S1P Deficiency Prolongs Endothelial Barrier Recovery After Fontan Operation

描述(申请人提供)：本提案的总体目标是通过对血管内皮细胞屏障功能的研究，解决Fontan手术后出现的一个普遍的临床问题，持续性胸腔积液。这一建议的基本假设是，接受Fontan手术的单心室生理学儿童经历了继发于体外循环(CPB)深刻炎症效应的血管通透性紊乱。因此，确定修复血管内皮细胞屏障的治疗靶点可能是改善Fontan手术后护理的关键。1-磷酸鞘氨醇(S1P)是一种磷脂，它与内皮细胞上的G蛋白偶联受体相互作用，介导内皮屏障功能的增强，从而降低血管通透性。S1P在调节体内血管通透性方面的关键作用已在动物模型中得到证实。例如，缺乏负责S1P生物合成的两种酶之一的小鼠，在脂多糖(LPS)或PAR-1激活诱导的炎性损伤时，表现出肺微血管通透性增加和更大程度的水肿形成。重要的是，在小鼠和狗身上静脉注射S1P已被证明通过减少肺血管渗漏和肺内分流而在急性肺损伤中起到保护作用。我们对Fontan患者血浆的初步分析显示，术后血浆高密度脂蛋白(HDL)水平下降，高密度脂蛋白是促进内皮屏障的鞘氨醇的主要载体，鞘氨醇-1-磷酸(S1P)。基于这些发现，我们假设，在体外循环后，儿童血浆高密度脂蛋白-S1P水平降低，并预示着持续性胸腔积液和较长的术后恢复时间。此外，我们假设，来自持续性胸腔积液的儿童的血液样本将显示出破坏屏障的倾向或缺乏促进屏障的能力的证据，这两种情况都可以通过外源性给予S1P来克服。为了解决这些相互关联的假设，有两个具体的目标：1)确定血浆S1P水平是否与Fontan手术后长期术后恢复(即持续性胸腔积液)的临床指标相关；2)确定Fontan手术患者血浆样本诱导的屏障完整性水平，并确定这是否与S1P水平和/或术后胸腔积液的严重程度相关。这些后续的研究还将确定外源性给予S1P是否可以增加样本在体外增强屏障的能力，达到使用术前样本所达到的水平。该应用的目的可能导致基于S1P的新策略，用于治疗Fontan患者和其他接受CPB的婴儿和儿童的血管通透性紊乱。

DESCRIPTION (provided by applicant): The overall goal of this proposal is to address a widespread clinical problem occurring after the Fontan operation, persistent pleural effusions, through the investigation of vascular endothelial barrier function. The foundational hypothesis of this proposal is that children with single ventricle physiology who undergo the Fontan operation experience derangements in vascular permeability secondary to the profound inflammatory effects of cardiopulmonary bypass (CPB). Therefore, identifying therapeutic targets that restore vascular endothelial barrier may be critical to improving post-operative care after the Fontan operation. Sphingosine 1-phosphate (S1P) is a phospholipid that interacts with G protein coupled receptors on endothelial cells and mediates enhancement of endothelial barrier function, thus reducing vascular permeability. The critical role of S1P in regulating vascular permeability in vivo has been illustrated in animal models. For example, mice deficient in one of the two enzymes responsible for S1P biosynthesis, sphingosine kinase-1, display increased pulmonary microvessel permeability and greater levels of edema formation in response to an inflammatory insult induced with lipopolysaccharide (LPS) or PAR-1 activation. Importantly, intravenous administration of S1P in mice and dogs has been shown to play a protective role in acute lung injury by reducing pulmonary vascular leakage and intrapulmonary shunting. Our preliminary analysis of plasma from Fontan patients has revealed a reduction in postoperative plasma levels of HDL, the principle carrier of the endothelial barrier- promoting sphingolipid, sphingosine-1-phosphate (S1P). Based on these findings we hypothesize that plasma HDL-S1P levels are lowered in children after exposure to CPB and are predictive of persistent pleural effusions and longer post-operative recovery time. Additionally, we hypothesize that blood samples from children with persistent pleural effusions will display evidence of either propensity to disrupt barrier or a lack of ability to promote barrier either of which can be overcome by exogenous administration of S1P. To address these inter-related hypotheses there are two specific aims: 1) to determine whether plasma levels of S1P correlate with clinical markers of prolonged postoperative recovery (i.e., persistent pleural effusions) after the Fontan operation, and 2) to determine the level of barrier integrity induced by plasma samples from Fontan operation patients and determine if this correlates with S1P levels and/or severity of post-operative pleural effusions. These later studies will also determine whether exogenous administration of S1P can increase the capacity of the samples to enhance barrier in vitro to levels achieved using preoperative samples. The aims of the application may lead to new S1P based strategies for treatment of derangements in vascular permeability in Fontan patients and other infants and children undergoing CPB.