冠状动脉支架再狭窄患者循环内皮祖细胞数量及活性的变化

目的 观察冠状动脉支架再狭窄患者循环内皮祖细胞(EPC)数量及活性的变化.方法 根据复查冠脉造影的结果将既往行冠脉支架置入术的研究对象分为两组:再狭窄组(15例)及对照组(17例),采用密度梯度离心法获取外周血总的单个核细胞,接种到包被有人纤维连接蛋白的24孔培养板,7 d后通过免疫荧光显微镜鉴定细胞表面特异性抗原CD34和KDR,并通过激光共聚焦显微镜鉴定EPC摄取DiI-acLDL及结合FITC-UEA-Ⅰ的能力.通过倒置显微镜进行计数,通过划痕试验测定EPC的迁移能力,通过MTT比色法测定EPC的增殖能力,并通过黏附试验测定EPC的黏附能力.结果 再狭窄组的EPC数量明显低于对照组(4.97±1.42比17.2±3.90,P=0.001),再狭窄组的EPC增殖能力明显低于对照组(增殖倍数1.37±0.32比2.01±0.62,P＜0.05),再狭窄组的EPC迁移能力明显低于对照组,但两组之间的EPC黏附能力无明显的统计学意义.结论 冠状动脉支架再狭窄患者的EPC数量及增殖能力、迁移能力明显下降,可能参与支架再狭窄的发生机制.

Activities of circulating endothelial progenitor cells in patients with in-stent restenosis

OBJECTIVE: To observe the number and activities of circulating endothelial progenitor cells (EPCs) in patients with in-stent restenosis. METHODS: Peripheral blood samples were collected from 15 patients with angiographically restenosis, and 17 baseline characteristics-matched patients without angiographically restenosis (control group). Mononuclear cells were isolated by Ficoll density-gradient centrifugation and plated on dishes coated with human fibronectin. After 7 days in culture, the nature of EPCs was characterized with anti-CD34 and anti-KDR, specific surface antibodies of EPC, and confirmed further with the use of fluorescein isothiocyanate-labeled ulex europaeus agglutinin-I (FITC-UEA-I) and DiI (1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine percolate)-labeled acetylated low-density lipoprotein (DiI-acLDL) by laser scanning confocal microscopy. The number of EPCs was counted in a blinded manner. EPCs were inoculated onto the culture plate and 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-2H-tetrazolium bromide assay was used to measure the A value by enzyme labeling instrument to evaluate the proliferation. The migration of EPCs was assayed by scratch assay. EPC adhesion was performed by replating cells on fibronectin-coated dishes and then counting the adherent cells. Results The number of EPCs of the patients with in-stent restenosis was 4.97 +/- 1.42/well, significantly lower than that of the control group (17.2 +/- 3.90/well, P = 0.001). MTT assay showed that the proliferative activities of the in-stent restenosis group was 1.37 +/- 0.32 times the baseline value, significantly lower than that of the control group (2.01 +/- 0.62, P < 0.05). The number of migrating EPCs of the in-stent restenosis group was remarkably lower than that of the control group. There was no significant difference in the adherent activity between the two groups. Conclusion The number, proliferation activity, and migration activity of the EPCs patients with in-stent restenosis are all significantly lower, which may contribute to the mechanism of in-stent restenosis.