Activation Of Autoreactive T-cells Research Articles

High-dimensional immune profiling using mass cytometry reveals IL-17A-producing γδ T cells as biomarkers in patients with T-cell-activated idiopathic severe aplastic anemia

Severe aplastic anemia (SAA) is a bone marrow failure syndrome characterized by activated T cells. Features of T-cell activation in the pathophysiology of SAA remain unknown. To understand T cell activation states, we investigated the atlas of peripheral immune cells and the secreted cytokine network with single cell mass cytometry analysis. We found decreased γδ T-cell frequencies in all patients with SAA, together with a significantly increased proportion of interleukin (IL)-17A-producing cell subsets. Cytokine network analysis of immune cells showed significant positive relationship between IL and 17A production from immune cells and disease severity of severe aplastic anemia. On separating SAA into two distinct subgroups based on T-cell activation stage, the proportion of γδ T cells tended to decrease in the T-cell-activated SAA group compared with non-T-cell-activated group. And the proportion of IL-17A-producing γδ T cells (γδT17) within γδ T cells was newly found to be significantly higher in the T-cell-activated SAA group, implying that IL-17A production by γδ T cells was associated with T-cell activation. Overall, our study revealed a role of γδT17 cells in mediating autoreactive T-cell activation in SAA and provided a novel diagnostic indicator for monitoring autoreactive T-cell activation status during the progression of aplastic anemia in the clinic.

The Hidden Enemy Within: Non-canonical Peptides in Virus-Induced Autoimmunity.

Viruses play a key role in explaining the pathogenesis of various autoimmune disorders, whose underlying principle is defined by the activation of autoreactive T-cells. In many cases, T-cells escape self-tolerance due to the failure in encountering certain MHC-I self-peptide complexes at substantial levels, whose peptides remain invisible from the immune system. Over the years, contribution of unstable defective ribosomal products (DRiPs) in immunosurveillance has gained prominence. A class of unstable products emerge from non-canonical translation and processing of unannotated mammalian and viral ORFs and their peptides are cryptic in nature. Indeed, high throughput sequencing and proteomics have revealed that a substantial portion of our genomes comprise of non-canonical ORFs, whose generation is significantly modulated during disease. Many of these ORFs comprise short ORFs (sORFs) and upstream ORFs (uORFs) that resemble DRiPs and may hence be preferentially presented. Here, we discuss how such products, normally “hidden” from the immune system, become abundant in viral infections activating autoimmune T-cells, by discussing their emerging role in infection and disease. Finally, we provide a perspective on how these mechanisms can explain several autoimmune disorders in the wake of the COVID-19 pandemic.

Current Understanding of the Roles of CD1a-Restricted T Cells in the Immune System.

Cluster of differentiation 1 (CD1) is a family of cell-surface glycoproteins that present lipid antigens to T cells. Humans have five CD1 isoforms. CD1a is distinguished by the small volume of its antigen-binding groove and its stunted A′ pocket, its high and exclusive expression on Langerhans cells, and its localization in the early endosomal and recycling intracellular trafficking compartments. Its ligands originate from self or foreign sources. There are three modes by which the T-cell receptors of CD1a-restricted T cells interact with the CD1a:lipid complex: they bind to both the CD1a surface and the antigen or to only CD1a itself, which activates the T cell, or they are unable to bind because of bulky motifs protruding from the antigen-binding groove, which might inhibit autoreactive T-cell activation. Recently, several studies have shown that by producing TH2 or TH17 cytokines, CD1a-restricted T cells contribute to inflammatory skin disorders, including atopic dermatitis, psoriasis, allergic contact dermatitis, and wasp/bee venom allergy. They may also participate in other diseases, including pulmonary disorders and cancer, because CD1a-expressing dendritic cells are also located in non-skin tissues. In this mini-review, we discuss the current knowledge regarding the biology of CD1a-reactive T cells and their potential roles in disease.

Autoantigen presentation by splenic dendritic cells is required for RBC-specific autoimmunity.

Failure of humoral tolerance to red blood cell (RBC) antigens may lead to autoimmune hemolytic anemia (AIHA), a severe and sometimes fatal disease. Previous studies have shown that although tolerance is robust in HOD mice, autoantibodies are generated upon adoptive transfer of OTII CD4+ T cells, which are specific for an epitope contained within the HOD antigen. These data imply that antigen-presenting cells (APCs) are presenting RBC-derived autoantigen(s) and are capable of driving T-cell activation. Given that multiple APCs participate in erythrophagocytosis, we used a transgenic approach to determine which cellular subsets were required for autoantigen presentation and subsequent autoreactive T-cell activation. HOD mice, which express an RBC-specific antigen consisting of hen egg lysozyme, ovalbumin, and human blood group molecule Duffy, were bred with IAbfl/fl and Cre-expressing transgenic animals to generate mice that lack I-Ab expression on particular cell subsets. OTII CD4+ T cell proliferation was assessed in vivo in HOD+ I-Abfl/fl xCre+ mice and in vitro upon coculture with sorted APCs. Analysis of HOD+ I-Abfl/fl xCre+ mice demonstrated that splenic conventional dendritic cells (DCs), but not macrophages or monocytes, were required for autoantigen presentation to OTII CD4+ T cells. Subsequent in vitro coculture experiments revealed that both CD8+ and CD8- DC subsets participate in erythrophagocytosis, present RBC-derived autoantigen and stimulate autoreactive T-cell proliferation. These data suggest that if erythrocyte T-cell tolerance fails, DCs are capable of initiating autoimmune responses. As such, targeting DCs may be a fruitful strategy for AIHA therapies.

Association of HLA class II (-DRB1,-DQB1,-DPB1) alleles and haplotypes on susceptibility to aplastic anemia in northern Chinese Han

The Human Leukocyte Antigen (HLA) genes, playing key roles in mediating the immune response, especially HLA class II alleles were suggested to play a role in the activation of autoreactive T-cells in aplastic anemia (AA). Previous studies in different ethnic groups have indicated that some of HLA-A,-B,-DRB1 alleles had a protective or susceptive association with the prevalence, pathogenesis and development of AA. HLA class II genes, especially HLA-DQB1 and -DPB1 alleles or haplotypes at high-resolution level associated with AA have not been fully identified in northern Chinese Han populations. The aim of this study was to identify association of the variations in HLA class II region with AA in northern Chinese Han population. A recent case-control study, including 96 AA patients and 824 healthy controls was performed. The high-resolution HLA genotyping was conducted by PCR-SBT, -SSO and NGS-ION S5TM platform. Based on genotypic data of the three loci, haplotype estimation was carried out. HLA-DRB1*15:01 (Pc = 2.87 × 10-3; OR = 2.11, 95% CI = 1.45–3.07) and HLA-DQB1*06:02 (Pc = 1.86 × 10-2; OR = 2.01, 95% CI = 1.32–3.06) were the risk and predisposition alleles to AA in northern Chinese Han after considering multiple testing. Moreover, the HLA-DRB1*15:01-DQB1*06:02 (Pc = 4.90 × 10-3; OR = 2.09, 95% CI = 1.37–3.19) and HLA-DRB1*14:05-DQB1*05:03 (Pc = 2.65 × 10-2; OR = 2.82, 95%CI = 1.45–5.50) haplotypes had direct strong relevance to AA and were the susceptible haplotypes. HLA-DPB1 alleles and 23 polymorphic amino acid residues spanning exon 2 ~ 4 of DPβ1 molecules have showed no statistically significant associations between AA and controls. The present findings establish a novel link between inherited HLA-DRB1,-DQB1,-DPB1 risk alleles and haplotypes in northern Chinese Han with AA, and open new avenues for development of targeted therapies to prevent or redirect immunopathology in AA.

Clinical Characteristics and Treatment of Immune-Related Adverse Events of Immune Checkpoint Inhibitors.

Immune checkpoint inhibitors (ICIs) have been changing the paradigm of cancer treatment. However, immune-related adverse effects (irAEs) have also increased with the exponential increase in the use of ICIs. ICIs can break up the immunologic homeostasis and reduce T-cell tolerance. Therefore, inhibition of immune checkpoint can lead to the activation of autoreactive T-cells, resulting in various irAEs similar to autoimmune diseases. Gastrointestinal toxicity, endocrine toxicity, and dermatologic toxicity are common side effects. Neurotoxicity, cardiotoxicity, and pulmonary toxicity are relatively rare but can be fatal. ICI-related gastrointestinal toxicity, dermatologic toxicity, and hypophysitis are more common with anti- CTLA-4 agents. ICI-related pulmonary toxicity, thyroid dysfunction, and myasthenia gravis are more common with PD-1/PD-L1 inhibitors. Treatment with systemic steroids is the principal strategy against irAEs. The use of immune-modulatory agents should be considered in case of no response to the steroid therapy. Treatment under the supervision of multidisciplinary specialists is also essential, because the symptoms and treatments of irAEs could involve many organs. Thus, this review focuses on the mechanism, clinical presentation, incidence, and treatment of various irAEs.

The role of regulatory T Cells in autoimmune orchitis.

Regulatory T cells (Tregs) mediate tolerance to self-antigens maintaining immune homeostasis. Defects in the number and function of Tregs lead to aberrant immune responses to autologous components, thereby causing autoimmune diseases. Male infertility as a result of immune testicular damage follows through auto-reactive T-cell activation by antigens or pathogens that disrupt testis tolerance mechanisms. In this review we summarise the main evidence on Treg behaviour in inflammatory testicular pathologies focusing on reports on experimental autoimmune orchitis. Increased numbers of different Treg phenotypes are observed in the chronically inflamed testis and in lymph nodes draining to it; however these cells are outnumbered by effector T cells. Distortion of the effector/regulatory cell balance in favour of a pro-inflammatory response is suspected to contribute to exacerbation of autoimmune disease. Under inflammatory conditions, effector T-cell subsets can overwhelm the inhibitory effect of Tregs, and pro-inflammatory cytokines may directly or indirectly affect the ability of Tregs to control autoimmunity. Therefore, Tregs alone may not be sufficient to limit excessive T-cell activation in autoimmune settings. Treg immunotherapy for autoimmune disease treatment aims to restore the normal balance of effector and Tregs in the inflamed tissue. Therapies combining the transfer of Tregs with Treg-stabilising drugs are expected to be the most effective to restrain autoimmune diseases.

Pathomechanism of Autoantibody Production in the Nervous System Diseases

Antibodies to different brain and peripheral nerve proteins have recently been found to be associated with several different autoimmune diseases. They can bind to either neuronal or non-neuronal antigens and may have a pathogenic role by themselves or in synergy with other inflammatory mediators after penetrating the blood-brain barrier or the blood-nerve barrier. In this review, we will describe the association with the impairment of immune tolerance, innate immunity, and autoantibody production of myasthenia gravis (MG), systemic lupus erythematosus (SLE), and Guillain-Barré syndrome (GBS). Impairment of central tolerance, which is characterized by the repertoire selection of immature T-lymphocytes in the thymus, is seen in patients with MG who are positive for anti-Ach R antibodies. Impairment of peripheral tolerance due to activation of autoreactive T-cells and suppression of regulatory T-cells is seen in SLE. In addition, molecular mimicry between the lipooligosaccharides of Campylobacter jejuni and gangliosides of the peripheral nerves results in the production of anti-gangliosides antibodies in GBS. Next, we will describe the antibody-mediated pathology in neuromyelitis optica and anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis. The binding of anti-aquaporin-4 antibodies or anti-NMDAR antibodies to their respective targets initiates target internalization and complement- or antibody-dependent cellular cytotoxicity of the target cells. Further understanding of antibody-mediated pathology may suggest novel therapeutic strategies.

Increased IRAK-M in peritoneal macrophages of NOD mice suggests in vivo exposure to bacterial endotoxins from the gut (INC6P.343)

Abstract Macrophages play an important role in innate immune reactions, and peritoneal macrophages guard the sterility of this compartment mainly against microbial threat from the gut. Type-1 Diabetes (T1D) is an autoimmune disease in which gut microbiota and gut immune system appear to play a role. We have observed increased permeability and free radical production of gut epithelium in nonobese diabetic (NOD) mice. Impaired barrier function could lead to bacterial endotoxin leakage to peritoneal cavity. To seek for extraintestinal consequences of impaired barrier function, we characterized peritoneal lavage cells from young NOD mice after weaning. We detected a rapid increase in the number of macrophages in 1-2 weeks from weaning in NOD mice compared to C57BL/6 and BALB/c mice, and to NOD mice fed an antidiabetogenic diet (ProSobee), which improves gut barrier function. Upon LPS stimulation ex vivo, macrophages produced low amounts of TNF-α and showed reduced expression of CD40. Expression of TLR signalling pathway negative regulator, Interleukin-1 Associated Kinase -M (IRAK-M) was increased, indicating prior in vivo exposure to TLR-4 ligand(s). Upon further phenotyping, increased levels of ARG1 were observed suggesting in vivo differentiation towards M2-like phenotype. Bacterial endotoxins in peritoneal cavity may contribute to autoreactive T-cell activation by increased production of cytokines and activation of antigen-presenting cells in pancreatic lymph nodes.

Innate immunity in myasthenia gravis thymus: Pathogenic effects of Toll-like receptor 4 signaling on autoimmunity

The thymus is the main site of immune sensitization to AChR in myasthenia gravis (MG). In our previous studies we demonstrated that Toll-like receptor (TLR) 4 is over-expressed in MG thymuses, suggesting its involvement in altering the thymic microenvironment and favoring autosensitization and autoimmunity maintenance processes, via an effect on local chemokine/cytokine network. Here, we investigated whether TLR4 signaling may favor abnormal cell recruitment in MG thymus via CCL17 and CCL22, two chemokines known to dictate immune cell trafficking in inflamed organs by binding CCR4. We also investigated whether TLR4 activation may contribute to immunodysregulation, via the production of Th17-related cytokines, known to alter effector T cell (Teff)/regulatory T cell (Treg) balance. We found that CCL17, CCL22 and CCR4 were expressed at higher levels in MG compared to normal thymuses. The two chemokines were mainly detected around medullary Hassall's corpuscles (HCs), co-localizing with TLR4+ thymic epithelial cells (TECs) and CCR4+ dendritic cells (DCs), that were present in higher number in MG thymuses compared to controls. TLR4 stimulation in MG TECs increased CCL17 and CCL22 expression and induced the production of Th17-related cytokines. Then, to study the effect of TLR4-stimulated TECs on immune cell interactions and Teff activation, we generated an in-vitro imaging model by co-culturing CD4+ Th1/Th17 AChR-specific T cells, naïve CD4+CD25+ Tregs, DCs and TECs from Lewis rats. We observed that TLR4 stimulation led to a more pronounced Teff activatory status, suggesting that TLR4 signaling in MG thymic milieu may affect cell-to-cell interactions, favoring autoreactive T-cell activation. Altogether our findings suggest a role for TLR4 signaling in driving DC recruitment in MG thymus via CCL17 and CCL22, and in generating an inflammatory response that might compromise Treg function, favoring autoreactive T-cell pathogenic responses.

Abstract 129: Suppressor of Cytokine Signaling 1 in Dendritic Cells Inhibits Myocardial Inflammation in Experimental Autoimmune Myocarditis

Introduction: Autoimmunity is considered to play an important role in the development of myocarditis and dilated cardiomyopathy. Recent reports have indicated that a subgroup of myocarditis patients may benefit from immune-targeted therapies. Suppressor of cytokine signaling1 (SOCS1) is an intracellular, cytokine-inducible protein that regulates the responses of immune cells to cytokines. We therefore hypothesized that overexpression of SOCS1 may inhibit the inflammation of myocarditis and cardiomyopathy. Methods and Results: Myocarditis was induced by subcutaneous immunization with cardiac specific peptides derived from α-myosin heavy chain in BALB/c mice on days 0 and 7. Plasmid DNA encoding SOCS1 (pSOCS1) was injected intraperitoneally into mice on days 0, 5 and 10. pSOCS1 treatment significantly decreased heart-to-body weight ratios and the number of infiltrating cells in the heart. Echocardiography showed preserved contractile function in pSOCS1-treated mice. Although autoimmune myocarditis is a CD4+ T cell-mediated disease, pSOCS1 treatment does not have a direct suppressive effect on autoreactive T-cell activation. The introduced pSOCS1 suppressed proinflammatory cytokine production and STAT1 phosphorylation in dendritic cells (DCs). In addition, the proliferative responses of autoreactive CD4+ T cells co-cultured with DCs from pSOCS1-treated mice were much weaker than those of cells cultured with DCs from control plasmid-injected mice. These results suggested that the inoculated pSOCS1 may have been transfected into DCs and impaired DC function in vivo. Conclusion: The administration of pSOCS1 protected mice from the development of experimental autoimmune myocarditis, which was mediated by the inhibition of DC function that in turn reduced the activation of autoreactive CD4+ T cells.

A Switch in Regulatory T-Cell Memory Phenotype Is Associated with Poor Survival in Lower Risk Myleodysplastic Syndrome Patients

Abstract 1703Myleodysplastic syndrome (MDS) is a heterogeneous group of clonal hematopoietic diseases marked by various cytopenias including neutropenia and anemia. Hematologic improvement in response to T-cell depleting agents and immunosuppressants in a subset of MDS patients suggests that hematopoietic impairment may result from autoreactive T-cell activation. Conversely, suppression of the T-cell compartment may be permissible to the development of malignant myeloid clones. The suspicion that T-cell homeostatic balance may be disrupted in MDS has spawned numerous studies investigating mechanisms of immune suppression and autoimmune regulation. T-cell autoimmunity is regulated in part by regulatory T-cells (Tregs), which express T cell receptors (TCRs) that recognize self-antigens and function to maintain immune homeostasis. Indeed, higher numbers of Tregs have been found in higher risk MDS patients, and those MDS subtypes with higher blast percentages. However, the role of Tregs in lower risk patients is less clear, but may still represent an important component of pathogenesis in early disease.Because of the autoimmune nature of low risk MDS, we hypothesized that alterations in the Treg compartment may play a role in lower risk MDS disease progression. To address this, we analyzed Treg phenotype and numbers in MDS patients largely classified as low risk by the International Prognostic Scoring System (IPSS) and 41 age-matched controls using cell surface markers as analyzed by flow cytometry. Tregs were identified as CD3+CD4+CD25+CD127dim. The use of CD127 as a Treg marker was validated by intracellular staining with FoxP3. Naïve and memory phenotypes within the Treg compartment were analyzed using CD45RA and CD27 expression. Naïve Tregs (TregN) were characterized as CD45RA+CD27+, central memory Tregs (TregCM) as CD45RA−CD27+, and effector memory Tregs (TregEM) as CD45RA−CD27−. Two distinct subgroups of MDS patients were identified: one with elevated total numbers of Tregs, and a second with a marked shift in Treg memory phenotype away from the common TregCM phenotype and toward the rare TregEM phenotype. These two subgroups of patients were distinct from each other, and appeared to be independent of MDS WHO subtype, IPSS score, karyotype, neutropenic state, and thrombocytopenic state. MDS patients with a shift toward TregEMwere associated with anemia (p=0.0433). This association was not found in MDS patients with total increases in Treg numbers.Next, we sought to determine if there are differences in overall survival (OS) of MDS patients with either increased total numbers of Tregs or in patients with a shift toward TregEM phenotype. Patients with greater than 50 Tregs/μl did have reduced OS compared to patients with less Tregs, but this difference was not statistically significant (p=0.1387). In contrast, a highly significant difference was observed in MDS patients with a shift toward the TregEM phenotype (p=0.0200), with patients having greater than 2.5TregEM/μl displaying the worst OS (Figure 1). [Display omitted] Lastly, because an increased TregEM compartment is associated with reduced OS in MDS patients, we sought to compare the suppressive capacities of TregN, TregCM, and TregEM cells to better understand the functional consequences of Treg memory shift. CD4+ T-cells were isolated by negative selection, and then TregN, TregCM, and TregEM cells were sorted using CD45RA and CD27 expression. Conventional CD4+ responder cells were then labeled with Carboxyfluorescein succinimidyl ester (CFSE) and co-cultured with increasing ratios of each isolated Treg population. Cellular division in response to CD3/CD28 stimulation was measured 5 days later by analyzing CFSE dilution. TregCM appear to be the least suppressive, while TregN and TregEM cells were more potently suppressive. These results indicate that a shift in Treg phenotype toward TregEM may result in a more suppressive Treg compartment without obvious increases in total Treg numbers. Taken together, this data suggests that in lower risk MDS, the phenotypic makeup of the Treg compartment may be more important than total Treg numbers. In addition, this data suggests that increased TregEM cells may be a novel prognostic indicator in low risk MDS. Disclosures:Komrokji:Celgene: Honoraria, Research Funding, Speakers Bureau. List:Celgene: Consultancy.

Role of TLR2 in the pathogenesis of autoimmune diabetes and its therapeutic implication

Recently, a couple of articles suggested the possibility that apoptosis of pancreatic β-cells induces inflammatory/immune responses to β-cells. Such a theory is based on the assumption that apoptotic cells can, under certain circumstances, induce immune responses, inflammatory and autoimmune disorders, which is in contrast to the dogma that apoptotic cells result in immunosuppression and necrotic cells provoke inflammation/immunity. We observed that late apoptotic β-cells with secondary necrosis elicited inflammatory responses in macrophages through the toll-like receptor 2 (TLR2)/MyD88/nuclear factor-κB signalling pathway. Late apoptotic cells also induced TLR2-dependent maturation of dendritic cells and then activation of autoreactive T-cells. TLR2 knockout mice showed defective priming of diabetogenic T-cells by apoptotic β-cells in the pancreatic lymph nodes. Furthermore, TLR2 deficiency conferred a significant protection against type 1 diabetes (T1D) and insulitis in T1D animal models. These findings present evidence suggesting that apoptosis of pancreatic β-cells could be one of the initial events in T1D and provide a novel strategy for therapeutic or preventive intervention in T1D.

CD19: a promising B cell target for rheumatoid arthritis

B-cell depletion with unconjugated CD20 monoclonal antibody (mAb) is used to treat rheumatoid arthritis and other autoimmune diseases. CD20-targeted immunotherapy depletes mature B cells through monocyte-mediated antibody-dependent cellular cytotoxicity, but does not effectively deplete pre-B or immature B cells, certain peripheral B cell subpopulations, most antibody-producing cells, or their malignant counterparts. As immature B cells expressing autoreactive antigen receptors are not depleted by anti-CD20 mAb, a new strategy for eliminating autoantigen-selected B cells and for treating early lymphoblastic leukemias and/or lymphomas was developed using CD19-specific mAbs that induce Fcgamma receptor-dependent and monocyte-dependent B-cell depletion. Preclinical studies using transgenic mice expressing human CD19 have shown that pre-B cells and their malignant counterparts, as well as pre-existing antibody-producing and autoantibody-producing cells, are depleted. Therefore, CD19-directed immunotherapy is expected to treat diverse pre-B-cell-related and plasmablast-related malignancies, and humoral transplant rejection. Moreover, in contrast to CD20-directed immunotherapies, CD19 mAbs could purge the B cell repertoire of autoreactive clones and reset the developmental clock to a point that curtails the extent of emerging self-reactivity, in addition to reducing autoreactive T-cell activation through the elimination of mature B cells. Humanized CD19 mAbs are expected to enter clinical trials in 2009, offering a new approach for the treatment of autoimmune disease that removes both immature B cells and antibodies with autoreactive specificities. CD19-directed immunotherapy could, therefore, offer a new horizon in B-cell depletion for the treatment of multiple autoimmune diseases.

An Apparent Role for Alox15 in the Pathogenesis of Diabetes in the NOD Mouse

Progenitors of the nonobese diabetic mouse strain (NOD/Shi) arose spontaneously in a colony at the Shionogi Research Laboratories in Aburahi, Japan in the 1970s. The strain has been used extensively in efforts to elucidate the pathogenesis of type 1 diabetes in humans. Both the mouse model and the human disease are characterized by the appearance of autoreactive T-cells targeting pancreatic islet antigens, the elaboration of anti-insulin autoantibodies, and the development of a β-cell–toxic inflammatory cellular infiltrate within the islets leading to insulin depletion and hyperglycemia. In both NOD mice and human type 1 diabetes, the breakdown of self-tolerance to β-cells is under polygenic control of major histocompatibility complex (MHC) class II alleles as well as non-MHC loci. An important difference between the mouse model and the human disease is that in NOD mice but not human type 1 diabetes, female subjects have a higher incidence of diabetes (75–100% by 30 weeks) than males (30–60%) (1). Because the development of diabetes in NOD mice depends on interacting genetic and environmental factors comparable with those interacting in human type 1 diabetes, genes that modify the NOD phenotype deserve close attention. The article by McDuffie et al. (2 in this issue identifies such a candidate molecule. Previous studies elucidating the critical steps in the development of autoimmune β-cell destruction in NOD mice have suggested several points of pathogenic relevance. β-Cell destruction in NOD mice is dependent on the production and unrestrained activation of autoreactive T-cells. Antibody-mediated elimination of T-cells and interventions that promote T-cell tolerance, such as the production of regulatory T-cells, effectively inhibit development of diabetes in …

Marginal-Zone B-Cells of Nonobese Diabetic Mice Expand With Diabetes Onset, Invade the Pancreatic Lymph Nodes, and Present Autoantigen to Diabetogenic T-Cells

B-cells are important for disease pathogenesis in the nonobese diabetic (NOD) mouse model of type 1 diabetes. Recent studies demonstrate that marginal-zone B-cells (MZBs), which connect innate with adaptive immune responses, are increased in NOD mice. However, beyond this, the contribution of different B-cell subsets to diabetes pathogenesis is poorly understood. To better understand the role of different B-cell subsets in the etiology of type 1 diabetes, we have examined the MZB compartment in NOD mice, with respect to their number, distribution, and function. We demonstrate that splenic MZB numbers in female NOD mice undergo a marked, approximately threefold expansion between approximately 12 and 16 weeks of age, coincident with the onset of frank diabetes. Functionally, NOD MZBs are hyperresponsive to toll-like receptor 9 ligation and CD40 ligation, as well as sphingosine-1-phosphate-dependent chemotactic cues, suggesting an increased sensitivity to selective innate- and activation-induced stimuli. Intriguingly, at 16 weeks of age, approximately 80% of female NOD mice present with MZB-like cells in the pancreatic lymph node (PLN). These MZB-like cells express major histocompatibility complex class II and high levels of CD80 and CD86, and their presence in the PLN is associated with an increased frequency of activated Vbeta4(+) CD4(+) T-cells. Significantly, we demonstrate that purified MZBs are able to present the autoantigen insulin to diabetogenic T-cells. These data are consistent with MZBs contributing to the pathogenesis of type 1 diabetes as antigen-presenting cells. By integrating innate-derived inflammatory signals with the activation of autoreactive T-cells, MZBs may help to direct T-cell responses against beta-cell self-constituents.

Dendritic cells pulsed with apoptotic cells activate self-reactive T-cells of lupus mice both in vitro and in vivo

Systemic lupus erythematosus (SLE) is characterized by the presence of autoantibodies (autoAbs) directed against the nuclear structure. Previous studies have demonstrated that dendritic cells (DCs) can process and present self-antigens (Ags) from apoptotic cells (ACs) in lupus. However, there is no direct evidence demonstrating that ACs provide self-Ags, such as histones, to stimulate autoreactive T-cells in lupus. AC-pulsed bone marrow-derived DCs (AC-BMDCs) were used to stimulate autoreactive T-cells in vitro and in vivo. In our study, we found that AC-BMDCs could induce the proliferation of CD4+ T-cells from unprimed NZB x NZW F1 (BWF1) mice, which spontaneously develop SLE, but not CD4+ T-cells, from non-autoimmune DBA-2 x NZW F1 (DWF1) mice. In addition, AC-BMDCs could induce significant proliferative responses to certain histone peptide-specific T-cells. Furthermore, these AC-BMDCs could induce a considerable anti-DNA Ab response in vivo after adoptive transfer into DWF1 mice, suggesting that AC-BMDCs can break tolerance in normal mice and initiate an autoimmune response. Our study provides a direct link between self-epitopes from ACs presented by DCs and autoreactive T-cell activation, and demonstrates that ACs are critical for the induction of autoimmunity in vivo.

Regulatory T‐cell physiology and application to treat autoimmunity

Endowed with the ability to actively suppress an immune response, regulatory T cells (Tregs) hold the promise of halting ongoing pathogenic autoimmunity and restoring self-tolerance in patients suffering from autoimmune diseases. Through many in vitro and in vivo studies, we have learned that Tregs can function in the lymph nodes as well as in the peripheral tissues. In vivo, Tregs act through dendritic cells to limit autoreactive T-cell activation, thus preventing their differentiation and acquisition of effector functions. By limiting the supply of activated pathogenic cells, Tregs prevent or slow down the progression of autoimmune diseases. However, this protective mechanism appears insufficient in autoimmune individuals, likely because of a shortage of Tregs cells and/or the development and accumulation of Treg-resistant pathogenic T cells over the long disease course. Thus, restoration of self-tolerance in these patients will likely require purging of pathogenic T cells along with infusion of Tregs with increased ability to control ongoing tissue injury. In this review, we highlight advances in dissecting Treg function in vivo in autoimmune settings and summarize multiple studies that have overcome the limitations of the low abundance of Tregs and their hypoproliferative phenotype to develop Treg-based therapies.

P0161 PP CD4+CD25+ T REGS MAINTAIN THE ABILITY TO SUPPRESS CD4 T CELL FUNCTION IN PATIENTS WITH AUTOIMMUNE LIVER DISEASE

Introduction: CD4+CD25+ T cells (T-regs) are central to immune regulation by preventing the activation of autoreactive T-cells. It has recently been reported that T-regs are decreased in number and impaired in their ability to expand in patients with autoimmune liver disease (AILD). Aim: to investigate the suppressor function of T-regs by testing their ability to inhibit IFN-gamma production by CD4+CD25- T-cells. Methods: Purified T-regs were obtained from PBMCs by CD4 negative selection followed by CD25 positive selection using immunomagnetic beads and added at ratios of 1/16 and 1/8 to CD4+CD25− cells. Percentage of IFN-gamma producing CD4+CD25− cells was determined after 5 days co-culture with or without T-regs by intracellular cytokine staining. Patients: Nine patients with AILD (8 ANA/SMA positive, 1 LKM1 positive) were studied at diagnosis when CD4+CD25+ T-cell frequency was the lowest. Six healthy laboratory workers served as controls. Results: CD4+ CD25+ T-cells were significantly fewer in patients than in controls (7.05 vs 2.17%; p=0.02). The frequency of IFN-gamma producing CD4 T-cells tended to be lower in patients than in controls (3.5% vs 8.8%; p=0.06). Their percentage was reduced to 2.5% and 2.0% in patients and to 8.4% and 7.2% in controls when autologous T-regs were added at a ratio of 1/16 and 1/8 respectively. The percentage of inhibition of IFN-gamma production was not significantly different in patients when compared to controls (p>0.05). Conclusion: Our data suggest that impairment of immune regulation leading to aggression in AILD is related to the number and not the function of regulatory T-cells.

The initiation of the autoimmune response in multiple sclerosis

Multiple sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system (CNS). Most evidence supports the autoimmune pathogenesis of the disease. According to this hypothesis, the activation of autoreactive T-cells is a central event in the development of autoimmune response in MS. We examined molecular events involved in the initiation of autoimmune response in MS. Recent studies in our laboratory have reported an unexpectedly high degree of T-cell receptor (TCR) degeneracy and molecular mimicry as a frequent phenomenon that might play a role in the initiation of autoimmune response in MS. This paper provides insights into the physiologic and pathologic role of autoreactive T-cells, and characterizes structurally and functionally the specific targets for new therapies of MS.