PDF(Pubmed)
Abstract：
Fate decisions in the embryo are controlled by a plethora of microenvironmental interactions in a three-dimensional niche. To investigate whether aspects of this microenvironmental complexity can be engineered to direct myogenic human-induced pluripotent stem cell (hiPSC) differentiation, we here screened murine cell types present in the developmental or adult stem cell niche in heterotypic suspension embryoids. We identified embryonic endothelial cells and fibroblasts as highly permissive for myogenic specification of hiPSCs. After two weeks of sequential Wnt and FGF pathway induction, these three-component embryoids are enriched in Pax7-positive embryonic-like myogenic progenitors that can be isolated by flow cytometry. Myogenic differentiation of hiPSCs in heterotypic embryoids relies on a specialized structural microenvironment and depends on MAPK, PI3K/AKT, and Notch signaling. After transplantation in a mouse model of Duchenne muscular dystrophy, embryonic-like myogenic progenitors repopulate the stem cell niche, reactivate after repeated injury, and, compared to adult human myoblasts, display enhanced fusion and lead to increased muscle function. Altogether, we provide a two-week protocol for efficient and scalable suspension-based 3D derivation of Pax7-positive myogenic progenitors from hiPSCs.
摘要：
胚胎中的命运决定由三维生态位中过多的微环境相互作用控制。为了研究这种微环境复杂性的各个方面是否可以被改造为指导生肌诱导的人多能干细胞（hiPSC）分化，我们在这里筛选了异型悬浮胚中发育或成体干细胞小生境中存在的鼠细胞类型。我们将胚胎内皮细胞和成纤维细胞鉴定为高度允许hiPSC的肌源性规范。在连续两周的Wnt和FGF途径诱导后，这些三组分胚状体富含Pax7阳性的胚胎样成肌祖细胞，可通过流式细胞术分离。异型胚状体中hiPSCs的成肌分化依赖于特殊的结构微环境，依赖于MAPK，PI3K/AKT,和Notch信号。在杜氏肌营养不良小鼠模型中移植后，胚胎样肌原祖细胞重新填充干细胞生态位，反复受伤后重新激活，and,与成人成肌细胞相比，显示增强的融合和导致增加的肌肉功能。总之,我们提供了一个为期两周的方案，用于从hiPSC中有效和可扩展的基于悬浮的Pax7阳性肌原祖细胞的3D衍生。
