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Neuromuscular junction formation in tissue-engineered skeletal muscle augments contractile function and improves cytoskeletal organization

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posted on 24.09.2015 by Neil Martin, Samantha L. Passey, Darren Player, Vivek Mudera, Keith Baar, Linda Greensmith, Mark Lewis
Neuromuscular and neurodegenerative diseases are conditions that affect both motor neurons and the underlying skeletal muscle tissue. At present, the majority of neuromuscular research utilizes animal models and there is a growing need to develop novel methodologies that can be used to help understand and develop treatments for these diseases. Skeletal muscle tissue-engineered constructs exhibit many of the characteristics of the native tissue such as accurate fascicular structure and generation of active contractions. However, to date, there has been little consideration toward the integration of engineered skeletal muscle with motor neurons with the aim of neuromuscular junction (NMJ) formation, which would provide a model to investigate neuromuscular diseases and basic biology. In the present work we isolated primary embryonic motor neurons and neonatal myoblasts from Sprague-Dawley rats, and cocultured the two cell types in three-dimensional tissue-engineered fibrin hydrogels with the aim of NMJ formation. Immunohistochemistry revealed myotube formation in a fascicular arrangement and neurite outgrowth from motor neuron cell bodies toward the aligned myotubes. Furthermore, colocalization of pre- and postsynaptic proteins and chemical inhibition of spontaneous myotube twitch indicated the presence of NMJs in the innervated constructs. When electrical field stimulation was employed to evoke isometric contractions, maximal twitch and tetanic force were higher in the constructs cocultured with motor neurons, which may, in part, be explained by improved myotube cytoskeletal organization in these constructs. The fabrication of such constructs may be useful tools for investigating neuromuscular pharmaceuticals and improving the understanding of neuromuscular pathologies.

Funding

This work was funded through the National Center for Replacement, Refinement, and Reduction (NC3R’s) of Animals in Research (Grant No. G0900762).

History

School

  • Sport, Exercise and Health Sciences

Published in

Tissue Engineering Part A: Tissue Engineering

Volume

21

Issue

19-20

Pages

? - ? (10)

Citation

MARTIN, N.R.W. ... et al., 2015. Neuromuscular junction formation in tissue-engineered skeletal muscle augments contractile function and improves cytoskeletal organization. Tissue Engineering Part A, 21 (19-20), pp. 2595-2604.

Publisher

© N.R.W. Martin et al. 2015, Published by Mary Ann Liebert, Inc.

Version

VoR (Version of Record)

Publisher statement

This work is made available according to the conditions of the Creative Commons Attribution-NonCommercial 4.0 International (CC BY 4.0) licence. Full details of this licence are available at: http://creativecommons.org/licenses/by/4.0/

Publication date

2015

Notes

This Open Access article is distributed under the terms of the Creative Commons Attribution Noncommercial License (http://creativecommons.org/licenses/by/4.0/) which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author(s) and the source are credited.

ISSN

1937-335X

Language

en

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