Sphingosine 1-phosphate receptors in vascular homeostasis and pathology

1-磷酸鞘氨醇（S1 P）是一种多功能脂质介质，通过S1 P受体（S1 PnR）发出信号。 调节血管系统的发育、稳态和表型变化。我们最近的研究 已经表明Gi-、Rac-和Akt-偶联的S1 P1 R在血管生成血管中被诱导，是肿瘤生长所需的。 血管生成，协调细胞-细胞粘附分子功能，促进血管成熟/稳定 并抑制血管渗透性。相反，我们发现激活Rho GT3的S1 P2 R， 肿瘤抑制磷酸酶PTEN的依赖性激活，抑制内皮细胞迁移，破坏 细胞-细胞连接，诱导血管通透性，炎症基因表达和病理 血管生成鉴于S1 P在血浆中丰富，S1 P的空间和时间调节表达 内皮细胞中的受体是给定血管床中S1 P生物反应的主要决定因素。 这项研究计划的长期目标是更好地理解S1 P信令的逻辑， 脉管系统由于第一代S1 P受体调节剂FTY 720目前正在进行临床试验， 多发性硬化症和移植排斥的临床试验（III期），这项研究计划的结果可能 提供可直接翻译到诊所的关键信息。 这一提议的中心假设是S1 P深刻地调节血管内皮细胞的表型。 通过S1 P1 R和S1 P2 R的时间和空间精确信号传导来调节内皮。 在第一个具体目标中，我们将测试S1 P1 R对生理性 血管生成S1 P1 R调节的信号通路促进周细胞募集和抑制 血管通透性和炎症将使用遗传小鼠模型和药理学方法进行研究。 工具.其次，我们将讨论内皮细胞中S1 P受体信号传导是如何被激活的。 考虑到配体在血浆中高度丰富的事实，其被减弱。我们将批判性地检验这一假设， S1 P1 R的选择性亚细胞靶向，即内体和质膜之间的再循环，或 基于泛素的靶向降解细胞器是调控和破译细胞周期的一个重要方面。 涉及的分子机制。第三，明确S1 P2 R信号通路在肿瘤病理过程中的意义。 血管生成我们将测试S1 PR 2与Rho/ROCK/PTEN和p38通路的偶联是否诱导血管生成， 渗透性和炎症基因表达。受体偶联的分子机制， 内皮连接完整性的调节和转录调节将得到解决。最后我们 提出S1 P1 R和S1 P2 R在血管稳态和发育中的协同信号转导。一 不内化的超形态S1 P1 R将被敲入小鼠基因组， 将表征血管生成、血管稳定性、血管渗透性和炎症。的能力 将通过改变S1 p2 r基因剂量来测试S1 P2 R调节S1 P1 R依赖性效应的能力。 S1 P1 R hypermorph的背景。 这些研究有望增强我们对S1 P在血管系统中功能的理解。这 这些信息可能有助于通过S1 P受体控制血管炎症和血管生成 调制器。叙事 这项拟议的研究将调查鞘氨醇1-磷酸受体1和2的作用， （S1 P1 R和S1 P2 R）对血管内皮细胞生物学和病理生理学的影响。概念 S1 P1 R和S1 P2 R介导相反的作用来调节生理和病理 将使用体外细胞培养系统详细检查血管生成，以研究 涉及的机制。视网膜中血管生成的小鼠模型将用于获得体内 相互关联最后，这两个受体的协调信号转导将被检查。一 我们将开发一种新的小鼠模型，使我们能够研究坐标信号传导 使用S1 P1 R的内化缺陷突变体。这些研究预计将产生关键的 深入了解S1 P受体的工作原理，这是重要的药物靶点，以控制 血管和免疫系统。

Sphingosine 1-phosphate (S1P), a multifunctional lipid mediator, signals via the S1P receptors (S1PnR) to regulate the development, homeostasis and phenotypic changes of the vascular system. Our recent studies have shown that the Gi-, Rac- and Akt-coupled S1P1R is induced in angiogenic vessels, is required for tumor angiogenesis, orchestrates cell-cell adhesion molecule function, promotes vascular maturation/stabilization and inhibits vascular permeability. In contrast, we found that S1P2R, which activates the Rho GTPase- dependent activation of the tumor suppressor phosphatase PTEN, inhibits endothelial cell migration, disrupts cell-cell junctions, induces vascular permeability, inflammatory gene expression and pathological angiogenesis. Given that S1P is abundant in plasma, spatially- and temporally-regulated expression of S1P receptors in endothelial cells is a major determinant of the biological response of S1P in a given vascular bed. The long-term goal of this research program is to better understand the logic of S1P signaling in the vasculature. Since the first generation S1P receptor modulator called FTY720 is currently undergoing clinical trials (phase III) for multiple sclerosis and transplant rejection, the findings from this research program may provide critical information that is directly translatable to the clinic. The central hypothesis of this proposal is that S1P profoundly regulates the phenotype of vascular endothelium by temporally- and spatially-precise signaling of S1P1R and S1P2R. In the first specific aim, we will test the hypothesis that S1P1R is essential for physiological angiogenesis. The ability of S1P1R-regulated signaling pathways to promote pericyte recruitment, and inhibit vascular permeability, and inflammation will be investigated using genetic mouse models and pharmacological tools. Secondly, we will address the question of how the S1P receptor signaling in the endothelium is attenuated given the fact that the ligand is highly abundant in plasma. We will critically test the hypothesis that selective subcellular targeting of S1P1R, namely, recycling between endosomes and plasma membrane or ubiquitin-based targeting to degradative organelle is an important aspect of regulation and decipher the molecular mechanisms involved. Thirdly, we will define the significance of S1P2R signaling in pathological angiogenesis. We will test if S1PR2 coupling to Rho/ROCK/PTEN and p38 pathways induces vascular permeability and inflammatory gene expression, respectively. Molecular mechanisms of receptor coupling, regulation of endothelial junctional integrity and transcriptional regulation will be addressed. And finally, we propose to define the coordinate signaling of S1P1R and S1P2R in vascular homeostasis and development. A hypermorphic S1P1R that does not internalize will be knocked into the mouse genome and the resultant tissue angiogenesis, vascular stabilization, vascular permeability and inflammation will be characterized. The ability of S1P2R to modulate S1P1R-dependent effects will be tested by varying the gene dosage of S1p2r in the context of the S1P1R hypermorph. These studies are anticipated to enhance our understanding of S1P function in the vasculature. This information may be useful in the control of vascular inflammation and angiogenesis by S1P receptor modulators. Narrative This proposed research will investigate the role of sphingosine 1-phosphate receptors 1 and 2 (S1P1R and S1P2R) on vascular endothelial cell biology and pathophysiology. The concept that S1P1R and S1P2R mediate opposing actions to regulate physiological and pathological angiogenesis will be examined in detail using in vitro cell culture systems to investigate the mechanisms involved. Mouse models of angiogenesis in the retina will be used to obtain in vivo correlates. Finally coordinate signal transduction of these two receptors will be examined. A novel mouse model that will allow us to examine the coordinate signaling will be developed using an internalization deficient mutant of S1P1R. These studies are expected to derive critical insights into the workings of S1P receptors, which are important drug targets to control the vascular and immune systems.