Production of transgenic mice expressing familial AD relevant genes

本研究致力于建立一种实验动物模型，证明大多数家族性阿尔茨海默病(FAD)的致病基因早老素1基因的错义突变导致AD病理损害之一的Aβ沉积的表型，并为分析参与该级联反应的蛋白质的加工提供工具。为此，我们试图培育出表达FAD相关蛋白的转基因小鼠。我们已经培育出表达野生型人类PS1(WtPS1)蛋白和表达错义PS1(密码子146 Met→Val)的TG小鼠，这是一种错义突变，被证明可以增加Aβ1-42的产生。此外，我们还培育出了在密码子717(伦敦类型的FAD)上表达突变APP的TG小鼠。我们制备了针对N端1~81个氨基酸序列和C端299~407个氨基酸序列的多克隆抗体，并将它们与GST融合。利用这些工具，我们将分析与Aβ沉积有关的机制。有报道表明，随着年龄的增长，突变的APP TG小鼠在大脑中会产生Aβ沉积(1.5年后)，而突变的PS1 TG小鼠不会在大脑中产生Aβ沉积。为了验证Aβ沉积的种子假说，对幼年APP717TG小鼠和M146VPS1TG小鼠进行了Aβ多肽的脑内注射。我们将观察这些动物体内注射的物质是否可以作为加速内源性小鼠Aβ沉积的种子。

This study is dedicated to establish an experimental animal model which allows to demonstrate the cascade that missense mutation in presenilin 1 gene identified as causative gene for the majority of familial Alzheimer's disease (FAD) leads to the phenotype of Aβ deposition, one of the pathological lesions of AD, and to provide a tool for analysis of processing of proteins involved in the cascade. For this purpose we attempted to produce trasgenic mice which express FAD relevant proteins. We have produced Tg mice which express wild type human PS1(wtPS1) protein and those which express missense PS1 (codon 146 Met → Val), one of the missense mutations shown to increase the production of Aβ1-42. In addition, we have produced Tg mice which express mutant APP at codon 717(London type of FAD). We have produced polyclonal antibodies generated against N-terminal 1 to 81 amino acid sequence and C-terminal 299 to 407 amino acids sequence, both of which were fused to GST. Using these tools we will analyze the mechanism involved in the deposition of Aβ. There have been reports showing that mutant APP Tg mice generate Aβ deposits in the brain with aging (after 1.5years), whereas mutant PS1 Tg mice fail to generate Aβ deposits in the brain. To demonstrate "the seeding hypothesis for Aβ deposition", young APP 717 Tg mice and M146V PS1 Tg mice are subjected to intracranial administration of Aβ peptide. We will see whether the injected material in these animals serve as a seed for accelerated deposition of endogenous murine Aβ.