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目的药物洗脱支架可抑制支架处血管内膜再内皮化,但对骨髓源性内皮修复作用的影响尚不清楚,为此观察了雷帕霉素对内皮祖细胞(endothelial progenitor cells,EPC)增殖、黏附及迁移的影响。方法采用密度梯度离心法从大鼠骨髓获取单个核细胞,将其接种在纤维连接素包被培养板,加入不同浓度雷帕霉素(0.01～100ng/ml),培养12天后,激光共聚焦显微镜鉴定FITC-UEA-Ⅰ和DiI-acLDL双染阳性细胞为正在分化的EPC,并在倒置荧光显微镜下计数。收集培养8天贴壁细胞,分别加入不同浓度雷帕霉素(0.1～200ng/ml)培养12～96h。然后分别采用MTT比色法、改良的Boyden小室和黏附能力测定实验观察EPC的增殖、迁移和黏附能力。结果雷帕霉素显著抑制骨髓单个核细胞分化为EPC,0.1ng/ml浓度雷帕霉素作用12天,EPC减少了(59.3±5.8)%(P<0.01)。雷帕霉素也显著抑制EPC增殖,其抑制作用随雷帕霉素浓度增加及作用时间延长而增加,1ng/ml浓度雷帕霉素作用24h使EPC数量减少(41.5±2.2)%(P<0.01)。雷帕霉素也显著抑制EPC的黏附和迁移能力。结论雷帕霉素可抑制骨髓单个核细胞分化为EPC,并抑制EPC的增殖及迁移能力。
The purpose of the drug-eluting stent can inhibit the endothelium re-endothelialization of vascular endothelium, but the effect of bone marrow-derived endothelial repair is not clear, for the observed rapamycin on endothelial progenitor cells (EPCs) proliferation , Adhesion and migration. Methods Mononuclear cells were obtained from bone marrow of rat by density gradient centrifugation. The cells were seeded onto fibronectin coated plates, and different concentrations of rapamycin (0.01-100ng / ml) were inoculated into the culture plates. After cultured for 12 days, confocal laser scanning microscopy Both FITC-UEA-I and DiI-acLDL double-stained positive cells were identified as differentiated EPCs and counted under an inverted fluorescence microscope. Adherent cells were harvested for 8 days and cultured for 12-96 h with different concentrations of rapamycin (0.1-200 ng / ml). Then the proliferation, migration and adhesion of EPC were observed by MTT assay, modified Boyden chamber and adhesion assay respectively. Results Rapamycin significantly inhibited the differentiation of bone marrow mononuclear cells into EPC. The effect of rapamycin at a concentration of 0.1ng / ml for 12 days was significantly reduced (59.3 ± 5.8%) (P <0.01). Rapamycin also significantly inhibited the proliferation of EPC. The inhibitory effect of rapamycin increased with the increase of rapamycin concentration and the prolongation of action time. The number of EPC decreased by 41.5 ± 2.2% at 1 ng / ml rapamycin for 24 h (P < 0.01). Rapamycin also significantly inhibited EPC adhesion and migration. Conclusion Rapamycin can inhibit the differentiation of bone marrow mononuclear cells into EPC and inhibit the proliferation and migration of EPCs.