消退素D1对肺泡巨噬细胞表型转化的影响及机制研究

Acute respiratory distress syndrome (ARDS) consisting of acute hypoxemic respiratory failure, occur with a high incidence and mortality. Resolvins are unique structures derived from DHA. Resolvins were the mediators recognized to have dual anti-inflammatory and pro-resolution activities, which have been described as the endogenous “braking signal” for inflammation. We showed that the resolvin D1 concentration and resolvin receptor expression decreased in ARDS patients. We also found that resolvin D1 not only promoted neutrophils apoptosis and efferocytosis, but also increased M1 transferring into M2 macrophages. Base on these findings, our hypothesis is that resolvin D1 may mediate M1 and M2 transformation to promote inflammation resolution. Therefore, LPS induced lung injury was established. Real-time PCR, flow cytometry, laser confocal microscope and western blot were used to measure the dynamic of resolvin D1, resolvin receptor and macrophage phenotype changes, discuss the effect of resolvin D1 on M1 and M2 phenotype. Finally, to gain a better understanding of the mechanisms, we also investigated the effect of agonist, antagonist and siRNA on macrophages phenotype in ARDS. The main aim of this grant is to reveal a novel mechanism of pro-resolution and provide experimental data explore the prevention and treatment of ARDS.

炎症消退调控障碍导致炎症失控是急性呼吸窘迫综合征（ARDS）重要发病机制。消退素D1是新近发现机体重要内源性抗炎促消退介质,为炎症反应“刹车信号”。在前期证实消退素D1促进肺水清除，保护肺损伤基础上，结合我们新近发现消退素D1抑制M1型肺泡巨噬细胞，促进M2型肺泡巨噬细胞增多，增强巨噬细胞吞噬凋亡中性粒细胞等线索。我们推测：消退素D1可能通过调控肺泡巨噬细胞表型转化促进炎症消退。为此，拟采用内毒素性大鼠ARDS模型，应用流式细胞仪、激光共聚焦显微镜、实时定量PCR、western blot等技术,动态观察肺泡巨噬细胞表型及消退素D1浓度、受体变化规律；探讨消退素D1对巨噬细胞M1型和M2型影响及转化机制；比较消退素D1与地塞米松诱导巨噬细胞表型转化异同性。进一步应用消退素受体及转导信号通路抑制剂、激动剂和siRNA技术验证我们假说。旨在揭示ARDS发病新机制并为探寻防治新策略提供实验依据。

炎症消退调控障碍导致炎症失控是急性呼吸窘迫综合征（ARDS）重要发病机制。消退素D1是新近发现机体重要的内源性抗炎促消退介质，被称为炎症反应的“刹车信号”。 我们在前期研究证实消退素D1能促进肺水的清除，减轻肺损伤的基础上，结合新近发现的消退素D1能减少M1型肺泡巨噬细胞，促进M2型肺泡巨噬细胞增多，增强巨噬细胞吞噬凋亡中性粒细胞等线索推测：消退素D1可能通过调控肺泡巨噬细胞表型转化从而促进炎症消退。为验证这一假说，我们采用内毒素性大鼠ARDS模型，探究消退素D1对巨噬细胞M1型和M2型的影响及转化机制；比较消退素D1与地塞米松诱导巨噬细胞表型转化异同性；进一步应用消退素受体及转导信号通路抑制剂、激动剂和siRNA技术进行验证。我们的研究结果表明，消退素D1不仅受体依赖性促进中性粒细胞凋亡，而且促进巨噬细胞吞噬凋亡的中性粒细胞，这一过程也依赖于消退素受体，使用BOC-2抑制剂后消退素的治疗作用被抑制；消退素D1增强肺组织常驻巨噬细胞表达CD206,减少CD86的表达，进而促进炎症消退；消退素D1通过抑制单核细胞浸润到肺组织，从而促进炎症的消退。我们的结果揭示了消退素D1对中性粒细胞凋亡的影响及巨噬细胞表型变化的影响，为探寻ARDS的防治新策略提供了新的靶点与实验依据。