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September 24, 2019

A stem-cell telephone for neurons to call muscle

The Megumu Saito laboratory reports a new method to cheaply and easily construct the neuromuscular junction from human iPS cells

The neuromuscular junction (NMJ) describes the point where neurons and muscles communicate to control our movements. Many diseases that kill neurons or muscle are the result of defective NMJ, leaving the neurons and muscles unused and to their eventual death. To study the normal development and diseases of the NMJ, CiRA Associate Professor Megumu Saito and colleagues show in a new study a relatively simple approach using iPS cells.

The NMJ describes a structure of motor neurons, myotubes, and Schwann cells. Motor neurons relay a message to the myotubes to contract or relax, while Schwann cells enhance the transmission of the signal and stabilize the NMJ.

"To build NMJs in the lab, scientists will coculture motor neurons and skeletal muscle cells, but this is technically difficult and time-consuming. We want a method in which we can use only iPS cells to construct the NMJ," says Saito.

To generate NMJs from human iPS cells, Saito's team exposed the cells to different differentiation cocktails; the first promoted the development of myotubes, and the second, implemented 10 days later, induced the development of motor neurons and Schwann cells.

The iPS cell-derived NMJs showed the gap junctions and acetylcholine receptors typical of normal NMJs. Furthermore, there was evidence that the development of the myotubes and neurons depended on the development of the other, implying each cell type secreted factors that benefited the other cell type.

One concern about NMJs made in the laboratory, however, is maturation, as stem cell-derived structures are generally immature compared to the structures found in the body. NMJs behave differently in a fetus than they do in an adult.

"The acetylcholine receptors shifted from gamma to epsilon type, which is an important hallmark of adult NMJs. Plus, the morphology of the NMJs including neuron density and synapse elimination suggested adult NMJs," observed Saito.

These changes were accompanied by myotube contractions that correlated with the control of calcium oscillations by the acetylcholine receptors.

To test whether their NMJs could be used to study disease, Dr. Chuang-Yu Lin, a scientist in the Saito lab, suppressed the expression of the gene SMN in iPS cells. Mutations in SMN are attributed to spinal muscular atrophy, a pediatric disease that is characterized by a loss of motor neurons and eventual death. Although classically spinal cord atrophy has been considered a disease of motor neurons, Saito's group had previously shown that the disease first begins in the NMJ.

"We found many characteristics consistent with spinal muscular atrophy. The area of the NMJ was much smaller, the mitochondria morphology had changed, and the area of contractions was smaller," Chuang-Yu says,

Saito notes that the easier construction of the NMJs in the new study has other benefits besides studying disease.

"Because our NMJs are easy to prepare and show the patterns of normal NMJ development, we believe they can be used to study related diseases and experimental drugs," he says.

Paper Details
  • Journal: JCI Insight
  • Title: iPSC-derived functional human neuromuscular junctions model pathophysiology of neuromuscular diseases
  • Authors: Chuang-Yu Lin1, Michiko Yoshida1,2, Li-Tzu Li3, Akihiro Ikenaka1, Shiori Oshima4, Kazuhiro Nakagawa4, Hidetoshi Sakurai1, Eriko Matsui4, Tatsutoshi Nakahata1 and Megumu K. Saito1
  • Author Affiliations:
    1. Department of Clinical Application, Center for iPS Cell Research and Application, Kyoto University, Kyoto, Japan
    2. Department of Pediatrics, Kyoto Prefectural University of Medicine, Kyoto, Japan
    3. Institute of Atomic and Molecular Sciences, Academia Sinica, Taipei, Taiwan
    4. Sony Imaging Products & Solutions Inc., Tokyo, Japan
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