摘要
目的：研究人外剧血中内皮祖细胞（endothelial progenitor cell，EPC）的培养及分离，并探索EPC生物学特性及诱导分化条件：方法：健康成人肘静脉血，用密度梯度离心法得到单个核细胞，接种于纤维连接蛋白包被的6孔板进行培养（含有人血管内皮细胞生长因子），观察细胞集落、梭形贴壁细胞的形成过程。应用激光共聚焦显微镜和流式细咆仪进行EPC的鉴定。结果：在细胞培养的第4天，人外周血来源的单个核细胞开始形成从中间向外剧放射的细胞集落；住第7天时形成典型的长梭形，首尾相连成条索状；培养第2周时，长悛形细胞渐消失，出现鹅卵石样的圆形椭圆形细胞；在第3周时，鹅卵石样细胞增殖旺盛。可以应用激光共聚焦显微镜成功鉴定EPC，并且VEGF和人纤维连接蛋白可以促进EPC的生长。结论：人外周血中的内皮祖细胞来源于单个核细胞，早期内皮祖细胞出现于细胞培养第4-7天，晚期内皮祖绌胞出现于细胞培养2-3周时。EPC任特定条件下可分化为内皮细胞，为EPC的进一步研究及临床应用奠定了基础。
Objective： To investigate the isolation and culture of endothelial progenitor cells （EPCs）from peripheral blood in healthy adults and explore the induction and differentiation of EPCs. Methods ： Mononuclear cells from adult venous blood were centrituged by density gradient centrifngation, then the cells were seeded onto a six-well plate which preeoated with human fibroneetin at a density of（2-3）×10^6 cells/cm^2 and cultured in medium-199 supplied with human vascular endothelial growth factor （VEGF）. The morphous and a- mount of cells were observed. Flow cytometry assay and laser scanning confocal mieroscope（LSCM） were performed to identify the adherent cells. Results ： On the forth day of culture, typical endothelial cell colony-forming units （CFU-EC） were observed. At day 7, large amount of spindle -shaped cells emerged. At the second week, the total number of spindle-shaped ceils began to decrease. Meanwhile, the cobble-stone shaped cells were displayed. At the third week, cobble-stone shaped cells proliferated rapidly. EPCs were identified sue- cessfully by LSCM among the adherent cells. VEGF and Human Fibroneetin were able to promote the growth of the EPCs. Conclusions：EPCs are derived from mononuelear cells in peripheral blood, which could be induced into endothelial cells under specific conditions. The attached mnnonuclear cells cultured about 4-7 days are the early EPCs, the late EPCs are observed after 2-3 weeks of culture. This study is a primary research about EPCs.
出处
《解剖与临床》
2009年第6期423-425,共3页
Anatomy and Clinics
关键词
内皮祖细胞
外周血
体外培养
Endothelial progenitor cell
Peripheral blood
Culture in vitro
