In vitro Neuromuscular Junction Induced from Human Induced Pluripotent Stem Cells
In vitro Neuromuscular Junction Induced from Human Induced Pluripotent Stem Cells
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DOI：
10.3791/61396
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发表时间：
2020-12-01
影响因子：
1.2
通讯作者：
Saito, Megumu K.
中科院分区：
文献类型：
--
作者：
Lin, Chuang-Yu;Yoshida, Michiko;Saito, Megumu K.
神经肌肉接头（NMJ）是一种专门的突触，将动作电位从运动神经元传递到骨骼肌进行机械运动。NMJ结构的构造影响神经元、肌肉的功能以及它们之间的相互作用。以前的研究报道了许多策略，通过共培养运动神经元和肌管，以产生NMJ在体外诱导过程复杂，培养时间长，但一直难以重现成熟的NMJ的形态和功能。我们的体外NMJ诱导系统通过在单个培养皿中分化人iPSC来构建。通过切换肌源性和神经源性诱导培养基进行诱导，在一个月的培养中，所得的NMJ含有突触前和突触后成分，包括运动神经元、骨骼肌和雪旺细胞。NMJ的功能测定也表明，Ca ~（++）可触发肌管收缩，而乙酰胆碱受体（AChR）抑制剂箭毒（curare）则可抑制肌管收缩，其中刺激信号通过NMJ传递。这种简单而稳健的方法成功地推导出了具有功能连接性的NMJ的复杂结构。这种体外人NMJ具有完整的结构和功能，对于研究病理机制和化合物筛选具有很大的潜力。
The neuromuscular junction (NMJ) is a specialized synapse that transmits action potentials from the motor neuron to skeletal muscle for mechanical movement. The architecture of the NMJ structure influences the functions of the neuron, the muscle and the mutual interaction. Previous studies have reported many strategies by co-culturing the motor neurons and myotubes to generate NMJ in vitro with complex induction process and long culture period but have struggled to recapitulate mature NMJ morphology and function. Our in vitro NMJ induction system is constructed by differentiating human iPSC in a single culture dish. By switching the myogenic and neurogenic induction medium for induction, the resulting NMJ contained pre- and postsynaptic components, including motor neurons, skeletal muscle and Schwann cells in the one month culture. The functional assay of NMJ also showed that the myotubes contraction can be triggered by Ca++ then inhibited by curare, an acetylcholine receptor (AChR) inhibitor, in which the stimulating signal is transmitted through NMJ. This simple and robust approach successfully derived the complex structure of NMJ with functional connectivity. This in vitro human NMJ, with its integrated structures and function, has promising potential for studying pathological mechanisms and compound screening.