Lipid Mediators in Pancreatic Islet Dysfunction

描述(由申请人提供)：自身免疫介导的胰岛β细胞损伤是1型糖尿病(T1 DM)及其在胰岛移植受者中复发的核心。促炎细胞因子和脂质介质是β细胞损伤的重要因素。我们重点研究了12-脂氧合酶(12LO)对花生四烯酸代谢在导致免疫介导的β细胞破坏中的作用。促炎症细胞因子可激活和诱导12LO。12LO代谢产物可启动细胞内信号级联反应，导致β细胞功能障碍和死亡。在这项提案中，我们将检验12LO通路在自身免疫介导的炎症反应中发挥关键作用的假设，该炎症反应导致T1 DM的β细胞抑制和死亡。其具体目的是：1)鉴定人胰岛和胰岛素释放的β细胞中12LO的激活和由促炎细胞因子诱导的特性。研究12LO通路在细胞因子诱导的β细胞功能障碍和死亡中的作用和机制。人类胰岛中的12LO的特殊类型将被描述。使用核酶或过度表达12LO的分子抑制对细胞因子信号和作用的影响将被测试。2)明确12LO在细胞因子介导的自身免疫性糖尿病β细胞破坏中的作用。12LO的表达和过氧化脂质的产生将在自身免疫性糖尿病NOD小鼠中得到监测。细胞因子敏感性和细胞因子信号传递过程将在12LO缺失的小鼠中确定。我们还将进一步研究在点头背景下生成的12-空鼠标。将测试糖尿病发病率、胰岛炎症的严重程度以及细胞因子对孤立胰岛的影响。12LO基因打靶减少自身免疫β细胞破坏的机制将被评估。我们还将确定这些小鼠在12LO耗尽时受到干扰的炎症信号通路。3)以自发性糖尿病NOD小鼠为受体，研究12LO基因打靶对提高胰岛移植物存活的作用。供体胰岛将来自12LO缺失的小鼠以及用12LO核酶处理的野生型小鼠胰岛。移植物存活和减少自身免疫损伤将被研究。这项完整的提案的结果将促进我们对导致自身免疫β细胞破坏的炎症过程的理解。这一发现将提供新的方法来保存β细胞团，最终导致新的治疗方式来预防T1 DM，并利用胰岛移植提高逆转T1 DM的能力。

DESCRIPTION (provided by applicant): Autoimmune-mediated pancreatic beta-cell damage is central to the development of type 1 diabetes (T1DM) and its recurrence in islet transplant recipients. Proinflammatory cytokines and lipid mediators are important contributors to beta-cell damage. We have focused on the role of arachidonic acid metabolism by 12-Lipoxygenase (12LO) in leading to immune-mediated beta-cell destruction. 12LO can be activated and induced by proinflammatory cytokines. 12LO metabolites can initiate intracellutar signaling cascades leading to beta-cell dysfunction and death. In this proposal, we will test the hypothesis that the 12LO pathway plays a key role in autoimmunity-mediated inflammatory responses leading to beta-cell inhibition and death in T1DM. The specific aims are: 1) to characterize 12LO activation and induction by proinflammatory cytokines in human islets and in insulin-releasing beta-cells. Study the role and mechanism of how the 12LO pathway mediates cytokine-induced beta-cell dysfunction and death. The particular type of 12LO in human islets will be characterized. The effect of molecular inhibition of 12LO using a ribozyme or over-expression on cytokine signaling and action will be tested. 2) to define the role of 12LO in cytokine-mediated beta-cell destruction in autoimmune diabetes. 12LO expression and lipid peroxide production will be monitored in autoimmune diabetic NOD mice. Cytokine-susceptibility and cytokine signaling processes will be determined in a 12LO-null mouse. We will also further study 12-null mouse generated on NOD background. Diabetes incidence, severeness of insulitis and cytokine effects on isolated islets will be tested. Mechanism of 12LO gene targeting reduces autoimmune beta-cell destruction will be evaluated. We will also identify the inflammatory signaling pathways disrupted upon 12LO depletion in these mice. 3) Effect of 12LO gene targeting on improving islet graft survival will be studied utilizing spontaneously diabetic NOD mice as recipients. Donor islets will be from 12LO null mice as well as the wild type mouse islets-treated with the 12LO ribozyme. Graft survival and reduction in autoimmune damage will be studied. The results from this completed proposal will advance our understanding of the inflammatory processes causing autoimmune beta-cell destruction. The findings should provide new methods to preserve beta-cell mass ultimately leading to new therapeutic modalities to prevent T1DM and improve ability to reverse T1DM using islet transplantation.