ROLE OF APP AND INTERACTING PROTEINS IN SYNAPTIC DAMAGE

阿尔茨海默病（AD）的特征在于存在β-淀粉样蛋白斑块和 大脑中的神经系统缠结越来越多的证据支持淀粉样β蛋白（Abeta）在老年斑中的生成和沉积是AD发病机制中的早期和可能的主要事件。根据这种“淀粉样蛋白假说”，Abeta可能首先靶向突触，最初引起功能紊乱，随后是物理损伤和突触丢失，随后是神经元损伤。Abeta引起突触丧失和神经元死亡的机制和细胞通路尚不清楚。我们最近在初步研究中证明，Abeta可以加速APP多聚化，这似乎增加了细胞死亡的易感性。该途径需要完整的APP胞质区域，尤其是靠近C-末端的半胱天冬酶切割位点。这些初步结果导致了工作假设，即APP胞质结构域在AD的突触丢失和神经元死亡中起重要作用。我们提出了一种模型，其中Abeta诱导的细胞死亡的一种途径涉及与APP的复合物形成，该模型与充分描述的FasL-Fas受体途径相似。重要的是，该模型的支持已经从新产生的APP转基因小鼠系的初步分析中获得，其中APP胞质尾部中的共有半胱天冬酶切割位点已经突变（D 664 A），使得其不能被切割。在这些动物的脑中产生高水平的Abeta以及Abeta沉积物，但是通过突触素免疫反应性确定没有可检测到的突触损失。这些体内观察结果与我们的模型完全一致，其中APP转基因小鼠中的突触损伤由涉及Abeta和APP胞质结构域的途径介导。在第一个特定目标中，我们的目标是详细描述这种新产生的APP D 664 A转基因小鼠系的形态学、电生理学和行为表型。在第二个具体目标中，我们将在人脑样本中启动研究，以将caspase裂解的APP水平与死亡前认知和AD病理学相关联。此外，我们将启动研究，以确定与APP在大脑中相互作用的蛋白质。总之，这些实验的结果应该为AD患者大脑中的突触丢失提供重要的机制性见解。

Alzheimer's disease (AD) is characterized by the presence of both beta-amyloid plaques and neurofibrillary tangles in brain. Increasing evidence favors the generation and deposition of amyloid beta-protein (Abeta) in senile plaques as an early and possibly primary event in the pathogenesis of AD. According to this "amyloid hypothesis", Abeta may first target the synapse, initially causing functional perturbations, followed by physical damage and synapse loss, and subsequently neuronal injury. The mechanisms and cellular pathways by which Abeta causes synapse loss and neuronal death are unclear. We have recently demonstrated in preliminary studies that Abeta can accelerate APP multimerization, an event that appears to increase the susceptibility to cell death. This pathway requires an intact APP cytoplasmic region, especially the caspase cleavage site near the C-terminus. These preliminary results have led to the working hypothesis that the APP cytoplasmic domain plays an important contributing role in synapse loss and neuronal death in AD. We propose a model in which one pathway of Abeta- induced cell death involves complex formation with APP, a model which has parallels to the well described FasL-Fas receptor pathway. Importantly, support of this model has been obtained from preliminary analyses of a newly generated line of APP transgenic mice in which the consensus caspase cleavage site in the APP cytosolic tail has been mutated (D664A) so that it cannot be cleaved. High levels of Abeta are generated in the brains of these animals together with Abeta deposits but there is no detectable loss of synapses as determined by synaptophysin immunoreactivity. Theses in vivo observations are entirely consistent with our model in which the synaptic damage in APP transgenic mice is mediated by a pathway that involves both Abeta and the APP cytoplasmic domain. In the first Specific Aim, our goal is to characterize in detail the morphological, electrophysiological, and behavioral phenotype of this newly generated APP D664A transgenic mouse line. In the second Specific Aim, we will initiate studies in human brain samples to correlate levels of caspase cleaved APP with premortem cognition and AD pathology. In addition, we will initiate studies to identify proteins that interact with APP in brain. In sum, results from these proposed experiments should provide important mechanistic insights into synapse loss in brain of AD individuals.