红霉素对香烟作用的沉默信息调节因子2相关酶sirtuin1调控的机制研究

The cigarette smoke-induced oxidative stress and inflammation are important pathogenesis of chronic obstructive pulmonary disease (COPD). Recent studies have shown that the expression of silent mating type information regulation 2 homolog 1 (SIRT1) in the lungs of COPD patients and smokers were down-regulated, which was associated with increasing of transcription factor activity and inflammatory mediators synthesis. These studies suggest that SIRT1 play an important role in inhibit oxidative stress and inflammation induced by cigarette smoke. Our previous studies have shown erythromycin can inhibit cigarette smoke-induced oxidative stress and inflammation. However, the mechanism is still unclear. Therefore, we designed this project. Human macrophages and rats will be treated with either cigarette smoke or erythromycin pretreatment. Reactive oxygen (ROS) will be assessed by using flow cytometer. Western-Blot and immune precipitation analysis will be used for SIRT1 and nuclear factor-КB (NF-КB) protein assays and the levels of NF-КB dependent proinflammatory cytokines the levels of was measured with by Luminex detection. This project will help us further explore the role of erythromycin, which how to inhibit the anti-inflammatory and antioxidant effects in cigarette smoke-caused oxidative stress and inflammatory microenvironment. Thus, these date will provide new ideas for the prevention and treatment of COPD.

吸烟导致的氧化应激和肺部炎症是慢性阻塞性肺疾病（COPD）的重要发病机制。最近研究表明，COPD 患者及吸烟人群肺部沉默信息调节因子2相关酶sirtuin1（SIRT1)表达下调，继而转录因子活性增加及炎症介质合成。为有效抑制氧化应激和肺部炎症，我们前期研究显示红霉素可抑制香烟烟雾诱导的氧化应激和炎症，但两者之间的相互联系及作用机制尚未明确。因此我们设计本课题，以香烟暴露的人巨噬细胞和COPD大鼠为切入点，以SIRT1活性为靶点，采用流式细胞仪检测氧化产物活性氧（ROS），通过Western-blot及免疫沉淀分析检测SIRT1，核因子-кB（NF-кB）蛋白表达及其相互关系，液相芯片技术检测NF-кB依赖的前炎症因子的变化，进一步探讨红霉素抑制香烟烟雾诱导的氧化应激和炎症作用与红霉素对SIRT1的调控是否相关及其相关机制，为临床防治COPD及慢性气道炎症提供新的思路。

背景：吸烟导致的氧化应激和肺部炎症是慢性阻塞性肺疾病（COPD）的重要发病机制。最近研究表明，COPD 患者及吸烟人群肺部沉默信息调节因子2相关酶sirtuin1（SIRT1)表达下调，继而转录因子活性增加及炎症介质合成。大环内酯类抗生素（红霉素）具有抗炎和抗氧化作用，但其具体作用机制尚不明确，沉默信息调节因子2相关酶sirtuins, 是一种 NAD+依赖性HDAC，共有7个成员（SIRT1-SIRT7）,其中SIRT1是目前研究最深入的一员，可以参与调节NF-κB乙酰化作用，进而使其蛋白表达受抑制，在COPD气道炎症与氧化应激中具有重要作用，研究内容：本实验研究内容在于探讨红霉素在烟草烟雾诱导的人巨噬细胞SIRT1-NF-κB通路和NF-κB依赖的炎症因子（IL-8，TNF-а, IL-6，MCP-1，MIP-2等）中的作用。研究结果：烟草烟雾提取物可以诱导人巨噬细胞ROS释放增加，减少SIRT1蛋白表达同时上调NF-κB蛋白表达。红霉素可以抑制烟草烟雾提取物导致的ROS释放增加，减轻氧化应激，增加SIRT1蛋白表达及抑制NF-κB表达进而减少炎症因子（IL-8，TNF-а, IL-6，MCP-1，MIP-2等）的释放。科学意义：红霉素能够通过减少烟草烟雾诱导的人巨噬细胞ROS释放而增加SIRT1表达，进而抑制了NF-κB蛋白表达，这样进一步使NF-κB依赖的炎症因子释放减少，而减轻炎症反应，这些数据为COPD的防治提供了新的理论基础。进一步探讨红霉素抑制香烟烟雾诱导的氧化应激和炎症作用与红霉素对SIRT1的调控是否相关及其相关机制。