AD LENS PATHOLOGY: BIOCHEMISTRY & DIAGNOSTIC IMAGING

描述（由申请人提供）：这项新研究者申请的目标是研究阿尔茨海默病（AD）在脑和眼透镜中的发病机制。阿尔茨海默病的主要特征是脑内B-淀粉样蛋白（AB）的积累。我们最近在人类晶状体中发现了AB，并在患有AD的老年人晶状体中检测到AB沉积、淀粉样蛋白病理学和共定位赤道核上性白内障（SNC），但在没有这种疾病的人中则没有。因为这些不寻常的白内障位于虹膜后面的透镜周边，所以这种表型既不是视觉残疾，也不是通过常规检查可观察到的。我们已经在Tg 2576 AD转基因小鼠晶状体中鉴定了类似的SNC和人AB过表达。AB定位于透镜纤维细胞的细胞质，在那里该肽与胞质透镜蛋白如AB-晶状体蛋白相互作用，AB-晶状体蛋白是在AD脑中上调的丰富的透镜结构蛋白/分子伴侣。AB促进金属依赖性透镜蛋白聚集和光散射。该数据支持AIS介导的透镜蛋白聚集增加核上光散射的模型，其最终表现为白内障。我们假设SNC表型及其分子前因可能可以作为AD疾病过程的早期器官特异性表达来检测。为了研究这一假设，我们将进行AB介导的透镜蛋白聚集的生化分析，并检查透镜AI 5作为推定的AD生物标志物的非侵入性体内监测的潜力。拟定研究将利用体外模型系统、离体人AD晶状体和Tg 2576转基因小鼠。目的1检验透镜AB蓄积与局部透镜蛋白质聚集、光散射和生物金属蓄积相关的假设。我们将确定人AD/对照和Tg 2576/WT小鼠晶状体中的AB和生物金属浓度、定位和组成。目的2验证AB与其他透镜蛋白相互作用通过金属蛋白氧化还原反应促进蛋白聚集的假设。我们将研究AB与胞质结构蛋白（a-、B-和γ-晶体蛋白）的结合，并研究在AB物质、金属螯合剂和气体（空气、O2、Ar）存在下透镜蛋白聚集。目的3检验AB介导的透镜蛋白聚集可作为外周可及AD生物标志物的假设。我们将构建专用的光学仪器，用于定量、非侵入性的体内检测AB相关透镜蛋白聚集。我们将利用这些仪器检查AD/非AD晶状体离体和Tg 2576/WT小鼠体内。我们将在四组小鼠（Tg 2576、Ctrl+/-Cu缺陷敲除、Tg 2576 x Ctrl+/-杂交、WT对照）中对透镜蛋白聚集进行纵向体内评估，并将首次透镜测量值与年龄、透镜/脑AB负荷、金属浓度和组织病理学相关联。由此产生的数据可以提供对AD相关蛋白质充气的基本机制的见解，并支持开发新的AD诊断和监测技术。

DESCRIPTION (provided by applicant): The goal of this new-investigator application is to study Alzheimer's disease (AD) pathogenesis in the brain and ocular lens. Age-dependent cerebral B-amyloid (AB) accumulation is a cardinal feature of Alzheimer's disease. We have recently identified AB in human lenses and detected AB deposition, amyloid pathology, and co-localizing equatorial supranuclear cataracts (SNC) in lenses from aged individuals with AD but not in those without the disorder. Because these unusual cataracts are located at the lens periphery behind the iris, this phenotype is neither visually disabling nor observable by routine examination. We have identified similar SNC and human AB overexpression in Tg2576 AD transgenic mouse lenses. AB localizes to the lens fiber cell cytoplasm, where this peptide interacts with cytosolic lens proteins such as AB-crystallin, an abundant lens structural protein/molecular chaperone that is upregulated in AD brain. AB promotes metal-dependent lens protein aggregation and light scattering. This data supports a model in which AIS-mediated lens protein aggregation increases supranuclear light scatter that ultimately manifests as cataracts. We hypothesize that the SNC phenotype and its molecular antecedents may be detectable as an early, organ-specific expression of the AD disease process. To investigate this hypothesis, we will conduct biochemical analyses of AB-mediated lens protein aggregation and examine the potential for non-invasive in vivo monitoring of lens AI5 as a putative AD biomarker. The proposed studies will utilize in vitro model systems, ex vivo human AD lenses, and Tg2576 transgenic mice. Aim 1 tests the hypothesis that lens AB accumulation is associated with localized lens protein aggregation, light scattering, and biometal accumulation. We will determine AB and biometal concentrations, localization, and composition in human AD/control and Tg2576/WT mice lenses. Aim 2 tests the hypothesis that AB interacts with other lens proteins to promote protein aggregation via metalloprotein redox reactions. We will investigate AB binding to cytosolic structural proteins (a-, b-, and y-crystallins) and study lens protein aggregation in the presence of AB species, metal chelators, and gases (air, 02, Ar). Aim 3 tests the hypothesis that AB-mediated lens protein aggregation may serve as a peripherally-accessible AD biomarker. We will construct purpose-built optical instruments for quantitative, non-invasive in vivo detection of AB-associated lens protein aggregation. We will utilize these instruments to examine AD/non-AD lenses ex vivo and Tg2576/WT mice in vivo. We will conduct a longitudinal in vivo assessment of lens protein aggregation in four groups of mice (Tg2576, Ctrl+/- Cu-deficit knockouts, Tg2576 x Ctrl+/- crosses, WT controls) and correlate index lens measurements with age, lens/brain AB burden, metal concentration, and histopathology. The resulting data may provide insights into fundamental mechanisms of AD-associated protein aeration and support development of novel AD diagnostic and monitoring technology.