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Title: Mechanisms of neurodegeneration in the transmissible spongiform encephalopathies
Author: Clarke, Gary James
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2005
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The primary aims of this thesis were firstly to analyse the role of caspase-independent apoptotic cell death pathways in TSEs, and secondly to develop an in vitro model of neuronal loss in order to asses the relationship between amyloid fibril structure and neurotoxocity. To analyse caspase-independent pathways in two well characterised murine scrapie models (ME7/CV and 87V/VM), whole or micro-dissected brain areas from terminal stage infected mice were separated into cytosolic and mitochondrial fractions, and their protein contents compared by Western blot to normal brain injected age-matched control animals. Micro-dissection proved to be a much more sensitive technique for the detection of apoptosis-related proteins in vivo. Caspase-independent AIF was found to translocate from the IMM to the cystosol in ME7/CV animals, but not in 87V/VM or in normal brain-injected control cohorts. The release of AIF into the cytosol is therefore specific to infection with the ME7 murine scrapie strain. A neuronal PC12 cell system was established to model TSE neurodegeneration in vitro. Murine PrP105-125, homologous to the neurotoxic human PrP106-126 synthetic peptide, was developed specifically for this research. Mature amyloid fibrils were created containing β-sheet structures, but different tertiary structures as revealed by electron microscopy, Thioflavin T binding assays and FT-IR techniques. The primary structure of the MoPrP105-125 peptide is critical for conferring the neurotoxicity, as a scrambled sequence is not toxic to neurons. Furthermore, the research within this thesis shows that morphology of mature amyloid fibrils does not have a significant effect on toxicity, suggesting that the intermediate soluble protofibrillar structures may prove to be the more toxic species.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available