Derivation and Characterization of Immortalized Human Muscle Satellite Cell Clones from Muscular Dystrophy Patients and Healthy Individuals
Jimmy Massenet,
Cyril Gitiaux,
Mélanie Magnan,
Sylvain Cuvellier,
Arnaud Hubas,
Patrick Nusbaum,
F Jeffrey Dilworth,
Isabelle Desguerre,
Bénédicte Chazaud
Affiliations
Jimmy Massenet
Institut NeuroMyoGène, Université Claude Bernard Lyon 1, Université de Lyon, CNRS 5310, INSERM U1217, 69008 Lyon, France
Cyril Gitiaux
Institut Cochin, INSERM U1016, CNRS UMR8104, Université Paris Descartes, Université Sorbonne Paris Cité, 75014 Paris, France
Mélanie Magnan
Institut Cochin, INSERM U1016, CNRS UMR8104, Université Paris Descartes, Université Sorbonne Paris Cité, 75014 Paris, France
Sylvain Cuvellier
Institut Cochin, INSERM U1016, CNRS UMR8104, Université Paris Descartes, Université Sorbonne Paris Cité, 75014 Paris, France
Arnaud Hubas
Laboratoire de Culture Cellulaire, Service de Génétique et Biologie Moléculaires—Hôpital Cochin, APHP.5, Assistance Publique-Hôpitaux de Paris, 75014 Paris, France
Patrick Nusbaum
Laboratoire de Culture Cellulaire, Service de Génétique et Biologie Moléculaires—Hôpital Cochin, APHP.5, Assistance Publique-Hôpitaux de Paris, 75014 Paris, France
F Jeffrey Dilworth
Sprott Center for Stem Cell Research, Ottawa Hospital Research Institute; Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, ON K1H 8M5, Canada
Isabelle Desguerre
Institut Cochin, INSERM U1016, CNRS UMR8104, Université Paris Descartes, Université Sorbonne Paris Cité, 75014 Paris, France
Bénédicte Chazaud
Institut NeuroMyoGène, Université Claude Bernard Lyon 1, Université de Lyon, CNRS 5310, INSERM U1217, 69008 Lyon, France
In Duchenne muscular dystrophy (DMD) patients, absence of dystrophin causes muscle wasting by impacting both the myofiber integrity and the properties of muscle stem cells (MuSCs). Investigation of DMD encompasses the use of MuSCs issued from human skeletal muscle. However, DMD-derived MuSC usage is restricted by the limited number of divisions that human MuSCs can undertake in vitro before losing their myogenic characteristics and by the scarcity of human material available from DMD muscle. To overcome these limitations, immortalization of MuSCs appears as a strategy. Here, we used CDK4/hTERT expression in primary MuSCs and we derived MuSC clones from a series of clinically and genetically characterized patients, including eight DMD patients with various mutations, four congenital muscular dystrophies and three age-matched control muscles. Immortalized cultures were sorted into single cells and expanded as clones into homogeneous populations. Myogenic characteristics and differentiation potential were tested for each clone. Finally, we screened various promoters to identify the preferred gene regulatory unit that should be used to ensure stable expression in the human MuSC clones. The 38 clonal immortalized myogenic cell clones provide a large collection of controls and DMD clones with various genetic defects and are available to the academic community.
