Effects of Angiopoietins on Shock-Induced Acute Lung Injury

项目总结（见说明）： 内皮细胞（EC）屏障功能的丧失在间接急性肺损伤（ALI）的发生发展中起重要作用。我们已经表明，在一种新的小鼠模型的出血（休克），随后脓毒症的挑战所造成的盲肠结扎和穿孔（CLP），中性粒细胞（PMN）与居民肺细胞的相互作用是中央这种病理。在ALI中，未解决的炎症引发了EC屏障完整性丧失和肺功能受损的病理过程。在生理条件下，EC生长因子血管生成素（Angiopoietin，Ang）-1和2通过与EC上表达的t3 T：腺苷激酶受体Tie 2竞争性相互作用来维持血管稳态。Ang-1/Tie 2结合已显示稳定血管并刺激下游促存活/抗炎信号传导，相反，从活化的EC的储存颗粒释放的Ang-2使血管不稳定。最近的研究结果报道，血浆Ang-2水平在发生ALI的患者中显著升高。我们在休克/CLP模型中发现了肺和血浆中类似的升高，并且我们发现休克前PMNs的消耗消除了Ang-2的升高。我们提出以下中心假设：Ang-2导致休克/CLP引起的ALI中肺EC屏障功能丧失，由EC与休克致敏的PMNs相互作用引发。我们提出了以下具体目标：目标1将确定Ang-1：Ang-2和Ang-2表达和再合成的变化动力学以及其与炎症指数变化的关系。我们将使用Ang-2（si）RNA抑制肺组织表达、Ang-2蛋白特异性抑制和Ang-1竞争性抑制Ang-2/Tie 2结合来评估休克引发中Ang-2释放对ALI发展的贡献。目的2将确定Ang-2改变肺EC表型/活化的机制，以响应来自ALI/ARDS小鼠的血浆。目的3将确定Ang-2介导与休克致敏的PMN共培养后培养的小鼠EC表型/活化变化的机制。本研究将为PMN相关的Ang-2介导的ALI机制提供新的见解，并将阐明具有治疗干预潜力的途径。

PROJECT SUMMARY (See instructions): Loss of endothelial cell (EC) barrier function is important in the development of indirect acute lung injury (ALI). We have shown, in a novel murine model of hemorrhage (shock) with a subsequent septic challenge caused by cecal ligation and performation (CLP), that neutrophil (PMN) interactions with resident pulmonary cells are central to this pathology. In ALI, unresolved inflammation elicits a pathological process with loss of EC barrier integrity and impaired lung function. EC growth factors, Angiopoietin (Ang)-i and 2, under physiological conditions, maintain vascular homeostasis through competitive interactions wdth the t3T:osine kinase receptor, Tie2, expressed on ECs. Ang-1/Tie2 binding has been shown to stabilize vessels and stimulate down stream pro-survival/anti-inflammatory signaling, in contrast, Ang-2, released from storage granules of activated ECs, destabilizes vessels. Recent findings report that plasma Ang-2 levels are significantly elevated in patients that develop ALI. We find similar elevation in the lungs and plasma in our shock/CLP model, and we have found that depletion of PMNs prior to shock abrogates Ang-2 elevation. We propose the following central hypothesis: Ang-2 causes loss of pulmonary EC barrier function in ALI due to shock/CLP, initiated by EC interaction with shock-primed PMNs. We propose the following specific aims: Aim 1 will determine the kinetics of change in Ang-1:Ang-2 and Ang-2 expression and re-synthesis as well as its relationship to changes in indices of inflammation. We will use Ang-2 (si)RNA to suppress lung tissue expression, Ang-2 protein specific inhibition, and Ang-1 competitive inhibition of Ang-2/Tie2 binding to assess the contribution of Ang-2 release in shock priming for the development of ALI. Aim 2 will determine mechanisms by which Ang-2 changes pulmonary EC phenotype/activation in response to plasma from mice with ALI/ARDS. Aim 3 will determine mechanisms by which Ang-2 mediates changes in EC phenotype /activation in cultured mouse ECs following co-culture with shock-primed PMNs. The studies in this proposal will provide novel insights into the mechanisms of PMN associated, Ang-2 mediated ALI and will elucidate pathways that hold potential for therapeutic intervention.