孤儿核受体Nur77抑制小鼠巨噬细胞脂质沉积  

Nuclear receptor Nur77 inhibits oxidized low density lipoprotein induced lipid loading in macrophages

作　　者：胡刘华[1] 何奔[1] 沈玲红[1] 周磊[1] 卜军[1] 江立生[1] 邵琴[1] 王力[2] 曾锦章[1] 

机构地区：[1]上海交通大学医学院附属仁济医院心内科,200001 [2]中科院上海生命科学研究院

出　　处：《中华心血管病杂志》2008年第11期1032-1036,共5页Chinese Journal of Cardiology

基　　金：国家自然科学基金资助项目（30670880）;上海市科委基础研究重点项目（05JC14037）

摘　　要：目的探讨孤儿核受体Nur77对巨噬细胞脂质沉积的影响及其机制。方法建立稳定表达绿色荧光蛋白（GFP）和GFP-Nur77质粒cDNA的小鼠巨噬细胞RAW264．7单克隆细胞系，40μg/ml氧化型低密度脂蛋白（ox-LDL）刺激24h后，用油红O染色定性观察细胞内脂质沉积，液相色谱-质谱联用法定量检测细胞内胆固醇酯，用荧光实时定量PCR（RT-PCR）检测CD36和腺苷三磷酸结合盒转运体A1（ABCA1）mRNA水平的变化，用流式细胞仪检测CD36蛋白表达，Westernblot方法检测ABCA1蛋白表达。结果Nur77能够显著减少ox-LDL诱导引起的巨噬细胞内脂质沉积。ox-LDL诱导前，GFP-RAW264．7组和GFP-Nur77-RAW264．7组细胞内总胆固醇含量分别为（38．66±0．13）μg／mg蛋白和（36．25±0．48）μg／mg蛋白，细胞内胆固醇酯含量为（18．85±0．03）μg／mg蛋白和（17．79±0．86）μg/mg蛋白。ox—LDL诱导24h后，GFP-RAW264．7组和GFP-Nur77-RAW264．7组细胞内总胆固醇含量分别为（68．98±1．68）μg／mg蛋白和（50．94±1．63）μg/mg蛋白，细胞内胆固醇酯含量为（35．92±2．28）μg/mg蛋白和（24．81±2．92）μg／mg蛋白。同时伴随CD36的mRNA及蛋白表达下降和ABCA1的mRNA及蛋白表达上调。结论孤儿核受体Nur77通过减少脂质摄取和增加脂质排出而减轻巨噬细胞内脂质沉积，这可能是其抗动脉粥样硬化的重要机制之一。Objective To observe the effect of Nur77 on lipid loading in macrophages exposed to 40μg/ml oxidized low density lipoprotein （ox-LDL）. Methods Stable RAW264.7 strain expressing green fluorescent protein （GFP） or GFP-Nur77 was established by G418 screening after transfection with corresponding plasmids and identified by Western blot. After 24 h stimulation with ox-LDL, intracellular lipid loading of each strain was observed by Oil Red O dyeing, and the intracellular cholesterol level was measured by liquid chromatographic-mass spectrometry （LC-MS）. The transcriptional changes of CD36 and ABCA1 were monitored by Real Time Quantitative-PCR, while the expressions of these two proteins were assayed by flow cytometry and Western blot, respectively. Results After 24 h stimulation with ox-LDL, intracellular total cholesterol and esterified cholesterol concentration in GFP-Nur77-RAW264. 7 were significantly dropped by 26. 15% and 30. 93% respectively （P 〈0. 05 vs. GFP-RAW264.7）. The transcription and expression of ABCA1 in GFP-Nur77-RAW264.7 were significantly increased while the transcription and expression of CD36 were significantly reduced （ all P 〈 0.05 vs. GFP-RAW264.7 ） . Conclusion Orphan nuclear receptor Nur77 reduced ox-LDL induced intracellular lipid loading in macrophages by inhibiting lipid influx and enhancing lipid efflux.

关 键 词：动脉粥样硬化 巨噬细胞 抗原 CD36 

分 类 号：R686[医药卫生—骨科学]