Metabolomic/lipidomic analysis of apoE isoform effects in AD-linked brain regions

描述（由申请人提供）：载脂蛋白E (APOE) ε4基因携带者患阿尔茨海默病（AD）的风险显著增加。尽管提出了许多理论，但这种联系的原因仍不清楚。最被广泛接受的观点是，APOE ε4携带者的AD病理加速是由于apoE4蛋白从大脑中清除a β的能力下降。然而，拥有APOE ε4基因也会导致一些与a β清除无关的其他神经功能缺陷，包括神经元结构改变、内嗅皮层（EC）层变薄和中风后预后较差，这表明可能有其他机制参与了这一过程。为了更全面地了解不同apoE亚型的表达如何影响大脑，以及这可能如何影响AD和其他与年龄相关的神经系统疾病的风险，我们建议使用质谱法鉴定脂质和小分子，其水平受apoE亚型表达变化的影响。为了实现这一目标，我们将首先从14个月大的小鼠和死后19-55岁表达不同载脂蛋白e亚型的个体的无病理EC和初级视觉皮层（PVC）组织中提取脂质、小分子和蛋白质。然后，从这些提取物中提取的脂质和小分子组分将用于进行靶向脂质组学和非靶向代谢组学，随后进行生物信息学分析和各种验证实验，以确定受大脑中apoE异构体表达影响的特定脂质、小分子和代谢途径。到目前为止，初步研究已经发现了能量代谢途径和几个重要的脂质亚类的重大变化，证明了这些技术在发现以前未知的apoE在大脑中的同种异构体特异性作用方面的力量。我们期望本文提出的完整研究将进一步揭示apoE亚型在脂质和小分子水平上的特异性变化，并将进一步了解apoE4如何影响阿尔茨海默病的病理，可能为阿尔茨海默病和其他受apoE4表达影响的神经系统疾病提供新的治疗策略。

DESCRIPTION (provided by applicant): Carriers of the apolipoprotein E (APOE) ε4 gene are at significantly increased risk for developing Alzheimer's disease (AD). Although numerous theories have been proposed, the cause of this association remains unclear. The most widely accepted view is that the accelerated AD pathology observed in APOE ε4 carriers is due to a decreased ability of the apoE4 protein to clear Aβ from the brain. However, possession of the APOE ε4 gene also results in a number of other neurological deficits unrelated to Aβ clearance, including alterations in neuronal structure, thinner entorhinal cortex (EC) layers and poorer outcomes after stroke, suggesting that there may be other mechanisms involved in this process. In order to gain a more comprehensive understanding of the how the expression of different apoE isoforms affects the brain and how this may impact the risk of developing AD and other age-related neurological illnesses, we propose to use mass spectrometry to identify lipids and small-molecules whose levels are affected by changes in apoE isoform expression. To accomplish this, we will first extract lipids, small-molecules and proteins from pathology-free EC and primary visual cortex (PVC) tissues obtained from 14-month old mice and postmortem 19-55 year old individuals expressing differing apoE isoforms. The lipid and small-molecule fractions from these extracts will then be used to perform targeted lipidomics and untargeted metabolomics, followed by bioinformatic analysis and a variety of validation experiments, in order to determine the specific lipids, small-molecules and metabolic pathways that are affected by alternative apoE isoform expression in the brain. Thus far, preliminary studies have uncovered significant changes in energy metabolism pathways and several important lipid subclasses, demonstrating the power of these techniques for discovering previously unknown isoform-specific effects of apoE in the brain. We expect that the full study proposed herein will uncover further apoE isoform-specific changes in lipid and small-molecule levels and will lead to a greater understanding of how apoE4 influences AD pathology, potentially leading to new therapeutic strategies for AD and other neurological illnesses influenced by apoE4 expression.