Direct conversion of human amniotic cells into endothelial cells without transitioning through a pluripotent state.
Direct conversion of human amniotic cells into endothelial cells without transitioning through a pluripotent state.
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DOI：
10.1038/nprot.2015.126
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发表时间：
2015-12
期刊：
影响因子：
14.8
通讯作者：
Rafii S
中科院分区：
文献类型：
--
作者：
Ginsberg M;Schachterle W;Shido K;Rafii S
内皮细胞（EC）在器官发育和再生中发挥重要作用，因此它们可用于再生疗法。然而，从多能干细胞生成丰富的功能性内皮细胞一直很困难，因为许多现有策略生成的 EC 的增殖潜力有限，并且表现出血管不稳定。后一个困难尤为重要，因为随着时间的推移失去其身份的细胞可能不适合治疗用途。在这里，我们描述了一个为期 3 周的平台，用于在不使用多能因子的情况下直接将人类妊娠中期谱系羊水衍生细胞 (AC) 转化为稳定且可扩展的血管 EC (rAC-VEC) 群体。通过瞬时表达 ETS 转录因子 ETV2 2 周和组成型表达 ETS 转录因子 FLI1 和 ERG1，同时抑制 TGF-β 3 周，上皮和间质 AC 高效转化为功能性 rAC-VEC。这些 rAC-VEC 在体外经过多次传代后仍保持其血管库和形态，并且在植入体内时形成功能性血管。植入几个月后，可以在受体小鼠中检测到 rAC-VEC。因此，rAC-VEC 可用于建立一个细胞平台，以揭示血管发育和异质性的分子决定因素，并可能代表治疗再生障碍的理想 EC。
Endothelial cells (ECs) have essential roles in organ development and regeneration, and therefore they could be used for regenerative therapies. However, generation of abundant functional endothelium from pluripotent stem cells has been difficult because ECs generated by many existing strategies have limited proliferative potential and display vascular instability. The latter difficulty is of particular importance because cells that lose their identity over time could be unsuitable for therapeutic use. Here, we describe a 3-week platform for directly converting human mid-gestation lineage-committed amniotic fluid–derived cells (ACs) into a stable and expandable population of vascular ECs (rAC-VECs) without using pluripotency factors. By transient expression of the ETS transcription factor ETV2 for 2 weeks and constitutive expression the ETS transcription factors FLI1 and ERG1, concomitant with TGF-β inhibition for 3 weeks, epithelial and mesenchymal ACs are converted, with high efficiency, into functional rAC-VECs. These rAC-VECs maintain their vascular repertoire and morphology over numerous passages in vitro, and they form functional vessels when implanted in vivo. rAC-VECs can be detected in recipient mice months after implantation. Thus, rAC-VECs can be used to establish a cellular platform to uncover the molecular determinants of vascular development and heterogeneity and potentially represent ideal ECs for the treatment of regenerative disorders.