Journal articles
Leete P, Oram R, McDonald T, Ziller C, Hattersley A, Richardson S, Morgan N (In Press). Studies of insulin and proinsulin in pancreas and serum support the existence of aetiopathological endotypes of type 1 diabetes associated with age at diagnosis. Diabetologia
Ifie E, Russell M, Dhayal S, Leete P, Sebastiani G, Nigi L, Dotta F, Marjomäki V, Eizirik D, Morgan N, et al (In Press). Unexpected subcellular distribution of a specific isoform of the Coxsackie and adenovirus receptor, CAR-SIV, in human pancreatic beta cells. Diabetologia
Torabi F, Vadakekolathu J, Wyatt R, Leete P, Tombs MA, Richardson CC, Boocock DJ, Turner MD, Morgan NG, Richardson SJ, et al (2023). Differential expression of genes controlling lymphocyte differentiation and migration in two distinct endotypes of type 1 diabetes.
Diabet Med,
40(9).
Abstract:
Differential expression of genes controlling lymphocyte differentiation and migration in two distinct endotypes of type 1 diabetes.
AIMS: Morphological studies of pancreas samples obtained from young people with recent-onset type 1 diabetes have revealed distinct patterns of immune cell infiltration of the pancreatic islets suggestive of two age-associated type 1 diabetes endotypes that differ by inflammatory responses and rates of disease progression. The objective of this study was to investigate whether these proposed disease endotypes are associated with pathological differences in immune cell activation and cytokine secretion by applying multiplexed gene expression analysis to pancreatic tissue from recent-onset type 1 diabetes cases. METHODS: RNA was extracted from samples of fixed, paraffin-embedded pancreas tissue from type 1 diabetes cases characterised by endotype and from controls without diabetes. Expression levels of 750 genes associated with autoimmune inflammation were determined by hybridisation to a panel of capture and reporter probes and these were counted as a measure of gene expression. Normalised counts were analysed for differences in expression between 29 type 1 diabetes cases and 7 controls without diabetes, and between the two type 1 diabetes endotypes. RESULTS: Ten inflammation-associated genes, including INS, were significantly under-expressed in both endotypes and 48 genes were more highly expressed. A different set of 13 genes associated with the development, activation and migration of lymphocytes was uniquely overexpressed in the pancreas of people developing diabetes at younger age. CONCLUSIONS: the results provide evidence that histologically defined type 1 diabetes endotypes differ in their immunopathology and identify inflammatory pathways specifically involved in disease developing at a young age, essential for a better understanding of disease heterogeneity.
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Boldison J, Hopkinson JR, Davies J, Pearson JA, Leete P, Richardson S, Morgan NG, Wong FS (2023). Gene expression profiling in NOD mice reveals that B cells are highly educated by the pancreatic environment during autoimmune diabetes.
Diabetologia,
66(3), 551-566.
Abstract:
Gene expression profiling in NOD mice reveals that B cells are highly educated by the pancreatic environment during autoimmune diabetes
Aims/hypothesis: B cells play an important role in driving the development of type 1 diabetes; however, it remains unclear how they contribute to local beta cell destruction during disease progression. Here, we use gene expression profiling of B cell subsets identified in inflamed pancreatic tissue to explore their primary functional role during the progression of autoimmune diabetes. Methods: Transcriptional profiling was performed on FACS-sorted B cell subsets isolated from pancreatic islets and the pancreatic lymph nodes of NOD mice. Results: B cells are highly modified by the inflamed pancreatic tissue and can be distinguished by their transcriptional profile from those in the lymph nodes. We identified both a discrete and a core shared gene expression profile in islet CD19+CD138– and CD19+CD138+ B cell subsets, the latter of which is known to have enriched autoreactivity during diabetes development. On localisation to pancreatic islets, compared with CD138– B cells, CD138+ B cells overexpress genes associated with adhesion molecules and growth factors. Their shared signature consists of gene expression changes related to the differentiation of antibody-secreting cells and gene regulatory networks associated with IFN signalling pathways, proinflammatory cytokines and Toll-like receptor (TLR) activation. Finally, abundant TLR7 expression was detected in islet B cells and was enhanced specifically in CD138+ B cells. Conclusions/interpretation: Our study provides a detailed transcriptional analysis of islet B cells. Specific gene signatures and interaction networks have been identified that point towards a functional role for B cells in driving autoimmune diabetes. Graphical abstract: [Figure not available: see fulltext.].
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Carr ALJ, Inshaw JRJ, Flaxman CS, Leete P, Wyatt RC, Russell LA, Palmer M, Prasolov D, Worthington T, Hull B, et al (2022). Circulating C-Peptide Levels in Living Children and Young People and Pancreatic β-Cell Loss in Pancreas Donors Across Type 1 Diabetes Disease Duration.
Diabetes,
71(7), 1591-1596.
Abstract:
Circulating C-Peptide Levels in Living Children and Young People and Pancreatic β-Cell Loss in Pancreas Donors Across Type 1 Diabetes Disease Duration
C-peptide declines in type 1 diabetes, although many long-duration patients retain low, but detectable levels. Histological analyses confirm that β-cells can remain following type 1 diabetes onset. We explored the trends observed in C-peptide decline in the UK Genetic Resource Investigating Diabetes (UK GRID) cohort (N = 4,079), with β-cell loss in pancreas donors from the network for Pancreatic Organ donors with Diabetes (nPOD) biobank and the Exeter Archival Diabetes Biobank (EADB) (combined N = 235), stratified by recently reported age at diagnosis endotypes (<7, 7–12, ≥13 years) across increasing diabetes durations. The proportion of individuals with detectable C-peptide declined beyond the first year after diagnosis, but this was most marked in the youngest age group (<1-year duration: age <7 years: 18 of 20 [90%], 7–12 years: 107 of 110 [97%], ≥13 years: 58 of 61 [95%] vs. 1–5 years postdiagnosis: <7 years: 172 of 522 [33%], 7–12 years: 604 of 995 [61%], ≥13 years: 225 of 289 [78%]). A similar profile was observed in β-cell loss, with those diagnosed at younger ages experiencing more rapid loss of islets containing insulin-positive (insulin+) β-cells <1 year postdiagnosis: age <7 years: 23 of 26 (88%), 7–12 years: 32 of 33 (97%), ≥13 years: 22 of 25 (88%) vs. 1–5 years postdiagnosis: <7 years: 1 of 12 (8.3%), 7–12 years: 7 of 13 (54%), ≥13 years: 7 of 8 (88%). These data should be considered in the planning and interpretation of intervention trials designed to promote β-cell retention and function.
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Krogvold L, Leete P, Mynarek IM, Russell MA, Gerling IC, Lenchik NI, Mathews C, Richardson SJ, Morgan NG, Dahl-Jørgensen K, et al (2022). Detection of Antiviral Tissue Responses and Increased Cell Stress in the Pancreatic Islets of Newly Diagnosed Type 1 Diabetes Patients: Results from the DiViD Study.
Frontiers in Endocrinology,
13Abstract:
Detection of Antiviral Tissue Responses and Increased Cell Stress in the Pancreatic Islets of Newly Diagnosed Type 1 Diabetes Patients: Results from the DiViD Study
Aims/hypothesisThe Diabetes Virus Detection (DiViD) study has suggested the presence of low-grade enteroviral infection in pancreatic tissue collected from six of six live adult patients newly diagnosed with type 1 diabetes. The present study aimed to compare the gene and protein expression of selected virally induced pathogen recognition receptors and interferon stimulated genes in islets from these newly diagnosed type 1 diabetes (DiViD) subjects vs age-matched non-diabetic (ND) controls.MethodsRNA was extracted from laser-captured islets and Affymetrix Human Gene 2.0 ST arrays used to obtain gene expression profiles. Lists of differentially expressed genes were subjected to a data-mining pipeline searching for enrichment of canonical pathways, KEGG pathways, Gene Ontologies, transcription factor binding sites and other upstream regulators. In addition, the presence and localisation of specific viral response proteins (PKR, MxA and MDA5) were examined by combined immunofluorescent labelling in sections of pancreatic tissue.ResultsThe data analysis and data mining process revealed a significant enrichment of gene ontologies covering viral reproduction and infectious cycles; peptide translation, elongation and initiation, as well as oxidoreductase activity. Enrichment was identified in the KEGG pathways for oxidative phosphorylation; ribosomal and metabolic activity; antigen processing and presentation and in canonical pathways for mitochondrial dysfunction, oxidative phosphorylation and EIF2 signaling. Protein Kinase R (PKR) expression did not differ between newly diagnosed type 1 diabetes and ND islets at the level of total RNA, but a small subset of β-cells displayed markedly increased PKR protein levels. These PKR+ β-cells correspond to those previously shown to contain the viral protein, VP1. RNA encoding MDA5 was increased significantly in newly diagnosed type 1 diabetes islets, and immunostaining of MDA5 protein was seen in α- and certain β-cells in both newly diagnosed type 1 diabetes and ND islets, but the expression was increased in β-cells in type 1 diabetes. In addition, an uncharacterised subset of synaptophysin positive, but islet hormone negative, cells expressed intense MDA5 staining and these were more prevalent in DiViD cases. MxA RNA was upregulated in newly diagnosed type 1 diabetes vs ND islets and MxA protein was detected exclusively in newly diagnosed type 1 diabetes β-cells.Conclusion/interpretationThe gene expression signatures reveal that pathways associated with cellular stress and increased immunological activity are enhanced in islets from newly diagnosed type 1 diabetes patients compared to controls. The increases in viral response proteins seen in β-cells in newly diagnosed type 1 diabetes provide clear evidence for the activation of IFN signalling pathways. As such, these data strengthen the hypothesis that an enteroviral infection of islet β-cells contributes to the pathogenesis of type 1 diabetes.
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Rodriguez-Calvo T, Chen Y-C, Verchere CB, Haataja L, Arvan P, Leete P, Richardson SJ, Morgan NG, Qian W-J, Pugliese A, et al (2021). Altered β-Cell Prohormone Processing and Secretion in Type 1 Diabetes.
Diabetes,
70(5), 1038-1050.
Abstract:
Altered β-Cell Prohormone Processing and Secretion in Type 1 Diabetes.
Analysis of data from clinical cohorts, and more recently from human pancreatic tissue, indicates that reduced prohormone processing is an early and persistent finding in type 1 diabetes. In this article, we review the current state of knowledge regarding alterations in islet prohormone expression and processing in type 1 diabetes and consider the clinical impact of these findings. Lingering questions, including pathologic etiologies and consequences of altered prohormone expression and secretion in type 1 diabetes, and the natural history of circulating prohormone production in health and disease, are considered. Finally, key next steps required to move forward in this area are outlined, including longitudinal testing of relevant clinical populations, studies that probe the genetics of altered prohormone processing, the need for combined functional and histologic testing of human pancreatic tissues, continued interrogation of the intersection between prohormone processing and autoimmunity, and optimal approaches for analysis. Successful resolution of these questions may offer the potential to use altered prohormone processing as a biomarker to inform therapeutic strategies aimed at personalized intervention during the natural history of type 1 diabetes and as a pathogenic anchor for identification of potential disease-specific endotypes.
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Leete P, Morgan NG (2021). Footprints of Immune Cells in the Pancreas in Type 1 Diabetes; to "B" or Not to "B": is That Still the Question?.
Front Endocrinol (Lausanne),
12Abstract:
Footprints of Immune Cells in the Pancreas in Type 1 Diabetes; to "B" or Not to "B": is That Still the Question?
Significant progress has been made in understanding the phenotypes of circulating immune cell sub-populations in human type 1 diabetes but much less is known about the equivalent populations that infiltrate the islets to cause beta-cell loss. In particular, considerable uncertainties remain about the phenotype and role of B-lymphocytes in the pancreas. This gap in understanding reflects both the difficulty in accessing the gland to study islet inflammation during disease progression and the fact that the number and proportion of islet-associated B-lymphocytes varies significantly according to the disease endotype. In very young children (especially those
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Russell M, Leete P (2019). Glucocorticoids: novel agents to stimulate beta-cell neogenesis?. Annals of Translational Medicine, 7(8).
Russell MA, Redick SD, Blodgett DM, Richardson SJ, Leete P, Krogvold L, Dahl-Jørgensen K, Bottino R, Brissova M, Spaeth JM, et al (2019). HLA Class II Antigen Processing and Presentation Pathway Components Demonstrated by Transcriptome and Protein Analyses of Islet β-Cells from Donors with Type 1 Diabetes.
Diabetes,
68(5), 988-1001.
Abstract:
HLA Class II Antigen Processing and Presentation Pathway Components Demonstrated by Transcriptome and Protein Analyses of Islet β-Cells from Donors with Type 1 Diabetes.
Type 1 diabetes studies consistently generate data showing islet β-cell dysfunction and T cell-mediated anti-β-cell-specific autoimmunity. To explore the pathogenesis, we interrogated the β-cell transcriptomes from donors with and without type 1 diabetes using both bulk-sorted and single β-cells. Consistent with immunohistological studies, β-cells from donors with type 1 diabetes displayed increased Class I transcripts and associated mRNA species. These β-cells also expressed mRNA for Class II and Class II antigen presentation pathway components, but lacked the macrophage marker CD68. Immunohistological study of three independent cohorts of donors with recent-onset type 1 diabetes showed Class II protein and its transcriptional regulator Class II MHC trans-activator protein expressed by a subset of insulin+CD68- β-cells, specifically found in islets with lymphocytic infiltrates. β-Cell surface expression of HLA Class II was detected on a portion of CD45-insulin+ β-cells from donors with type 1 diabetes by immunofluorescence and flow cytometry. Our data demonstrate that pancreatic β-cells from donors with type 1 diabetes express Class II molecules on selected cells with other key genes in those pathways and inflammation-associated genes. β-Cell expression of Class II molecules suggests that β-cells may interact directly with islet-infiltrating CD4+ T cells and may play an immunopathogenic role.
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Vecchio F, Lo Buono N, Stabilini A, Nigi L, Dufort MJ, Geyer S, Rancoita PM, Cugnata F, Mandelli A, Valle A, et al (2018). Abnormal neutrophil signature in the blood and pancreas of presymptomatic and symptomatic type 1 diabetes.
JCI Insight,
3(18).
Abstract:
Abnormal neutrophil signature in the blood and pancreas of presymptomatic and symptomatic type 1 diabetes.
BACKGROUND: Neutrophils and their inflammatory mediators are key pathogenic components in multiple autoimmune diseases, while their role in human type 1 diabetes (T1D), a disease that progresses sequentially through identifiable stages prior to the clinical onset, is not well understood. We previously reported that the number of circulating neutrophils is reduced in patients with T1D and in presymptomatic at-risk subjects. The aim of the present work was to identify possible changes in circulating and pancreas-residing neutrophils throughout the disease course to better elucidate neutrophil involvement in human T1D. METHODS: Data collected from 389 subjects at risk of developing T1D, and enrolled in 4 distinct studies performed by TrialNet, were analyzed with comprehensive statistical approaches to determine whether the number of circulating neutrophils correlates with pancreas function. To obtain a broad analysis of pancreas-infiltrating neutrophils throughout all disease stages, pancreas sections collected worldwide from 4 different cohorts (i.e. nPOD, DiViD, Siena, and Exeter) were analyzed by immunohistochemistry and immunofluorescence. Finally, circulating neutrophils were purified from unrelated nondiabetic subjects and donors at various T1D stages and their transcriptomic signature was determined by RNA sequencing. RESULTS: Here, we show that the decline in β cell function is greatest in individuals with the lowest peripheral neutrophil numbers. Neutrophils infiltrate the pancreas prior to the onset of symptoms and they continue to do so as the disease progresses. of interest, a fraction of these pancreas-infiltrating neutrophils also extrudes neutrophil extracellular traps (NETs), suggesting a tissue-specific pathogenic role. Whole-transcriptome analysis of purified blood neutrophils revealed a unique molecular signature that is distinguished by an overabundance of IFN-associated genes; despite being healthy, said signature is already present in T1D-autoantibody-negative at-risk subjects. CONCLUSIONS: These results reveal an unexpected abnormality in neutrophil disposition both in the circulation and in the pancreas of presymptomatic and symptomatic T1D subjects, implying that targeting neutrophils might represent a previously unrecognized therapeutic modality. FUNDING: Juvenile Diabetes Research Foundation (JDRF), NIH, Diabetes UK.
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Shields B, McDonald T, Oram R, Hill A, Hudson M, Leete P, Pearson E, Richardson S, Morgan N, Hattersley A, et al (2018). C-peptide decline in type 1 diabetes has two phases: an initial exponential fall and a subsequent stable phase. Diabetes Care
Colli ML, Hill JLE, Marroquí L, Chaffey J, Dos Santos RS, Leete P, Coomans de Brachène A, Paula FMM, Op de Beeck A, Castela A, et al (2018). PDL1 is expressed in the islets of people with type 1 diabetes and is up-regulated by interferons-α and-γ via IRF1 induction.
EBioMedicine,
36, 367-375.
Abstract:
PDL1 is expressed in the islets of people with type 1 diabetes and is up-regulated by interferons-α and-γ via IRF1 induction.
BACKGROUND: Antibodies targeting PD-1 and its ligand PDL1 are used in cancer immunotherapy but may lead to autoimmune diseases, including type 1 diabetes (T1D). It remains unclear whether PDL1 is expressed in pancreatic islets of people with T1D and how is it regulated. METHODS: the expression of PDL1, IRF1, insulin and glucagon was evaluated in samples of T1D donors by immunofluorescence. Cytokine-induced PDL1 expression in the human beta cell line, EndoC-βH1, and in primary human pancreatic islets was determined by real-time RT-PCR, flow cytometry and Western blot. Specific and previously validated small interference RNAs were used to inhibit STAT1, STAT2, IRF1 and JAK1 signaling. Key results were validated using the JAK inhibitor Ruxolitinib. FINDINGS: PDL1 was present in insulin-positive cells from twelve T1D individuals (6 living and 6 deceased donors) but absent from insulin-deficient islets or from the islets of six non-diabetic controls. Interferons-α and -γ, but not interleukin-1β, induced PDL1 expression in vitro in human islet cells and EndoC-βH1 cells. Silencing of STAT1 or STAT2 individually did not prevent interferon-α-induced PDL1, while blocking of JAKs - a proposed therapeutic strategy for T1D - or IRF1 prevented PDL1 induction. INTERPRETATION: These findings indicate that PDL1 is expressed in beta cells from people with T1D, possibly to attenuate the autoimmune assault, and that it is induced by both type I and II interferons via IRF1.
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Leete P, Mallone R, Richardson SJ, Sosenko JM, Redondo MJ, Evans-Molina C (2018). The Effect of Age on the Progression and Severity of Type 1 Diabetes: Potential Effects on Disease Mechanisms.
Current Diabetes Reports,
18(11).
Abstract:
The Effect of Age on the Progression and Severity of Type 1 Diabetes: Potential Effects on Disease Mechanisms
Purpose of Review: to explore the impact of age on type 1 diabetes (T1D) pathogenesis. Recent Findings: Children progress more rapidly from autoantibody positivity to T1D and have lower C-peptide levels compared to adults. In histological analysis of post-mortem pancreata, younger age of diagnosis is associated with reduced numbers of insulin containing islets and a hyper-immune CD20hi infiltrate. Moreover compared to adults, children exhibit decreased immune regulatory function and increased engagement and trafficking of autoreactive CD8+ T cells, and age-related differences in β cell vulnerability may also contribute to the more aggressive immune phenotype observed in children. To account for some of these differences, HLA and non-HLA genetic loci that influence multiple disease characteristics, including age of onset, are being increasingly characterized. Summary: the exception of T1D as an autoimmune disease more prevalent in children than adults results from a combination of immune, metabolic, and genetic factors. Age-related differences in T1D pathology have important implications for better tailoring of immunotherapies.
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Hodik M, Anagandula M, Fuxe J, Krogvold L, Dahl-Jørgensen K, Hyöty H, Sarmiento L, Frisk G, Atkinson M, Pugliese A, et al (2016). Coxsackie-adenovirus receptor expression is enhanced in pancreas from patients with type 1 diabetes.
BMJ Open Diabetes Research and Care,
4(1).
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Coxsackie-adenovirus receptor expression is enhanced in pancreas from patients with type 1 diabetes
Objectives: One of the theories connecting enterovirus (EV) infection of human islets with type 1 diabetes (T1D) is the development of a fertile field in the islets. This implies induction of appropriate proteins for the viral replication such as the coxsackie- adenovirus receptor (CAR). The aim of this study was to investigate to what extent CAR is expressed in human islets of Langerhans, and what conditions that would change the expression. Design: Immunohistochemistry for CAR was performed on paraffin-embedded pancreatic tissue from patients with T1D (n=9 recent onset T1D, n=4 long-standing T1D), islet autoantibody-positive individuals (n=14) and non-diabetic controls (n=24) individuals. The expression of CAR was also examined by reverse transcription PCR on microdissected islets (n=5), exocrine tissue (n=5) and on explanted islets infected with EV or exposed to chemokines produced by EV-infected islet cells. Results: an increased frequency of patients with T1D and autoantibody-positive individuals expressed CAR in the pancreas (p
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Leete P, Willcox A, Krogvold L, Dahl-Jørgensen K, Foulis AK, Richardson SJ, Morgan NG (2016). Differential Insulitic Profiles Determine the Extent of β-Cell Destruction and the Age at Onset of Type 1 Diabetes.
Diabetes,
65(5), 1362-1369.
Abstract:
Differential Insulitic Profiles Determine the Extent of β-Cell Destruction and the Age at Onset of Type 1 Diabetes.
Type 1 diabetes (T1D) results from a T cell-mediated destruction of pancreatic β-cells following the infiltration of leukocytes (including CD8(+), CD4(+), and CD20(+) cells) into and around pancreatic islets (insulitis). Recently, we reported that two distinct patterns of insulitis occur in patients with recent-onset T1D from the U.K. and that these differ principally in the proportion of infiltrating CD20(+) B cells (designated CD20Hi and CD20Lo, respectively). We have now extended this analysis to include patients from the Network for Pancreatic Organ Donors with Diabetes (U.S.) and Diabetes Virus Detection (DiViD) study (Norway) cohorts and confirm that the two profiles of insulitis occur more widely. Moreover, we show that patients can be directly stratified according to their insulitic profile and that those receiving a diagnosis before the age of 7 years always display the CD20Hi profile. By contrast, individuals who received a diagnosis beyond the age of 13 years are uniformly defined as CD20Lo. This implies that the two forms of insulitis are differentially aggressive and that patients with a CD20Hi profile lose their β-cells at a more rapid rate. In support of this, we also find that the proportion of residual insulin-containing islets (ICIs) increases in parallel with age at the onset of T1D. Importantly, those receiving a diagnosis in, or beyond, their teenage years retain ∼40% ICIs at diagnosis, implying that a functional deficit rather than an absolute β-cell loss may be causal for disease onset in these patients. We conclude that appropriate patient stratification will be critical for correct interpretation of the outcomes of intervention therapies targeted to islet-infiltrating immune cells in T1D.
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Richardson SJ, Rodriguez-Calvo T, Gerling IC, Mathews CE, Kaddis JS, Russell MA, Zeissler M, Leete P, Krogvold L, Dahl-Jørgensen K, et al (2016). Islet cell hyperexpression of HLA class I antigens: a defining feature in type 1 diabetes.
Diabetologia,
59(11), 2448-2458.
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Islet cell hyperexpression of HLA class I antigens: a defining feature in type 1 diabetes.
AIMS/HYPOTHESIS: Human pancreatic beta cells may be complicit in their own demise in type 1 diabetes, but how this occurs remains unclear. One potentially contributing factor is hyperexpression of HLA class I antigens. This was first described approximately 30 years ago, but has never been fully characterised and was recently challenged as artefactual. Therefore, we investigated HLA class I expression at the protein and RNA levels in pancreases from three cohorts of patients with type 1 diabetes. The principal aims were to consider whether HLA class I hyperexpression is artefactual and, if not, to determine the factors driving it. METHODS: Pancreas samples from type 1 diabetes patients with residual insulin-containing islets (n = 26) from the Network for Pancreatic Organ donors with Diabetes (nPOD), Diabetes Virus Detection study (DiViD) and UK recent-onset type 1 diabetes collections were immunostained for HLA class I isoforms, signal transducer and activator of transcription 1 (STAT1), NLR family CARD domain containing 5 (NLRC5) and islet hormones. RNA was extracted from islets isolated by laser-capture microdissection from nPOD and DiViD samples and analysed using gene-expression arrays. RESULTS: Hyperexpression of HLA class I was observed in the insulin-containing islets of type 1 diabetes patients from all three tissue collections, and was confirmed at both the RNA and protein levels. The expression of β2-microglobulin (a second component required for the generation of functional HLA class I complexes) was also elevated. Both 'classical' HLA class I isoforms (i.e. HLA-ABC) as well as a 'non-classical' HLA molecule, HLA-F, were hyperexpressed in insulin-containing islets. This hyperexpression did not correlate with detectable upregulation of the transcriptional regulator NLRC5. However, it was strongly associated with increased STAT1 expression in all three cohorts. Islet hyperexpression of HLA class I molecules occurred in the insulin-containing islets of patients with recent-onset type 1 diabetes and was also detectable in many patients with disease duration of up to 11 years, declining thereafter. CONCLUSIONS/INTERPRETATION: Islet cell HLA class I hyperexpression is not an artefact, but is a hallmark in the immunopathogenesis of type 1 diabetes. The response is closely associated with elevated expression of STAT1 and, together, these occur uniquely in patients with type 1 diabetes, thereby contributing to their selective susceptibility to autoimmune-mediated destruction.
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Krogvold L, Edwin B, Buanes T, Frisk G, Skog O, Anagandula M, Korsgren O, Undlien D, Eike MC, Richardson SJ, et al (2015). Detection of a low-grade enteroviral infection in the islets of langerhans of living patients newly diagnosed with type 1 diabetes.
Diabetes,
64(5), 1682-1687.
Abstract:
Detection of a low-grade enteroviral infection in the islets of langerhans of living patients newly diagnosed with type 1 diabetes.
The Diabetes Virus Detection study (DiViD) is the first to examine fresh pancreatic tissue at the diagnosis of type 1 diabetes for the presence of viruses. Minimal pancreatic tail resection was performed 3-9 weeks after onset of type 1 diabetes in six adult patients (age 24-35 years). The presence of enteroviral capsid protein 1 (VP1) and the expression of class I HLA were investigated by immunohistochemistry. Enterovirus RNA was analyzed from isolated pancreatic islets and from fresh-frozen whole pancreatic tissue using PCR and sequencing. Nondiabetic organ donors served as controls. VP1 was detected in the islets of all type 1 diabetic patients (two of nine controls). Hyperexpression of class I HLA molecules was found in the islets of all patients (one of nine controls). Enterovirus-specific RNA sequences were detected in four of six patients (zero of six controls). The results were confirmed in various laboratories. Only 1.7% of the islets contained VP1(+) cells, and the amount of enterovirus RNA was low. The results provide evidence for the presence of enterovirus in pancreatic islets of type 1 diabetic patients, which is consistent with the possibility that a low-grade enteroviral infection in the pancreatic islets contributes to disease progression in humans.
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Arif S, Leete P, Nguyen V, Marks K, Nor NM, Estorninho M, Kronenberg-Versteeg D, Bingley PJ, Todd JA, Guy C, et al (2015). Erratum. Blood and Islet Phenotypes Indicate Immunological Heterogeneity in Type 1 Diabetes. Diabetes 2014;63:3835-3845.
Diabetes,
64(9).
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Leete P, Richardson SJ, Krogvold L, Willcox A, Dahl-Jorgensen K, Foulis AK, Morgan NG (2015). Low B-lymphocyte numbers within the insulitic lesion correlates with an elevated proportion of persisting beta cells and later onset of disease in patients with Type 1 diabetes.
DIABETIC MEDICINE,
32, 63-63.
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Arif S, Leete P, Nguyen V, Marks K, Nor NM, Estorninho M, Kronenberg-Versteeg D, Bingley PJ, Todd JA, Guy C, et al (2014). Blood and islet phenotypes indicate immunological heterogeneity in type 1 diabetes.
Diabetes,
63(11), 3835-3845.
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Blood and islet phenotypes indicate immunological heterogeneity in type 1 diabetes.
Studies in type 1 diabetes indicate potential disease heterogeneity, notably in the rate of β-cell loss, responsiveness to immunotherapies, and, in limited studies, islet pathology. We sought evidence for different immunological phenotypes using two approaches. First, we defined blood autoimmune response phenotypes by combinatorial, multiparameter analysis of autoantibodies and autoreactive T-cell responses in 33 children/adolescents with newly diagnosed diabetes. Multidimensional cluster analysis showed two equal-sized patient agglomerations characterized by proinflammatory (interferon-γ-positive, multiautoantibody-positive) and partially regulated (interleukin-10-positive, pauci-autoantibody-positive) responses. Multiautoantibody-positive nondiabetic siblings at high risk of disease progression showed similar clustering. Additionally, pancreas samples obtained post mortem from a separate cohort of 21 children/adolescents with recently diagnosed type 1 diabetes were examined immunohistologically. This revealed two distinct types of insulitic lesions distinguishable by the degree of cellular infiltrate and presence of B cells that we termed "hyper-immune CD20Hi" and "pauci-immune CD20Lo." of note, subjects had only one infiltration phenotype and were partitioned by this into two equal-sized groups that differed significantly by age at diagnosis, with hyper-immune CD20Hi subjects being 5 years younger. These data indicate potentially related islet and blood autoimmune response phenotypes that coincide with and precede disease. We conclude that different immunopathological processes (endotypes) may underlie type 1 diabetes, carrying important implications for treatment and prevention strategies.
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Richardson SJ, Leete P, Dhayal S, Russell MA, Oikarinen M, Laiho JE, Svedin E, Lind K, Rosenling T, Chapman N, et al (2014). Detection of enterovirus in the islet cells of patients with type 1 diabetes: what do we learn from immunohistochemistry? Reply to Hansson SF, Korsgren S, Pontén F et al [letter].
Diabetologia,
57(3), 647-649.
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Leete P, Richardson SJ, Bone AJ, Foulis AK, Morgan NG (2014). Differential expression of anti-apoptotic B-cell lymphoma-2 (Bcl-2) family proteins by the islet endocrine cell subtypes in human pancreas.
DIABETIC MEDICINE,
31, 47-48.
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Richardson SJ, Leete P, Dhayal S, Russell MA, Oikarinen M, Laiho JE, Svedin E, Lind K, Rosenling T, Chapman N, et al (2014). Evaluation of the fidelity of immunolabelling obtained with clone 5D8/1, a monoclonal antibody directed against the enteroviral capsid protein, VP1, in human pancreas.
Diabetologia,
57(2), 392-401.
Abstract:
Evaluation of the fidelity of immunolabelling obtained with clone 5D8/1, a monoclonal antibody directed against the enteroviral capsid protein, VP1, in human pancreas.
AIMS/HYPOTHESIS: Enteroviral infection has been implicated in the development of islet autoimmunity in type 1 diabetes and enteroviral antigen expression has been detected by immunohistochemistry in the pancreatic beta cells of patients with recent-onset type 1 diabetes. However, the immunohistochemical evidence relies heavily on the use of a monoclonal antibody, clone 5D8/1, raised against an enteroviral capsid protein, VP1. Recent data suggest that the clone 5D8/1 may also recognise non-viral antigens; in particular, a component of the mitochondrial ATP synthase (ATP5B) and an isoform of creatine kinase (CKB). Therefore, we evaluated the fidelity of immunolabelling by clone 5D8/1 in the islets of patients with type 1 diabetes. METHODS: Enteroviral VP1, CKB and ATP5B expression were analysed by western blotting, RT-PCR and immunocytochemistry in a range of cultured cell lines, isolated human islets and human tissue. RESULTS: Clone 5D8/1 labelled CKB, but not ATP5B, on western blots performed under denaturing conditions. In cultured human cell lines, isolated human islets and pancreas sections from patients with type 1 diabetes, the immunolabelling of ATP5B, CKB and VP1 by 5D8/1 was readily distinguishable. Moreover, in a human tissue microarray displaying more than 80 different cells and tissues, only two (stomach and colon; both of which are potential sites of enterovirus infection) were immunopositive when stained with clone 5D8/1. CONCLUSIONS/INTERPRETATION: When used under carefully optimised conditions, the immunolabelling pattern detected in sections of human pancreas with clone 5D8/1 did not reflect cross-reactivity with either ATP5B or CKB. Rather, 5D8/1 is likely to be representative of enteroviral antigen expression.
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Morgan NG, Leete P, Foulis AK, Richardson SJ (2014). Islet inflammation in human type 1 diabetes mellitus.
IUBMB Life,
66(11), 723-734.
Abstract:
Islet inflammation in human type 1 diabetes mellitus.
Type 1 diabetes mellitus (T1DM) is caused by the selective deletion of pancreatic β-cells in response to an assault mounted within the pancreas by infiltrating immune cells. However, this apparently clear and focussed annunciation conceals a stark reality in which the cellular and molecular events leading to β-cell loss remain poorly understood in humans. This reflects the difficulty of studying these processes in living individuals and the fact that, using pathological specimens, islet inflammation has been analysed in fewer than 200 recent-onset cases of T1DM worldwide, over the past century. Nevertheless, insights have been gained and the composition of the islet infiltrate is being disclosed. This is shown to be primarily lymphocytic in nature, with populations of both CD8+ and CD4+ T cells displaying an autoreactivity against specific islet antigenic peptides. The T cells are often accompanied by influent CD20+ B cells, although new data imply that the proportions of these individual cell types vary and that patients fall into at least two distinct categories having either a hyper-immune (CD20Hi) or a pauci-immune (CD20Lo) phenotype. The overall rate of β-cell decline appears to correlate with these two phenotypes such that hyper-immune patients lose β-cells more quickly and tend to develop disease at an earlier age than those with the pauci-immune profile. In this article, we review the evidence which underpins our current understanding of the aetiology of T1DM and highlight both the established features as well as areas of on-going ambiguity and debate.
Abstract.
Author URL.
Richardson SJ, Leete P, Bone AJ, Foulis AK, Morgan NG (2013). Expression of the enteroviral capsid protein VP1 in the islet cells of patients with type 1 diabetes is associated with induction of protein kinase R and downregulation of Mcl-1.
Diabetologia,
56(1), 185-193.
Abstract:
Expression of the enteroviral capsid protein VP1 in the islet cells of patients with type 1 diabetes is associated with induction of protein kinase R and downregulation of Mcl-1.
AIMS/HYPOTHESIS: Immunohistochemical staining reveals that the enteroviral capsid protein VP1 is present at higher frequency in the insulin-containing islets of patients with recent-onset type 1 diabetes than in controls. This is consistent with epidemiological evidence suggesting that enteroviral infection may contribute to the autoimmune response in type 1 diabetes. However, immunostaining of VP1 is not definitive since the antibody widely used to detect the protein (Clone 5D8/1) might also cross-react with additional proteins under some conditions. Therefore, we sought to verify that VP1 immunopositivity correlates with additional markers of viral infection. METHODS: Antigen immunoreactivity was examined in formalin-fixed, paraffin-embedded, pancreases from two different collections of type 1 diabetes and control cases: a historical collection from the UK and the nPOD (network of Pancreatic Organ donors with Diabetes) cohort from the USA. RESULTS: VP1 immunoreactivity was present in ~20% of insulin-containing islets of both cohorts under stringent conditions but was absent from insulin-deficient islets. The presence of VP1 was restricted to beta cells but only a minority of these contained the antigen. The innate viral sensor, protein kinase R (PKR) was upregulated selectively in beta cells that were immunopositive for VP1. The anti-apoptotic protein myeloid cell leukaemia sequence-1 (Mcl-1) was abundant in beta cells that were immunonegative for VP1 but Mcl-1 was depleted in cells containing VP1. CONCLUSIONS/INTERPRETATION: the presence of immunoreactive VP1 within beta cells in type 1 diabetes is associated with a cellular phenotype consistent with the activation of antiviral response pathways and enhanced sensitivity to apoptosis. However, definitive studies confirming whether viral infections are causal to beta cell loss in human diabetes are still awaited.
Abstract.
Author URL.
Lind K, Richardson SJ, Leete P, Morgan NG, Korsgren O, Flodström-Tullberg M (2013). Induction of an Antiviral State and Attenuated Coxsackievirus Replication in Type III Interferon Treated Primary Human Pancreatic Islets. Journal of virology
