Molecular Genetic Dissection of Fenestrated Vascular Development in the Choroid Plexus

项目总结/摘要 脑血管疾病仍然是造成严重长期残疾的主要原因。许多这些疾病 至少部分起源于影响血管发育和成熟的遗传缺陷， 中枢神经系统（CNS）和脑膜。尽管我们对中枢神经系统的了解取得了实质性进展， 血管生成和屏障形成，我们对高渗透性或有孔微血管系统的了解 在脉络丛（CP）和脑室周围器官中形成的血管仍然有限。这个项目的总体目标是 我们的建议是通过了解遗传病的分子遗传机制来填补科学知识的这一空白。 有孔脑血管发育，特别关注CP。我们最近发现CP 斑马鱼的血管化需要独特的血管生成机制， 生长因子（Vegf）直向同源物，特别是Vegfab和Vegfc，起着功能冗余的作用。令人惊奇的是， 这种机制是CP血管化所特有的，因为邻近的无穿孔脑血管系统是 在VEGFab中不受影响;VEGFC双突变体。这些结果使我们制定了一个范式转变， 假设有窗孔CP和无窗孔脑血管发育需要不同的 Vegf/Vegfr代码和信令。鉴于Vegfs在不同的病理生理过程中的关键作用， 而且在任何脊椎动物物种中没有报道驱动CP血管形成的分子线索， 本研究的结果将对CP生理学和血管领域产生重大的积极影响， CP血管化之外的生物学。我们进一步的遗传学、药理学和表达数据提出了一种新的方法。 模型中，来自CP上皮细胞的旁分泌Vegfab和来自内皮细胞的自分泌Vegfc一起 通过PI 3 K和ERK信号通路控制CP血管化。以下两个具体目标是 来检验我们的假设目的1将检验有窗脑区和无窗脑区 血管发育需要不同的Vegf/Vegfr编码和信号传导。目标2将检验以下假设： 来自CP上皮细胞的旁分泌Vegfab和来自 内皮共同驱动有孔CP血管发育。我们将通过以下方式实现这些目标： 新的斑马鱼分子遗传工具，先进的成像技术和遗传镶嵌分析。的 拟议的工作是一个创新的分子遗传学研究在这一领域，因为它利用了各种优势， 斑马鱼模型克服了技术限制，并解决了以下基本过程：1） 有窗CP维管发育的细胞和分子基础; 2）发育机制 形成异质性脑血管网络的背后。从这个收集到的机械信息 这项工作将为CP血管发病机制中潜在的分子途径提供新的线索， 畸形，并且还有助于设计用于预防或治疗CNS血管疾病的治疗剂。

PROJECT SUMMARY/ABSTRACT Cerebrovascular diseases remain a leading cause of serious long-term disability. Many of these diseases originate, at least in part, from genetic defects that impact the development and maturation of blood vessels in the central nervous system (CNS) and meninges. Despite substantial progress in our understanding of CNS angiogenesis and barriergenesis, our knowledge of the highly permeable, or fenestrated, microvasculature formed in the choroid plexus (CP) and circumventricular organs remains limited. The overall goal of this proposal is to fill this gap in scientific knowledge by understanding the molecular genetic mechanisms of fenestrated brain vascular development, specifically focusing on the CP. We recently found that CP vascularization in zebrafish requires a unique angiogenic mechanism whereby zebrafish vascular endothelial growth factor (Vegf) orthologs, specifically Vegfab and Vegfc, play a functionally redundant role. Surprisingly, this mechanism is specific to CP vascularization, since the neighboring non-fenestrated brain vasculature is not affected in the vegfab;vegfc double mutants. These results led us to formulate a paradigm-shifting hypothesis that fenestrated CP and non-fenestrated brain vascular development requires distinct sets of the Vegf/Vegfr codes and signaling. Given the critical roles for Vegfs in diverse pathophysiological processes and also the fact that no molecular cue that drives CP vascularization has been reported in any vertebrate species, the results of this study will have a significant positive impact on the fields of CP physiology and vascular biology beyond CP vascularization. Our further genetic, pharmacological, and expression data propose a model in which paracrine Vegfab from CP epithelial cells and autocrine Vegfc from the endothelium together controls CP vascularization via PI3K and ERK signaling pathways. The following two Specific Aims are proposed to test our hypotheses. Aim 1 will test the hypothesis that fenestrated CP and non-fenestrated brain vascular development requires distinct Vegf/Vegfr codes and signaling. Aim 2 will test the hypothesis that the spatiotemporal coordination of paracrine Vegfab from CP epithelial cells and autocrine Vegfc from the endothelium together drives fenestrated CP vascular development. We will pursue these aims by combining novel zebrafish molecular genetic tools with advanced imaging technologies and genetic mosaic analyses. The proposed work is an innovative molecular genetics study in this field because it exploits various strengths of the zebrafish model to overcome technical limitations and address the following fundamental processes: 1) the cellular and molecular basis of fenestrated CP vascular development; and 2) the developmental mechanisms behind the formation of heterogeneous brain vascular networks. Mechanistic information gleaned from this work will shed new light on potential molecular pathways involved in the pathogenesis of CP vascular malformations and also help to design therapeutics for preventing or treating CNS vascular diseases.