Flow Cytometry-Defined CD49d Expression in Circulating T-Lymphocytes Is a Biomarker for Disease Progression in Duchenne Muscular Dystrophy

  • Wilson Savino
  • Fernanda Pinto-Mariz
  • Vincent Mouly
Part of the Methods in Molecular Biology book series (MIMB, volume 1687)


Duchenne muscular dystrophy (DMD) affects 1:3500–1:5000 male births, and is caused by X-linked mutations in the dystrophin gene, manifested by progressive muscle weakness and wasting due to the absence of dystrophin protein, leading to degeneration of skeletal muscle. DMD patients are clinically heterogeneous and the functional phenotype often cannot be correlated with the genotype. Therefore, defined reliable noninvasive biomarkers aiming at predicting if a given DMD child will progress more or less rapidly will be instrumental to better design inclusion of defined patients for future therapeutic assays. We recently showed that CD49d expression levels in blood-derived T-cell subsets can predict disease progression in DMD patients. Herein we describe in detail the methodology to be applied for defining, through four-color flow cytometry, the membrane expression levels of the CD49d (the α4 chain of the integrins α4β1 and α4β7) in circulating CD4+ and CD8+ T cell subsets. Since we have also shown that this molecule can also be placed as a potential target for therapeutics in DMD, we also describe the cell migration functional assay that can be applied to test potential CD49d inhibitors that can modulate their ability to cross endothelial or extracellular matrix (ECM) barriers.

Key words

Muscular dystrophy Integrins Flow cytometry Inflammation VLA-4 T lymphocytes Biomarker Immunotherapy 



This work was funded by the following institutions or funding agencies: Fiocruz, Faperj, CNPq, Capes (Brazil), Association Française contre les Myopathies (AFM), UPMC, Inserm and CNRS (France); Faperj-Sorbonne Conjoint Research Program (Brazil-France); FOCEM (Mercosur), United Parent’s Project for Muscular Dystrophies, Agence Nationale de Recherche (Genopath INAFIB), MyoAge (EC 7th framework program).


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Copyright information

© Springer Science+Business Media LLC 2018

Authors and Affiliations

  • Wilson Savino
    • 1
    • 2
    • 3
  • Fernanda Pinto-Mariz
    • 4
  • Vincent Mouly
    • 2
    • 3
    • 5
  1. 1.Laboratory on Thymus Research, Oswaldo Cruz InstituteOswaldo Cruz FoundationRio de JaneiroBrazil
  2. 2.Fiocruz-Inserm-UPMC Franco-Brazilian International Laboratory on Cell Therapy and ImmunotherapyPierre and Marie Curie University, Sorbonne UniversitiesParisFrance
  3. 3.Brazilian National Institute of Science and Technology on Neuroimmunomodulation (INCT-NIM)Rio de JaneiroBrazil
  4. 4.Institute of PediatricsFederal University of Rio de JaneiroRio de JaneiroBrazil
  5. 5.Center for Research in MyologySorbonne Universités, UPMC Université Paris 06, INSERM UMRS974ParisFrance

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