Reverse engineering zonation-specific and age-specific iPSC-derived cerebrovascular models based on transcriptomic profiling of the human brain

基于人脑转录组分析的逆向工程分区特异性和年龄特异性 iPSC 衍生脑血管模型

• 批准号: 10321473
• 负责人: Myriam Heiman
• 金额: $ 83.31万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-15 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10321473
• 关键词: 3-Dimensional; Age; Aging; Automobile Driving; Biochemical Process; Blood; Blood Vessels; Blood capillaries; Blood flow; Brain; Brain Diseases; Cell Line; Cell Nucleus; Cells; Cerebrovascular Disorders; Cerebrovascular system; Collaborations; Cues; Development; Disease; Disease Progression; Engineering; Functional disorder; Gene Expression; Gene Expression Profiling; Genetic Screening; Genomics; Geometry; Goals; Health; Human; Human Engineering; In Vitro; Individual; Intrinsic factor; Libraries; Maintenance; Modeling; Molecular; Neurodegenerative Disorders; Nutrient; Pathology; Perfusion; Phase; Phenotype; Play; Process; Resected; Role; Serum; Small Nuclear RNA; Source; Supporting Cell; Tissue Engineering; Tissues; Translations; Western Blotting; age related; aged; arteriole; base; brain endothelial cell; brain health; cellular transduction; cerebrovascular; induced pluripotent stem cell; next generation; physical process; protein expression; response; shear stress; stem cell technology; trafficking; transcription factor; transcriptome sequencing; transcriptomics; venule; wasting

Project Summary Cerebrovascular dysfunction is emerging as a common pathology in many diseases of the brain, including neurodegenerative diseases, cerebrovascular diseases, as well as in aging. Therefore, understanding the role of cerebrovascular dysfunction in disease progression and aging will be key to long-term maintenance of brain health. With developments in tissue engineering and stem cell technology, in vitro cerebrovascular models can play an important role in understanding the role of cerebrovascular dysfunction in disease progression and aging. Next-generation cerebrovascular models should take into account three key factors: (1) differences in phenotype of brain microvascular endothelial cells along the arterio-venous axis, (intrinsic factors), (2) differences in microenvironmental cues along the arterio-venous axis (extrinsic factors), and (3) changes in zonation-specific cerebrovascular phenotype during aging and in response to aged serum. Therefore, the overall goal of this project is to use zonation- and age-specific intrinsic and extrinsic cues to reverse engineer human cerebrovascular models, and to use these models to understand cerebrovascular phenotype during aging. We will first perform a pooled genetic screen to identify transcription factor combinations that are capable of driving source cells towards gene expression profiles of human brain microvascular endothelial cells along the arterio-venous axis (Aim 1). Three candidate induced brain microvascular endothelial cells (iBMECs) for each zone will be generated using lentiviral transduction. The top candidate for each zone will then be selected from analysis of gene and protein expression profiles (Aim 2). We will then use the three iBMECs to demonstrate zonation-specific cerebrovascular phenotype in zonation-specific models (arteriole, capillary, venule) (Aim 3). Next, we will assess the influence of young and old serum on cerebrovascular phenotype in the zonation-specific models (Aim 4). Finally, we will use the same approach to create an aged cerebrovascular model in one zone. We will create iBMECs that match the transcription factor profile of human brain microvascular endothelial cells in the aged cerebrovasculature, and then assess the role of microenvironmental cues and young/old serum on cerebrovascular phenotype. This project is a collaboration between the Searson group (JHU) with expertise in tissue-engineered microvascular models, and the Heiman group (MIT) with expertise in genomics and molecular mechanisms of neurodegenerative disease. This project builds upon key recent accomplishments from the two labs: (1) the creation of a library of zonation-specific transcription factor profiles for brain microvascular endothelial cells from the human brain, (2) identification of key transcription factors to enable reverse engineering of zonation- specific human brain microvascular endothelial cells, and (3) tissue-engineered platforms for zonation-specific cerebrovascular models.

项目概要 脑血管功能障碍正在成为许多脑部疾病的常见病理，包括 神经退行性疾病、脑血管疾病以及衰老。因此，了解角色 脑血管功能障碍在疾病进展和衰老中的作用将是长期维持大脑功能的关键 健康。随着组织工程和干细胞技术的发展，体外脑血管模型可以 在理解脑血管功能障碍在疾病进展中的作用方面发挥着重要作用 老化。 下一代脑血管模型应考虑三个关键因素：（1） 沿动静脉轴的脑微血管内皮细胞的表型，（内在因素），（2） 沿动静脉轴的微环境线索的差异（外在因素），以及（3） 衰老过程中的分区特异性脑血管表型以及对衰老血清的反应。因此， 该项目的总体目标是利用分区和年龄特定的内在和外在线索来进行逆向工程 人类脑血管模型，并利用这些模型来了解脑血管表型 老化。我们将首先进行汇总遗传筛选，以确定转录因子组合 能够驱动源细胞朝向人脑微血管内皮细胞的基因表达谱 沿动静脉轴（目标 1）。三种候选诱导脑微血管内皮细胞（iBMEC） 每个区域将使用慢病毒转导生成。每个区域的最佳候选人将是 从基因和蛋白质表达谱分析中选择（目标 2）。然后我们将使用三个 iBMEC 在分区特异性模型（小动脉、毛细血管、 小静脉）（目标 3）。接下来，我们将评估年轻和老年血清对脑血管表型的影响 特定区域模型（目标 4）。最后，我们将使用相同的方法来创建一个老化的 一个区域的脑血管模型。我们将创建与人类转录因子谱相匹配的 iBMEC 脑微血管内皮细胞在老化的脑血管中的作用，然后评估 微环境线索和年轻/年老血清对脑血管表型的影响。 该项目是具有组织工程专业知识的 Season 集团 (JHU) 之间的合作 微血管模型，以及在基因组学和分子机制方面拥有专业知识的 Heiman 小组（麻省理工学院） 神经退行性疾病。该项目建立在两个实验室最近取得的主要成就的基础上：(1) 创建脑微血管内皮细胞分区特异性转录因子谱库 来自人脑，（2）识别关键转录因子以实现分区的逆向工程- 特定的人脑微血管内皮细胞，以及（3）针对分区特异性的组织工程平台 脑血管模型。