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Induced Formation and Maturation of Acetylcholine Receptor Clusters in a Defined 3D Bio-Artificial Muscle

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Abstract

Dysfunction of the neuromuscular junction is involved in a wide range of muscular diseases. The development of neuromuscular junction through which skeletal muscle is innervated requires the functional modulation of acetylcholine receptor (AchR) clustering on myofibers. However, studies on AchR clustering in vitro are mostly done on monolayer muscle cell culture, which lacks a three-dimensional (3D) structure, a prominent limitation of the two-dimensional (2D) system. To enable a better understanding on the structure–function correlation underlying skeletal muscle innervation, a muscle system with a well-defined geometry mimicking the in vivo muscular setting is needed. Here, we report a 3D bio-artificial muscle (BAM) bioengineered from green fluorescent protein-transduced C3H murine myoblasts as a novel in vitro tissue-based model for muscle innervation studies. Our cell biological and molecular analysis showed that this BAM is structurally similar to in vivo muscle tissue and can reach the perinatal differentiation stage, higher than does 2D culture. Effective clustering and morphological maturation of AchRs on BAMs induced by agrin and laminin indicate the functional activity and plasticity of this BAM system toward innervation. Taken together, our results show that the BAM provides a favorable 3D environment that at least partially recapitulates real physiological skeletal muscle with regard to innervation. With a convenience of fabrication and manipulation, this 3D in vitro system offers a novel model for studying mechanisms underlying skeletal muscle innervation and testing therapeutic strategies for relevant nervous and muscular diseases.

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Acknowledgments

We are grateful to Dr. Justin Fallon for providing mouse agrin. This research was supported by NIH grants R41 AR053386 and R43 AG029705. The authors confirm that no competing financial interests exist, and there has been no financial support for this research could have influenced its outcome.

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Authors and Affiliations

  1. Medical Research Center, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China

    Lin Wang

  2. Center for Tissue Engineering and Regenerative Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China

    Lin Wang

  3. Department of Molecular Pharmacology, Physiology and Biotechnology, Brown University, Providence, RI, USA

    Lin Wang

  4. Department of Pathology and Laboratory Medicine, Miriam Hospital, Providence, RI, USA

    Janet Shansky & Herman Vandenburgh

  5. Department of Pathology and Laboratory Medicine, Brown University, Providence, RI, USA

    Herman Vandenburgh

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  1. Lin Wang
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  2. Janet Shansky
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  3. Herman Vandenburgh
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Correspondence to Herman Vandenburgh.

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Wang, L., Shansky, J. & Vandenburgh, H. Induced Formation and Maturation of Acetylcholine Receptor Clusters in a Defined 3D Bio-Artificial Muscle. Mol Neurobiol 48, 397–403 (2013). https://doi.org/10.1007/s12035-013-8412-z

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  • DOI: https://doi.org/10.1007/s12035-013-8412-z

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