Derivation of mesenchymal stromal cells from pluripotent stem cells through a neural crest lineage using small molecule compounds with defined media.
Derivation of mesenchymal stromal cells from pluripotent stem cells through a neural crest lineage using small molecule compounds with defined media.
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DOI：
10.1371/journal.pone.0112291
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Toguchida J
中科院分区：
文献类型：
--
作者：
Fukuta M;Nakai Y;Kirino K;Nakagawa M;Sekiguchi K;Nagata S;Matsumoto Y;Yamamoto T;Umeda K;Heike T;Okumura N;Koizumi N;Sato T;Nakahata T;Saito M;Otsuka T;Kinoshita S;Ueno M;Ikeya M;Toguchida J
神经嵴细胞（NCC）是一种胚胎迁移细胞群，具有分化为多种细胞类型的能力，这些细胞类型有助于颅面骨骼，角膜，周围神经系统和皮肤色素沉着。这种能力表明NCC作为基于细胞的治疗来源的有希望的作用。尽管已经使用了几种方法来从人多能干细胞（hPSC）（例如胚胎干细胞（ESC）和诱导的多能干细胞（iPSC））诱导人NCC（hNCC），但是需要进一步的修改来改善这些方法的稳健性、功效和简单性。化学成分确定的培养基（CDM）用作诱导和维持步骤中的基础培养基。通过优化培养条件，GSK 3 β抑制剂和TGF β抑制剂与最小生长因子（胰岛素）的组合非常有效地从hPSC诱导hNCC（70 - 80%）。诱导的hNCC表达颅NCC相关基因，并在补充有EGF和FGF2的CDM中稳定增殖，最多至少10代，而没有观察到主要基因表达谱的变化。外周神经元、神经胶质细胞、黑素细胞和角膜内皮细胞的分化特性得到证实。此外，使用对人MSC特异性的CDM从hNCC诱导具有类似于多能间充质基质细胞（MSC）的分化特征的细胞。我们使用小分子化合物与限定培养基的简单而稳健的诱导方案使得能够产生hNCC作为中间材料，产生用于基于细胞的创新药物的终末分化细胞。
Neural crest cells (NCCs) are an embryonic migratory cell population with the ability to differentiate into a wide variety of cell types that contribute to the craniofacial skeleton, cornea, peripheral nervous system, and skin pigmentation. This ability suggests the promising role of NCCs as a source for cell-based therapy. Although several methods have been used to induce human NCCs (hNCCs) from human pluripotent stem cells (hPSCs), such as embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), further modifications are required to improve the robustness, efficacy, and simplicity of these methods. Chemically defined medium (CDM) was used as the basal medium in the induction and maintenance steps. By optimizing the culture conditions, the combination of the GSK3β inhibitor and TGFβ inhibitor with a minimum growth factor (insulin) very efficiently induced hNCCs (70–80%) from hPSCs. The induced hNCCs expressed cranial NCC-related genes and stably proliferated in CDM supplemented with EGF and FGF2 up to at least 10 passages without changes being observed in the major gene expression profiles. Differentiation properties were confirmed for peripheral neurons, glia, melanocytes, and corneal endothelial cells. In addition, cells with differentiation characteristics similar to multipotent mesenchymal stromal cells (MSCs) were induced from hNCCs using CDM specific for human MSCs. Our simple and robust induction protocol using small molecule compounds with defined media enabled the generation of hNCCs as an intermediate material producing terminally differentiated cells for cell-based innovative medicine.
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