MOLECULAR CHANGES IN NEURODEGENERATION

我们实验室过去几年的研究表明， APP的生理功能之一是调节细胞功能 和生存 特别地，APP已被发现是营养分子 在体外和突触中调节神经突延伸 在体内形成/结构。 考虑到在许多情况下， 因子保护神经元免受各种损伤引起的损伤，APP是 一个突出的候选人干预神经元损伤。 因此，我们建议在几个模型中研究APP的参与 神经元和突触的退化和再生。 具体目标 1、我们将确定APP在兔脊髓中的参与程度， 脊髓缺血模型（RSCIM）。 我们的假设是APP的合成和 降解受缺血的影响，作为保护的一部分， 对抗缺血引起的损伤的机制。 app异构型 在缺血之前和之后将被量化和定位，其中 可能的，在蛋白质和mRNA水平上使用各种技术 包括Western blotting、免疫组化、北方印迹、狭缝 印迹、RT-PCR技术和原位杂交。这项工作服务于 作为具体目标2的基础，我们将优化程序 用于使用RSCIM的APP-肽疗法。 这项研究是基于我们的 最近发现，一个代表活性的短肽片段 APP营养结构域拮抗兔脑缺血损伤 脊髓 我们将使用临床， 生物化学、分子生物学和形态学标准。 形态学标准包括神经元和突触计数。 在特定 目标3，我们将研究一个潜在的APP参与萌芽 穿通后齿状回分子层的反应 路径（PP）病变。 我们会问哪些神经元产生APP来响应 PP病变，并且如果输注干扰PP病变的肽， APP的作用是抑制发芽。 第4章测试 假设NGF和β-FGF的作用拯救了中间隔 穹窿海马伞切断后神经元变性 部分是通过激活假定的APP途径。 我们将 研究APP肽灌注对大鼠中隔神经元的影响 FF病变后。 然后，我们将尝试拮抗神经生长因子的作用， 和bFGF，通过共输注一种肽，该肽与APP一起完成并阻断其 功能 最后，在具体目标5中，我们将测试潜在影响 APP激动剂和拮抗剂在生长因子保护 谷氨酸诱导的神经元毒性，采用体外培养模型。

Research for the last few years in our laboratory has revealed that one of the physiological functions of APP is to regulate cellular functions and survival. In particular, APP has been found to be a trophic molecule that regulates neurite extension in vitro and synaptic formation/structure in vivo. Considering that in many instances trophic factors protect neurons from damages caused by various injuries, APP is a prominent candidate for the intervention of neuronal injuries. Therefore, we propose to study the involvement of APP in several models of neuronal and synaptic degeneration and regeneration. In Specific Aim 1, we will determine the degree of APP involvement in the rabbit spinal cord ischemia model (RSCIM). Our hypothesis is that APP synthesis and degradation are affected by ischemia as a part of the protective mechanisms to counteract the damage induced by ischemia. APP isoforms before and after ischemia will be quantified and localized, where possible, at both protein and mRNA levels using various techniques including Western blotting, immunohistochemistry, Northern blotting, slot blotting, RT-PCR technique, and in situ hybridization. This work serves as a foundation for Specific Aim 2 where we will optimize the procedure for the APP-peptide therapy using RSCIM. This study is based on our recent finding that a short peptide fragment representing the active trophic domain of APP antagonizes the ischemia-induced damage in rabbit spinal cords. We will evaluate ischemic injury using clinical, biochemical, molecular biological, and morphological criteria. Morphological criteria include neuronal and synaptic counts. In Specific Aim 3, we will examine a potential APP involvement in the sprouting reactions in the molecular layer of the dentate gyrus after the perforant path (PP) lesion. We will ask which neurons produce APP in response to PP lesion, and also if the infusion of a peptide which interferes with the function of APP inhibits the sprouting. Specific Aim 4 will test the hypothesis that the effect of NGF and beta-FGF to rescue medioseptal neurons from degeneration after the transection of fimbria-fornix (FF) is partially through the activation of putative APP pathways. We will investigate the effect of APP peptide infusion on the medioseptal neurons after FF lesion. Then, we will attempt to antagonize the effects of NGF and bFGF by co-infusing a peptide that completes with APP and blocks its function. Finally, in Specific Aim 5, we will test the potential effect of APP agonists and antagonists in the protection by growth factors from glutamate-induced neuronal toxicity employing the in vitro culture model.