Dataset Information

Islet-Derived eATP Fuels Autoreactive CD8⁺ T Cells and Facilitates the Onset of Type 1 Diabetes.

ABSTRACT: Extracellular ATP (eATP) activates T cells by engaging the P2X7R receptor. We identified two loss-of-function P2X7R mutations that are protective against type 1 diabetes (T1D) and thus hypothesized that eATP/P2X7R signaling may represent an early step in T1D onset. Specifically, we observed that in patients with newly diagnosed T1D, P2X7R is upregulated on CD8⁺ effector T cells in comparison with healthy control subjects. eATP is released at high levels by human/murine islets in vitro in high-glucose/inflammatory conditions, thus upregulating P2X7R on CD8⁺ T cells in vitro. P2X7R blockade with oxidized ATP reduces the CD8⁺ T cell-mediated autoimmune response in vitro and delays diabetes onset in NOD mice. Autoreactive CD8⁺ T-cell activation is highly dependent upon eATP/P2X7R-mediated priming, while a novel sP2X7R recombinant protein abrogates changes in metabolism and the autoimmune response associated with CD8⁺ T cells. eATP/P2X7R signaling facilitates the onset of autoimmune T1D by fueling autoreactive CD8⁺ cells and therefore represents a novel targeted therapeutic for the disorder.

SUBMITTER: Tezza S

PROVIDER: S-EPMC6905486 | biostudies-literature | 2018 Oct

REPOSITORIES: biostudies-literature

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Islet-Derived eATP Fuels Autoreactive CD8<sup>+</sup> T Cells and Facilitates the Onset of Type 1 Diabetes.

Tezza Sara S Ben Nasr Moufida M D'Addio Francesca F Vergani Andrea A Usuelli Vera V Falzoni Simonetta S Bassi Roberto R Dellepiane Sergio S Fotino Carmen C Rossi Chiara C Maestroni Anna A Solini Anna A Corradi Domenico D Giani Elisa E Mameli Chiara C Bertuzzi Federico F Pezzolesi Marcus G MG Wasserfall Clive H CH Atkinson Mark A MA Füchtbauer Ernst-Martin EM Ricordi Camillo C Folli Franco F Di Virgilio Francesco F Pileggi Antonello A Dhe-Paganon Sirano S Zuccotti Gian Vincenzo GV Fiorina Paolo P Fiorina Paolo P

Diabetes 20180731 10

Extracellular ATP (eATP) activates T cells by engaging the P2X7R receptor. We identified two loss-of-function P2X7R mutations that are protective against type 1 diabetes (T1D) and thus hypothesized that eATP/P2X7R signaling may represent an early step in T1D onset. Specifically, we observed that in patients with newly diagnosed T1D, P2X7R is upregulated on CD8<sup>+</sup> effector T cells in comparison with healthy control subjects. eATP is released at high levels by human/murine islets in vitro ...[more]

PMID: 30065030

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Project description:The notion that T cell insulitis increases as type 1 diabetes (T1D) develops is unsurprising, however, the quantitative analysis of CD4+ and CD8+ T cells within the islet mass is complex and limited with standard approaches. Optical microscopy is an important and widely used method to evaluate immune cell infiltration into pancreatic islets of Langerhans for the study of disease progression or therapeutic efficacy in murine T1D. However, the accuracy of this approach is often limited by subjective and potentially biased qualitative assessment of immune cell subsets. In addition, attempts at quantitative measurements require significant time for manual analysis and often involve sophisticated and expensive imaging software. In this study, we developed and illustrate here a streamlined analytical strategy for the rapid, automated and unbiased investigation of islet area and immune cell infiltration within (insulitis) and around (peri-insulitis) pancreatic islets. To this end, we demonstrate swift and accurate detection of islet borders by modeling cross-sectional islet areas with convex polygons (convex hulls) surrounding islet-associated insulin-producing β cell and glucagon-producing α cell fluorescent signals. To accomplish this, we used a macro produced with the freeware software ImageJ equipped with the Fiji Is Just ImageJ (FIJI) image processing package. Our image analysis procedure allows for direct quantification and statistical determination of islet area and infiltration in a reproducible manner, with location-specific data that more accurately reflect islet areas as insulitis proceeds throughout T1D. Using this approach, we quantified the islet area infiltrated with CD4+ and CD8+ T cells allowing statistical comparison between different age groups of non-obese diabetic (NOD) mice progressing towards T1D. We found significantly more CD4+ and CD8+ T cells infiltrating the convex hull-defined islet mass of 13-week-old non-diabetic and 17-week-old diabetic NOD mice compared to 4-week-old NOD mice. We also determined a significant and measurable loss of islet mass in mice that developed T1D. This approach will be helpful for the location-dependent quantitative calculation of islet mass and cellular infiltration during T1D pathogenesis and can be combined with other markers of inflammation or activation in future studies.

Project description:Aims/hypothesisThe aim of this study was to characterise islet autoantibody profiles and immune cell phenotypes in slow progressors to type 1 diabetes.MethodsImmunological variables were compared across peripheral blood samples obtained from slow progressors to type 1 diabetes, individuals with newly diagnosed or long-standing type 1 diabetes, and healthy individuals. Polychromatic flow cytometry was used to characterise the phenotypic attributes of B and T cells. Islet autoantigen-specific B cells were quantified using an enzyme-linked immunospot (ELISpot) assay and islet autoantigen-specific CD8+ T cells were quantified using peptide-HLA class I tetramers. Radioimmunoassays were used to detect islet autoantibodies. Sera were assayed for various chemokines, cytokines and soluble receptors via ELISAs.ResultsIslet autoantibodies were lost over time in slow progressors. Various B cell subsets expressed higher levels of CD95 in slow progressors, especially after polyclonal stimulation, compared with the corresponding B cell subsets in healthy donors (p < 0.05). The phenotypic characteristics of CD4+ and CD8+ T cells were similar in slow progressors and healthy donors. Lower frequencies of CD4+ T cells with a central memory phenotype (CD27int, CD127+, CD95int) were observed in slow progressors compared with healthy donors (mean percentage of total CD4+ T cells was 3.00% in slow progressors vs 4.67% in healthy donors, p < 0.05). Autoreactive B cell responses to proinsulin were detected at higher frequencies in slow progressors compared with healthy donors (median no. of spots was 0 in healthy donors vs 24.34 in slow progressors, p < 0.05) in an ELISpot assay. Islet autoantigen-specific CD8+ T cell responses were largely absent in slow progressors and healthy donors. Serum levels of DcR3, the decoy receptor for CD95L, were elevated in slow progressors compared with healthy donors (median was 1087 pg/ml in slow progressors vs 651 pg/ml in healthy donors, p = 0.06).Conclusions/interpretationIn this study, we found that slow progression to type 1 diabetes was associated with a loss of islet autoantibodies and a distinct B cell phenotype, consistent with enhanced apoptotic regulation of peripheral autoreactivity via CD95. These phenotypic changes warrant further studies in larger cohorts to determine their functional implications.

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Islet-Derived eATP Fuels Autoreactive CD8⁺ T Cells and Facilitates the Onset of Type 1 Diabetes.

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Islet-Derived eATP Fuels Autoreactive CD8<sup>+</sup> T Cells and Facilitates the Onset of Type 1 Diabetes.

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Islet-Derived eATP Fuels Autoreactive CD8+ T Cells and Facilitates the Onset of Type 1 Diabetes.

Publications

Islet-Derived eATP Fuels Autoreactive CD8<sup>+</sup> T Cells and Facilitates the Onset of Type 1 Diabetes.

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OmicsDI is part of the ELIXIR infrastructure

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Islet-Derived eATP Fuels Autoreactive CD8⁺ T Cells and Facilitates the Onset of Type 1 Diabetes.