McCloskey Kara E, Gilroy Meghan E, Nerem Robert M
The Parker H. Petit Institute for Bioengineering and Bioscience, Georgia Institute of Technology, Atlanta, GA 30332, USA.
Tissue Eng. 2005 Mar-Apr;11(3-4):497-505. doi: 10.1089/ten.2005.11.497.
Embryonic stem (ES) cells could potentially serve as an excellent cell source for various applications in regenerative medicine and tissue engineering. Our laboratory is particularly interested in generating a reproducible endothelial cell source for the development of prevascularized materials for tissue/organ reconstruction. After developing methods to isolate highly purified (>96%) proliferating populations of endothelial cells from mouse embryonic stem cells, we tested their ability to form three-dimensional (3-D) vascular structures in vitro. The ES cell-derived endothelial cells were embedded in 3-D collagen gel constructs with rat tail collagen type I (2 mg/mL) at a concentration of 10(6) cells/mL of gel. The gels were observed daily with a phase-contrast microscope to analyze the time course for endothelial cell assembly. The first vessels were observed between days 3 and 5 after gel construct formation. The number and complexity of structures steadily increased, reaching a maximum before beginning to regress. By 2 weeks, all vessel-like structures had regressed back to single cells. Histology and fluorescent images of the vessel-like structures verified that tube structures were multicellular and could develop patent lumens. We have shown that endothelial cells derived, purified and expanded in vitro from ES cells sustain an important endothelial cell function, the ability to undergo vasculogenesis in collagen gels, indicating that endothelial products derived in vitro from stem cells could be useful in regenerative medicine applications.
胚胎干细胞有潜力成为再生医学和组织工程中各种应用的优质细胞来源。我们实验室尤其感兴趣的是生成一种可重复的内皮细胞来源，用于开发用于组织/器官重建的预血管化材料。在开发出从小鼠胚胎干细胞中分离高度纯化（>96%）的增殖性内皮细胞群体的方法后，我们测试了它们在体外形成三维（3-D）血管结构的能力。将胚胎干细胞衍生的内皮细胞以10(6)个细胞/mL凝胶的浓度包埋在含有I型大鼠尾胶原（2 mg/mL）的三维胶原凝胶构建物中。每天用相差显微镜观察凝胶，分析内皮细胞组装的时间进程。在凝胶构建物形成后的第3至5天观察到第一批血管。结构的数量和复杂性稳步增加，在开始退化之前达到最大值。到2周时，所有血管样结构都退化为单个细胞。血管样结构的组织学和荧光图像证实，管状结构是多细胞的，并且可以形成有功能的管腔。我们已经表明，从胚胎干细胞体外衍生、纯化和扩增的内皮细胞维持着重要的内皮细胞功能，即在胶原凝胶中进行血管生成的能力，这表明从干细胞体外衍生的内皮细胞产物可能在再生医学应用中有用。
