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

阿尔茨海默病（AD）的脑微血管系统发生改变，但很少有方法用于治疗AD。 研究活组织中的细胞机制活的脑微血管（MV）可以从整个脑组织中分离出来。 在UW神经病理学中进行快速尸检（死后<12小时）的受试者的脑组织 核心最近，该技术已经被调整，使得3维（3D）矩阵中的MV具有 将存活力延长至分离后至少10天（d），并且可扩展性以产生用于 问题研究受试者对于痴呆或非痴呆的临床诊断具有良好的特征，并且所有 随后对标本进行广泛的神经病理学评估。 MV的血脑屏障（BBB）是调节物质进入脑内的运动的界面。 从大脑中取出BBB的主要细胞成分是脑内皮细胞（BEC）， 周细胞和星形胶质细胞足突。Pericytes与BEC密切相关，支持维护 BBB地产重要的是，BEC和周细胞有助于糖萼，并受其影响， 和基底膜（BM）细胞外基质（ECM），其被AD显著改变。 3D培养系统中MV的研究允许相关细胞和蛋白质的空间分析 件.延长的MV活力允许评价干预措施和潜在机制， 影响MV健康。目的是建立这些MV，以获得微血管的基础信息 AD的变化。有2个具体目标，每个受试者的MV作为其自己的基线对照。在 目的1，假设MV来自年龄和性别匹配的受试者，有和无临床诊断 将保持保真度（细胞活力，BBB功能蛋白质水平和ECM成分） 将在3D中测试其初始隔离MV的长时间（长达10天）。所有样本将具有 分析他们的供体整块脑标本，以确定3D中的MV是否将维持细胞， BBB蛋白和脑MV的ECM表型进行原位组织学检查。 在受AD神经病理学影响的脑区域的MV中，周细胞损失加速。缺乏 已知周细胞导致BBB功能的变化，但对ECM的影响尚未得到充分研究。目的2 将确定周细胞损失对来自无痴呆受试者脑的MV的影响， BBB ECM。在目标2中，假设周细胞损失导致ECM含量的改变，例如MV 将测试来自未患痴呆症的人的ECM将类似于来自患有痴呆症的人的MV ECM。旨在 1和2将同时进行，以优化脑标本的利用。综合来看，来自 这些目标将推进AD中脑MV的研究，为额外的介入工作创建平台。

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.