Viable Microvessels for the Study of the Microvasculature in Alzheimer's Disease

阿尔茨海默病(AD)患者的脑微血管结构会发生改变，但几乎没有治疗方法 研究活组织中的细胞机制。存活的脑微血管(MV)可以从整体分离出来 UW神经病理学中快速尸检受试者的脑组织 核心。最近，该技术已经被修改，使得在3维(3D)矩阵中MV具有 将生存能力延长到隔离后至少10天(D)，以及生成大量副本的可扩展性 学习。受试者对于痴呆症或非痴呆症的临床诊断具有很好的特征，而且 随后对标本进行广泛的神经病理学评估。 MV的血脑屏障(BBB)是调节物质进入 并从大脑中流出。血脑屏障的主要细胞成分是脑内皮细胞(BEC)， 周细胞和星形胶质细胞的足突。周细胞与BEC紧密相对，支持维持。 血脑屏障的属性。重要的是，BEC和周细胞促进并受这两者的影响。 基底膜(BM)和细胞外基质(ECM)，AD使其发生显著改变。 3D培养系统中MVS的研究使相关细胞和蛋白质的空间分析成为可能 组件。延长的MV存活率允许对干预措施和潜在机制进行评估 影响MV健康。目标是建立这些MV以获得关于微血管基础信息 AD中的变化。有两个特定的目标，每个受试者的MV作为他们自己的基线对照。在……里面 目的1、假设MV来自年龄和性别匹配的受试者，有无临床诊断 对于痴呆症，将保持保真度(细胞活力、血脑屏障功能蛋白水平和细胞外基质成分) 长时间(长达10d)的初始隔离MV将在3D中进行测试。所有的标本都会有 分析他们的供体整体脑组织样本，以确定3D MV是否能维持细胞， BBB蛋白和脑MV的ECM表型，进行组织学原位检测。 在受AD神经病理影响的脑区，周细胞丢失加速。缺乏 周细胞可导致血脑屏障功能改变，但对细胞外基质的影响尚未得到很好的研究。目标2 将确定周细胞丢失对非痴呆受试者脑部MV的影响 BBB ECM。在目标2中，假设周细胞丢失会导致细胞外基质含量的变化，从而导致MV 非痴呆症患者的ECM将类似于痴呆症患者的MV ECM，将进行测试。目标 1和2将同时进行，以优化脑标本的利用。综合来看，数据来自 这些目的将促进AD脑MV的研究，为进一步的干预工作创造一个平台。

The brain microvasculature is altered in Alzheimer’s disease (AD), but there are few methods for studying cellular mechanisms in living tissue. Viable brain microvessels (MV) can be isolated from whole brain tissue of subjects undergoing rapid autopsy (<12 hours post mortem) in the UW Neuropathology Core. Recently, this technique has been adapted such that MV in a 3 dimensional (3D) matrix have extended viability to at least 10 days (d) post-isolation, and scalability to generate numerous replicates for studies. Subjects are well characterized for the clinical diagnosis of dementia or no dementia, and all specimens undergo subsequent extensive neuropathologic assessment. The blood-brain barrier (BBB) of the MV is the interface that regulates movement of substances into and out of the brain. The principle cellular components of the BBB are brain endothelial cells (BEC), pericytes, and astrocyte foot processes. Pericytes are closely apposed to BEC and support maintenance of BBB properties. Importantly, BEC and pericytes contribute to, and are affected by, both the glycocalyx and basement membrane (BM) extracellular matrix (ECM), which is altered significantly by AD. Studies of MVs in 3D culture systems allow for spatial analysis of relevant cellular and protein components. Extended MV viability allows for evaluation of interventions and potential mechanisms that impact MV health. The goal is to establish these MV to obtain foundational information on microvascular changes in AD. There are 2 specific aims, with each subject’s MV serving as their own baseline control. In Aim 1, the hypothesis that MV from age and sex matched subjects, with and without the clinical diagnosis of dementia, will maintain fidelity (cellular viability, levels of BBB functional proteins, and ECM components) to their initially isolated MV for prolonged periods (up to 10d) in 3D will be tested. All specimens will have analysis of their donor en bloc brain specimens, to determine whether MV in 3D will maintain the cellular, BBB proteins, and ECM phenotype of their brain MV that are examined histologically in situ. Pericyte loss is accelerated in the MV of brain regions affected by AD neuropathology. A lack of pericytes is known to result in changes in BBB function, but effects on ECM are not well studied. Aim 2 will determine the effects of pericyte loss on MV derived from brains of subjects without dementia on the BBB ECM. In Aim 2, the hypothesis that pericyte loss results in alterations in ECM content, such that MV ECM from those without dementia will resemble MV ECM from those with dementia, will be tested. Aims 1 and 2 will be performed concurrently to optimize utilization of brain specimens. Taken together, data from these Aims will advance the study of brain MV in AD to create a platform for additional interventional work.