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Biological Significance of Oxidized Neprilysin

氧化脑啡肽酶的生物学意义

基本信息

批准号：
7273587
负责人：
DENGSHUN WANG
金额：
$ 6.28万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-09-01 至 2008-08-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): The long-term objective of this study is to understand the biological significance of oxidative modification of neprilysin on amyloid deposition in aging and Alzheimer's disease (AD). Amyloid deposition in the brain parenchyma and in vessels is a common phenomenon with advanced age and one of the diagnostic hallmarks for AD. Two possible explanations for excessive beta-amyloid (AB) deposition in aging and AD brains are overproduction and decreased degradation. Although the mechanisms involved in AB production have been extensively studied and the excessive production of AB has been confirmed to play a critical role in the pathogenesis of familial AD cases, there is little evidence so far to suggest that increased brain AB production is important in aging and sporadic AD. Recently, the role of AB degradation has been increasingly studied and several enzymes have been described with a range of abilities to degrade AB. Among them, neprilysin (NEP) has been considered by some a pivotal enzyme for AB degradation. Previous studies have shown that decreased NEP may contribute to the accumulation of AB in AD. Recent data from our group indicated that brain NEP is subject to oxidative modification in both AD and aging. To understand the biological significance of oxidized-NEP it is very important to know if oxidization decreases NEP enzymatic activity. It is also important to know the HNE modification site(s) of NEP to design therapeutic reagents that can block oxidative modification of NEP. Our working hypothesis is that initial amyloid accumulation will lead to HNE production with progressive inhibitory NEP, starting a feed-forward cycle of increasing amyloid accumulation, oxidative stress, and NEP inhibition. To test this hypothesis, as initial steps we propose three specific aims in this RO3 proposal: 1. To determine if human NEP expressed in cultured cells can be modified by exogenous HNE or endogenous HNE induced by synthetic AB; 2. To discover if HNE-modified NEP expressed in cultured cells has decreased enzymatic activity. 3. To identify critical HNE-modification sites which alter NEP activity. Our goal through this RO3 mechanism is to develop enough preliminary data for a future RO1 application that will further explore this hypothesis in animal models and in humans.

描述（由申请人提供）：本研究的长期目标是了解脑啡肽酶氧化修饰对衰老和阿尔茨海默病（AD）中淀粉样蛋白沉积的生物学意义。脑实质和血管中淀粉样蛋白沉积是高龄人群的常见现象，也是 AD 的诊断标志之一。衰老和 AD 大脑中 β-淀粉样蛋白 (AB) 沉积过多的两种可能解释是生产过剩和降解减少。尽管AB产生的机制已被广泛研究，并且AB的过量产生已被证实在家族性AD病例的发病机制中发挥关键作用，但迄今为止几乎没有证据表明大脑AB产生的增加在衰老和散发性AD中具有重要作用。最近，人们对 AB 降解的作用进行了越来越多的研究，并且已经描述了几种具有多种降解 AB 能力的酶。其中，脑啡肽酶（NEP）被一些人认为是AB降解的关键酶。先前的研究表明，NEP 的减少可能会导致 AD 中 AB 的积累。我们小组的最新数据表明，大脑 NEP 在 AD 和衰老过程中都会受到氧化修饰。要了解氧化 NEP 的生物学意义，了解氧化是否会降低 NEP 酶活性非常重要。了解 NEP 的 HNE 修饰位点对于设计可以阻止 NEP 氧化修饰的治疗试剂也很重要。我们的工作假设是，最初的淀粉样蛋白积累将导致 HNE 产生，并逐渐抑制 NEP，开始增加淀粉样蛋白积累、氧化应激和 NEP 抑制的前馈循环。为了检验这一假设，作为初步步骤，我们在 RO3 提案中提出了三个具体目标： 1. 确定培养细胞中表达的人类 NEP 是否可以通过合成 AB 诱导的外源 HNE 或内源 HNE 进行修饰； 2. 观察培养细胞中表达的HNE修饰的NEP是否降低了酶活性。 3. 确定改变NEP活性的关键HNE修饰位点。我们通过这种 RO3 机制的目标是为未来的 RO1 应用开发足够的初步数据，从而在动物模型和人类中进一步探索这一假设。