CXCR4 signaling in lung epithelial repair

描述（由申请人提供）：社区获得性肺炎（CAP）是急性呼吸窘迫综合征（ARDS）的最常见原因，ARDS是一种严重的急性肺损伤，是进入重症监护室的最常见原因之一。几乎没有治疗选择，死亡率很高。辅助供氧和机械通气的支持治疗是必不可少的，但呼吸机可能会引起额外的损伤，称为呼吸机诱导的肺损伤（VILI）。上皮修复对疾病至关重要 解决和生存，但我们对修复的潜在机制以及机械拉伸如何影响这些机制的知识有限。该项目的长期目标是增加我们对上皮修复机制的理解，以及肺上皮细胞过度扩张如何导致VILI和适应不良的修复机制。我们以前确定了自分泌作用的趋化因子CXCL 12在肺泡上皮修复涉及其受体CXCR 4。我们现在有初步的数据显示，在支气管肺泡灌洗液中有高水平CXCL 12的CAP诱导的ARDS患者的机械通气持续时间较短，死亡率较低。根据额外的初步数据，我们提出CXCR 4与一系列信号分子相互作用，包括粘着斑激酶（FAK）和调节上皮修复的凋亡信号调节激酶-1（ASK 1）。本申请的中心假设是CXCL 12促进上皮修复，但机械拉伸导致抑制细胞扩散、迁移和修复的CXCR 4-FAK-ASK 1信号传导中断。我们将首先通过测量支气管肺泡灌洗液和血浆中的CXCL 12来检查CXCL 12是否是经历适应性修复的ARDS患者的生物标志物。此外，我们将使用来自ARDS患者的尸检样本来评估CXCR 4和磷酸化（活化）ASK 1的表达。在第二个目标中，我们将研究CXCR 4，FAK和ASK 1之间的相互作用，从LPS引起的肺损伤恢复过程中作为肺炎模型。我们将使用在肺上皮中CXCR 4条件性缺失的小鼠以及ASK 1敲除小鼠。我们还将研究这些信号分子在培养的肺泡上皮细胞在划痕模型的生化相互作用。在第三个目标中，我们将研究在肺炎（LPS）和机械通气联合模型中，高拉伸机械通气或培养细胞的周期性拉伸如何在修复过程中破坏这些信号通路。这些研究将阐明参与肺泡上皮修复的新信号通路以及机械拉伸如何破坏修复过程。

DESCRIPTION (provided by applicant): Community-acquired pneumonia (CAP) is the most common cause of acute respiratory distress syndrome (ARDS), a severe form of acute lung injury that is one of the most frequent causes of admission into the intensive care unit. Few therapeutic options are available, and mortality is high. Supportive therapy with supplemental oxygen and mechanical ventilation are essential, but additional injury can be caused by the ventilator, termed ventilator-induced lung injury (VILI). Epithelial repair is critical for disease resolution and survival, but we have limited knowledge of the underlying mechanisms of repair and how mechanical stretch impacts these mechanisms. The long term objective of this project is to increase our understanding of the mechanisms of epithelial repair and how overdistention of pulmonary epithelial cells contributes to VILI and maladaptive repair mechanisms. We previously identified an autocrine role for the chemokine CXCL12 in alveolar epithelial repair involving its receptor CXCR4. We now have preliminary data showing that patients with CAP-induced ARDS that had high levels of CXCL12 in their bronchoalveolar lavage fluid had shorter duration of mechanical ventilation and lower mortality. Based upon additional preliminary data, we propose that CXCR4 interacts with a complex of signaling molecules including focal adhesion kinase (FAK) and apoptosis signal-regulating kinase-1 (ASK1) that regulates epithelial repair. The central hypothesis of this application is that CXCL12 promotes epithelial repair, but mechanical stretch causes disruption of CXCR4-FAK-ASK1 signaling that inhibits cell spreading, migration, and repair. We will first examine whether CXCL12 is a biomarker for ARDS patients undergoing adaptive repair by measuring CXCL12 in banked samples of bronchoalveolar lavage fluid and plasma. In addition we will use autopsy samples from ARDS patients to evaluate expression of CXCR4 and phosphorylated (activated) ASK1. In the second aim we will investigate the interactions between CXCR4, FAK, and ASK1 during recovery from lung injury caused by LPS as a model of pneumonia. We will use mice with conditional deletion of CXCR4 in lung epithelial as well as ASK1 knockout mice. We will also examine the biochemical interactions of these signaling molecules in cultured alveolar epithelial cells in a scratch wound model. In the third aim we will investigate how high stretch mechanical ventilation or cyclic stretch of cultured cells disrupts these signaling pathways during repair in combined model of pneumonia (LPS) and mechanical ventilation. These studies will elucidate new signaling pathways involved in alveolar epithelial repair and how mechanical stretch disrupts the repair processes.