EPA和DHA经Src 激酶调控中性粒细胞趋化反应介导的抗炎机制研究

项目名称： EPA和DHA经Src 激酶调控中性粒细胞趋化反应介导的抗炎机制研究

项目编号： No.81502799

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 医药、卫生

项目作者： 楚心唯

作者单位： 南方医科大学

项目金额： 18万元

中文摘要： 近年来中性粒细胞(PMNs)成为抗炎的新靶标，健康人补充鱼油（主要成分EPA和DHA）抑制其趋化反应，但机制未明。本研究以EPA和DHA为受试物，结合体内外实验，首先观察PMNs趋化能力和胞内聚合F-actin的分布，检测Src家族激酶pSFKs,pLyn,pFgr,pHck,和GTP-Rac2蛋白表达水平，分析PMNs趋化反应中EPA和DHA对Src家族激酶磷酸化的影响。其次用confocal和超高速冷冻离心法观察和检测SFKs,Lyn,Fgr和Hck在脂筏中的表达水平，分析PMNs趋化反应中EPA和DHA对Src家族激酶膜定位的影响。最后建立GFP标记PMNs的转基因斑马鱼炎症模型，定量炎症部位PMNs数量并检测pSKFs蛋白表达水平，验证PMNs是鱼油抗炎的靶标。本研究系统阐明EPA和DHA经Src激酶调控PMNs趋化反应的分子机制，为鱼油防治PMNs介导的炎性疾病提供新的理论依据。

中文关键词： 二十碳五烯酸；二十二碳六烯酸；中性粒细胞；趋化；Src家族激酶

英文摘要： The last few years, novel and unexpected roles of neutrophils have been discovered in diverse aspects of immunity and inflammation,and evidence is now accumulating that suggests considering them as therapeutic targets.A number of studies on fish oil supplementation demonstrated a decrease in chemotaxis of human neutrophils towards various chemoattractants. The omega-3 fatty acids (FA) found in these oils are mainly eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). However, the mechanism of their targets and molecular mechanisms are not fully understood. In this study, we will focus on investigating the role of the Src family kinases (SFKs) in EPA and DHA-mediated neutrophil chemotaxis. First, we will observe the effect of EPA and DHA treatment on neutrophil chemotaxis using Zigmond chamber and Transwell. The impact of dietary (n-3) PUFA on both SFKs (Lyn, Fgr and Hck) will be examined in neutrophil chemotaxis using immunofluorescence stain, GST pull down and western blot.Secondly, we will explore the association between SFKs and lipid rafts in EPA and DHA regulating neutrophil polarization and chemotaxis by means of confocal microscopy and sucrose gradient ultracentrifugation At last,we will confirm the effect of EPA and DHA on neutrophil chemotaxis use transgenic zebrafish(Tg:mpx-GFP) ,in which neutrophil were labeled by green fluorescence protein(GFP).The present study will not only provide a new possible immunomodulatory mechanism by EPA and DHA during neutrophil chemotaxis, but also a novel scientific evidence for the treatment of neutrophil-mediated autoimmune disorders by fish oil.

英文关键词： Eicosapentaenoic acid;Docosahexaenoic acid;Neutrophil;Chemotaxis;Src family kinase