Florence Susan Wong

Type 1 diabetes (T1D) is an autoimmune disease characterized by T cell–mediated destruction of pancreatic β cells, in which antigen presentation is essential. Although professional antigen-presenting cells initiate autoreactive T cell responses, whether endocrine cells contribute to antigen presentation within inflamed islets remains unclear. Here, we show that pancreatic α cells acquire major histocompatibility complex (MHC) class II (MHC-II)–dependent antigen-presenting capacity during T1D progression. Reanalysis of single-cell RNA sequencing datasets from nonobese diabetic (NOD) mice and human T1D islets revealed progressive up-regulation of MHC-II associated genes in α cells. Immunostaining validated the increased of MHC-II level both in NOD mice and patients with T1D. Mechanistically, inflammatory cytokines activated the Janus kinase–signal transducer and activator of transcription (JAK-STAT) pathway in α cells, leading to JAK-STAT–dependent MHC-II induction. Functionally, activated α cells internalized, processed, and presented exogenous antigens, acquired expression of costimulatory molecules, and promoted antigen-specific CD4 ⁺ T cell activation. These findings identify pancreatic α cells as nonprofessional antigen-presenting cells and expand the current understanding of antigen presentation in autoimmune diabetes.

Identifying and monitoring autoreactive T cells that drive β cell destruction remains a major obstacle to developing effective immunotherapies for type 1 diabetes (T1D). These cells are extremely rare in peripheral blood and cannot be accessed directly from the pancreas. We used intradermal injection of glutamic acid decarboxylase (GAD)-alum to recruit GAD-specific T cells to accessible sites in the skin and skin-draining lymph nodes (LNs), sampled by skin suction blisters and ultrasound-guided LN aspiration. Peripheral blood samples obtained before GAD injection were restimulated with GAD in vitro to detect reactive CD4 ⁺ T cells. Single-cell RNA sequencing (scRNA-seq) followed by re-expression of selected T cell receptors (TCRs) confirmed antigen specificity. Up to 70% of T cells at the skin injection site were clonally expanded and 4 of 14 (28%) re-expressed TCRs were GAD reactive. In LNs, 1 of 14 (4%) clonally expanded TCRs was GAD reactive, representing approximately 0.08% of all T cells. GAD-reactive cells across compartments displayed Th1- and Th17-associated transcription signatures. These results demonstrate that intradermal autoantigen challenge and scRNA-seq enable direct identification and molecular profiling of autoreactive T cells in vivo. This minimally invasive approach provides a powerful platform for tracking antigen-specific T cells to monitor disease activity and evaluate immune interventions in T1D.

Aims/hypothesis: The chemokine receptor C-X-C chemokine receptor type 3 (CXCR3) is a key chemoattractant molecule that facilitates the migration of activated T cells to the pancreas, leading to beta cell death. In this study, we investigated CXCR3 responses in B cells during type 1 diabetes progression. Methods: Peripheral blood samples were obtained from individuals with recent-onset and long-duration type 1 diabetes, who were age- and sex-matched to non-diabetic donors. We isolated peripheral blood mononuclear cells (PBMCs) and examined changes in CXCR3 expression on lymphocytes from donors, performing multiparameter flow cytometry and functional cell culture assays. Human post-mortem pancreatic tissue was obtained from the Exeter Archival Diabetes Biobank. Immunofluorescence staining was used to assess CXCR3 expression in pancreatic tissues. Results: We observed reduced CXCR3 expression on antigen-experienced B cells in individuals with a long duration of type 1 diabetes, although B cells remained responsive to IFNγ. In individuals who were recently diagnosed, IFNγ treatment resulted in increased CXCR3 expression compared with B cells from non-diabetic donors. B cells in pancreases that were recovered post-mortem from young recent-onset donors lacked CXCR3 expression, but co-staining to detect CD8 ⁺ T cells revealed a CXCR3 ⁺ CD20 ⁺ CD8 ⁺ T cell population, with their circulating counterpart showing increased CXCR3 expression. Conclusions/interpretation: We conclude that the CXCR3 response in antigen-experienced B cells is dysregulated during the progression of type 1 diabetes. CXCR3 expression is limited in CD20 ⁺ B cells in pancreases from recent-onset individuals diagnosed with type 1 diabetes under 7 years of age, but evident on CD8 ⁺ T cells that express CD20. Graphical

The NOD mouse is the best-known, although by no means the only, murine model of type 1 diabetes. In this review, we provide a historical perspective, particularly highlighting areas of progress in understanding aspects of aetiology and immune pathogenesis, and utility in helping to shape the immunotherapeutic landscape. We introduce points of interest where the NOD mouse, a much-studied model, has signposted discovery and knowledge. We discuss genetics, pancreatic islet beta cell stress, innate and adaptive immunity and autoantigens, and also focus on immunotherapeutic agents that have been tested in NOD mice and in humans. Some therapies, particularly those that are non-antigen-specific, have been more effectively signposted, while others, which include antigen-specific therapies, have not. There is an inevitable divergence between mice and humans that illustrates the need to use models appropriately. We suggest how to make use of this and other models effectively in order to maximise information and knowledge, and suggest not dismissing this important resource because of inappropriate comparisons or unrealistic expectations. Graphical Abstract

Introduction: In this study, we report a novel therapeutic approach redirecting antigen-specific CD4⁺ T cells recognizing a hybrid insulin peptide (BDC2.5 T cell receptor (TCR) transgenic CD4⁺ T cells) to attract and suppress islet-specific CD8⁺ T cells T cells in the non-obese diabetic (NOD) mouse model, and prevent the development of autoimmune diabetes.Methods: Purified BDC2.5 CD4⁺ T cells were induced to differentiate into regulatory T cells (Tregs). The Tregs were then electroporated with mRNA encoding chimeric human β₂ microglobulin (hβ₂m) covalently linked to insulin B chain amino acids 15-23 (designated INS-eTreg) or islet-specific glucose-6-phosphatase related protein (IGRP) peptide 206-214 (designated IGRP-eTreg). Immunoregulatory functions of these engineered regulatory T cells (eTregs) were tested by in vitro assays and in vivo co-transfer experiments with β-cell-antigen-specific CD8⁺ T cells in NOD.Scid mice or by adoptive transfer into young, pre-diabetic NOD mice.Results: These eTregs were phenotyped by flow cytometry, and shown to have high expression of FoxP3, as well as other markers of Treg function, including IL-10. They suppressed polyclonal CD4⁺ T cells and antigen-specific CD8⁺ T cells (recognizing insulin or IGRP), decreasing proliferation and increasing exhaustion and regulatory markers in vitro. In vivo, eTregs reduced diabetes development in co-transfer experiments with pathogenic antigen-specific CD8⁺ T cells (INS-CD8⁺ or IGRP-CD8⁺ cells) into NOD.Scid mice. Finally, when the eTreg were injected into young NOD mice, they reduced insulitis and prevented spontaneous diabetes in the recipient mice.Conclusion: Our results suggest a novel therapeutic strategy to protect NOD mice by targeting antigen-specific cytotoxic CD8⁺ T cells, using redirected antigen-specific CD4⁺ Treg cells, to suppress autoimmune diabetes. This may suggest an innovative therapy for protection of people at risk of development of type 1 diabetes.

Introduction: In this study, we report a novel therapeutic approach using B lymphocytes to attract islet-specific T cells in the non-obese diabetic (NOD) mouse model and prevent the development of autoimmune diabetes. Rather than using the antibody receptor of B cells, this approach utilizes their properties as antigen-presenting cells to T cells.Methods: Purified splenic B cells were treated with lipopolysaccharide, which increases regulatory B (Breg) cell function, then electroporated with mRNA encoding either chimeric MHC-I or MHC-II molecules covalently linked to antigenic peptides. Immunoregulatory functions of these engineered B cells (e-B cells) were tested by in vitro assays and in vivo co-transfer experiments with beta-cell-antigen-specific CD8⁺ or CD4⁺ T cells in NOD.Scid mice, respectively.Results: The e-B cells expressing chimeric MHC-I-peptide inhibited antigen-specific CD8⁺ T-cell cytotoxicity in vitro. The e-B cells expressing chimeric MHC-II-peptide induced antigen-specific CD4⁺ T cells to express the regulatory markers, PD-1, ICOS, CTLA-4, Lag3, and Nrp1. Furthermore, e-B cells encoding the chimeric MHC-I and MHC-II peptide constructs protected NOD.Scid mice from autoimmune diabetes induced by transfer of antigen-specific CD8⁺ and CD4⁺ T cells.Discussion: MHC–peptide chimeric e-B cells interacted with pathogenic T cells, and protected the host from autoimmune diabetes, in a mouse model. Thus, we have successfully expressed MHC–peptide constructs in B cells that selectively targeted antigen-specific cells, raising the possibility that this strategy could be used to endow different protective cell types to specifically regulate/remove pathogenic cells.

PDF file - 72K, Figure S1. Binding of nivolumab to human T cells.

PDF file - 22K, Figure S3. Nivolumab inhibits PD-1 interaction with PD-L1 and PD-L2.