activity. These data suggest that myeloid cell-derived Wnt ligands drive early Wnt/ß-catenin signaling that curbs IFN-γ responses, but that, subsequently, Wnt ligands sustain IFN-γ expression independent of ß-catenin activity. Our analyses in ICG001-treated mice confirmed a role for ß-catenin activity in driving early IL-4 responses by liver NKT cells. However, neither pharmacologic nor genetic perturbation of Wnt production affected the IL-4 response, suggesting that IL-4 production by NKT cells in response to α-GalCer is not driven by released Wnt ligands. Collectively, these data reveal complex temporal roles of Wnt ligands and ß-catenin signaling in the regulation of liver NKT cell activation, and highlight Wnt-dependent and -independent contributions of ß-catenin to NKT cell functions.
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Interferon gama/imunologia , Células T Matadoras Naturais/imunologia , Via de Sinalização Wnt/imunologia , beta Catenina/imunologia , Animais , Benzotiazóis/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Feminino , Interleucina-4/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Camundongos , Pirimidinonas/farmacologia , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Receptores Acoplados a Proteínas G/imunologia , Via de Sinalização Wnt/efeitos dos fármacosRESUMO
Detecting naïve antigen-specific B cells can be challenging. Use of multiple, complementary tetramers with different fluorochromes enhances sensitivity and specificity allowing naïve antigen-specific B cells to be readily distinguished within a polyclonal repertoire. Activated, affinity-matured B cells, however, can be detected effectively using a single tetramer.
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Subpopulações de Linfócitos B/imunologia , Linfócitos B/imunologia , Citometria de Fluxo/métodos , Receptores de Antígenos de Linfócitos B/imunologia , Afinidade de Anticorpos , Antígenos/metabolismo , Células Cultivadas , Epitopos , Corantes Fluorescentes/metabolismo , Antígenos HLA/metabolismo , Humanos , Ativação Linfocitária , Ligação Proteica , Receptores de Antígenos de Linfócitos B/genética , Sensibilidade e EspecificidadeRESUMO
Purpose: Allogeneic bone marrow transplantation (BMT) provides curative therapy for leukemia via immunologic graft-versus-leukemia (GVL) effects. In practice, this must be balanced against life threatening pathology induced by graft-versus-host disease (GVHD). Recipient dendritic cells (DC) are thought to be important in the induction of GVL and GVHD.Experimental Design: We have utilized preclinical models of allogeneic BMT to dissect the role and modulation of recipient DCs in controlling donor T-cell-mediated GVHD and GVL.Results: We demonstrate that recipient CD8α+ DCs promote activation-induced clonal deletion of allospecific donor T cells after BMT. We compared pretransplant fms-like tyrosine kinase-3 ligand (Flt-3L) treatment to the current clinical strategy of posttransplant cyclophosphamide (PT-Cy) therapy. Our results demonstrate superior protection from GVHD with the immunomodulatory Flt-3L approach, and similar attenuation of GVL responses with both strategies. Strikingly, Flt-3L treatment permitted maintenance of the donor polyclonal T-cell pool, where PT-Cy did not.Conclusions: These data highlight pre-transplant Flt-3L therapy as a potent new therapeutic strategy to delete alloreactive T cells and prevent GVHD, which appears particularly well suited to haploidentical BMT where the control of infection and the prevention of GVHD are paramount. Clin Cancer Res; 24(7); 1604-16. ©2018 AACR.
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Antígenos CD8/imunologia , Ciclofosfamida/farmacologia , Células Dendríticas/imunologia , Doença Enxerto-Hospedeiro/imunologia , Efeito Enxerto vs Leucemia/imunologia , Proteínas de Membrana/imunologia , Linfócitos T/imunologia , Animais , Transplante de Medula Óssea/métodos , Células Dendríticas/efeitos dos fármacos , Feminino , Doença Enxerto-Hospedeiro/prevenção & controle , Efeito Enxerto vs Leucemia/efeitos dos fármacos , Leucemia/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Linfócitos T/efeitos dos fármacos , Doadores de Tecidos , Transplante Homólogo/métodosRESUMO
Respiratory allergies represent a significant disease burden worldwide affecting up to 300 million people globally. Medication and avoidance of known triggers do not address the underlying pathology. Traditional immunotherapies for allergy aim to reinstate immune homeostasis but require years of treatment and have poor long-term efficacy. Novel approaches, such as gene-engineered hematopoietic stem cell transplantation, induce profound antigen-specific tolerance in autoimmunity. Recent evidence shows this approach may also have therapeutic utility for allergy. Here, we review the mechanisms of antigen-specific tolerance and the potential of stem cell-mediated gene therapy to induce tolerance in allergic respiratory diseases.
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Terapia Genética , Transplante de Células-Tronco Hematopoéticas , Hipersensibilidade Respiratória/terapia , Animais , Antígenos/imunologia , Dessensibilização Imunológica , Humanos , Tolerância Imunológica , Memória Imunológica , Hipersensibilidade Respiratória/imunologiaRESUMO
AIMS/HYPOTHESIS: The aim of this study was to determine whether therapy with the cytokine IL-22 could be used to prevent the development of, or treat, autoimmune diabetes in the NOD mouse. METHODS: Six-week-old NOD mice were administered bi-weekly either recombinant mouse IL-22 (200 ng/g) or PBS (vehicle control) intraperitoneally until overt diabetes was diagnosed as two consecutive measurements of non-fasting blood glucose ≥ 11 mmol/l. At this time, NOD mice in the control arm were treated with LinBit insulin pellets and randomised to bi-weekly therapeutic injections of either PBS or IL-22 (200 ng/g) and followed until overt diabetes was diagnosed, as defined above. RESULTS: IL-22 therapy did not delay the onset of diabetes in comparison with the vehicle-treated mice. We did not observe an improvement in islet area, glycaemic control, beta cell residual function, endoplasmic reticulum stress, insulitis or macrophage and neutrophil infiltration as determined by non-fasting blood glucose, C-peptide and histological scoring. Therapeutic administration of IL-22 did not reduce circulating lipopolysaccharide, a marker of impaired gut mucosal integrity. CONCLUSIONS/INTERPRETATION: Our study suggests that, at this dosing regimen introduced either prior to overt diabetes or at diagnosis of diabetes, recombinant mouse IL-22 therapy cannot prevent autoimmune diabetes, or prolong the honeymoon period in the NOD mouse.
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Diabetes Mellitus Tipo 1/tratamento farmacológico , Interleucinas/uso terapêutico , Animais , Bioensaio , Diabetes Mellitus Tipo 1/imunologia , Feminino , Técnicas In Vitro , Camundongos , Camundongos Endogâmicos NOD , Interleucina 22RESUMO
Type 1 diabetes (T1D) results from autoimmune destruction of insulin-producing pancreatic ß cells. Therapies need to incorporate strategies to overcome the genetic defects that impair induction or maintenance of peripheral T-cell tolerance and contribute to disease development. We tested whether the enforced expression of an islet autoantigen in antigen-presenting cells (APC) counteracted peripheral T-cell tolerance defects in autoimmune-prone NOD mice. We observed that insulin-specific CD8+ T cells transferred to mice in which proinsulin was transgenically expressed in APCs underwent several rounds of division and the majority were deleted. Residual insulin-specific CD8+ T cells were rendered unresponsive and this was associated with TCR downregulation, loss of tetramer binding and expression of a range of co-inhibitory molecules. Notably, accumulation and effector differentiation of insulin-specific CD8+ T cells in pancreatic lymph nodes was prominent in non-transgenic recipients but blocked by transgenic proinsulin expression. This shift from T-cell priming to T-cell tolerance exemplifies the tolerogenic capacity of autoantigen expression by APC and the capacity to overcome genetic tolerance defects.
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Células Apresentadoras de Antígenos/imunologia , Autoantígenos/imunologia , Linfócitos T CD8-Positivos/imunologia , Diabetes Mellitus Tipo 1/imunologia , Ilhotas Pancreáticas/imunologia , Proinsulina/imunologia , Animais , Autoimunidade , Células Cultivadas , Humanos , Tolerância Imunológica , Ativação Linfocitária , Masculino , Camundongos , Camundongos Endogâmicos NODRESUMO
CD4+CD8+ double-positive (DP), mature, peripheral T cells are readily detectable in a variety of species and tissues. Despite a common association with autoimmune and malignant skin disorders, however, little is understood about their role or function. Herein, we show that DP T cells are readily detectable in the blood, spleen, and peripheral lymph nodes of naïve C57BL/6 mice. DP T cells were also present in Jα18-/- and CD1d-/- mice, indicating that these cells are not NK-T cells. After skin administration of CASAC adjuvant, but not Quil A adjuvant, both total DP T cells and skin-infiltrating DP T cells increased in number. We explored the possibility that DP T cells could represent aggregates between CD4+ and CD8+ single-positive T cells and found strong evidence that a large proportion of apparent DP T cells were indeed aggregates. However, the existence of true CD4+CD8+ DP T cells was confirmed by Amnis ImageStream (Millipore Sigma, Billerica, MA, USA) imaging. Multiple rounds of FACS sorting separated true DP cells from aggregates and indicated that conventional analyses may lead to â¼10-fold overestimation of DP T cell numbers. The high degree of aggregate contamination and overestimation of DP abundance using conventional analysis techniques may explain discrepancies reported in the literature for DP T cell origin, phenotype, and function.