Autoantibodies and associated T-cell responses to determinants within the 831-860 region of the autoantigen IA-2 in Type 1 diabetes
ABSTRACT B-cells influence T-cell reactivity by facilitating antigen presentation, but the role of autoantibody-secreting B-cells in regulating T-cell responses in Type 1 diabetes is poorly defined. The aims of this study were to characterise epitopes on the IA-2 autoantigen for three monoclonal antibodies from diabetic patients by amino acid substitutions of selected residues of IA-2, establish contributions of these epitopes to binding of serum antibodies in Type 1 diabetes and relate B- and T-cell responses to overlapping determinants on IA-2. The monoclonal antibodies recognised overlapping epitopes, with residues within the 831-860 region of IA-2 contributing to binding; substitution of Glu836 inhibited binding of all three antibodies. Monoclonal antibody Fab fragments and substitution of residues within the 831-836 region blocked serum antibody binding to an IA-2 643-937 construct. IL-10-secreting T-cells responding to peptides within the 831-860 region were detected by cytokine-specific ELISPOT in diabetic patients and responses to 841-860 peptide were associated with antibodies to the region of IA-2 recognised by the monoclonal antibodies. The study identifies a region of IA-2 frequently recognised by antibodies in Type 1 diabetes and demonstrates that these responses are associated with T-cells secreting IL-10 in response to a neighbouring determinant.
- SourceAvailable from: Deborah Karen Shoemark
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- "To identify more clearly the epitope region affected by substitutions of the core cysteine, we investigated the effect of lysine substitution at E836. This substitution was previously shown to abolish binding of monoclonal antibody 96/3 and reduce binding of sera from patients with type 1 diabetes (23,24). The mutation E836K in IA-2ic and IA-2 PTP resulted in modest reductions in binding by patient sera (median 10%, range −0.8–93%, P < 0.001; and 18%, range −16–98%, P < 0.001, respectively) (Fig. 4). "
ABSTRACT: Cysteines are thought integral to conformational epitopes of islet antigen-2 (IA-2) autoantibodies (IA-2A), possibly through disulfide bond formation. We therefore investigated which cysteines are critical to IA-2A binding in patients with newly diagnosed type 1 diabetes. All 10 cysteines in the intracellular domain of IA-2 were modified to serine by site-directed mutagenesis, and the effects of these changes on autoantibody binding in comparison with wild-type control were investigated by radiobinding assay. Mutation of the protein tyrosine phosphatase (PTP) core cysteine (C909) in IA-2 caused large reductions in autoantibody binding. In contrast, little or no reduction in binding was seen following substitution of the other cysteines. Modification of the core cysteine (C945) in IA-2β also greatly reduced autoantibody binding. Lysine substitution of glutamate-836 in IA-2 or glutamate-872 in IA-2β resulted in modest reductions in binding and identified a second epitope region. Binding to IA-2 PTP and IA-2β PTP was almost abolished by mutation of both the core cysteine and these glutamates. The core cysteine is key to the major PTP conformational epitope, but disulfide bonding contributes little to IA-2A epitope integrity. In most patients, at disease onset, >90% of antibodies binding to the PTP domain of IA-2 recognize just two epitope regions.Diabetes 09/2012; 62(1). DOI:10.2337/db11-1590 · 8.47 Impact Factor
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- "The evidence for molecular mimicry between an IA-2 (and GAD) epitope and the rotavirus VP7 protein has been further detailed by showing strong binding of both autoantigen and viral peptides to HLA-DRB1*04 and cross recognition by IA-2-specific T cells . The 831–860 region of IA-2 frequently recognized by autoantibodies has been shown to be recognized by IL-10-secreting T cells from T1D patients . Following identification of DRA1*0101/DRB1*0401-retricted IGRP23-35 and IGRP247-259 and DRA1*0101/DRB1*0301-retricted IGRP13-25 and IGRP226-238 epitopes, IGRP-specific CD4+ T cells have been detected in more than 80% of DRB1*0401 or DRB1*0301 healthy and T1D subjects . "
ABSTRACT: Type 1 diabetes (T1D) is an autoimmune disease driven by the activation of lymphocytes against pancreatic β-cells. Among β-cell autoantigens, preproinsulin has been ascribed a key role in the T1D process. The successive steps that control the activation of autoreactive lymphocytes have been extensively studied in animal models of T1D, but remains ill defined in man. In man, T lymphocytes, especially CD8(+) T cells, are predominant within insulitis. Developing T-cell assays in diabetes autoimmunity is, thus, a major challenge. It is expected to help defining autoantigens and epitopes that drive the disease process, to pinpoint key functional features of epitope-specific T lymphocytes along the natural history of diabetes and to pave the way towards therapeutic strategies to induce immune tolerance to β-cells. New T-cell technologies will allow defining autoreactive T-cell differentiation programs and characterizing autoimmune responses in comparison with physiologically appropriate immune responses. This may prove instrumental in the discovery of immune correlates of efficacy in clinical trials.Clinical and Developmental Immunology 07/2011; 2011:513210. DOI:10.1155/2011/513210 · 2.93 Impact Factor
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ABSTRACT: An autoimmune response can be induced with the application of dendritic cells (DCs), offering a viable tumor vaccine for cancer immunotherapy. Previous studies have shown that DC-based tumor vaccines in animal tumor models can inhibit tumor growth and induce autoantibodies transiently without clinical or histological features of autoimmunity. The present study aimed to investigate the role of immune response induced by dendritic cells-based therapy, especially anti-ds DNA antibodies in tumor inhibition. In this study, high titers of anti-ds DNA antibodies were induced after injecting syngeneic dendritic cells into BALB/c mice. In addition, mice immunized with DCs showed the inhibition of RL male symbol 1, BALB/c leukemia cell line, tumor growth, and prolonged survival times of tumor mice but no significant difference was found in specific CTL response and NK cell activity when compared to those of the control group. Anti-ds DNA monoclonal antibodies can recognize RL male symbol 1 cells but not normal cells by FACS analysis. Monoclonal anti-ds DNA antibody was demonstrated to be able to lyse tumor cells via complement mediated reaction in vitro and also exhibits the anti-tumor effects when the antibody was injected into tumor-implanted mice. The data suggested that immunization with dendritic cells can induce autoimmune response, which might exert anti-tumor activity in vivo.Journal of Autoimmunity 10/2009; 34(4):364-70. DOI:10.1016/j.jaut.2009.08.013 · 7.02 Impact Factor