DNA methylation profiling identifies epigenetic dysregulation in pancreatic islets from type 2 diabetic patientsThe first genome-scale DNA methylation study on pancreatic islets from type 2 diabetic patients identifies disease-associated DNA methylation pattern that translate into aberrant gene expression in novel factors relevant for β-cell function and survival.

Cytokines produced by islet-infiltrating immune cells induce

OBJECTIVECytokines contribute to pancreatic β-cell death in type 1 diabetes. This effect is mediated by complex gene networks that remain to be characterized. We presently utilized array analysis to define the global expression pattern of genes, including spliced variants, modified by the cytokines interleukin (IL)-1β + interferon (IFN)-γ and tumor necrosis factor (TNF)-α + IFN-γ in primary rat β-cells.RESEARCH DESIGN AND METHODSFluorescence-activated cell sorter–purified rat β-cells were exposed to IL-1β + IFN-γ or TNF-α + IFN-γ for 6 or 24 h, and global gene expression was analyzed by microarray. Key results were confirmed by RT-PCR, and small-interfering RNAs were used to investigate the mechanistic role of novel and relevant transcription factors identified by pathway analysis.RESULTSNearly 16,000 transcripts were detected as present in β-cells, with temporal differences in the number of genes modulated by IL-1β + IFNγ or TNF-α + IFN-γ. These cytokine combinations induced differential expression of inflammatory response genes, which is related to differential induction of IFN regulatory factor-7. Both treatments decreased the expression of genes involved in the maintenance of β-cell phenotype and growth/regeneration. Cytokines induced hypoxia-inducible factor-α, which in this context has a proapoptotic role. Cytokines also modified the expression of >20 genes involved in RNA splicing, and exon array analysis showed cytokine-induced changes in alternative splicing of >50% of the cytokine-modified genes.CONCLUSIONSThe present study doubles the number of known genes expressed in primary β-cells, modified or not by cytokines, and indicates the biological role for several novel cytokine-modified pathways in β-cells. It also shows that cytokines modify alternative splicing in β-cells, opening a new avenue of research for the field.

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.

Environmental factors such as diets rich in saturated fats contribute to dysfunction and death of pancreatic β-cells in diabetes. Endoplasmic reticulum (ER) stress is elicited in β-cells by saturated fatty acids. Here we show that palmitate-induced β-cell apoptosis is mediated by the intrinsic mitochondrial pathway. By microarray analysis, we identified a palmitate-triggered ER stress gene expression signature and the induction of the BH3-only proteins death protein 5 (DP5) and p53-upregulated modulator of apoptosis (PUMA). Knockdown of either protein reduced cytochrome c release, caspase-3 activation, and apoptosis in rat and human β-cells. DP5 induction depends on inositol-requiring enzyme 1 (IRE1)–dependent c-Jun NH2-terminal kinase and PKR–like ER kinase (PERK)–induced activating transcription factor (ATF3) binding to its promoter. PUMA expression is also PERK/ATF3-dependent, through tribbles 3 (TRB3)–regulated AKT inhibition and FoxO3a activation. DP5−/− mice are protected from high fat diet–induced loss of glucose tolerance and have twofold greater pancreatic β-cell mass. This study elucidates the crosstalk between lipotoxic ER stress and the mitochondrial pathway of apoptosis that causes β-cell death in diabetes.