银杏叶提取物对外周血内皮祖细胞数量和功能的影响

Effects of Ginkgo biloba extract on number and activity of endothelial progenitor cells from peripheral blood

目的 观察银杏叶提取物对外周血内皮祖细胞 (endothelialprogenitorcell,EPC)数量和功能的影响。方法 密度梯度离心法获取外周血单个核细胞 ,培养 7d后 ,收集贴壁细胞并加入银杏叶提取物 (10 ,2 5和 5 0mg·L- 1 )干预一定时间 (6 ,12 ,2 4和 4 8h)。激光共聚焦显微镜和流式细胞仪鉴定EPC ,分别观察EPC的增殖、迁移、粘附和体外血管生成能力。结果 银杏叶提取物促进外周血EPC扩增 ,2 5mg·L- 1 银杏叶提取物作用 2 4h对EPC数量的影响最为显著 (较对照组增加了 1倍 ,P <0 .0 1)。银杏叶提取物也显著改善了外周血EPC的粘附、迁移、增殖和体外血管生成能力。

Aim To investigate whether Ginkgo biloba extract can augment endothelial progenitor cell (EPC) number, and promote EPC proliferation, migration and adhesion. Methods Total mononuclear cells (MNCs) were isolated from peripheral blood by Ficoll density gradient centrifugation, and then the cells were plated on fibronectin-coated culture dishes. After 7 days of culture, attached cells were stimulated with Ginkgo biloba extract (10, 25 and 50 mg·L -1) or vehicle control for the respective time points (6, 12, 24 and 48 h). EPC were characterized as adherent cells double positive for DiLDL-uptake and lectin binding by direct fluorescent staining under a laser scanning confocal microscope. EPC were further documented by demonstrating the expression of CD34, VEGFR-2 and AC133 with flow cytometry. EPC proliferation, migration and in vitro vasculogenesis activity were assayed with MTT assay, modified Boyden chamber assay and in vitro vasculogenesis kit, respectively. EPCs adhesion assay was performed by replating MNCs on fibronectin-coated dishes, and then counting adherent cells. Results Incubation of isolated human MNCs with Ginkgo biloba extract increased the number of EPC, maximum at 25 mg·L -1, 24 hours (approximately 1-fold increase, P<0.01). In addition, Ginkgo biloba extract promotes EPC proliferative, migratory, adhesive and in vitro vasculogenesis capacity. Conclusion Ginkgo biloba may promote EPC augmentation and enhance its functional activity.