Abstract
The characterization of the membrane repair machinery in human skeletal muscle has become crucial, since it has been shown that some muscular dystrophies result from a defect of this fundamental physiological process. Deciphering membrane repair mechanism requires the development of methodologies allowing studying the response of skeletal muscle cells to sarcolemma damage and identifying candidate proteins playing a role in the membrane repair machinery. Here, we describe a protocol that is based on the creation of cell membrane disruption by infrared laser irradiation in human myotubes. Membrane disruption and repair are assayed by monitoring the incorporation into myotubes of the membrane probe FM1-43. This methodology has recently enabled us to show that Annexin-A5 is required for membrane repair in human skeletal muscle cells (Carmeille et al., Biochim Biophys Acta 1863:2267–2279, 2016).
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Acknowledgment
This work was supported by the AFM-Telethon (Research program 17140 to A.B..). The platform for immortalization of human cells of the Center of Research in Myology (Paris, France) is acknowledged for the gift of human immortalized LHCN-M2 myoblasts. The expert help of Christel Poujol, Philippe Legros, Laure Malicieux, Sébastien Marais, and Fabrice Cordelières at the Bordeaux Imaging Center, a service unit of the CNRS-INSERM and Bordeaux University, member of the national infrastructure France BioImaging, is acknowledged.
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Carmeille, R., Croissant, C., Bouvet, F., Bouter, A. (2017). Membrane Repair Assay for Human Skeletal Muscle Cells. In: Ryall, J. (eds) Skeletal Muscle Development. Methods in Molecular Biology, vol 1668. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7283-8_14
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DOI: https://doi.org/10.1007/978-1-4939-7283-8_14
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