小径聚氨酯人工血管内皮化种子细胞的体外诱导分化及种植

[目的]探讨体外诱导骨髓单个核细胞分化成为内皮祖细胞,为小径聚氨酯人工血管内皮化提供合适的种子细胞的可行性.[方法]收集健康成人骨髓单个核细胞,置于纤维连接蛋白预衬的DMEM培养基中,用血管内皮生长因子(vascular endothelial growth factor,VEGF)和碱性成纤维细胞生长因子(basicfibroblast growth factor,bFGF)加以诱导,通过荧光显微镜和免疫组化分析等方法观察和鉴定诱导后的细胞.将诱导分化的内皮祖细胞种植到聚氨酯小径人工血管表面,用扫描电镜观察.[结果]在VEGF、bFGF等诱导因子存在的条件下,骨髓单个核细胞分化成为内皮祖细胞,倒置荧光显微镜下呈典形的"纺锤样"梭形细胞,单层细胞贴壁生长融合时呈铺路石样排列,免疫组化示VWF和CD34抗体染色阳性.扫描电镜下,未种植细胞的聚氨酯小径人工血管表面呈典型的多孔蜂窝状结构,孔径大小比较适合内皮祖细胞爬行.种植细胞后,聚氨酯小径人工血管表面有大量的内皮祖细胞黏附、爬行及铺展生长,有时可见内皮祖细胞长入蜂窝状孔径内.[结论]体外诱导骨髓单个核细胞分化为内皮祖细胞可作为小径聚氨酯人工血管内皮化的种子细胞.

In Vitro Induction, Differentiation, and Seeding of Source Cells for Endothelialization of Polyurethane Small Diameter Artificial Blood Vessel

[Objective]The study was designed to induce bone-marrow mononuclear cells (MNCS) differentiating into endothelial progenitor cells (EPCs) in vitro and to seed them on polyurethane small diameter artificial blood vessel, in order to provide source cells for the endothelization of small diameter artificial vessel. [Methods] The bone marrow mononuclear cells of healthy adult were acquired and put in DMEM culture which was coated with fibronectin, and induced by vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). The differentiated endothelial cells were observed and identified by fluorescent microscope and immunohistochemical analysis, seeded on the polyurethane small-diameter artificial vessels, and observed by scanning electronic microscope. [Results] Under the induction factors such as VEGF and bFGF, bone marrow mononuclear cells differentiated into EPCs. They presented typical "spindle-shaped" appearance and formed a monolayer that arrayed in "cobblestone-like" morphology. Identifications of immunohistochemical analysis indicated that they were positively stained for von Willebrand factor (VWF) and CD34 antigen. The unseeded polyurethane small-diameter artificial vessel showed polyporous honeycomb structure under observation of scanning electronic microscope. The size of the hole suited the crawling of EPCs. After being seeded with the EPCs, the cells showed adhesion, crawling, and spreading on the polyurethane vessel surface. Some cells grew into the honeycomb-like holes.[Conclusion]The present study demonstrates that the EPCs may be used as source cells for the endo-thelization of small diameter artificial vessels.