Microenvironmental control of the Wnt pathway for the establishment of barrier heterogeneity in the CNS vasculature

血脑屏障(BBB)是由内皮细胞、周细胞、星形胶质细胞、神经元和小胶质细胞组成的神经血管单位形成的，对神经功能至关重要。我们已经证明，在脑血管生成过程中，Wnt/β-catenin信号通路在血管内皮细胞中是活跃的，通过调节屏障相关基因促进血脑屏障的发展。最近，我们提供了证据表明，经典的Wnt信号的内皮激活通过调节Pdgfb使胶质瘤血管正常化，从而减少瘤周水肿。自最近以来，其他几个途径，如声波刺猬，维甲酸已被证明参与诱导，成熟和维持内皮细胞的屏障属性。同时，很明显，内皮屏障的特征在不同的大脑区域并不相同。最明显的例子是脑室周围器官(CVO)，由于它们的神经分泌和/或神经感觉功能，缺乏内皮BBB。我们收集的初步数据支持这样的解释，即Wnt/β-catenin途径的活性在脑室周围器官的血管中特异性地缺失。这表明在CVO中，在胚胎发育期间建立了特定的血管分化和重塑的微环境，阻止了Wnt途径的激活。内皮β-连环蛋白信号的主导激活导致屏障的上调和通透性相关基因claudin-5和MECA-32的下调。这一观察结果表明，CVO中的血管能够分化为屏障内皮细胞。尽管一些研究描述和讨论了中枢神经系统的不同区域表现出不同的血脑屏障特性，但对其发展以及介导不同内皮屏障表型形成的分子线索知之甚少。基于我们已发表和未发表的结果，我们假设中枢神经系统中的血管异质性是通过微环境对Wnt/β-catenin信号的影响而实现的，这在CVO中最为明显。为了验证这一假说，我们的目标是：1.详细描述CVO血管的发育，屏障和通透性标志物的表达，以及Wnt通路的组成部分及其与其他通路的相互作用。利用转基因的三苯氧胺可诱导的动物模型，研究β-连环素在体内内皮特异性功能增强的后果。这些方法将有助于理解屏障异质性的基本原理，以及内皮屏障特性在调节中枢神经系统水稳态和免疫反应中的功能参与。

The blood-brain barrier (BBB) is formed by the neurovascular unit, comprising endothelial cells, pericytes, astrocytes, neurons and microglia, and is crucial for neural function. We have shown that Wnt/beta-catenin signalling - canonical pathway - is active in endothelial cells during brain angiogenesis, contributing to the development of the BBB by regulating barrier-associated genes. More recently, we have provided evidence that endothelial activation of canonical Wnt signalling normalizes glioma vessels by regulating Pdgfb, leading to reduced peritumoral oedema. Since recently, several other pathways like sonic hedgehog, retinoic acid have been shown be involved in the induction, maturation and maintenance of barrier properties in endothelial cells. In parallel it became evident that endothelial barrier characteristics are not identical in different brain regions. The most striking examples are the circumventricular organs (CVOs) that, due to their neurosecretory and/or neurosensory function, lack an endothelial BBB.We have collected preliminary data that support the interpretation that Wnt/beta-catenin pathway activity is specifically absent in vessels of the circumventricular organs. This suggests that in the CVOs a specific microenvironment of vascular differentiation and remodelling becomes established during embryonic development, prohibiting Wnt pathway activation.Dominant activation of endothelial beta-catenin signalling led to up-regulation of the barrier- and down-regulation of the permeability-related genes claudin-5 and MECA-32, respectively. This observation suggests that vessels in the CVOs are capable of differentiating into barrier endothelia. Although several studies describe and discuss that different areas of the CNS exhibit different BBB properties, nothing is known about the development and about the molecular cues that mediate the formation of differential endothelial barrier phenotypes. Based on our published and unpublished results, we hypothesize that vascular heterogeneity in the CNS is achieved by micro-environmental impact on Wnt/beta-catenin signalling, most evident in the CVOs. To test this hypothesis, we aim to: 1. characterize in detail the development of the CVO vessels, the expression of barrier and permeability markers, as well as Wnt pathway components and their interaction with other pathways.2. investigate the consequences of endothelial-specific gain-of-function of beta-catenin in vivo, making use of transgenic, tamoxifen-inducible animal models3. characterize the physiological and pathological consequences of beta-catenin activation, focussing on water homeostasis and on neuroinflammatory conditions.These approaches will help to understand basic principles of barrier heterogeneity, as well as functional involvement of endothelial barrier properties in the regulation of water homeostasis and immune responses in the CNS.