CNS angiogenesis and blood-brain barrier regulation by the Wnt inhibitor Apcdd1.

描述（由申请人提供）：血管生成与获得不同器官血管内皮细胞的特定特性的协调对于其正常功能是必不可少的。这一原理的一个重要例子是在脑内皮中发现的，即血脑屏障（BBB），它通过在内皮细胞之间形成高阻力紧密连接来限制分子在细胞旁扩散到大脑。尽管它对中枢神经系统（CNS）功能很重要，但调节血管生成和这一屏障发育的机制仍不清楚。我们之前已经发现，在中枢神经系统血管生成和屏障形成过程中，Wnt/¿-catenin信号在大脑中活跃，而在肝内皮中不活跃。此外，该途径对中枢神经系统血管生成和内皮细胞获得某些屏障特性至关重要。此外，Apcdd1是Wnt/¿-catenin信号的下游效应物，当内皮细胞获得血脑屏障特性时，Apcdd1在血管生成后的CNS内皮细胞中高度表达。Apcdd1在中枢神经系统中选择性表达，但在外周内皮细胞中不表达。Apccd1在出生后20天（P20）前存在于中枢神经系统内皮中，但当血管生成完成、血脑屏障完全形成时，Apccd1在成年中枢神经系统血管中消失。该蛋白定位于质膜和分泌通路内，当在接受Wnt信号传导的细胞中过度表达时，它以细胞自主的方式抑制Wnt/¿-catenin信号的激活。我们提出Apcdd1抑制中枢神经系统内皮中的Wnt/¿-catenin信号，使细胞成熟并获得血脑屏障特性。在本研究中，我们将探讨Apcdd1在中枢神经系统血管生成和血脑屏障形成中的作用。我们将首先检查Apcdd1是否与诱导内皮细胞屏障特性的Wnt配体（例如Wnt7a/7b）相互作用。然后，我们将测试Apcdd1在体外诱导内皮细胞血管生成和屏障特性的各个方面是否必要和充分。我们利用基因靶向方法培育了Apcdd1敲除小鼠，并在CNS内皮细胞中诱导过表达Apcdd1。这些小鼠将允许我们在体内测试Apcdd1对中枢神经系统血管生成和血脑屏障形成的需求和充足性。了解中枢神经系统血管生成和血脑屏障的发展将揭示中枢神经系统内皮细胞内紧密连接形成的机制，与中枢神经系统血管生成异常和血脑屏障破坏相关的病理中枢神经系统疾病的病因学，并有助于开发新的治疗方法来调节这些过程。

DESCRIPTION (provided by applicant): The coordination of angiogenesis with the acquisition of specific properties of endothelial cells that line the blood vessels in distinct organs is essenial for their proper function. An important example of this principle is found in the brain endothelium, namely the blood-brain barrier (BBB), that restricts paracellular diffusion of molecules into the brain by forming high resistance tight junctions between endothelial cells. Despite its importance for the central nervous system (CNS) function, the mechanisms that regulate angiogenesis and development of this barrier remain poorly characterized. We have previously identified that Wnt/¿-catenin signaling is active in brain but not liver endothelium during CNS angiogenesis and barrier formation. Moreover, this pathway is essential for CNS angiogenesis and acquisition of some barrier properties by endothelial cells. In addition, Apcdd1, a downstream effector of Wnt/¿-catenin signaling, is highly expressed in CNS endothelial cells after angiogenesis when endothelial cells acquire BBB properties. Apcdd1 is selectively expressed in CNS, but not, peripheral endothelial cells. Apccd1 is present in CNS endothelium until postnatal day 20 (P20), but it is extinguished in the adult CNS blood vessels when angiogenesis is complete and BBB is fully formed. The protein is localized within the plasma membrane and the secretory pathway and it inhibits the activation of Wnt/¿-catenin signaling in a cell-autonomous manner when overexpressed in cells that receive Wnt signaling. We propose that Apcdd1 inhibits Wnt/¿-catenin signaling in CNS endothelium to allow cells to mature and acquire BBB properties. In this proposal, we will investigate the role of Apcdd1 in CNS angiogenesis and BBB formation. We will first examine if Apcdd1 interacts with Wnt ligands (e.g. Wnt7a/7b) that induce barrier properties in endothelial cells. Then we will test if Apcdd1 is necessary and sufficient to induce various aspects of angiogenesis and barrier properties in endothelial cells in vitro. We have generated Apcdd1 knockout mice using gene targeting methods and mice that overexpress Apcdd1 in CNS endothelial cells in an inducible manner. These mice will allow us to test the requirement and sufficiency of Apcdd1 for CNS angiogenesis and BBB formation in vivo. Understanding the development of CNS angiogenesis and BBB will shed light on the mechanisms of tight junction formation within CNS endothelial cells, the etiology of pathological CNS conditions associated with abnormal CNS angiogenesis and BBB breakdown, and help to develop novel therapeutic approaches to regulate these processes.