羟丁基壳聚糖携载CD133抗体温敏膜对内皮祖细胞的影响  

Influence of hydroxybutyl-chitosan carrying CD133 antibody thermosensitive membrane on the endothelial progenitor cells

作　　者：张翠娟 王向真[2] 于帮旭[1] 安毅[3] 李丹[3] ZHANG Cui-juan;WANG Xiang-zhen;YU Bang-xu;AN Yi;LI Dan(Department of Critical Care Medicine,The Affiliated Hospital of Qingdao University,Qingdao,Shandong 266000,China;Department of Anesthesia,The Affiliated Hospital of Qingdao University,Qingdao,Shan-dong 266000,China;Department of Cardiology,The Affiliated Hospital of Qingdao University,Qingdao,Shandong 266000,China)

机构地区：[1]青岛大学附属医院重症医学科,山东青岛266000 [2]青岛大学附属医院麻醉医学科,山东青岛266000 [3]青岛大学附属医院心血管内科,山东青岛266000

出　　处：《岭南心血管病杂志》2021年第5期581-586,共6页South China Journal of Cardiovascular Diseases

基　　金：山东省自然科学基金面上项目(项目编号:ZR2016HP17)。

摘　　要：目的研究羟丁基壳聚糖携载CD133抗体温敏膜对外周血内皮祖细胞(endothelial progenitor cells,EPCs)生长、黏附、迁移及分化的影响。方法(1)用密度梯度离心法取人外周血单个核细胞,培养7 d后收集贴壁细胞进行Dil-乙酰化低密度脂蛋白(acetylated low-density lipoprotein,ac-LDL)和FITC-荆豆凝集素I(UEA-I)双荧光染色鉴定;(2)鉴定后的细胞分3组,分别接种到铺有裸钴铬合金片膜、雷帕霉素药物涂层钴铬合金片膜、复合羟丁基壳聚糖携载CD133抗体温敏膜钴铬合金片膜的培养板中培养,分别记为裸金属组、雷帕霉素组及抗体温敏膜组;(3)取培养7 d的3组细胞计数并采用黏附能力测定实验、transwell小室法分别测定3组细胞的黏附和迁移能力的差异;(4)取培养14 d的3组细胞用实时聚合酶链反应(real-time polymerase chain reaction,RT-PCR)法测定细胞内内皮型一氧化氮合成酶(endothelial nitric oxide synthase,eNOS)基因含量的表达差异。结果(1)培养7 d的贴壁细胞CD133表达率为(20.65±2.70)%,CD34表达率为(33.90±3.31)%,KDR表达率为(62.98±3.18)%,细胞摄取ac-LDL显红色荧光,与UEA-Ⅰ结合显绿色荧光,双阳性细胞为EPCs,显黄色。(2)EPCs在抗体温敏膜上生长良好,形态正常。与裸金属组、雷帕霉素组相比,抗体温敏膜组EPCs的细胞个数、黏附数量、迁移细胞数量增高,差异有统计学意义(P<0.01)。(3)RT-PCR示抗体温敏膜上的EPCs eNOS mRNA表达水平增高,差异有统计学意义(P<0.01),是裸金属组基因含量的2.43倍,是雷帕霉素组基因含量的5.90倍。结论羟丁基壳聚糖携载CD133抗体温敏膜能促进EPCs生长、黏附、迁移及分化,为理想的生物学材料,促进了药物洗脱支架的进展。Objectives To investigate the influence of hydroxybutyl-chitosan carrying CD133 antibody thermosensitive membrane on growth,adlhesion,migration and differentiation of the endothelial progenitor cells in peripheral blood.Methods(1)Human peripheral blood mononuclear cells isolated by density gradient centrifugation were cultured for 7 days,then the adherent cells were identified by Dil-ac-LDL and FITC-UEA-I.(2)The identified cells were inoculated into three kinds of culture plate,the first covered with bare cobalt-chromium alloy film(bare mental group),the second with rapamycin drug-eluting cobalt-chromium alloy film(rapamycin group),the third with hydroxybutyl chitosan carrying CD133 antibody thermosensitive membrane cobalt-chromium alloy film(antibody thermosensitive group).(3)When the three groups of cells had cultured for 7 days,taking cells counting and using adhesion experiments and transwell chamber to assay the capability differences of adhesion and migration.(4)Using real-time polymerase chain reaction(PT-PCR)to test the expression of endothelial nitric oxide synthase(eNOS)in cells of the three groups after the cells had cultured for 14 days.Results(1)In 7 days cultured adherent cell,CD133 expression was 20.65%±2.70%,CD34 expression was 33.90%±3.31%,KDR expression was 62.98%±3.18%,and the cells which uptook ac-LDL showed red fluorescence,which binded with UEA-I showed green fluorescence,double positive cells which showed yellow fluorescence were identified as endothelial progenitor cells.(2)Endothelial progenitor cells grew well in antibody thermosensitive group.Compared with the other two groups,the cells count and the number of adhesion and migrating cells of endothelial progenitor cells in antibody thermosensitive group both increased(P<0.01);(3)RT-PCR showed eNOS mRNA expression level was higher than those of the other two groups,which was 2.43 times as much as bare mental group,was 5.90 times as much as rapamycin group.Conclusions Hydroxybutyl-chitosan carrying CD133 antibody thermosensitive membra

关 键 词：羟丁基壳聚糖 CD133 内皮祖细胞 雷帕霉素 

分 类 号：R33[医药卫生—人体生理学]