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AbstractAcute pancreatitis (AP) is a disease characterized by an acute inflammatory phase followed by a convalescent phase. Diabetes mellitus (DM) was historically felt to be a transient phenomenon related to acute inflammation; however, it is increasingly recognized as an important late and chronic complication. There are several challenges that have prevented precisely determining the incidence rate of DM after AP and understanding the underlying mechanisms. The DREAM (Diabetes RElated to Acute Pancreatitis and its Mechanisms) Study is a prospective cohort study designed to address these and other knowledge gaps to provide the evidence needed to screen for, prevent, and treat DM after AP. In the following article, we summarize literature regarding the epidemiology of DM after AP and provide the rationale and an overview of the DREAM study.

The pool of beta cell-specific CD8+ T cells in type 1 diabetes (T1D) sustains an autoreactive potential despite having access to a constant source of antigen. To investigate the long-lived nature of these cells, we established a DNA methylation-based T cell 'multipotency index' and found that beta cell-specific CD8+ T cells retained a stem-like epigenetic multipotency score. Single-cell assay for transposase-accessible chromatin using sequencing confirmed the coexistence of naive and effector-associated epigenetic programs in individual beta cell-specific CD8+ T cells. Assessment of beta cell-specific CD8+ T cell anatomical distribution and the establishment of stem-associated epigenetic programs revealed that self-reactive CD8+ T cells isolated from murine lymphoid tissue retained developmentally plastic phenotypic and epigenetic profiles relative to the same cells isolated from the pancreas. Collectively, these data provide new insight into the longevity of beta cell-specific CD8+ T cell responses and document the use of this methylation-based multipotency index for investigating human and mouse CD8+ T cell differentiation.

Islet autoantibodies are predominantly measured by radioassay to facilitate risk assessment and diagnosis of type 1 diabetes. However, the reliance on radioactive components, large sample volumes and limited throughput renders radioassay testing costly and challenging. We developed a multiplex analysis platform based on antibody detection by agglutination-PCR (ADAP) for the sample-sparing measurement of GAD, IA-2 and insulin autoantibodies/antibodies in 1 μL serum. The assay was developed and validated in 7 distinct cohorts (n = 858) with the majority of the cohorts blinded prior to analysis. Measurements from the ADAP assay were compared to radioassay to determine correlation, concordance, agreement, clinical sensitivity and specificity. The average overall agreement between ADAP and radioassay was above 91%. The average clinical sensitivity and specificity were 96% and 97%. In the IASP 2018 workshop, ADAP achieved the highest sensitivity of all assays tested at 95% specificity (AS95) rating for GAD and IA-2 autoantibodies and top-tier performance for insulin autoantibodies. Furthermore, ADAP correctly identified 95% high-risk individuals with two or more autoantibodies by radioassay amongst 39 relatives of T1D patients tested. In conclusion, the new ADAP assay can reliably detect the three cardinal islet autoantibodies/antibodies in 1μL serum with high sensitivity. This novel assay may improve pediatric testing compliance and facilitate easier community-wide screening for islet autoantibodies.

Hyaluronan (HA), an extracellular matrix glycosaminoglycan, is implicated in the pathogenesis of both type 1 diabetes (T1D) as well as type 2 diabetes (T2D) and has been postulated to be increased in these diseases due to hyperglycemia. We have examined the serum and tissue distribution of HA in human subjects with T1D and T2D and in mouse models of these diseases and evaluated the relationship between HA levels and glycemic control. We found that serum HA levels are increased in T2D but not T1D independently of hemoglobin-A1c, C-peptide, body mass index, or time since diabetes diagnosis. HA is likewise increased in skeletal muscle in T2D subjects relative to non-diabetic controls. Analogous increases in serum and muscle HA are seen in diabetic db/db mice (T2D), but not in diabetic DORmO mice (T1D). Diabetes induced by the β-cell toxin streptozotozin (STZ) lead to an increase in blood glucose but not to an increase in serum HA. These data indicate that HA levels are increased in multiple tissue compartments in T2D but not T1D independently of glycemic control. Given that T2D but not T1D is associated with systemic inflammation, these patterns are consistent with inflammatory factors and not hyperglycemia driving increased HA. Serum HA may have value as a biomarker of systemic inflammation in T2D.

Context: The presence of islet autoimmunity identifies individuals likely to progress to clinical type 1 diabetes (T1D). In clinical research studies, autoantibody screening followed by regular metabolic monitoring every 6 months reduces incidence of diabetic ketoacidosis (DKA) at diagnosis. Objective: We hypothesized that DKA reduction can be achieved on a population basis with a reduced frequency of metabolic monitoring visits. We reasoned that prolonged time between the development of T1D and the time of clinical diagnosis ("undiagnosed time") would more commonly result in DKA and thus that limiting undiagnosed time would decrease DKA. Methods: An analysis was conducted of data from TrialNet's Pathway to Prevention (PTP), a cross-sectional longitudinal study that identifies and follows at-risk relatives of people with T1D. PTP is a population-based study enrolling across multiple countries. A total of 6193 autoantibody (AAB)-positive individuals participated in PTP from March 2004 to April 2019. We developed models of progression to clinical diagnosis for pediatric and adult populations with single or multiple AAB, and summarized results using estimated hazard rate. An optimal monitoring visit schedule was determined for each model to achieve a minimum average level of undiagnosed time for each population. Results: Halving the number of monitoring visits usually conducted in research studies is likely to substantially lower the population incidence of DKA at diagnosis of T1D. Conclusion: Our study has clinical implications for the metabolic monitoring of at-risk individuals. Fewer monitoring visits would reduce the clinical burden, suggesting a path toward transitioning monitoring beyond the research setting. [ABSTRACT FROM AUTHOR]

The early to mid-1980s were an inflection point in the history of type 1 diabetes research. Two landmark events occurred: the initiation of immune-based interventions seeking to prevent type 1 diabetes and the presentation of an innovative model describing the disorder's natural history. Both formed the basis for hundreds of subsequent studies designed to achieve a dramatic therapeutic goal-a means to prevent and/or reverse type 1 diabetes. However, the need to screen large numbers of individuals and prospectively monitor them using immunologic and metabolic tests for extended periods of time suggested such efforts would require a large collaborative network. Hence, the National Institutes of Health formed the landmark Diabetes Prevention Trial-Type 1 (DPT-1) in the mid-1990s, an effort that led to Type 1 Diabetes TrialNet. TrialNet studies have helped identify novel biomarkers; delineate type 1 diabetes progression, resulting in identification of highly predictable stages defined by the accumulation of autoantibodies (stage 1), dysglycemia (stage 2), and disease meeting clinical criteria for diagnosis (stage 3); and oversee numerous clinical trials aimed at preventing disease progression. Such efforts pave the way for stage-specific intervention trials with improved hope that a means to effectively disrupt the disorder's development will be identified. [ABSTRACT FROM AUTHOR]

Aims/hypothesis: Validated biomarkers are needed to monitor the effects of immune intervention in individuals with type 1 diabetes. Despite their importance, few options exist for monitoring antigen-specific T cells. Previous reports described a combinatorial approach that enables the simultaneous detection and quantification of multiple islet-specific CD8+ T cell populations. Here, we set out to evaluate the performance of a combinatorial HLA-A2 multimer assay in a multi-centre setting.Methods: The combinatorial HLA-A2 multimer assay was applied in five participating centres using centralised reagents and blinded replicate samples. In preliminary experiments, samples from healthy donors were analysed using recall antigen multimers. In subsequent experiments, samples from healthy donors and individuals with type 1 diabetes were analysed using beta cell antigen and recall antigen multimers.Results: The combinatorial assay was successfully implemented in each participating centre, with CVs between replicate samples that indicated good reproducibility for viral epitopes (mean %CV = 33.8). For beta cell epitopes, the assay was very effective in a single-centre setting (mean %CV = 18.4), but showed sixfold greater variability across multi-centre replicates (mean %CV = 119). In general, beta cell antigen-specific CD8+ T cells were detected more commonly in individuals with type 1 diabetes than in healthy donors. Furthermore, CD8+ T cells recognising HLA-A2-restricted insulin and glutamate decarboxylase epitopes were found to occur at higher frequencies in individuals with type 1 diabetes than in healthy donors.Conclusions/interpretation: Our results suggest that, although combinatorial multimer assays are challenging, they can be implemented in multiple laboratories, providing relevant T cell frequency measurements. Assay reproducibility was notably higher in the single-centre setting, suggesting that biomarker analysis of clinical trial samples would be most successful when assays are performed in a single laboratory. Technical improvements, including further standardisation of cytometry platforms, will likely be necessary to reduce assay variability in the multi-centre setting. [ABSTRACT FROM AUTHOR]

The authors respond to comments on a 2019 article by Sims et al on proinsulin secretion in type 1 diabetes. They state that replicates and concentrations tested across standard curves for proinsulin, proinsulin split products, C-peptide, and insulin are required to be able to respond adequately to comments. Also noted are the authors' openness for participation in future collaborative efforts on testing differences in assays and comparison of results.

Although most patients with type 1 diabetes (T1D) retain some functional insulin-producing islet β cells at the time of diagnosis, the rate of further β cell loss varies across individuals. It is not clear what drives this differential progression rate. CD8+ T cells have been implicated in the autoimmune destruction of β cells. Here, we addressed whether the phenotype and function of autoreactive CD8+ T cells influence disease progression. We identified islet-specific CD8+ T cells using high-content, single-cell mass cytometry in combination with peptide-loaded MHC tetramer staining. We applied a new analytical method, DISCOV-R, to characterize these rare subsets. Autoreactive T cells were phenotypically heterogeneous, and their phenotype differed by rate of disease progression. Activated islet-specific CD8+ memory T cells were prevalent in subjects with T1D who experienced rapid loss of C-peptide; in contrast, slow disease progression was associated with an exhaustion-like profile, with expression of multiple inhibitory receptors, limited cytokine production, and reduced proliferative capacity. This relationship between properties of autoreactive CD8+ T cells and the rate of T1D disease progression after onset make these phenotypes attractive putative biomarkers of disease trajectory and treatment response and reveal potential targets for therapeutic intervention. [ABSTRACT FROM AUTHOR]

Highlights • Proinsulin specific T cells are predominant in peripheral blood. • Subjects with type 1 diabetes exhibit different patterns of T cell reactivity. • The breadth of responses targeted by CD8+ and CD4+ T cells are correlated. Abstract CD4+ T cell responses are thought to play a role in type 1 diabetes (T1D). However, detection and characterization of T cells that respond to beta cell epitopes in subjects with T1D has been limited by technical obstacles, including the inherently low frequencies in peripheral blood and variable responsiveness of individual subjects to single epitopes. We implemented a multicolor staining approach that allows direct ex vivo characterization of multiple CD4+ T cell specificities in a single sample. Here we demonstrate and apply that multicolor approach to directly measure the frequency and phenotype of beta cell specific CD4+ T cells in T1D patients and HLA matched controls. For this work we utilized five DR0401 restricted peptides from proinsulin, GAD65, IA-2, and IGRP, which were previously reported as disease relevant epitopes. Surprisingly, although responses to each of these peptides can be readily detected after in vitro expansion, our results indicated that only proinsulin specific T cells were consistently detectable at moderate frequencies in subjects with T1D. Characterization of beta cell specific CD4+ T cells revealed only modest differences between subjects with T1D and healthy controls. Subjects with T1D did have higher proportions of CD45RA negative epitope specific T cells than controls. In patients epitope specific T cells were often CXCR3 positive and a substantial proportion were CCR7 negative, suggesting a Th1-like effector phenotype. Finally, we demonstrated that our multicolor staining approach is compatible with class I multimer analysis, facilitating the characterization of self-reactive CD4+ and CD8+ T cells using a single sample. [ABSTRACT FROM AUTHOR]