Astrocyte-specific molecular cues guiding retinal angiogenesis

星形胶质细胞特异性分子线索指导视网膜血管生成

基本信息

批准号：
10676468
负责人：
Juan Valdez-Lopez
金额：
$ 7.43万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-07-01 至 2026-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10676468
关键词：
Adopted Alleles Angiogenic Factor Astrocytes Autocrine Communication Automobile Driving Axon Binding Bioinformatics Biological Assay Blindness Blood Vessels Blood capillaries Cell surface Cells Child Childhood Coculture Techniques Cues Data Databases Defect Development Disease Endothelial Cells Event Future Genes Human Hypoxia In Vitro Knowledge Mediating Microglia Molecular Molecular Target Mus Nerve Fibers Nervous System Neurophysiology - biologic function Optic Nerve Outcome Pathogenicity Pathology Pathway interactions Pattern Peptides Porifera Positioning Attribute Pregnancy Process Production Property RAMP2 Radial Regulation Retina Retinal Ganglion Cells Retinopathy of Prematurity Risk Signal Transduction Source Testing VEGFA gene Vascularization Work adrenomedullin adrenomedullin receptor angiogenesis calcitonin receptor-like receptor cell growth cell type developmental disease ganglion cell in vivo insight nervous system development new therapeutic target novel prevent receptor receptor-activity-modifying protein recruit retinal angiogenesis retinal nerve fiber layer selective expression single-cell RNA sequencing

项目摘要

ABSTRACT Angiogenesis is a central event in nervous system development that requires further elucidation within the context of the retina. In normal development, retinal angiogenesis initiates from the optic nerve and proceeds along the innermost layer of retina consisting of retinal nerve fibers and the developing astrocytic network. Astrocytic development occurs prior to angiogenesis and is critical for guiding the developing vasculature. Specialized endothelial cells called tip cells crawl along the astrocytes as the angiogenic wavefront advances, so that the vasculature matches the pattern of astrocyte arbors. It is well understood that astrocytes secrete VEGF-A in hypoxic zones to initiate angiogenesis, but the signals that advance the wavefront, and guide tip cells along the astrocyte template, are unknown. The objective of this proposed study is to identify novel astrocyte- derived molecular cues that guide retinal vascularization. The central hypothesis is that astrocytes provide molecular cues that promote angiogenesis by 1) enhancing tip cell production; and 2) guiding tip cell growth so that vasculature adopts the pattern of the astrocyte network. The rationale for this work is to provide a deeper understanding of glial-mediated regulation of angiogenesis that may be applicable to the entire nervous system while also providing candidate molecules to target in retinal pathologies such as retinopathy of prematurity (ROP). The specific aims are: 1) Identify astrocyte-derived cues driving progression of the angiogenic wavefront. Adrenomedullin (ADM) is a secreted peptide that was previously found to mediate angiogenesis via its receptor, calcitonin receptor-like receptor (CLR) in conjunction with receptor activity modifying protein, RAMP2, which are expressed by endothelial cells. Preliminary single-cell RNA-seq data shows that the adrenomedullin gene is highly expressed by immature astrocytes. To test this, its angiogenic properties will be tested utilizing a functional assay that consists of analyzing vasculature wavefront progression in cultured mouse retinal explants that shall be co-cultured with ADM-expressing HEK293 cells. 2) Identify astrocyte-derived cues that pattern growing vessels. Genes selectively expressed by astrocytes are candidates to guide tip cell growth and angiogenesis. To identify such molecules, each cell type in the nerve fiber layer will be purified for scRNA-seq. Astrocyte-specific genes encoding cell-surface or secreted molecules will be identified bioinformatically. As in Aim 1, a functional assay using retinal explants will be used to test angiogenic capabilities of putative tip cell-guiding astrocyte-specific genes. Completion of this work will be significant because it will reveal novel components and drivers of angiogenesis expressed preferentially by astrocytes in the retina. Such information will elucidate basic mechanisms of retinal angiogenesis, while also pinpointing important molecular cues that may be involved in angiogenesis during pathogenic conditions such as ROP.

抽象的血管生成是神经系统发育中的一个中心事件，需要进一步阐明视网膜的背景。在正常发育中，视网膜血管生成从视神经开始并进行沿着视网膜神经纤维和发展中的星形胶质细胞网络的视网膜最终层。星形胶质细胞发育发生在血管生成之前，对于指导发展的脉管系统至关重要。随着血管生成波前的进展，称为尖端细胞的专门内皮细胞沿星形胶质细胞爬行，使脉管系统与星形胶质细胞轴的模式相匹配。众所周知，星形胶质细胞会分泌低氧区域中的VEGF-A启动血管生成，但是前进波前的信号和指导尖端细胞沿着星形胶质细胞模板是未知的。这项拟议的研究的目的是确定新型的星形胶质细胞引导的分子提示，指导视网膜血管化。中心假设是星形胶质细胞提供通过1）增强尖端细胞的产生来促进血管生成的分子提示； 2）引导尖端细胞生长如此该脉管系统采用星形胶质细胞网络的模式。这项工作的理由是提供更深的了解可能适用于整个神经系统的胶质介导的血管生成调节同时还可以在视网膜病理中提供候选分子，例如早产性视网膜病变（ROP）。具体目的是：1）确定驱动血管生成的星形胶质细胞衍生的提示波前。肾上腺素蛋白（ADM）是一种分泌的肽，以前被发现通过它的受体，降钙素受体样受体（CLR）与受体活性改变了蛋白质， RAMP2，由内皮细胞表达。初步的单细胞RNA-seq数据表明肾上腺肾上腺素基因由未成熟的星形胶质细胞高度表达。为了测试这一点，其血管生成特性将是利用功能测定法进行了测试，该功能测定包括分析培养的小鼠脉管波前的进展视网膜外植体应与表达ADM的HEK293细胞共培养。 2）确定星形胶质细胞衍生提示这种生长的容器。星形胶质细胞选择性表达的基因是指导尖端细胞的候选者生长和血管生成。为了识别此类分子，将纯化神经纤维层中的每种细胞类型 scrna-seq。将确定编码细胞表面或分泌分子的星形胶质细胞特异性基因生物信息。与AIM 1一样，使用视网膜外植体的功能测定将用于测试血管生成能力假定的尖端诱导星形胶质细胞特异性基因。这项工作的完成将是重要的，因为它将揭示了视网膜中星形胶质细胞优先表达血管生成的新型成分和驱动因素。这样的信息将阐明视网膜血管生成的基本机制，同时还指出了重要的分子在病原条件（例如ROP）期间可能参与血管生成的线索。