AQP1调控巨噬细胞炎症小体NLRP3活化在急性肺损伤中的作用及机制研究

Clinical events, such as lung transplantation, trauma, shock and pulmonary embolism, are associated with Lung Ischemia Reperfusion Injury (LIRI) and likely contribute to the high mortality rates in these patients. Lung ischemia-reperfusion injury resolution, an active process that requires a series of integrated steps including restoration of endothelial integrity, transitions in the balance of pro- and anti-inflammatory cytokines, as well as clearance of neutrophils from sites of inflammation, is essential after ischemia-reperfusion injury. The water channel Aquaporin-1 (AQP1) facilitates water transport in lung injury and pulmonary edema. The applicant have investigated the important role of AQP1 in the resolution of LIRI via a mechanism involving increased stability of hypoxia-inducible factors 2α, which had been published by Am J Respir Cell Mol Biol, 2016. Our preliminary study found that AQP1 expression decreased in LIRI at early points, and AQP1-/- mice exhibited a significantly decreased survival when compared to that of wild type mice. NLRP3 inflammasome downstream factors IL-1β and IL-18 secretion were also increased in AQP1-/- mice. The project intends to investigate the role of AQP1 in LIRI inflammation and the regulation of NLRP3 inflammasome activation using wild type and AQP1-/- mice LIRI model. Besides, primary alveolar macrophages from wild type and AQP1-/- mice bronchoalveolar lavage fluid were stimulated under hypoxia reoxygenation, and β-catenin phosphorylation, NF-κB transcription activity and NLRP3 inflammasome activation levels were detected to elucidate the potential mechanism of AQP1. This study will further reveal the mechanisms of inflammatory regulation in LIRI, and provide some theoretical and experimental basis for clinical treatment of LIRI.

肺缺血再灌注损伤（LIRI）在肺移植、创伤、休克、肺栓塞等临床事件中发生率及死亡率极高，在肺损伤中水通道蛋白1（AQP1）参与肺水转运及液体管理。项目申请人发现AQP1通过HIF-2α促进血管生成参与LIRI后期修复(Am J Respir Cell Mol Biol, 2016)。AQP1在LIRI早期表达下调，AQP1敲除小鼠LIRI早期死亡率较野生小鼠高，NLRP3炎症小体效应分子IL-1β和IL-18分泌增高。本项目在前期工作基础上应用AQP1敲除和野生小鼠LIRI模型研究AQP1对LIRI炎症反应及NLRP3炎症小体的活化调节作用；应用巨噬细胞缺氧复氧模型观察β-catenin磷酸化、NF-κB转录活性及NLRP3炎症小体活化水平，阐明AQP1经由β-catenin调控NLRP3炎症小体活化的机制。本研究可丰富AQP1在LIRI过程中的作用，为寻求LIRI防治新途径提供实验依据。

肺损伤表现为微血管通透性增加、肺水肿、肺血管阻力增加及肺动脉高压。探索其发生机制并及早干预非常重要。我们关注介导跨膜水转运的水通道蛋白1在肺损伤发生发展进程中发挥的作用。在本项目资助下，项目负责人围绕介导跨膜水转运的水通道蛋白1在肺损伤发生发展进程中发挥的作用开展研究工作并取得一系列研究成果。在SCI发表相关文章2篇、核心期刊发表相关文章4篇，待发表SCI文章3篇；依托本项目培养研究生3名；项目研究成果参加国内外学术交流3次。.肺损伤早期刺激巨噬细胞活化并释放一系列促炎因子作用于中性粒细胞、内皮细胞等效应细胞导致炎症级联反应，内皮细胞损伤及微血管通透性增加。我们首次发现肺损伤早期阶段，水通道蛋白1（AQP1）通过NLRP3炎症小体调控巨噬细胞的活化，参与急性肺损伤的早期炎症反应；肺损伤晚期包括内皮细胞完整性修复，炎性细胞因子和抗炎细胞因子重新获得平衡，以及炎症细胞从损伤局部清除等修复过程。我们首次发现肺损伤修复阶段，水通道蛋白（AQP1）通过调节缺氧诱导因子（HIF-2α）稳定性参与急性肺损伤修复过程。本项目全面研究了AQP1在急性肺损伤过程始终的重要作用，为急性肺损伤诊治奠定理论基础。