干扰素调节因子5调控巨噬细胞胞葬作用介导肺炎症消退的机制研究

Acute lung injury（ALI） is characterized by overwhelming lung inflammatory response involving the recruitment and activation of polymorphonuclear neutrophils. The resolution of lung inflammation depends on the apoptosis and subsequent clearance of neutrophils. Phagocytosis of apoptotic cells by macrophages, efferocytosis, is recently considered as the key mechanism orchestrating resolution of lung inflammation. Our preliminary results demonstrated that efferocytosis by alveolar macrophages (AMs) was impaired in patients with ALI. Meanwhile, there is a high expression of interferon rgulatory factor 5 (IRF5) in AMs. We then found that IRF5 knockdown led to enhanced efferocytosis in vitro, indicating that IRF5 may be involved in the resolution of lung inflammation via regulating efferocytosis. However, the underlying mechanism remains elusive. In this project, therefore, we will explore the effect of IRF5 on macrophage efferocytosis which might play a crucial role in resolution of lung inflammation. We will measure the changes in efferocytosis and lung inflammation, through silencing of IRF5 by siRNA, to investigate whether IRF5 knockdown could promote resolution of lung inflammation via increasing macrophage efferocytosis. Next, using a anti-CD36 monoclonal antibody, we will determine whether CD36 serves the key molecule by which IRF5 regulates macrophage efferocytosis. At last, the underlying mechanism regarding how IRF5 is involved in regulation of CD36 expression would be determined after stimulation by tumor necrosis factor alpha and/or interleukin-10 and depletion of IRF5. ..Persistent lung inflammation during ALI will lead to acute respiratory distress syndrome, which will further lead to unresolved inflammation and impairment of lung function. Promoting the resolution of pulmonary inflammation will be of help to attenuate the damage and inhibit the exacerbation of lung injury. We believe that our research will illustrate and determine the regulatory mechanism of inflammation resolution and macrophage efferocytosis. Our project will provide new theoretical and experimental findings to further understand the underlying mechanism for resolution of lung inflammation in future.

以中性粒细胞浸润为特点的肺炎症反应介导着急性肺损伤（ALI）的发生，而炎症的消退依赖于所募集中性粒细胞的凋亡与清除。巨噬细胞参与的凋亡细胞清除,即胞葬作用，是肺炎症消退的关键。我们前期研究发现，巨噬细胞胞葬作用障碍的ALI患者伴有干扰素调节因子5(IRF5)表达增加，而下调IRF5可增强胞葬作用，提示IRF5可能通过调控巨噬细胞胞葬作用参与肺炎症消退过程。为此，本研究拟下调巨噬细胞IRF5表达，通过观察巨噬细胞胞葬作用及肺炎症变化，研究下调IRF5是否增强胞葬作用而促进肺炎症消退；通过阻断胞葬受体CD36，研究IRF5是否通过CD36调控胞葬作用；通过TNF-a和/或IL-10处理巨噬细胞并下调IRF5，研究IRF5影响 CD36表达的机制。.持续的肺炎症反应会导致ALI进一步恶化，而促进肺炎症消退将会阻断ALI进展。本课题为深刻理解肺炎症消退机制，并为开发特效药物提供新的途径和实验依据。

急性肺损伤(acute lung injury, ALI)是临床常见的危重疾病，年均发病率为79/10万例。随着ALI病情进展，约50%-75%患者发展为急性呼吸窘迫综合征(ARDS)，死亡率高达40%，且远期预后堪忧。ALI/ARDS临床治疗的困难凸显深入研究其发病机制与病理生理过程的迫切性和重要性。目前证据显示，以中性粒细胞浸润为特点的肺部炎症反应介导ALI/ARDS的发生，而炎症的消退依赖所募集中性粒细胞的凋亡与清除。巨噬细胞密切参与凋亡细胞的清除过程(胞葬作用)，是肺炎症消退的关键。IRF5是干扰素调节因子（interferon rgulatory factors,IRFs）家族的重要成员,表达于包括巨噬细胞在内的多种免疫细胞，新近研究提示IRF5可能是调节巨噬细胞功能的关键分子。.本项目以巨噬细胞的胞葬作用为着眼点，深入研究了IRF5对巨噬细胞胞葬作用的调控，及其在ALI炎症消退中的意义与相关机制。本研究中，我们首先在体外使用siRNA沉默巨噬细胞IRF5表达，发现下调巨噬细胞IRF5表达使其对凋亡细胞的胞葬作用显著增强；动物研究中，我们利用纳米载体双靶向siIRF5递送系统（FW-LP/siIRF5复合物）,于ALI造模两天后尾静脉注射，选择性下调小鼠巨噬细胞中IRF5基因表达，1天后检测肺部炎症变化，研究IRF5对ALI炎症消退的影响。结果发现，与对照组相比，FW-LP/siIRF5治疗后肺泡灌洗液蛋白含量以及肺组织病理损伤评分均有降低，提示IFR5可调控巨噬细胞的胞葬作用并在LPS诱发的ALI炎症消退中具有重要治疗价值。此外，体外研究进一步发现，沉默IRF5后巨噬细胞CD36mRNA及蛋白表达均显著升高，提示CD36可能为IRF5调控巨噬细胞胞葬作用的关键因子。综合上述结果和分析，本研究认为IFR5对巨噬细胞胞葬功能的调控及ALI的炎症消退具有重要作用，是有潜力的治疗靶标，值得进一步深入研究；此外，本研究中使用纳米载体FW-LP/siIRF5复合体促进ALI炎症消退，也为开发特效药物提供了新的途径和实验依据。