Vascular mural cells in the development of the blood brain barrier

血脑屏障发育中的血管壁细胞

• 批准号: 10687558
• 负责人: Elizabeth Erin Crouch
• 金额: $ 138.89万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10687558
• 关键词: Adult; Age; Astrocytes; Atlases; Blood - brain barrier anatomy; Blood Cells; Blood Vessels; Blood flow; Brain; Cell physiology; Cells; Cerebrovascular system; Consumption; Data; Detection; Development; Disease; Effector Cell; Elements; Embryo; Endothelial Cells; Endothelium; Fibroblasts; Fluorescence-Activated Cell Sorting; Future; Genetic; Goals; Heterogeneity; Human; Knowledge; Lead; Microglia; Mitotic; Molecular; Mus; Neonatal; Nervous System Physiology; Neurons; Pathologic; Patients; Pericytes; Physiological; Play; Proteins; RNA; Reporting; Role; Second Pregnancy Trimester; Signal Transduction; Smooth Muscle Myocytes; Time; Tissues; brain cell; experimental study; neurovascular; neurovascular unit; prenatal; progenitor; response; single-cell RNA sequencing; spatiotemporal; stem cells; transcriptomics; treatment strategy

PROJECT ABSTRACT/SUMMARY Brain blood vessel cells play key roles in both physiological and pathological states. Endothelial and mural cells compose the main structural and functional elements of the vasculature. Situated between the endothelial cell lumen and the surrounding astrocytes, microglia, and neurons, mural cells serve as a critical signaling hub of the neurovascular unit. Mural cells are required for the development of the blood brain barrier and regulate blood flow in response to neuronal activity. However, most studies of brain blood vessels use adult mice and therefore little is known about the developing vasculature or human brain blood vessels at any age. These knowledge gaps impact patients. During development, both genetic and sporadic disorders of the vasculature can have a severe impact on long-term neurological function, and we currently have no treatments. To molecularly define the stages and subtypes of mural cells in the developing brain, we have developed an integrated approach to enrich for vascular mural cells, profile them, and functionally characterize their effects. In the prenatal human brain, Fluorescence Activated Cell Sorting (FACS) followed by single cell RNA sequencing (scRNAseq) identified four different types of mural cells during the second trimester: mitotic, classic pericyte, fibroblast, and smooth muscle cells. Trajectory analysis of our transcriptomic data showed smooth muscle cells as potential progenitors that give rise to classic pericytes and fibroblasts. The four mural cell subtypes had recently been reported in the adult human brain, and the presence of their full repertoire at such a young age suggests a highly dynamic and time-consuming maturation process for these cells. In the embryonic mouse brain, previous studies had not been able to define subtypes of mural cells. Our application of FACS followed by scRNAseq at embryonic day 13.5 uncovered the same subtypes as the prenatal human brain. In sum, the above experiments lead me to hypothesize that smooth muscle cells are evolutionarily conserved mural stem cells in the developing brain which give rise to classic pericytes, the differentiated effector cells of the blood brain barrier. My short-term goals for the proposed DP2 are the following: 1. Construct a spatiotemporal atlas of brain mural cells during development using scRNAseq and RNA and protein detection in tissue. 2. Perform lineage tracing with mouse and human cells to interrogate stage-specific roles in blood brain barrier development. As a neuroscientist, vascular biologist, and neonatologist, I am uniquely suited to pioneer our understanding of brain vascular development and the application of this knowledge towards future therapy.

项目摘要/总结 脑血管细胞在生理和病理状态下都发挥着关键作用。 内皮细胞和壁细胞构成了人体的主要结构和功能元素 脉管系统。位于内皮细胞腔和周围星形胶质细胞之间， 小胶质细胞和神经元、壁细胞是神经血管的关键信号中枢 单元。壁细胞是血脑屏障发育和调节血液所必需的 响应神经元活动的流动。然而，大多数脑血管研究都使用成年小鼠 因此，人们对发育中的脉管系统或人脑血管知之甚少 任何年龄。这些知识差距会影响患者。在发育过程中，遗传性和散发性 脉管系统疾病会对长期神经功能产生严重影响，我们 目前没有治疗方法。 为了从分子上定义发育中大脑中壁细胞的阶段和亚型，我们 开发了一种综合方法来富集血管壁细胞，对其进行分析，并 从功能上表征它们的效果。在产前人类大脑中，荧光激活细胞 分选 (FACS) 随后进行单细胞 RNA 测序 (scRNAseq) 鉴定出四种不同的类型 妊娠中期壁细胞的组成：有丝分裂、经典周细胞、成纤维细胞和平滑肌 细胞。我们的转录组数据的轨迹分析显示平滑肌细胞具有潜力 产生经典周细胞和成纤维细胞的祖细胞。四种壁细胞亚型 最近在成人大脑中被报道，并且它们的全部功能都存在于这样的情况下 年轻时表明这些细胞的成熟过程高度动态且耗时。 在胚胎小鼠大脑中，之前的研究未能定义壁画的亚型 细胞。我们在胚胎第 13.5 天应用 FACS 和 scRNAseq 发现了相同的结果 亚型如产前人类大脑。总之，上述实验让我假设 平滑肌细胞是发育中进化上保守的壁干细胞 产生经典周细胞的大脑，周细胞是血脑的分化效应细胞 障碍。我对拟议的 DP2 的短期目标如下： 1. 构建一个 使用 scRNAseq 和 RNA 绘制发育过程中脑壁细胞的时空图谱 组织中的蛋白质检测。 2. 用小鼠和人类细胞进行谱系追踪来询问 血脑屏障发育中的阶段特定作用。作为神经科学家、血管生物学家， 作为新生儿科医生，我非常适合开拓我们对脑血管的理解 这些知识的发展和应用在未来的治疗中。