Aims/hypothesis IL-1β and TNF-α contribute to pancreatic beta cell death in type 1 diabetes. Both cytokines activate the transcription factor nuclear factor-κB (NF-κB), but recent observations suggest that NF-κB blockade prevents IL-1β + IFN-γ-but not TNF-α + IFN-γ-induced beta cell apoptosis. The aim of the present study was to compare the effects of IL-1β and TNF-α on cell death and the pattern of NF-κB activation and global gene expression in beta cells. Methods Cell viability was measured after exposure to IL-1β or to TNF-α alone or in combination with IFN-γ, and blockade of NF-κB activation or protein synthesis. INS-1E cells exposed to IL-1β or TNF-α in time course experiments were used for IκB kinase (IKK) activation assay, detection of p65 NF-κB by immunocytochemistry, realtime RT-PCR and microarray analysis. Results Blocking NF-κB activation protected beta cells against IL-1β + IFNγ-or TNFα + IFNγ-induced apoptosis. Blocking de novo protein synthesis did not increase TNF-α-or IL-1β-induced beta cell death, in line with the observations that cytokines induced the expression of the anti-apoptotic genes A20, Iap-2 and Xiap to a similar extent. Microarray analysis of INS-1E cells treated with IL-1β or TNF-α showed similar patterns of gene expression. IL-1β, however, induced a higher rate of expression of NF-κB target genes putatively involved in beta cell dysfunction and death and a stronger activation of the IKK complex, leading to an earlier translocation of NF-κB to the nucleus. Conclusions/interpretation NF-κB activation in beta cells has a pro-apoptotic role following exposure not only to IL-1β but also to TNF-α. The more marked beta cell death induced by IL-1β is explained at least in part by higher intensity NF-κB activation, leading to increased transcription of key target genes.

type 1 diabetes mellitus (t1d) is characterized by severe insulin deficiency resulting from chronic and progressive destruction of pancreatic beta-cells by the immune system. the triggering of autoimmunity against the beta-cells is probably caused by environmental agent(s) acting in the context of a predisposing genetic background. once activated, the immune cells invade the islets and mediate their deleterious effects on beta-cells via mechanisms such as Fas/Fasl, perforin/granzyme, reactive oxygen and nitrogen species and pro-inflammatory cytokines. Binding of cytokines to their receptors on the beta-cells activates MAP-kinases and the transcription factors stAt-1 and nFκ-B, provoking functional impairment, endoplasmic reticulum stress and ultimately apoptosis. this review discusses the potential mediators and mechanisms leading to beta-cell destruction in t1d. Keywords: Pancreatic beta-cell; diabetes mellitus; Apoptosis; cytokines; endoplasmic reticulum; er stress RESuMOMecanismos de Destruição e Morte da Célula-beta Pancreática no Diabetes. o diabetes melito tipo 1 (dM1) tem como característica uma grave deficiência de insulina que resulta da destruição da célula-beta, crônica e progressiva, pelo sistema imune. o desencadeamento da autoimunidade contra a célula-beta é causado, provavelmente, por agentes ambientais que atuam quando existe predisposição genética. uma vez ativadas, células imunes invadem as ilhotas, e os efeitos deletérios sobre as células-beta são mediados por mecanismos relacionados a Fas/Fasl, perforina/granzima, espécies reativas de oxigênio e nitrogênio, e a citocinas pró-inflamatórias. A ligação de citocinas a seus receptores na célula-beta ativa MAP-quinase e fatores de transcrição stAt-1 e nFkB, provocando prejuízo funcional, estresse de retículo endoplasmático e, por fim, apoptose. esta revisão discute os mecanismos e os mediadores potenciais que levam à destruição da célula-beta no dM1. Descritores: célula-beta pancreática; diabetes melito; Apoptose; citocinas; estresse do retículo endoplasmático thE GENEtic BAcKGROuND Of typE 1 DiABEtES MEllituS (t1D)

Aims/hypothesis Pancreatic beta cells respond to endoplasmic reticulum (ER) stress by activating the unfolded protein response. If the stress is prolonged, or the adaptive response fails, apoptosis is triggered. We used a 'homemade' microarray specifically designed for the study of beta cell apoptosis (the APOCHIP) to uncover mechanisms regulating beta cell responses to ER stress. Materials and methods A time course viability and microarray analysis was performed in insulin-producing INS-1E cells exposed to the reversible ER stress inducer cyclopiazonic acid (CPA). Modification of selected genes was confirmed by real-time RT-PCR, and the observed inhibition of expression of the insulin-1 (Ins1) and insulin-2 (Ins2) genes was further characterised in primary beta cells exposed to a diverse range of agents that induce ER stress. Results CPA-induced ER stress modified the expression of 183 genes at one or more of the time points studied. The expression of most of these genes returned to control levels after a 3 h recovery period following CPA removal, with all cells surviving. Two groups of genes were particularly affected by CPA, namely, those related to cellular responses to ER stress, which were mostly upregulated, and those related to differentiated beta cell functions, which were downregulated. Levels of Ins1 and Ins2 mRNAs were severely decreased in response to CPA treatment as a result of degradation, and there was a concomitant increase in the level of IRE1 activation. Conclusions/interpretation In this study we provide the first global analysis of beta cell molecular responses to a severe ER stress, and identify the early degradation of mRNA transcripts of the insulin genes as an important component of this response.

Endoplasmic reticulum stress-mediated apoptosis may play an important role in the destruction of pancreatic ␤-cells, thus contributing to the development of type 1 and type 2 diabetes. One of the regulators of endoplasmic reticulum stress-mediated cell death is the CCAAT/enhancer binding protein (C/EBP) homologous protein (Chop). We presently studied the molecular regulation of Chop expression in insulin-producing cells (INS-1E) in response to three pro-apoptotic and endoplasmic reticulum stress-inducing agents, namely the cytokines interleukin-1␤ ؉ interferon-␥, the free fatty acid palmitate, and the sarcoendoplasmic reticulum pump Ca

CPA-induced endoplasmic reticulum stress and apoptosis is enhanced in IFN-gamma-treated beta cells. These effects are mediated via downregulation of the expression of genes involved in beta cell defence against endoplasmic reticulum stress.

Aims/hypothesis Pro-inflammatory cytokines involved in the pathogenesis of type 1 diabetes deplete endoplasmic reticulum (ER) Ca 2+ stores, leading to ER-stress and beta cell apoptosis. However, the cytokine-induced ER-stress response in beta cells is atypical and characterised by induction of the pro-apoptotic PKR-like ER kinase (PERK)-C/EBP homologous protein (CHOP) branch of the unfolded protein response, but defective X-box binding protein 1 (XBP1) splicing and activating transcription factor 6 activation. The purpose of this study was to overexpress spliced/active Xbp1 (XBP1s) to increase beta cell resistance to cytokine-induced ER-stress and apoptosis.Methods Xbp1s was overexpressed using adenoviruses and knocked down using small interference RNA in rat islet cells. In selected experiments, Xbp1 was also knocked down in FACS-purified rat beta cells and rat fibroblasts. Expression and production of XBP1s and key downstream genes and proteins was measured and beta cell function and viability were evaluated.

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