氧化型低密度脂蛋白协同机械牵张力促进巨噬细胞ERK1/2活化
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国家自然科学基金(30570762、30871023、81070124);;广东省自然科学基金(8151008901000044);;高等学校博士学科点专项科研基金(20090171110049)


Oxidized Low Density Lipoprotein and Stretch Stress Synergistically Promote the Activition of ERK1/2 in Macrophages
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    摘要:

    目的探讨机械牵张力、氧化型低密度脂蛋白单独或共同存在时对巨噬细胞ERK1/2磷酸化的影响。方法用墨汁染色法对体外培养的巨噬细胞株RAW 264.7进行鉴定;用硫酸铜氧化天然低密度脂蛋白获得氧化型低密度脂蛋白,并用琼脂糖凝胶电泳进行鉴定和定量。体外培养静息状态的巨噬细胞分别给予氧化型低密度脂蛋白、机械牵张力以及机械牵张力+氧化型低密度脂蛋白处理不同时间和强度,收集的细胞蛋白用免疫印迹法检测细胞内ERK1/2磷酸化水平。结果巨噬细胞株RAW 264.7可吞噬墨汁,镜下细胞内有墨汁斑块或墨汁颗粒存在,或者有淡染呈云雾状的黑色物质存在。硫酸铜可氧化天然低密度脂蛋白成氧化型低密度脂蛋白,琼脂糖凝胶电泳显示天然低密度脂蛋白和氧化型低密度脂蛋白泳道在血浆β脂蛋白处出现单一的条带,并且氧化型低密度脂蛋白的迁移率明显较天然低密度脂蛋白高。氧化型低密度脂蛋白和机械牵张力刺激可分别引起细胞内ERK1/2活性增加,呈时间和强度依赖性;与氧化型低密度脂蛋白组和机械牵张力组相比,氧化型低密度脂蛋白和机械牵张力共同处理可使ERK1/2磷酸化明显增加。结论氧化型低密度脂蛋白、机械牵张力单独作用均可激活细胞内ERK1/2,而两者共同存在时可协同促进细胞内ERK1/2信号。为高血压时巨噬细胞如何参与动脉粥样硬化形成的机制探讨提供有用资料。

    Abstract:

    Aim To explore effects of oxidized low density lipoprotein(ox-LDL) and mechanical stretch stress(SS) on activation of extracellular regulated kinase(ERK1/2) in macrophage RAW264.7 cells. Methods The cultured RAW264.7 cells were identified via ink staining.Agarose gel electrophoresis was employed to identify and quantitate ox-LDL which was oxidated from n-LDL with copper sulfate.The identified RAW264.7 cells were subjected to treatment with ox-LDL and SS,respectively or jointly,for different dose/elongation and time duration,and then the phosphorylation of ERK1/2 in the macrophages was detected by Western blotting. Results RAW264.7 cells could phagocytize ink,forming cloudy ink plaque or black particles in cytoplasm.N-LDL could be oxidized into ox-LDL by copper sulfate,since a single lane about ox-LDL could be seen in agarose gel electrophoresis,and the electrophoretic mobility of ox-LDL was higher than that of n-LDL,indicating successful ox-LDL preparation.Ox-LDL and SS could induce ERK1/2 phosphorylation,respectively,in a time and dose dependent manner,and dramatical increase of ERK1/2 phosphorylation was observed when the cells were co-treated with SS and ox-LDL. Conclusions Ox-LDL and SS could induce ERK1/2 phosphorylation,and combined treatment of ox-LDL and SS could synergistically promote ERK1/2 phosphorylation in macrophages.This study could provide useful information for exploring the roles of macrophages and its mechanism in hypertension-related atherosclerosis.

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宁粉,汪照静,张征宇,张敏,刘树迎,李宇煌,黄锦桃,李朝红.氧化型低密度脂蛋白协同机械牵张力促进巨噬细胞ERK1/2活化[J].中国动脉硬化杂志,2010,18(12):925~930.

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  • 收稿日期:2010-09-27
  • 最后修改日期:2010-11-07
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