Abstract
Aims/hypothesisSlow progressors to type 1 diabetes are individuals positive for multiple pancreatic islet autoantibodies who have remained diabetes-free for at least 10 years; regulation of the autoimmune response is understudied in this group. Here, we profile CD4+ regulatory T cells (Tregs) in a small but well-characterised cohort of extreme slow progressors with a median age 43 (range 31–72 years), followed up for 18–32 years.MethodsPeripheral blood samples were obtained from slow progressors (n = 8), age- and sex-matched to healthy donors. One participant in this study was identified with a raised HbA1c at the time of assessment and subsequently diagnosed with diabetes; this donor was individually evaluated in the analysis of the data. Peripheral blood mononuclear cells (PBMCs) were isolated, and to assess frequency, phenotype and function of Tregs in donors, multi-parameter flow cytometry and T cell suppression assays were performed. Unsupervised clustering analysis, using FlowSOM and CITRUS (cluster identification, characterization, and regression), was used to evaluate Treg phenotypes.ResultsUnsupervised clustering on memory CD4+ T cells from slow progressors showed an increased frequency of activated memory CD4+ Tregs, associated with increased expression of glucocorticoid-induced TNFR-related protein (GITR), compared with matched healthy donors. One participant with a raised HbA1c at the time of assessment had a different Treg profile compared with both slow progressors and matched controls. Functional assays demonstrated that Treg-mediated suppression of CD4+ effector T cells from slow progressors was significantly impaired, compared with healthy donors. However, effector CD4+ T cells from slow progressors were more responsive to Treg suppression compared with healthy donors, demonstrated by increased suppression of CD25 and CD134 expression on effector CD4+ T cells.Conclusions/interpretationsWe conclude that activated memory CD4+ Tregs from slow progressors are expanded and enriched for GITR expression, highlighting the need for further study of Treg heterogeneity in individuals at risk of developing type 1 diabetes.Graphical abstract
Highlights
Progression rates from the first appearance of pancreatic islet autoantibodies to development of clinical symptoms of type 1 diabetes are well described in childhood, with 70% of multiple islet autoantibody-positive children developing diabetes within 10 years of seroconversion, which increases to 84% for those followed for 15 years [1]
Slow progressors have significantly increased proportions of CD4+ effector memory Tregs We investigated the frequency of Tregs present in the peripheral blood of slow progressors (SP group), compared with age- and sex-matched healthy donors (HD group)
Surface markers used for analysis included CD25, CD127, forkhead box P3 (FOXP3), HLA-DR, CD39, cytotoxic T-lymphocyte-associated protein 4 (CTLA4), glucocorticoid-induced TNF receptor (TNFR)-related protein (GITR), CD49b and LAG3, and FlowSOM clustering was performed on down-sampled CD45RA− cells
Summary
Progression rates from the first appearance of pancreatic islet autoantibodies to development of clinical symptoms of type 1 diabetes are well described in childhood, with 70% of multiple islet autoantibody-positive children developing diabetes within 10 years of seroconversion, which increases to 84% for those followed for 15 years [1]. A staging system is increasingly widely utilised [2] to define progression to type 1 diabetes; individuals enter stage 1 at development of multiple islet autoantibodies, stage 2 with dysglycaemia, and stage 3 at onset of symptoms. Some multiple islet autoantibody-positive individuals, in stage 1 and 2, progress more slowly and develop adult-onset type 1 diabetes. We showed that islet autoantigen-specific CD8+ T cell responses were largely absent in slow progressors but were readily detectable in individuals with recent-onset and longstanding diabetes [4]. This might suggest that regulation of the autoimmune response is enhanced in these individuals compared with those who progress. No change was observed in the levels of activation via anti-CD3/CD28 beads
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