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中血管内皮细胞特定特性的协调对于其正常功能至关重要。该原理的一个重要实例发现于脑内皮，即血脑屏障（BBB），其通过在内皮细胞之间形成高阻力紧密连接来限制分子向脑中的细胞旁扩散。尽管其对中枢神经系统（CNS）功能的重要性，但调节血管生成和这种屏障发展的机制仍然缺乏特征。我们以前已经确定，在CNS血管生成和屏障形成过程中，Wnt/β-连环蛋白信号在脑内皮中是活跃的，但在肝内皮中不是。此外，该途径对于CNS血管生成和内皮细胞获得某些屏障特性是必需的。此外，当内皮细胞获得BBB特性时，血管生成后，Apcdd 1（Wnt/β-catenin信号传导的下游效应子）在CNS内皮细胞中高度表达。Apcdd 1选择性表达于中枢神经系统，而不是外周血管内皮细胞。Apccd 1在出生后20天（P20）之前存在于CNS内皮中，但是当血管生成完成并且BBB完全形成时，Apccd 1在成人CNS血管中消失。该蛋白定位于质膜和分泌途径内，并且当在接受Wnt信号传导的细胞中过表达时，其以细胞自主方式抑制Wnt/β-连环蛋白信号传导的活化。我们认为Apcdd 1抑制中枢神经系统内皮细胞中的Wnt/â-连环蛋白信号传导，使细胞成熟并获得BBB特性。在这个提议中，我们将研究Apcdd 1在CNS血管生成和BBB形成中的作用。我们将首先检查Apcdd 1是否与诱导内皮细胞屏障特性的Wnt配体（例如Wnt 7a/7 b）相互作用。然后，我们将测试Apcdd 1是否是必要的和足够的诱导血管生成和屏障特性的各个方面在体外内皮细胞。我们已经产生了Apcdd 1基因敲除小鼠使用基因打靶方法和小鼠过表达Apcdd 1在中枢神经系统内皮细胞中的诱导方式。这些小鼠将使我们能够测试体内CNS血管生成和BBB形成对Apcdd 1的需求和充分性。了解CNS血管生成和BBB的发展将揭示CNS内皮细胞内紧密连接形成的机制，与异常CNS血管生成和BBB破坏相关的病理性CNS疾病的病因，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.