Browsing by keyword "pancreatic islets"
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Coxsackievirus-Induced Proteomic Alterations in Primary Human Islets Provide Insights for the Etiology of DiabetesEnteroviral infections have been associated with the development of type 1 diabetes (T1D), a chronic inflammatory disease characterized by autoimmune destruction of insulin-producing pancreatic beta cells. Cultured human islets, including the insulin-producing beta cells, can be infected with coxsackievirus B4 (CVB4) and thus are useful for understanding cellular responses to infection. We performed quantitative mass spectrometry analysis on cultured primary human islets infected with CVB4 to identify molecules and pathways altered upon infection. Corresponding uninfected controls were included in the study for comparative protein expression analyses. Proteins were significantly and differentially regulated in human islets challenged with virus compared with their uninfected counterparts. Complementary analyses of gene transcripts in CVB4-infected primary islets over a time course validated the induction of RNA transcripts for many of the proteins that were increased in the proteomics studies. Notably, infection with CVB4 results in a considerable decrease in insulin. Genes/proteins modulated during CVB4 infection also include those involved in activation of immune responses, including type I interferon pathways linked to T1D pathogenesis and with antiviral, cell repair, and inflammatory properties. Our study applies proteomics analyses to cultured human islets challenged with virus and identifies target proteins that could be useful in T1D interventions.
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Innate Immune Signaling Drives Pathogenic Events Leading to Autoimmune DiabetesType 1 diabetes (T1D) is a chronic autoimmune disease characterized by the immune-mediated destruction of insulin-producing beta-cells of pancreatic islets, culminating in critical insulin deficiency. Both genetic and environmental factors likely orchestrate an immune-mediated functional loss of beta cell mass, leading to the clinical manifestation of disease and lifelong dependence on insulin therapy. Additional evidence suggests the role of innate and adaptive immune mechanisms leading to inflammation in beta cells mediated by proinflammatory cytokines and chemokines, activation of beta-cell-reactive T cells,and failure of immune tolerance. Viral infections have been proposed as causal determinants or initiating triggers for T1D but remain unproven. Understanding the relationship between viral infections and the development of T1D is essential for T1D prevention. Importantly, virus-induced innate immune responses, particularly type I interferon (IFN-I, IFN-a/b), have been implicated in the initiation of islet autoimmunity and development of T1D. The goal of my thesis project is to investigate how the IFN-I signaling pathway affects the development of T1D using the LEW.1WR1 rat model of autoimmune diabetes. My hypothesis is that disrupting IFN-Isignaling via functional deficiency of the IFN-I interferon receptor (IFNAR) prevents or delays the development of virus-induced diabetes.For this purpose, I generated IFNAR subunit 1(IFNAR1)-deficient LEW.1WR1 rats using CRISPR-Cas9 genome editing and confirmed the functional disruption of IFNAR1. The absence of IFNAR1 results in a significant delay in onset and frequency of diabetes following poly I:C challenge and reduces the incidence of insulitis after poly I:C treatment. The frequency of diabetes induced by Kilham rat virus (KRV) is also reduced in IFNAR1-deficient LEW.1WR1 rats. Furthermore, I observe a decrease in CD8+T cells in spleens from KRV-infected IFNAR1-deficient rats relative to that in KRV-infected wild-type rats. While splenic regulatory T cells are depleted in WT rats during KRV-infection, no such decrease is observed in KRV-infected IFNAR1-deficient rats. A comprehensive bulk RNA-seq analysis reveals a decrease of interferon-stimulated genes and inflammatory gene expression in IFNAR1-deficient rats relative to wild-type rats following KRV challenge. Collectively, the results from these studies provided mechanistic insights into the essential role of virus-induced, IFN-I-initiated innate immune responses in the early phase of autoimmune diabetes pathogenesis.
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NLRP3 inflammasome mediates oxidative stress-induced pancreatic islet dysfunctionInflammasomes are multiprotein inflammatory platforms that induce caspase-1 activation and subsequently interleukin (IL)-1beta and IL-18 processing. The NLRP3 inflammasome is activated by different forms of oxidative stress, and, based on the central role of IL-1beta in the destruction of pancreatic islets, it could be related to the development of diabetes. We therefore investigated responses in wild-type C57Bl/6 (WT) mice, NLRP3(-/-) mice, and mice deficient in apoptosis-associated speck-like protein containing a caspase-recruitment domain (ASC) after exposing islets to short-term hypoxia or alloxan-induced islet damage. NLRP3-deficient islets compared with WT islets had preserved function ex vivo and were protected against hypoxia-induced cell death. Furthermore, NLRP3 and ASC-deficient mice were protected against oxidative stress-induced diabetes caused by repetitive low-dose alloxan administration, and this was associated with reduced beta-cell death and reduced macrophage infiltration. This suggests that the beneficial effect of NLRP3 inflammasome deficiency on oxidative stress-mediated beta-cell damage could involve reduced macrophage infiltration and activation. To support the role of macrophage activation in alloxan-induced diabetes, we injected WT mice with liposomal clodronate, which causes macrophage depletion before induction of a diabetic phenotype by alloxan treatment, resulting in improved glucose homeostasis in WT mice. We show here that the NLRP3 inflammasome acts as a mediator of hypoxia and oxidative stress in insulin-producing cells, suggesting that inhibition of the NLRP3 inflammasome could have beneficial effects on beta-cell preservation.
