Title

Muscle cells of sporadic amyotrophic lateral sclerosis patients secrete neurotoxic vesicles

Authors

Laura Le Gall, Northern Ireland Center for Stratified Medicine, Biomedical Sciences Research Institute, Londonderry, UK.
William J. Duddy, Northern Ireland Center for Stratified Medicine, Biomedical Sciences Research Institute, Londonderry, UK.
Cecile Martinat, I-Stem, INSERM/UEVE UMR 861, I-STEM, AFM, Paris, France.
Virginie Mariot, NIHR Biomedical Research Centre, University College London, Great Ormond Street Institute of Child Health and Great Ormond Street Hospital NHS Trust, London, UK.
Owen Connolly, Northern Ireland Center for Stratified Medicine, Biomedical Sciences Research Institute, Londonderry, UK.
Vanessa Milla, Northern Ireland Center for Stratified Medicine, Biomedical Sciences Research Institute, Londonderry, UK.
Ekene Anakor, Northern Ireland Center for Stratified Medicine, Biomedical Sciences Research Institute, Londonderry, UK.
Zamalou G. Ouandaogo, Sorbonne Université, Institut National de la Santé et de la Recherche Médicale, Association Institut de Myologie, Centre de Recherche en Myologie, Paris, France.
Stephanie Millecamps, Inserm U1127, CNRS UMR7225, Sorbonne Universités, UPMC Univ, Paris, France.
Jeanne Lainé, Sorbonne Université, Institut National de la Santé et de la Recherche Médicale, Association Institut de Myologie, Centre de Recherche en Myologie, Paris, France.
Udaya Geetha Vijayakumar, Northern Ireland Center for Stratified Medicine, Biomedical Sciences Research Institute, Londonderry, UK.
Susan Knoblach, Genetic Medicine, Children's National Medical Center, George Washington University, Washington, DC, USA.
Cedric Raoul, The Neuroscience Institute of Montpellier, Inserm UMR1051, Univ Montpellier, Saint Eloi Hospital, Montpellier, France.
Olivier Lucas, The Neuroscience Institute of Montpellier, Inserm UMR1051, Univ Montpellier, Saint Eloi Hospital, Montpellier, France.
Jean Philippe Loeffler, Mécanismes Centraux et Périphériques de la Neurodégénérescence, Université de Strasbourg, INSERM UMR_S 1118, Strasbourg, France.
Peter Bede, Computational Neuroimaging Group, Academic Unit of Neurology, Trinity College Dublin, Dublin, Ireland.
Anthony Behin, APHP, Centre de Référence des Maladies Neuromusculaires Nord/Est/Ile de France, Institut de Myologie, Hôpital Pitié-Salpêtrière, Paris, France.
Helene Blasco, Laboratoire de Biochimie et Biologie Moléculaire, Hôpital Bretonneau, CHRU de Tours, Tours, France.
Gaelle Bruneteau, Sorbonne Université, Institut National de la Santé et de la Recherche Médicale, Association Institut de Myologie, Centre de Recherche en Myologie, Paris, France.
Maria Del Mar Amador, APHP, Département de Neurologie, Hôpital Pitié-Salpêtrière, Centre référent SLA, Paris, France.
David Devos, INSERM U1171, Pharmacologie Médicale & Neurologie Université, Faculté de Médecine, CHU de Lille, Lille, France.
Alexandre Henriques, Mécanismes Centraux et Périphériques de la Neurodégénérescence, Université de Strasbourg, INSERM UMR_S 1118, Strasbourg, France.
Adele Hesters, APHP, Département de Neurologie, Hôpital Pitié-Salpêtrière, Centre référent SLA, Paris, France.
Lucette Lacomblez, CNRS, INSERM, Laboratoire d'Imagerie Biomédicale, Sorbonne Université, Paris, France.
Pascal Laforet, Département de Neurologie, Centre de Référence Maladies Neuromusculaires Paris-Est, Hôpital Raymond-Poincaré, Garches, France.
Timothee Langlet, APHP, Département de Neurologie, Hôpital Pitié-Salpêtrière, Centre référent SLA, Paris, France.
Pascal Leblanc, Laboratory of Molecular Biology of the Cell, Ecole Normale Supérieure de Lyon, Lyon, France.
Nadine Le Forestier, APHP, Département de Neurologie, Hôpital Pitié-Salpêtrière, Centre référent SLA, Paris, France.
Thierry Maisonobe, APHP, Département de Neurologie, Hôpital Pitié-Salpêtrière, Centre référent SLA, Paris, France.
Vincent Meininger, Hôpital des Peupliers, Ramsay Générale de Santé, Paris, France.
Laura Robelin, Mécanismes Centraux et Périphériques de la Neurodégénérescence, Université de Strasbourg, INSERM UMR_S 1118, Strasbourg, France.
Francois Salachas, APHP, Département de Neurologie, Hôpital Pitié-Salpêtrière, Centre référent SLA, Paris, France.

Document Type

Journal Article

Publication Date

2-22-2022

Journal

Journal of cachexia, sarcopenia and muscle

DOI

10.1002/jcsm.12945

Keywords

Cell-cell communication; MND; Secreted vesicles; sporadic ALS

Abstract

BACKGROUND: The cause of the motor neuron (MN) death that drives terminal pathology in amyotrophic lateral sclerosis (ALS) remains unknown, and it is thought that the cellular environment of the MN may play a key role in MN survival. Several lines of evidence implicate vesicles in ALS, including that extracellular vesicles may carry toxic elements from astrocytes towards MNs, and that pathological proteins have been identified in circulating extracellular vesicles of sporadic ALS patients. Because MN degeneration at the neuromuscular junction is a feature of ALS, and muscle is a vesicle-secretory tissue, we hypothesized that muscle vesicles may be involved in ALS pathology. METHODS: Sporadic ALS patients were confirmed to be ALS according to El Escorial criteria and were genotyped to test for classic gene mutations associated with ALS, and physical function was assessed using the ALSFRS-R score. Muscle biopsies of either mildly affected deltoids of ALS patients (n = 27) or deltoids of aged-matched healthy subjects (n = 30) were used for extraction of muscle stem cells, to perform immunohistology, or for electron microscopy. Muscle stem cells were characterized by immunostaining, RT-qPCR, and transcriptomic analysis. Secreted muscle vesicles were characterized by proteomic analysis, Western blot, NanoSight, and electron microscopy. The effects of muscle vesicles isolated from the culture medium of ALS and healthy myotubes were tested on healthy human-derived iPSC MNs and on healthy human myotubes, with untreated cells used as controls. RESULTS: An accumulation of multivesicular bodies was observed in muscle biopsies of sporadic ALS patients by immunostaining and electron microscopy. Study of muscle biopsies and biopsy-derived denervation-naïve differentiated muscle stem cells (myotubes) revealed a consistent disease signature in ALS myotubes, including intracellular accumulation of exosome-like vesicles and disruption of RNA-processing. Compared with vesicles from healthy control myotubes, when administered to healthy MNs the vesicles of ALS myotubes induced shortened, less branched neurites, cell death, and disrupted localization of RNA and RNA-processing proteins. The RNA-processing protein FUS and a majority of its binding partners were present in ALS muscle vesicles, and toxicity was dependent on the expression level of FUS in recipient cells. Toxicity to recipient MNs was abolished by anti-CD63 immuno-blocking of vesicle uptake. CONCLUSIONS: ALS muscle vesicles are shown to be toxic to MNs, which establishes the skeletal muscle as a potential source of vesicle-mediated toxicity in ALS.

Department

Genomics and Precision Medicine

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