人心脏微血管内皮细胞血管细胞黏附分子1及核因子κB在缺氧再复氧损伤后的表达与黄芪多糖的干预  

Intervention of astragalus polysaccharides on the expression of vascular cell adhesion molecule-1 and nuclear factor-kappa B in human cardiac microvascular endothelial cells after hypoxia and reoxygenation injury

作　　者：朱海燕[1] 陈立新 朱陵群[3] 

机构地区：[1]北京中医药大学东直门医院中医内科学教育部重点实验室,北京市100700 [2]北京王府中西医结合医院心血管内科,北京市102209 [3]北京中医药大学东直门医院中医脑病研究室,北京市100700

出　　处：《中国组织工程研究与临床康复》2007年第41期8255-8258,共4页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：国家自然科学基金资助项目(30572391)~~

摘　　要：目的:前期对整体动物的研究发现,黄芪多糖具有抗心肌缺血再灌注损伤的作用。此次主要定量测定人心脏微血管内皮细胞血管细胞黏附分子1及核因子κB mRNA的表达,探讨黄芪多糖治疗心肌缺血再灌注损伤的机制。方法:实验于2007-01/03在北京中医药大学东直门医院中医内科学教育部重点实验室完成。①实验材料:原代人心脏微血管内皮细胞(ScienCell公司:从人心肌组织中分离后立即冻存)。②实验过程及分组:以体外缺氧再复氧复制缺血再灌注损伤模型。将细胞分为4组:正常对照组、再灌注损伤组、黄芪多糖组(分为100,50,25mg/L3个亚组,在缺氧/复氧的同时加入相应浓度的药物)及吡咯烷二硫代氨基甲酸组(50μmol/L,于缺氧前加入,预处理30min后吸弃)。③实验评估:应用实时荧光定量聚合酶链反应检测血管细胞黏附分子1和核因子κB mRNA表达的变化。结果:①人心脏微血管内皮细胞的培养:复苏16h细胞散在分布,胞体肥厚、透明,呈菱形或多角形;36h形成多个小岛样克隆;48～72h细胞约85%融合,呈铺路石样生长。②实时荧光定量聚合酶链反应:扩增动力曲线平滑,每条曲线均有较明显的指数扩增期;扩增回归曲线的回归系数均>0.990,起始模版浓度与Ct值之间呈良好的线性关系。与正常对照组比,缺氧再复氧可明显增加人心脏微血管内皮细胞中血管细胞黏附分子1和核因子κB mRNA的表达(P<0.05,P<0.01)。黄芪多糖各剂量组均明显抑制缺氧再复氧后人心脏微血管内皮细胞上血管细胞黏附分子1mRNA的表达(P<0.01);核因子κB mRNA的表达在黄芪多糖50mg/L剂量组中降低(P<0.05)。结论:黄芪多糖能够抑制再灌注损伤的人心脏微血管内皮细胞中血管细胞黏附分子1及核因子κB mRNA的表达,从而减轻内皮细胞的炎症反应及黏附作用,改善心肌缺血再灌注损伤。AIM： The early researches on general animals suggest that astragalus polysaccharides （APS） could inhibit myocardial injury after ischemia/reperfusion. In this study, the expression of vascular cell adhesion molecule-1 （VCAM-1） and nuclear factor-KB （NF-KB） mRNA in human cardiac microvascular endothelial cells （HCMEC） was quantitatively detected, so as to explore the mechanism of APS on reducing myocardial injury after ischemia/reperfusion. METHODS： The experiment was performed at Key Laboratory of Chinese Internal Medicine of Ministry of Education, Dongzhimen Hospital of Beijing University of Chinese Medicine from January to March 2007. （1）Primary cultured HCMEC was immediately froze after isolated from human myocardial tissue （ScienCell）. Ischemia/reperfusion （I/R） injury model was established by oxygen deprivation and reoxygenation method. HCMECs were divided into control group, I/R injury group, 100, 50 and 25 μg/mL APS groups （besides oxygen deprivation and reoxygenation） and pyrrolidine dithiocarbamate group （50 μmol/L added before hypoxia, and disposed after 30 minutes preconditioning）. （2）The mRNA concentrations of VCAM-1 and NF-κB were detected by real time fluorescence quantitative polymerase chain reaction. RESULTS： （1）Cultivation of HCMEC： Cells distributed separately after recovered for 16 hours. The cell bodies were pachynsis and transparent and showed in rhombus and polygon shapes; several islet liking clones were formed after 36 hours culture; about 85% cells were confluent and showed a cobblestone appearance after 48 to 72 hours. （2）Flu orescence quantitative polymerase chain reaction showed that amplifiable dynamic curves were smooth, and the stage of index amplification was obvious; regression coefficient of amplifiable regression curves were larger than 0.990 and a well linear relationship was presented between initiation template concentration and Ct value. Compared with control group, the expression of VCAM-1 and NF-KB mRNA was increa

关 键 词：黄芪多糖 微血管内皮细胞 缺氧 胞间粘附分子1 核因子ΚB 组织构建 

分 类 号：R542.2[医药卫生—心血管疾病]