氨氯地平拮抗ox-LDL损伤大鼠骨髓源性内皮祖细胞血管样结构形成及机制  

Antagonistic Effect of Amlodipine on Vascular Tube-like Information of Oxidized Low Density Lipoprotein Injured Rat Bone Marrow Derived Endothelial Progenitor Cells

作　　者：童海[1] 雷建军[2] 王仁[2] 孟军[1] 张凯[2] 冯祺论 吴灿[2] 王佐[2] 

机构地区：[1]南华大学附属第一医院 [2]南华大学心血管疾病研究所动脉硬化学湖南省重点实验室,湖南省衡阳市421001

出　　处：《中国动脉硬化杂志》2015年第3期231-236,共6页Chinese Journal of Arteriosclerosis

基　　金：国家自然科学基金项目(81070221);江苏省博士后科研资助基金(1302095B);湖南省卫生厅课题(B2013-034;B2011-054)资助

摘　　要：目的探索氨氯地平拮抗氧化型低密度脂蛋白(ox-LDL)损伤大鼠骨髓源性内皮祖细胞(EPC)血管样结构形成及其作用机制。方法实验分为对照组、ox-LDL组(50 mg/L ox-LDL)和氨氯地平组(50 mg/L ox-LDL+0.5μmol/L氨氯地平)。差异贴壁法培养分离EPC,ac-LDL摄取和结合UEA-1鉴定EPC,Transwell测定细胞迁移,Matrigel法测血管生成,Western blot和RT-PCR分别检测内皮型一氧化氮合酶(e NOS)的蛋白水平和mRNA表达情况,DCFH-DA法检测活性氧(ROS)水平,Griess法测定一氧化氮(NO)含量。结果 EPC经50 mg/L的ox-LDL处理后,细胞的迁移能力下降近3倍,血管样结构形成能力明显下降,用0.5μmol/L氨氯地平干预可部分恢复EPC的迁移能力,并显著恢复EPC的血管样结构形成能力(P<0.05)。机制研究发现,ox-LDL下调EPC e NOS mRNA和蛋白表达水平,显著减少NO的含量(P<0.01),氨氯地平显著拮抗ox-LDL的上述作用;EPC经50 mg/L的ox-LDL处理后,细胞内ROS含量显著增加(P<0.01),氨氯地平显著减少胞内ROS的水平(P<0.05)。结论氨氯地平对ox-LDL损伤EPC血管样结构的形成有显著拮抗作用,其作用与上调e NOS表达和降低细胞内ROS水平有关。Aim To investigate the antagonistic effect of amlodipine on oxidized low density lipoprotein( oxLDL) induced rat marrow endothelial progenitor cells( EPC) vascular tube-like information injury and related mechanisms.Methods EPC was isolated from rat marrow,and was divided into three groups,control( 200 μmol / L EDTA),ox-LDL group( 50 mg / L ox-LDL) and amlodipine group( 50 mg / L ox-LDL + 0. 5 μmol / L amlodipine). Differential adhesion rate was used to isolate EPC,endocytosis ac-LDL and binding UEA-1 were used to identify EPC,and transwell for migration,matrigel for tube information,Western blot and RT-PCR were used to measure the protein and mRNA level of endothelial nitric oxide synthase( e NOS),and DCFH-DA dyeing for reactive oxygen species( ROS),Griess Reagent for nitric oxide( NO),respectively. Results The migration capacity of EPC was reduced almost 3 times by 50 mg / L ox-LDL,and this inhibitory effect can be recovered by 0. 5 μmol / L amlodipine; The tube-like information was reduced by 50 mg / L ox-LDL remarkably( P < 0. 01),and it was partly recovered by administrating 0. 5 μmol / L amlodipine. Both mRNA and protein of e NOS were reduced by 50 mg / L ox-LDL,also the NO level was reduced( 11. 44 ± 1. 15 μmol / L vs 24. 35 ±4. 62 μmol / L),and it can be restored by administrating 0. 5 μmol / L amlodipine significantly( P < 0. 05); The ROS level was increased by 50 mg / L ox-LDL in EPC( 1. 00 ± 0. 25 vs 2. 53 ± 0. 32),and it was decreased by administrating 0. 5μmol /L amlodipine significantly( P < 0. 05). Conclusions Injury effect of ox-LDL on tube-like information of EPC can be antagonized by amlodipine,and it was related to up-regulating expression of e NOS and reducing ROS generation.

关 键 词：内皮祖细胞 氧化型低密度脂蛋白 氨氯地平 血管样生成 

分 类 号：R965[医药卫生—药理学]