Pathogenesis of Chronic Wasting Disease in Transgenic Mice

项目摘要/摘要： 这项建议使用慢性消耗性疾病(CWD)的转基因小鼠模型来解决序列 Pron的发病机制和骨髓在Pron转运和神经侵袭中的作用。世界银行的目标是 建议的研究如下：(1)在转基因CWD中表征CWD发病机制的多个方面 表达PrP的(Tg[CerPrP])小鼠通过评估PrP的组织趋向性和序列模式 神经病理学；和(2)探讨骨髓及其细胞后代在Pron转运中的作用 以及通过相互移植研究的发病机制。我们将利用单-和双-的组合- 标记免疫组织化学(IHC)、免疫印迹(WB)和免疫荧光共聚焦显微镜 结合神经病理标记物检测错折叠蛋白(PrPres)的累积。至 定义骨髓在Pron运输和血源性神经侵袭中的作用，我们将使用倒数 过继骨髓移植创造两个嵌合小鼠谱系，将表达PrPc 仅限于造血室或非造血室。我们将通过流动来描述这些老鼠的特征 流式细胞术，并记录PrPres通过IHC和WB的顺序积累。我们预计CWD感染者 小鼠将模仿自然感染的宫颈在易感性、临床疾病、显微镜下 神经病理学和PrPres分布。我们假设/预期CWD感染的小鼠将揭示出新的， Pron病的神经病理特征，包括胶质细胞增生标志物的表达增加，神经元丢失 PrPres聚集区附近的细胞凋亡。在过继骨髓移植实验中，我们 假设感染CWD的TG[CerPrP]小鼠的骨髓中将含有感染性普恩病毒，这将是 与髓系细胞有关。此外，我们假设这些细胞会将PrPre运送到大脑 作为细胞更新和贩运模式的一部分。这些研究的结果将加深对 Pron病相关神经病理学的机制，有助于为抗Pron病的治疗策略提供信息， 并探索一条未被充分研究的普恩病毒传播途径。利用提议的实验，我的短文- 学期目标是在2010年夏天完成我的博士学位；然后我的长期目标将是致力于 我想成为一名成功的独立研究人员.研究事业的关键要素 为实现这些目标而设计的发展计划包括：(1)建立咨询/指导 帮助建立和监测职业表现里程碑的小组；(2)确定和成功 在一个专注于研究的部门和机构担任调查病理学家，具有良好的- 建立传染病和/或神经退行性疾病方案；(3)参与校外研究 在现有的普恩病毒和神经退行性疾病中心提供培训机会；(4)提交同行-- 审查手稿和校外赠款申请，以及(5)扩大责任和参与 实验室、部门和国家/国际会议和期刊俱乐部。

Project Summary/Abstract: This proposal employs a transgenic murine model of chronic wasting disease (CWD) to addresses sequential prion pathogenesis and the role of bone marrow in prion trafficking and neuroinvasion. The goals of the proposed studies are: (1) to characterize the multiple aspects of the pathogenesis of CWD in transgenic, cervid PrP-expressing (Tg[CerPrP]) mice by evaluation of sequential patterns of prion tissue tropism and neuropathology; and (2) to address the role of the bone marrow and its cellular progeny in the prion trafficking and pathogenesis through reciprocal transplantation studies. We will utilize a combination of single- and dual- labeling immunohistochemistry (IHC), western blotting (WB), and immunofluorescent confocal microscopy to detect accumulations of misfolded prion protein (PrPres) in conjunction with markers of neuropathology. To define the role of the bone marrow in prion trafficking and hematogenous neuroinvasion, we will use reciprocal adoptive bone marrow transfer to create two lineages of chimeric mice, which will have PrPc expression restricted to hematopoietic or non-hematopoietic compartments. We will characterize these mice through flow cytometry, and document sequential PrPres accumulation via IHC and WB. We expect that the CWD-infected mice will mimic naturally-infected cervids with respect to susceptibility, clinical disease, microscopic neuropathology, and PrPres distribution. We hypothesize/anticipate that CWD-infected mice will reveal novel, neuropathologic features of prion disease, including increased expression of markers of gliosis, neuronal loss and apoptosis adjacent to areas of PrPres accumulation. In the adoptive bone marrow transfer experiments, we hypothesize that marrow of CWD-infected Tg[CerPrP] mice will contain infectious prions, which will be associated with myeloid lineage cells. Additionally, we hypothesize that these cells will ferry PrPres to the brain as part of cell renewal and trafficking patterns. The results of these studies will enhance understanding of the mechanisms of prion disease-associated neuropathology, help inform anti-prion disease therapeutic strategies, and explore an under-investigated pathway of prion dissemination. Using the proposed experiments, my short- term objective will be to complete my PhD in the summer of 2010; my long term goal will then be to devote myself to a becoming a successful independent researcher. The key elements of the research career development plan designed to accomplish these goals include: (1) the establishment of an advisory/mentoring panel to help establish and monitor career performance milestones; (2) the identification and successful employment as an investigative pathologist in a research-focused department and institution with a well- established infectious disease and/or neurodegenerative program; (3) participation in extramural research training opportunities at established centers of prion and neurodegenerative disease; (4) submission of peer- reviewed manuscripts and extramural grant applications, and (5) extended responsibility and participation in laboratory, departmental, and national/international meetings and journal clubs.