Amyloid precursor protein : cellular studies and animal models

Abstract

Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterised by a number of neuropathological features, including extracellular deposits composed primarily of amyloid-β (Aβ) peptides. Aβ is derived from proteolysis of the amyloid precursor protein (APP) by consecutive action of β and γ-secretases. APP is a ubiquitously expressed type I transmembrane protein with a large N-terminal extracellular domain, a single transmembrane span and a short C-terminal cytoplasmic tail. Its physiological function is still unknown, but the existing data suggest that APP may function by linking extracellular cues, such as ligand- or substratum-binding, to intracellular signalling pathways. Because the natural ligand of APP is unknown, several studies have used antibodies against different epitopes in APP extracellular domain to mimic the ligand-receptor interaction.

In our study (paper I), using the antibody approach, we examined the role of APP in activation of gene transcription, and found that antibody-bound APP upregulates expression of ornithine decarboxylase (ODC), which is the initial and rate-controlling enzyme in polyamine biosynthesis. The induced ODC expression was rapid and biphasic, resembling growth-factor stimulated signalling events. This APP signalling did not require γ-secretase cleavage, as it was independent of the presence of presenilin-1 and -2. In paper II, we investigated the localisation and levels of ODC protein in AD brain. Specifically, we examined the ODC immunoreactivity in three different brain regions; in hippocampus, frontal cortex and cerebellum taken from control, possible and definite AD cases. Qualitative and quantitative analyses of these results demonstrated that ODC translocates in AD, from the nuclear compartment towards the cytoplasmic. Western blotting of frontal cortex homogenates showed that the levels of ODC in AD ar c increased. Both the translocation and change in ODC levels occur early in the disease process, i.e. in possible AD.

Animal models of AD are extremely valuable for the discovery and development of new treatments. Most of these models have been generated in mice. However, for decades the rat has been the preferred model for pharmacological and behavioural studies. In paper III and IV, we report the establishment and characterization of transgenic rats expressing human APP695 with the Swedish double mutation (tgAPPswe). These rats were generated by pronuclear injection. using a construct that contained human APPswe cDNA driven by the ubiquitin promoter paper III). The highest expression of the transgene was observed in cortex, hippocampus and cerebellum. Immunohistochemical examination of brain tissue revealed extracellular Aβ42 staining, either as cerebrovascular deposits or very rare diffuse plaques in the deep layers of the cortex, but the amyloid pathology was limited and occurred at a high age, above 15 months (paper III). Western blot analysis of hippocampal and cortical brain hornogenates showed hyperphosphorylated tau, but no neuronal and synaptic loss Taper IV).

We further characterised the tgAPPswe rats using behavioural tests, finding that these rats were more active (in the open-field) and showed impaired acquisition of learning (in the Morris water maze) when compared to the controls (paper IV). We also investigated the hippocampus and lateral ventricles of transgenic rats by in vivo MRI at the age of 16 months (paper IV). Both visual examination of the MR images and quantitative determination of the areas of these two structures suggest hippocampal atrophy and enlargement of lateral ventricles in transgenic rats compared to agematched controls. We believe that the tgAPPswe rats represent a unique model of early AD).