Detection and Characterization of Blood-borne Prions

描述（由申请人提供）：人类变异型克雅氏病（vCJD）、宫颈慢性消耗性疾病（CWD）和羊痒病之间的共性非常显著。在人类和动物传染性海绵状脑病（tse）中重现了与淋巴网状系统（LRS）相关的血液朊病毒。特别是，vCJD、CWD和痒病的特点都是疾病的长时间无症状期，传染性血源性朊病毒可以有效地从一个宿主传播到另一个宿主。由于这些传染性脑病在易感宿主内的传播方式相似，因此利用动物朊病毒疾病为评估血源性朊病毒的生物学及其传播提供了独特的机会。我实验室这项研究的长期目标是制定干预策略，以检测、减轻和预防血朊病毒的传播。作为实现这一目标的一步，我们建议使用动物tse来更好地定义和理解与血源性朊病毒的贩运、传播和传播相关的机制。使用动物的tse将允许在感染vCJD的人身上做没有也不能做的事情，即从感染到绝症的几分钟内收集连续的纵向血液样本。这项工作的统一假设是，血源性朊病毒在感染后不久以特定细胞表型在宿主内循环，并负责宿主内的早期传播和宿主之间的隐蔽传播。为了验证这一假设，我们：1)采用快速、敏感和经济有效的体外转化试验检测血源性朊病毒；2)建立了一个独特的纵向血液样本库，收集了自然和实验感染疯牛病的动物、啮齿动物和灵长类动物的血液样本；3)建立有能力在动物和啮齿动物体内模型中容纳和评估tse的设施和合作者。我们建议解决两个特定的目标，1)确定感染TSE的宿主中朊病毒血症的时间状态，2)定量确定哪些血室和细胞表型窝藏朊病毒。我们将使用自然感染和实验感染动物（cwd感染的奶牛，实验性vCJD， BSE和cwd感染的灵长类动物和tme感染的仓鼠）的血液来研究我们最近建立的PrP转化试验，全血实时振动诱导转化（wbRT- QuIC）的有效性。这项工作的结果将建立朊病毒血症的时间状态，以及辨别哪些隔室和特定的细胞表型窝藏朊病毒。在适当的情况下，这些结果将与我实验室的蛋白质错误折叠循环扩增和生物测定方法进行比较。这些目标将提供在朊病毒领域长期寻求的东西；阐明朊病毒在宿主体内的贩运和传播机制。这将允许及时检测，从而更有效地治疗人类tse和其他人类蛋白质错误折叠疾病。

DESCRIPTION (provided by applicant): The commonalities between human variant Creutzfeldt-Jakob Disease (vCJD), cervid chronic wasting disease (CWD) and sheep scrapie are quite remarkable. Hematogenous prions exhibiting association with the lymphoreticular system (LRS) are recapitulated in both human and animal transmissible spongiform encephalopathies (TSEs). In particular, vCJD, CWD and scrapie are all characterized by a long asymptomatic phase of disease from which infectious blood-borne prions efficiently transmit from one host to another. Because these TSEs traffic and disseminate similarly within susceptible hosts, the use of animal prion diseases provides a unique opportunity to evaluate the biology of blood-borne prions and their spread. The long-term goal of this research in my laboratory is to develop intervention strategies to detect, mitigate and prevent hematogenous prion spread. As a step towards this goal, we propose the use of animal TSEs to better define and understand the mechanisms associated with trafficking, dissemination and transmission of blood-borne prions. The use of animal TSEs will allow what has not- and cannot- be done with humans infected with vCJD, namely collect serial longitudinal blood samples from minutes after infection to terminal disease. The unifying hypothesis for this work is that blood-borne prions circulate within the host on/in specific cell phenotypes soon after infection and are responsible for early dissemination within the host and covert transmission between hosts. To test this hypothesis, we: 1) adapted fast, sensitive and cost effective in vitro conversion assays for the detection of blood-borne prions; 2) assembled a unique repository of longitudinal blood samples from natural and experimental TSE-infected cervids, rodents and primates; and 3) established facilities and collaborators with the capacity to house and assess TSEs in cervid and rodent in vivo models. We propose to address two specific aims, to 1) determine the temporal status of prionemia in TSE- infected hosts and 2) quantitatively determine which blood compartments and cell phenotypes harbor prions. We will employ blood, from naturally and experimentally infected animals (CWD-infected cervids, experimentally vCJD, BSE and CWD-infected primates and TME-infected hamsters) to investigate the efficacy of our recently established PrP conversion assay, whole blood real-time quaking induced conversion (wbRT- QuIC). The outcome of this work will establish the temporal status of prionemia as well as discern which compartments and specific cell phenotypes harbor prions. These results will be compared, where appropriate, to protein misfolding cyclic amplification and bioassay methods in place in my laboratory. These aims will provide what has long been sought in the prion field; elucidating the mechanisms of prion trafficking and dissemination throughout the host. This will permit timely detection, and thus, more effective therapies for human TSEs and other human protein misfolding diseases.