Protein microarray analysis as a tool for monitoring cellular autoreactivity in type 1 diabetes patients and their relatives

Vrabelová, Zuzana; Koloušková, Stanislava; Böhmová, Kristýna; Faresjö, Maria; Šumnı́k, Zdenĕk; Pechová, M; Kverka, Miloslav; Chudoba, D; Zacharovová, K.; Stadlerova, Gabriela; Piťhová, Pavlína; Hladíková, Marie; Stechova, Katerina

doi:10.1111/j.1399-5448.2007.00308.x

Cited by 10 publications

(12 citation statements)

References 18 publications

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“…Nevertheless, the presence of IL-4Rs on normal human pancreatic β-cells is significant since it implies that, as in rats [16], these receptors may play a protective role against cell damage mediated by pro-inflammatory cytokines. Moreover, others have reported that elevated IFNγ/IL-4 ratios correlate with the presence of destructive insulitis and impaired Th2 cell cytokine production from cells that infiltrate the pancreas of diabetic patients [19][20][21]. In this context, it is interesting to note that a relative lack of IL-4 production has been reported in patients with Type 1 diabetes in many studies.…”

Section: Discussionmentioning

confidence: 92%

Human and rodent pancreatic β-cells express IL-4 receptors and IL-4 protects against β-cell apoptosis by activation of the PI3K and JAK/STAT pathways

et al. 2009

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Secretion of pro-inflammatory cytokines is associated with loss of pancreatic beta-cell viability and cell death. IL-4 (interleukin-4) has been reported to mediate a protective effect against the loss of pancreatic beta-cells, and IL-4 receptors have been found in rat pancreatic beta-cells at both the RNA and the protein level. The aim of the present study was to investigate IL-4 receptor expression in human islet cells and to examine the signalling pathways by which IL-4 exerts its effects using the rat beta-cell lines, BRIN-BD11 and INS-1E. By means of immunohistochemistry, it was demonstrated that IL-4 receptors are present on human islet cells. Using a flow cytometric method for evaluating cell death, it was confirmed that incubating beta-cells with IL-4 attenuated cell death induced by IL-1beta and interferon-gamma by approx. 65%. This effect was abrogated by the presence of the PI3K (phosphoinositide 3-kinase) inhibitor, wortmannin, suggesting that activation of the PI3K pathway is involved. In support of this, Western blotting revealed that incubation of cells with IL-4 resulted in increased phosphorylation of Akt (also called protein kinase B), a downstream target of PI3K. Increased tyrosine phosphorylation of STAT6 (signal transducer and activator of transcription 6) also occurred in response to IL-4 and a selective JAK3 (Janus kinase 3) inhibitor reduced the cytoprotective response. Both effects were prevented by overexpression of the tyrosine phosphatase, PTP-BL (protein tyrosine phosphatase-BL). We conclude that IL-4 receptors are functionally competent in pancreatic beta-cells and that they signal via PI3K and JAK/STAT pathways. These findings may have implications for future therapeutic strategies for the management of diabetes.

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Section: Discussionmentioning

confidence: 92%

Human and rodent pancreatic β-cells express IL-4 receptors and IL-4 protects against β-cell apoptosis by activation of the PI3K and JAK/STAT pathways

et al. 2009

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“…21 After 72 h, cells were washed and subjected to RNA extraction and supernatants were frozen at À801C until used for cytokine analysis.…”

Section: Immune Cell Isolationmentioning

confidence: 99%

Case report: type 1 diabetes in monozygotic quadruplets

et al. 2011

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Type 1 diabetes (T1D) is an autoimmune disease characterized by the lack of insulin due to an autoimmune destruction of pancreatic beta cells. Here, we report a unique case of a family with naturally conceived quadruplets in which T1D was diagnosed in two quadruplets simultaneously. At the same time, the third quadruplet was diagnosed with the pre-diabetic stage. Remarkably, all four quadruplets were positive for anti-islet cell antibodies, GAD65 and IA-A2. Monozygotic status of the quadruplets was confirmed by testing 14 different short tandem repeat polymorphisms. Serological examination confirmed that all quadruplets and their father suffered from a recent enteroviral infection of EV68-71 serotype. To assess the nature of the molecular pathological processes contributing to the development of diabetes, immunocompetent cells isolated from all family members were characterized by gene expression arrays, immune-cell enumerations and cytokine-production assays. The microarray data provided evidence that viral infection, and IL-27 and IL-9 cytokine signalling contributed to the onset of T1D in two of the quadruplets. The propensity of stimulated immunocompetent cells from non-diabetic members of the family to secrete high level of IFN-a further corroborates this conclusion. The number of T regulatory cells as well as plasmacytoid and/or myeloid dendritic cells was found diminished in all family members. Thus, this unique family is a prime example for the support of the so-called 'fertile-field' hypothesis proposing that genetic predisposition to anti-islet autoimmunity is 'fertilized' and precipitated by a viral infection leading to a fully blown T1D.

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“…These methods have also been used in T1D research to improve the prediction of T1D and increase the general knowledge of T1D pathogenesis [16, 17]. …”

Section: Introductionmentioning

confidence: 99%

The Effect of Diabetes-Associated Autoantigens on Cell Processes in Human PBMCs and Their Relevance to Autoimmune Diabetes Development

Včeláková

Blatny

Halbhuber

et al. 2013

Journal of Diabetes Research

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Type 1 Diabetes (T1D) is considered to be a T-helper- (Th-) 1 autoimmune disease; however, T1D pathogenesis likely involves many factors, and sufficient tools for autoreactive T cell detection for the study of this disease are currently lacking. In this study, using gene expression microarrays, we analysed the effect of diabetes-associated autoantigens on peripheral blood mononuclear cells (PBMCs) with the purpose of identifying (pre)diabetes-associated cell processes. Twelve patients with recent onset T1D, 18 first-degree relatives of the TD1 patients (DRL; 9/18 autoantibody positive), and 13 healthy controls (DV) were tested. PBMCs from these individuals were stimulated with a cocktail of diabetes-associated autoantigens (proinsulin, IA-2, and GAD65-derived peptides). After 72 hours, gene expression was evaluated by high-density gene microarray. The greatest number of functional differences was observed between relatives and controls (69 pathways), from which 15% of the pathways belonged to “immune response-related” processes. In the T1D versus controls comparison, more pathways (24%) were classified as “immune response-related.” Important pathways that were identified using data from the T1D versus controls comparison were pathways involving antigen presentation by MHCII, the activation of Th17 and Th22 responses, and cytoskeleton rearrangement-related processes. Genes involved in Th17 and TGF-beta cascades may represent novel, promising (pre)diabetes biomarkers.

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Protein microarray analysis as a tool for monitoring cellular autoreactivity in type 1 diabetes patients and their relatives

Abstract: By multiplex analysis, it may be possible, for example, to define the risk immunological response pattern among relatives or to monitor the immune response in patients on immune modulation therapy.

Cited by 10 publications

References 18 publications

Human and rodent pancreatic β-cells express IL-4 receptors and IL-4 protects against β-cell apoptosis by activation of the PI3K and JAK/STAT pathways

Human and rodent pancreatic β-cells express IL-4 receptors and IL-4 protects against β-cell apoptosis by activation of the PI3K and JAK/STAT pathways

Case report: type 1 diabetes in monozygotic quadruplets

The Effect of Diabetes-Associated Autoantigens on Cell Processes in Human PBMCs and Their Relevance to Autoimmune Diabetes Development

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