Project 3 - Animal Models Examining Neurovasculature

项目3--项目摘要/摘要 血脑屏障和神经血管单位的每个细胞成分的功能障碍 -血管细胞、神经胶质细胞和神经元-在实验中被认为与阿尔茨海默病(AD)的演变有关， 影像、病理学和流行病学研究。这些发现导致了一种正在出现的“神经血管” 阿尔茨海默病的假说，认为脑血管功能障碍导致认知功能减退和痴呆 在公元后。项目3支持P01的总体主题，重点是载脂蛋白E(APOE)基因，主要遗传基因 阿尔茨海默病的危险因素，通过在细胞、分子上实验性地询问血脑屏障/神经血管假说 和系统级别，并以特定于异构体和特定性别的方式使用新的APOE3和APOE4敲击 在(KI)FLOX/FLOX小鼠(E3F；E4F)中，星形胶质细胞和周细胞(主要来源)apoE缺失或不缺失 血脑屏障相关的载脂蛋白E)，并有和没有Aβ和tau病理。根据我们的试点和公布的数据，我们 假设1)破坏了血脑屏障(BBB)蛋白质-蛋白质相互作用(PPI)信号网络 潜在的内皮屏障破坏和血脑屏障功能障碍将先于和预测突触和神经元 新的APOE KIF系中APOE4相对APOE3的功能障碍和行为改变 和AD病理；以及2)3K3A激活蛋白C(APC)，一种APC的细胞信号类似物，将正确 调节失调的BBB和PSD95突触PPI信号网络，将保护BBB和神经元功能，以及 减轻AD病理，从而延缓APOE/AD模型的发病和疾病进展。至 测试我们的假设，我们将使用新的E3F和E4F小鼠，并与Cre品系杂交， 在星形胶质细胞和周细胞中表达Cre重组酶以确定细胞特异性缺失Key的效果 血脑屏障相关载脂蛋白E源(S和II)与APP/PS1-21小鼠和P301S tau小鼠的相互作用 有β和tau通路。为了评估血脑屏障/血管功能障碍，我们将使用：1)高级分子 同时检测脑脊液中血脑屏障/NVU细胞特异性标志物及Aβ和Tau生物标志物评价血脑屏障功能 和血浆；2)新的BBB蛋白质组学分析；3)高级MRI对BBB和CBF的评估；以及4) 通过组织分析验证生物流体生物标记物。为了评估突触和神经元功能障碍，我们将 用途：5)PSD95相互作用分析；6)病毒追踪Papez环路/DMN中的海马区通路； 6)行为和7)神经病理学分析。我们将评估apoE等位基因对血脑屏障和 生物流体，组织和成像生物标记物，血脑屏障和PSD95 PPI分析，海马神经元功能障碍 不同年龄apoE kif鼠(雄性、雌性)中apoE缺失和不缺失的病毒追踪和行为 来自星形胶质细胞和周细胞(AIM 1)，与APP/PS1-21和P301S tau小鼠杂交与AD病理相关 (目标2)，并将评估3K3A-APC在不同年龄的APOE KIF小鼠(雄性、雌性)中的治疗效果，单独和 与APP/PS1-21和P301S tau小鼠杂交(AIM 3)。项目3将增进我们对 阿尔茨海默病在细胞和分子水平上的发病机制与载脂蛋白E亚型和性别有关。

PROJECT 3 – PROJECT SUMMARY/ ABSTRACT Dysfunctionin the blood-brain barrier (BBB) and each of the cellular components of the neurovascular unit (NVU) – vascular cells, glial cells, and neurons – has been linked to Alzheimer’s disease (AD) evolution in experimental, imaging, pathological, and epidemiological studies. These findings have led to an emerging ‘neurovascular hypothesis’ of AD which holds that cerebrovascular dysfunction contributes to cognitive decline and dementia in AD. Project 3 supports the overall theme of P01 focused on apolipoprotein E (APOE) gene, the major genetic risk factor for AD, by interrogating the BBB/neurovascular hypothesis experimentally at the cellular, molecular and systems levels and in an isoform-specific and gender-specific fashion using novel APOE3 and APOE4knock in (KI)flox/flox mice (E3F; E4F) with and without apoE deletion from astrocytes and pericytes (the key sources of BBB-associated apoE), and with and without Aβ and tau pathology. Based on our pilot and published data we hypothesize that 1) Disrupted blood-brain barrier (BBB) protein-protein interaction (PPI) signaling networks underlying endothelial barrier disruption and BBB dysfunction will precede and predict synaptic and neuronal dysfunction and behavioral changes driven by APOE4 relative to APOE3 in new APOE KIF lines both without and with AD pathology; and 2) 3K3A-activated protein C (APC), a cell signaling analog of APC, will correct dysregulated BBB and PSD95 synaptic PPI signaling networks, will protect BBB and neuronal function, and diminish AD pathology, thereby delaying onset and slowing down disease progression in APOE/AD models. To test our hypothesis, we will use novel E3F and E4F mice alone and crossed i) with Cre lines that specifically express Cre recombinase in astrocytes and pericytes to determine the effects of cell-specific deletion of the key BBB-associated apoE source(s) and ii) with APP/PS1-21 mice and P301S tau mice to investigate interactions with Aβ and tau pathways. To evaluate BBB/vascular dysfunction we will use: 1) advanced molecular assessment of the BBB by simultaneously measured BBB/NVU cell-specific and Aβ and tau biomarkers in CSF and plasma; 2) a novel BBB proteomics analysis; 3) advanced MRI assessment of BBB and CBF; and 4) validation of biofluid biomarkers by tissue analysis. To evaluate synaptic and neuronal dysfunction we will use: 5) PSD95 interactomeanalysis; and 6) viral tract-tracing of hippocampal pathways in the Papez circuit/DMN; and 6) behavior and 7) neuropathology analysis. We will evaluate apoE-allele-specific effects on BBB and neuronal dysfunction by biofluid, tissue and imaging biomarkers, BBB and PSD95 PPI analysis, hippocampal viral tract-tracing, and behavior in APOE KIF mice(male, female) at different ages with and without apoE deletion from astrocytes and pericytes (AIM 1), crossed with APP/PS1-21 and P301S tau micein relation to AD pathology (AIM 2), and will evaluate 3K3A-APC therapy in APOE KIF mice (male, female) at different ages, alone and crossed with APP/PS1-21 and P301S tau mice (AIM 3). Project 3 will advance our understanding of the pathogenesis of AD at the cellular and molecular level in an apoE isoform-specific and gender-specific fashion.