CWD: Role of the Lymphoid Tissue Phase in Prion Disease

The immediate goal of the candidate, Christina J. Sigurdson, DVM, is to complete a mentored training program in prion pathogenesis research which will culminate in the PhD degree and the opportunity for postdoctoral study. Dr. Sigurdson's long term goal is to become a principal investigator employing animal models to investigate the diagnosis and pathogenesis of transmissible spongiform encephalopathies (TSEs). The research training program will be conducted under the guidance of the sponsor Dr. Edward Hoover at Colorado State University. The training environment has a strong record of extramural funding and post-DVM scientists trained. The candidate's plan emphasizes primary laboratory training in contemporary research methods applied to an animal model of prion pathogenesis. Proposed are studies of the early prion protein accumulation in lymphoid tissue, an intriguing, topical, and important feature of several TSEs, including variant Creutzfeldt-Jakob disease (vCJD) in humans, scrapie in sheep, and chronic wasting disease (CWD) in deer. Understanding mechanisms and pathways of prion transmucosal entry and subsequent neuroinvasion is critical for development of diagnostic assays and intervention strategies; thus, we propose to study CWD of deer as a model for vCJD/prion pathogenesis. CWD PrPres is abundant in lymphoid tissue of infected deer, therefore, may potentially be more readily detected in blood and circulating leukocytes.
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We propose three aims which explore basic issues in prion biology: <ol> <li>In the first aim, we will seek to identify PrPres-bearing cell phenotypes in lymphoid tissues by using established PrPres- specific dual immunohisto/cytochemistry and dual immunofluorescence assays. We will localize the PrPres subcellularly within lymphoid tissue using confocal and immunogold electron microscopy.
<li>Relatively little is known of the initial prion trafficking pathways to lymphoid tissue or the CNS. In aim 2, we will use PrPres-specific antibodies, flow cytometry, dual immunocytochemistry, and in vivo infectivity to seek potential cell vectors for prion dissemination in blood and lymph of CWD-infected deer. Detection of PrPres in the blood has dramatic implications for blood recipients.
<li>Dendritic cells (DC) are suspected to serve as the primary accumulator of PrPres in lymphoid tissue. In aim 3, we will isolate monocyte-derived DC from blood, transfect these cells with an epitope-tagged deer PrPc-expressing vector, expose the DC to exogenous CWD PrPres, and use immunostaining followed by confocal and electron microscopy to determine whether in situ PrPres conversion has occurred. The results of these studies will contribute to understanding PrP trafficking in the lymphoid system and could provide a basis for development of blood-based diagnostic assays and intervention strategies.</ol> </p>