NR AXLN
AU Eigenbrod,S.; Karmi,M.; Maringer,M.; Bertsch,U.; Fuhrmann,M.; Pace,C.; Piening,N.; Herms,J.; Pilz,C.; Windl,O.; Kretzschmar,H.A.
TI Monitoring of Scrapie Infection in a New EGFP-PrP Transgenic Mouse Model
QU International Conference - Prion 2007 (26.-28.9.2007) Edinburgh International Conference Centre, Edinburgh, Scotland, UK - Book of Abstracts: Pathology and Pathogenesis P03.95
IA http://www.prion2007.com/pdf/Prion Book of Abstracts.pdf
PT Konferenz-Poster
AB
Background: The immediate cellular processes and pathogenetic mechanisms leading to prion (PrP) diseases following infection are not well understood.
Objectives: The aim of this study is to further enlighten these pathogenic mechanisms.
Methods: We generated enhanced green fluorescent protein (EGFP)-PrP transgenic mice by pronucleus injection, carrying the coding sequence for an EGFP-PrP fusion protein including a L42-tag under the regulatory elements of the half-genomic PrP locus. The resulting mouse lines were analyzed regarding PrP expression, F35phenotype rescue and scrapie-infection experiments using confocal microscopy, Western Blot and immunohistochemistry. In addition, the EGFP-PrP fusion protein was tested in an in vitro conversion assay.
Results: EGFP-PrP expression was observed in all mouse lines with some variation in the expression pattern, most likely due to integration effects of the transgene. EGFP-
PrP failed in a functional test regarding disease phenotype rescue in F35-transgenic mice and also was not convertible to the proteinase K (PK)-resistant form (PrPsc) in vitro. However, Western Blot analysis of EGFP-PrP mice on a wildtype-PrP background which were infected intracerebraly with RML scrapie clearly detected PKresistant EGFP-PrP using specific L42 antibody. This result was confirmed by immunohistochemistry showing a staining pattern comparable to that of wildtype PrP. Confocal microscopy showed EGFP-positive aggregates only in the scrapie infected mice, but not in the Mock-controls, most likely corresponding to EGFP-PrPsc.
Conclusion: In summary, those lines are ideal models to monitor early changes in the localization of the EGFP-PrP fusion protein during scrapie infection. Experiments to identify the first target cells after oral infection are in progress.
AD S. Eigenbrod, M. Karmi, M. Maringer, U. Bertsch, M. Fuhrmann, C. Pace, N. Piening, J. Herms, C. Pilz, O. Windl, H. Kretzschmar, Ludwig-Maximilians-University, Center for Neuropathology and Prion Research, Germany
SP englisch
PO Schottland