Ormiston Mark L, Toshner Mark R, Kiskin Fedir N, Huang Christopher J Z, Groves Emily, Morrell Nicholas W, Rana Amer A
Department of Medicine, University of Cambridge;
Papworth Hospital.
J Vis Exp. 2015 Dec 23(106):e53384. doi: 10.3791/53384.
Historically, the limited availability of primary endothelial cells from patients with vascular disorders has hindered the study of the molecular mechanisms underlying endothelial dysfunction in these individuals. However, the recent identification of blood outgrowth endothelial cells (BOECs), generated from circulating endothelial progenitors in adult peripheral blood, may circumvent this limitation by offering an endothelial-like, primary cell surrogate for patient-derived endothelial cells. Beyond their value to understanding endothelial biology and disease modeling, BOECs have potential uses in endothelial cell transplantation therapies. They are also a suitable cellular substrate for the generation of induced pluripotent stem cells (iPSCs) via nuclear reprogramming, offering a number of advantages over other cell types. We describe a method for the reliable generation, culture and characterization of BOECs from adult peripheral blood for use in these and other applications. This approach (i) allows for the generation of patient-specific endothelial cells from a relatively small volume of adult peripheral blood and (ii) produces cells that are highly similar to primary endothelial cells in morphology, cell signaling and gene expression.
从历史上看，血管疾病患者的原代内皮细胞获取有限，这阻碍了对这些个体内皮功能障碍潜在分子机制的研究。然而，最近发现的血液来源内皮细胞（BOECs），由成人外周血中循环的内皮祖细胞生成，可为患者来源的内皮细胞提供类似内皮的原代细胞替代物，从而可能克服这一限制。除了在理解内皮生物学和疾病建模方面的价值外，BOECs在内皮细胞移植治疗中也有潜在用途。它们也是通过核重编程生成诱导多能干细胞（iPSC）的合适细胞底物，与其他细胞类型相比具有许多优势。我们描述了一种从成人外周血中可靠地生成、培养和鉴定BOECs的方法，用于这些及其他应用。这种方法（i）能够从相对少量的成人外周血中生成患者特异性内皮细胞，并且（ii）所产生的细胞在形态、细胞信号传导和基因表达方面与原代内皮细胞高度相似。
