AMD3100对apoE^-/-小鼠骨髓源性内皮祖细胞增殖、迁移和黏附的影响

The Effect of AMD3100 on The Proliferation, Migration and Adhesion of apoE^(-/-) Mice Bone Marrow Endothelial Progenitor Cells

探讨AMD3100对apoE-/-小鼠骨髓内皮祖细胞的动员作用及其增殖、迁移和黏附的影响.12只8周龄雄性apoE-/-小鼠随机分为AMD3100组(2.5mg/(kg·2d))和对照组(PBS0.1ml/2d),高脂高胆固醇饲料喂养12周后,差速贴壁法结合微孔法分离培养小鼠骨髓细胞,免疫荧光鉴定CD133/VEGFR-2双阳性细胞为内皮祖细胞;MTT比色法、Transwell、黏附试验分别检测细胞的增殖、迁移和黏附能力;通过计数典型内皮祖细胞克隆形成单位,观察次级集落单位的大小及细胞密度,检测各组内皮祖细胞的克隆形成能力;RT-PCR和Westernblot检测内皮祖细胞上CXCR4mRNA和蛋白质表达水平.与对照组比较,AMD3100组骨髓源性内皮祖细胞的增殖、迁移、黏附和克隆形成能力均显著低于对照组,其CXCR4mRNA和蛋白质表达均显著低于对照组.结果表明:持续注射AMD3100可抑制骨髓源内皮祖细胞的增殖、迁移、黏附和克隆形成能力,并下调CXCR4的表达.

To study the effect of AMD3100 on the mobilization, proliferation, migration and adhesion of endothelial progenitor cells （EPC）, EPC was isolated from apoE^-/- mice bone marrow, 12 male apoE^-/- mice, with 8 weeks old,were randomly divided into two groups, AMD3100 group （2.5mg / （kg·2d）） and control group（PBS, 0.1 ml/2d）. After feeding western （high fat and cholesterol） for 12 weeks, the bone marrow cells were isolated and cultured by way of differential-speed-adherence and Micropore-Method. CD 133＋ VEGFR-2＋ bone marrow cell was identified as endothelial progenitor cells by immunofluorescence. The proliferation, migration and adhesion of endothelial progenitor cells were detected by MTT chromometry, transwell and adhesion test, respectively. By counting the typical endothelial progenitor cells-colony forming units （EPC-CFUs） and observing the size and cell density of second EPC-CFUs, the clonality of endothelial progenitor cells was determined. The expression of CXCR4 mRNA and protein were measured by RT-PCR and Western blot. As a result, the proliferation, migration, adhesion and clonality of endothelial progenitor cells derived from AMD3100 group were attenuated in comparison to the control group; the expression of CXCR4 mRNA and protein of AMD3100 group were also lower to control group. It can be concluded that, lasting administration of AMD3100 inhibits the proliferation, migration, adhesion and clonality of bone marrow endothelial progenitor cells and down-regulate the expression of CXCR4 on endothelial progenitor cells.