Advertisement

Methods of Isolation and Analysis of TREG Immune Infiltrates from Injured and Dystrophic Skeletal Muscle

  • Dalia Burzyn
  • Amy Wagers
  • Diane Mathis
  • Massimiliano CerlettiEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1899)

Abstract

The immune infiltrate present in acutely injured or dystrophic skeletal muscle has been shown to play an important role in the process of muscle regeneration. Our work has described, for the first time, muscle regulatory T cells (Tregs), a unique population in phenotype and function capable of promoting skeletal muscle repair. Herein, we describe the methods we have optimized to study muscle Tregs, including their isolation from injured muscle, immuno-labeling for analysis/separation by flow cytometry, and measurement of their proliferation status.

Key words

Regulatory T cell Skeletal muscle infiltrate Tissue-resident leukocytes Treg proliferation Skeletal muscle injury Muscular dystrophy 

References

  1. 1.
    Josefowicz SZ, Lu LF, Rudensky AY (2012) Regulatory T cells: mechanisms of differentiation and function. Annu Rev Immunol 30:531–564CrossRefGoogle Scholar
  2. 2.
    Burzyn D, Benoist C, Mathis D (2013) Regulatory T cells in nonlymphoid tissues. Nat Immunol 14:1007–1013CrossRefGoogle Scholar
  3. 3.
    Feuerer M, Herrero L, Cipolletta D, Naaz A, Wong J, Nayer A, Lee J, Goldfine AB, Benoist C, Shoelson S, Mathis D (2009) Lean, but not obese, fat is enriched for a unique population of regulatory T cells that affect metabolic parameters. Nat Med 15:930–939CrossRefGoogle Scholar
  4. 4.
    Shi X, Garry DJ (2006) Muscle stem cells in development, regeneration, and disease. Genes Dev 20:1692–1708CrossRefGoogle Scholar
  5. 5.
    McKinnell IW, Parise G, Rudnicki MA (2005) Muscle stem cells and regenerative myogenesis. Curr Top Dev Biol 71:113–130CrossRefGoogle Scholar
  6. 6.
    Seale P, Asakura A, Rudnicki MA (2001) The potential of muscle stem cells. Dev Cell 1:333–342CrossRefGoogle Scholar
  7. 7.
    Sherwood RI, Christensen JL, Conboy IM, Conboy MJ, Rando TA, Weissman IL, Wagers AJ (2004) Isolation of adult mouse myogenic progenitors: functional heterogeneity of cells within and engrafting skeletal muscle. Cell 119:543–554CrossRefGoogle Scholar
  8. 8.
    Montarras D, Morgan J, Collins C, Relaix F, Zaffran S, Cumano A, Partridge TA, Buckingham M (2005) Direct isolation of satellite cells for skeletal muscle regeneration. Science 309:2064–2067CrossRefGoogle Scholar
  9. 9.
    Collins CA, Olsen I, Zammit PS, Heslop L, Petrie A, Partridge TA, Morgan JE (2005) Stem cell function, self-renewal, and behavioral heterogeneity of cells from the adult muscle satellite cell niche. Cell 122:289–301CrossRefGoogle Scholar
  10. 10.
    Cerletti M, Jurga S, Witczak CA, Hirshman MF, Shadrach JL, Goodyear LJ, Wagers AJ (2008) Highly efficient, functional engraftment of skeletal muscle stem cells in dystrophic muscles. Cell 134:37–47CrossRefGoogle Scholar
  11. 11.
    Sacco A, Doyonnas R, Kraft P, Vitorovic S, Blau HM (2008) Self-renewal and expansion of single transplanted muscle stem cells. Nature 456:502–506CrossRefGoogle Scholar
  12. 12.
    Tidball JG, Villalta SA (2010) Regulatory interactions between muscle and the immune system during muscle regeneration. Am J Physiol Regul Integr Comp Physiol 298:1173–1187CrossRefGoogle Scholar
  13. 13.
    Arnold L, Henry A, Poron F, Baba-Amer Y, Van Rooijen N, Plonquet A, Gherardi RK, Chazaud B (2007) Inflammatory monocytes recruited after skeletal muscle injury switch into anti-inflammatory macrophages to support myogenesis. J Exp Med 204:1057–1069CrossRefGoogle Scholar
  14. 14.
    Burzyn D, Kuswanto W, Kolodin D, Shadrach JL, Cerletti M, Jang Y, Sefik E, Tan TG, Wagers AJ, Benoist C, Mathis D (2013) A special population of regulatory T cells potentiates muscle repair. Cell 155:12821895CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Dalia Burzyn
    • 1
  • Amy Wagers
    • 2
  • Diane Mathis
    • 1
  • Massimiliano Cerletti
    • 3
    Email author
  1. 1.Division of Immunology, Department of Microbiology and ImmunobiologyHarvard Medical SchoolBostonUSA
  2. 2.Department of Stem Cell and Regenerative BiologyHarvard Stem Cell Institute, Harvard UniversityCambridgeUSA
  3. 3.UCL Research Department for Surgical Biotechnologies & Institute for Immunity and TransplantationUniversity College LondonLondonUK

Personalised recommendations