Intracellular S1P & signaling in lung endothelial cells

描述（由申请人提供）：鞘氨醇-1-磷酸（S1 P）是一种生物活性脂质介质，不仅作为G蛋白偶联的SIP 1-5受体（以前称为内皮分化基因受体）的细胞外配体，而且作为细胞内第二信使参与Ca 2+动员、增殖和细胞凋亡抑制。 早先，我们和其他人已经证明血小板衍生的和外源性添加的S1 P增强内皮屏障功能和趋化性。我们的初步实验表明，人肺内皮细胞（EC）迅速转换外源性和血浆来源的S1 P细胞内的S1 P，但是，目前还不清楚细胞内S1 P信号的分子机制，调节EC的关键功能。最近，几个脂质磷酸磷酸酶（LPPs）和两个鞘氨醇激酶（Sphks）已被克隆和哺乳动物细胞中的特点，可以调节细胞外和细胞内的S1 P水平之间的动态平衡。该建议将解决LPPs和Sphks在调节内皮细胞钙稳态，增殖和细胞骨架重组中的作用。据推测，“LPPs和Sphks调节人肺EC中S1 P的细胞内产生和信号传导。具体目标1将表征LPP及其在调节内皮细胞内S1 P产生中的作用。特异性目的2将确定Sphks 1和2在催化由LPP作用于外源性S1 P而衍生的鞘氨醇磷酸化中的作用和调节。具体目标3将解决细胞内S1 P的细胞靶点和功能。总之，这些实验将揭示LPPs和Sphks调节循环血浆中细胞内S1 P产生的新机制，并确定参与肺内皮细胞中钙稳态、分泌和粘附分子表达、趋化性和伤口愈合的S1 P细胞内靶点。更好地了解LPPs和Sphks在介导循环中S1 P细胞内形成中的调节分子机制，将为S1 P在正常和病理条件下（如动脉粥样硬化和哮喘）在肺内皮细胞中作为细胞内第二信使的可能生理作用提供新的见解。

DESCRIPTION (provided by applicant): Sphingosine-1-phosphate (S1P), a bioactive lipid mediator, not only acts as an extra-cellular ligand for the G-protein coupled SIP 1-5 receptors (formerly known as endothelial differentiation gene receptors), but also functions as an intra-cellular second messenger involved in Ca2+ mobilization, proliferation and suppression of apoptosis. Earlier, we and others have demonstrated that platelet derived and exogenously added S1P enhance endothelial barrier function and chemotaxis. Our preliminary experiments suggest that human lung endothelial cells (ECs) rapidly convert exogenous and plasma-derived S1P to intra-cellular S1P; however, it is unclear the molecular mechanisms of intra-cellular S1P signaling that regulate key EC functions. Recently, several lipid phosphate phosphatases (LPPs) and two sphingosine kinases (Sphks) have been cloned and characterized in mammalian cells which may regulate the dynamic balance between extra- and intra-cellular S1P levels. This proposal will address the role of LPPs and Sphks in regulating endothelial calcium homeostasis, proliferation and cytoskeletal reorganization. It is postulated that "LPPs and Sphks regulate intra-cellular generation and signaling of S1P in human lung ECs. Specific Aim 1 will characterize the LPPs and their role in regulating intra-cellular S1P production in the endothelium. Specific Aim 2 will determine the role and regulation of Sphks 1 and 2 in catalyzing the phosphorylation of sphingosine derived by the action of the LPPs on exogenous S1P. Specific Aim 3 will address the cellular targets and functions of intra-cellular S1P. Taken together, these experiments will uncover novel mechanisms by which LPPs and Sphks regulate production of intra-cellular S1P from circulating plasma and also identify intra-cellular targets of S1P involved in calcium homeostasis, secretion, and expression of adhesion molecules, chemotaxis, and wound healing in lung endothelial cells. A better understanding of molecular mechanisms of regulation of LPPs and Sphks in mediating intra-cellular formation of S1P from circulation will provide new insights into possible physiological role of S1P as an intra-cellular second messenger in lung endothelial cells under normal and pathological conditions such as atherosclerosis and asthma.