Biochemical Mechanism of Beta-Cell Destruction

自身免疫性糖尿病的特征在于胰岛内和周围的炎症反应，随后是产生胰岛素的β细胞的选择性破坏。传统观点认为，在胰岛炎症期间释放的细胞因子通过直接损害β细胞功能和减少β细胞质量而促进自身免疫性糖尿病的发展。为了支持这一假设，用单独的IL-1或与IFN-γ和/或TNF组合的IL-1处理胰岛导致胰岛素分泌和氧化代谢的抑制、DNA损伤的诱导和β细胞活力的丧失，这是由iNOS表达和β细胞产生一氧化氮介导的。30多年来，人们一直专注于确定IL-1损伤β细胞的机制，但几乎没有直接证据支持IL-1在自身免疫性糖尿病发展中的作用。胰腺β细胞终末分化，自我更新能力有限，但产生生物体生存所必需的激素（胰岛素）。IL-1是一种致热性细胞因子，众所周知其在感染和损伤期间诱导发热和炎症。如果β-细胞对IL-1的反应仅是破坏性的，那么大多数个体将对糖尿病高度敏感，因为进入胰腺的血液体积的90%穿过胰岛（其代表胰腺湿重的1%），使得β-细胞在感染和损伤期间将沐浴在IL-1中。本申请将检验以下假设：β细胞中的IL-1信号传导具有生理作用，旨在保护这些细胞免受即将发生的危险或损伤。为了支持这一假说，我们提供了初步证据表明，一氧化氮减弱DNA损伤诱导的β细胞凋亡。通过了解细胞因子对β细胞功能和存活的损伤和保护作用之间的微妙平衡，我们希望阐明IL-1信号在β细胞中的生理和病理生理作用。有两个具体目标。1.为了检验β细胞保持强大的氧化防御系统的假设，该系统提供对破坏性活性氮和氧的保护。2.检验细胞因子处理后产生的一氧化氮通过调节DNA损伤反应（DDR）的转导激酶的激活来减弱与细胞凋亡相关的DNA损伤的假设。许多生物化学，分子，免疫学，细胞生物学和转基因技术将被用于研究细胞途径，通过该途径一氧化氮及其反应中间体参与保护β细胞免受损伤。希望从这些研究中获得的对控制细胞因子刺激后β细胞中激活的保护性应答的机制的见解将影响治疗策略的设计，所述治疗策略旨在基于激活β细胞中的保护性途径作为维持功能性β细胞群并减弱移植环境中糖尿病发展或糖尿病复发的机制。

Autoimmune diabetes is characterized by an inflammatory reaction in and around pancreatic islets that is followed by the selective destruction of insulin producing β-cells. The conventional wisdom is that cytokines, released during islet inflammation, contribute to the development of autoimmune diabetes by directly impairing β-cell function and reducing β-cell mass. In support of this hypothesis, treatment of islets with IL-1 alone, or in combination with IFN-γ and/or TNF, results in an inhibition of insulin secretion and oxidative metabolism, induction of DNA damage and a loss of β-cell viability that is mediated by iNOS expression and the production of nitric oxide by β-cells. For over 30 years there has been an intense focus on determining the mechanisms by which IL-1 damages β-cells, yet there is little direct evidence supporting a role for IL-1 in the development of autoimmune diabetes. Pancreatic β-cells are terminally differentiated with a limited capacity for self-renewal, yet produce a hormone (insulin) that is essential for organismal survival. IL-1 is a pyrogenic cytokine that is well known to induce fever and inflammation during infection and injury. If the β-cell response to IL-1 were solely damaging, then most individuals would be highly susceptible to diabetes, as 90 % of the volume of blood that enters the pancreas travels through islets (which represents 1% of the wet weight of the pancreas) such that β-cells would be bathed in IL-1 during infection and injury. This application will examine the hypothesis that there is a physiological role for IL-1 signaling in β-cells that is designed to protect these cells from impending danger or insult. In support of this hypothesis, we provide preliminary evidence showing that nitric oxide attenuates DNA damage induced β-cell apoptosis. By understanding the delicate balance between the damaging and protective actions of cytokines on β-cell function and survival, we hope to elucidate the physiological and pathophysiological roles of IL-1 signaling in β-cells. There are two specific aims. 1. To test the hypothesis that β-cells maintain a robust oxidant defense system that provides protection against damaging reactive nitrogen and oxygen species. 2. To test the hypothesis that nitric oxide, produced following cytokine treatment, attenuates DNA damage associated apoptosis by regulating the activation of transducing kinases of the DNA damage response (DDR). A number of biochemical, molecular, immunological, cell biological, and transgenic techniques will be utilized to investigate the cellular pathways through which nitric oxide and its reactive intermediates participate in the protection of β-cells from damage. It is hoped that insights into the mechanisms controlling the protective responses activated in β-cells following cytokine stimulate that are gained from these studies will influence the design of therapeutic strategies aimed that are based on activating protective pathways in β-cell as a mechanism to maintain functional β-cell mass and attenuate the development of diabetes or recurrence of diabetes in the transplantation setting.