Mutated human genes associated with human neurodegenerative conditions were used to develop both <i>in vitro</i> and <i>in vivo</i> models to study cellular pathology and disease progression. The <i>in vitro</i> models employed adenoviral vectors for the gene delivery into primary rat hippocampal neurones. Introduction of both APP and Tau transgenes reduced neuronal viability, with the latter leading to accelerated toxicity and faster onset of cell death. Time-lapsing imaging analyses revealed apoptosis-like features for APP-positive neurones, while Tau-positive neuronal death appeared more necrotic. Interestingly, a direct correlation between cell death and protein content in the APP-transduced neurones was not confirmed. A comparison between viral gene delivery and electroporation with the same transgenic constructs confirmed the cellular toxicity of APP and Tau but also showed that with a lower amount of transgene expressing cells per culture dish Tau-induced toxicity was no longer as aggressive as with the viral model. Therefore, electroporation may allow single-cell investigations of functional parameters whereas the large amount of transgene-positive neurones in viral transductions allows faster quantification of cell death. These methods complement each other and thus offer <i>in vitro </i>models suitable for mechanistic studies and drug screening. Accordingly, initial testing of inhibitors of Tau aggregation and amyloid formation were found to ameliorate the transgene-induced damage as proof of principle for our novel <i>in vitro</i> models. Further testing revealed caffeine to be a very promising drug candidate in AD treatment, since it improved viability in both APP and Tau transduced neurones. The transgenic PLB1 mouse knock-in model harbours the same mutated APP and Tau genes as the viral models. A triple transgenic line (PLB1<sub>Triple</sub>) has been generated with additional mutated presenilin 1 for increased APP processing and accelerated pathology. The mRNA expression of both APP and Tau transgenes was stable over the investigated time (6 and 12 months) with about 2 to 3-fold higher APP over Tau mRNA levels. This expression was specific to the forebrain and negligible in the cerebellum and thus targets a brain region that is vulnerable in AD. The model showed progressive accumulation of AD-linked histopathological features as well as memory- and activity-related symptoms. This low transgene expression in conjunction with a progressive phenotype is advantageous over other aggressive animals models for studying early disease-related pathology.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:521240 |
| Date | January 2010 |
| Creators | Stoppelkamp, Sandra |
| Publisher | University of Aberdeen |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://digitool.abdn.ac.uk:80/webclient/DeliveryManager?pid=136906 |
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