Albini Sonia, Puri Pier Lorenzo
Muscle Development and Regeneration Program, Sanford-Burnham Institute for Medical Research;
Muscle Development and Regeneration Program, Sanford-Burnham Institute for Medical Research; IRCCS Fondazione Santa Lucia;
J Vis Exp. 2014 Jun 21(88):e51243. doi: 10.3791/51243.
Generation of a homogeneous and abundant population of skeletal muscle cells from human embryonic stem cells (hESCs) is a requirement for cell-based therapies and for a "disease in a dish" model of human neuromuscular diseases. Major hurdles, such as low abundance and heterogeneity of the population of interest, as well as a lack of protocols for the formation of three-dimensional contractile structures, have limited the applications of stem cells for neuromuscular disorders. We have designed a protocol that overcomes these limits by ectopic introduction of defined factors in hESCs--the muscle determination factor MyoD and SWI/SNF chromatin remodeling complex component BAF60C--that are able to reprogram hESCs into skeletal muscle cells. Here we describe the protocol established to generate hESC-derived myoblasts and promote their clustering into tridimensional miniaturized structures (myospheres) that functionally mimic miniaturized skeletal muscles.
从人类胚胎干细胞（hESC）中生成同质且数量丰富的骨骼肌细胞群体，是基于细胞的疗法以及人类神经肌肉疾病“培养皿中的疾病”模型所必需的。主要障碍，如目标细胞群体的低丰度和异质性，以及缺乏形成三维收缩结构的方案，限制了干细胞在神经肌肉疾病中的应用。我们设计了一种方案，通过在hESC中异位引入特定因子——肌肉决定因子MyoD和SWI/SNF染色质重塑复合体组分BAF60C，来克服这些限制，这些因子能够将hESC重编程为骨骼肌细胞。在此，我们描述了所建立的方案，用于生成hESC衍生的成肌细胞，并促进它们聚集成功能上模拟小型化骨骼肌的三维微型结构（肌球）。
