Unfolded Protein Response: Regulator of Human beta-cells

描述（由申请人提供）： 促炎细胞因子被认为参与了功能性胰岛质量的丧失以及移植后的移植物排斥。使人胰岛移植复杂化的还有缺乏足够的胰岛质量，部分原因是分离后胰岛细胞凋亡。本研究的广泛目标是阐明细胞因子介导胰腺β细胞破坏的生化机制，并确定从尸体供体分离后介导胰岛质量损失的机制。内质网（ER）应激激活了已知可诱导细胞凋亡的未折叠蛋白反应（UPR）。最近，我们已经表明，一氧化氮，白细胞介素-1（IL-1）和干扰素-γ（IFN-γ）对啮齿动物和人类β细胞功能的抑制作用的主要介质，是β细胞中UPR的激活剂。该提议将解决以下假设：在人胰岛分离期间响应于细胞因子或一氧化氮处理或应激的延长的UPR激活导致胰腺β细胞的凋亡损失。有两个具体目标： 1.为了验证一氧化氮激活人胰岛中的UPR（或ER应激）通路以及该通路的长期激活导致β-细胞凋亡丢失的假设。提出的实验将评估细胞因子和一氧化氮供体对UPR活化的影响，并确定UPR抑制是否保护β细胞免于精氨酸介导的死亡。 2.为了验证隔离期间胰岛损伤诱导ER应激和UPR激活的假设，以及UPR激活是隔离期间胰岛质量损失的一种机制。拟议的实验将检查人胰岛分离对UPR激活的影响，并确定UPR抑制是否阻止或减弱分离后胰岛的损失。 许多生物化学、分子生物学、免疫学和组织化学技术将被用于研究UPR活化作为细胞因子处理后或人胰岛分离后β细胞凋亡的一种潜在介质的作用。人们希望，从这些研究中获得的对精氨酸介导的β-细胞凋亡机制的了解，以及与分离后胰岛质量损失相关的机制，将影响旨在减轻胰岛移植排斥反应和增加可用于移植的胰岛功能质量的治疗策略的设计。

DESCRIPTION (provided by applicant): Proinflammatory cytokines are believed to participate in the loss of functional islet mass as well as graft rejection following transplantation. Also complicating human islet transplantation is the lack of sufficient islet mass due, in part, to islet cell apoptosis following isolation. The broad goals of this research are to elucidate the biochemical mechanisms by which cytokines mediate pancreatic beta-cell destruction and to identify the mechanisms that mediate the loss of islet mass following isolation from cadaver donors. Endoplasmic reticulum (ER) stress activates an unfolded protein response (UPR) that is known to induce apoptosis. Recently, we have shown that nitric oxide, a primary mediator of the inhibitory actions of interleukin-1 (IL-1) and interferon-gamma(IFN-gamma) on rodent and human beta-cell function, is an activator of the UPR in beta-cells. This proposal will address the hypothesis that prolonged UPR activation in response to cytokine or nitric oxide treatment or stress during human islet isolation results in the apoptotic loss of pancreatic beta-cells. There are two specific aims: 1. To test the hypothesis that nitric oxide activates the UPR (or ER stress) pathway in human islets and that prolonged activation of this pathway results in the apoptotic loss of (-cells. Experiments proposed will evaluate the effects of cytokines and nitric oxide donors on UPR activation and determine if UPR inhibition protects beta-cells from cytokine-mediated death. 2. To test the hypothesis that islet damage during isolation induces ER stress and UPR activation, and that UPR activation is one mechanism responsible for the loss of islet mass during isolation. Proposed experiments will examine the effects of human islet isolation on UPR activation and determine whether UPR inhibition prevents or attenuates the loss of islets following isolation. A number of biochemical, molecular biological, immunological, and histochemical techniques will be utilized to investigate the role of UPR activation as one potential mediator of beta-cell apoptosis following cytokine treatment or following human islet isolation. It is hoped that insights into the mechanisms of cytokine-mediated beta-cell apoptosis, and the mechanisms associated with the loss of islet mass following isolation gained from these studies will influence the design of therapeutic strategies aimed at the attenuation of islet graft rejection and increase the functional mass of islets available for transplantation.