Isolation of Skeletal Muscle Stem Cells for Phenotypic Screens for Modulators of Proliferation

Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1787)

Abstract

Adult skeletal muscle contains a population of resident stem cells known as muscle stem cells (MuSC) or satellite cells. This population of cells is required for regeneration of functional myofibers after damage. Aging reduces the proliferative response of satellite cells post-injury. This deficient response is thought to contribute to slowed recovery of muscle function after damage in the elderly and may also contribute to age-related loss of muscle function (sarcopenia). Numerous techniques are now available for the isolation of highly purified satellite cells from mice and humans (Sherwood, et al. Cell 119:543–554, 2004; Cerletti, et al. Cell 134:37–47; 2008; Conboy, et al. Methods Mol Biol 621:165–173, 2010; Bareja, et al. PLoS One 9:e90398; 2014; Castiglioni et al. Stem Cell Rep 2:92–106, 2014; Charville, et al. Stem Cell Rep 5:621–632, 2015; Liu et al. Nat Protoc 10:1612–1624, 2015; Sincennes et al. Methods Mol Biol 1556:41–50, 2017), thus opening an opportunity to use satellite cells in phenotypic screens for regulators of satellite cell proliferation and differentiation. In this chapter, we describe a technique for the prospective isolation of mouse satellite cells that we have recently used in a phenotypic screen of a focused set of small molecules.

Key words

Skeletal muscle Satellite cells Cell sorting Screen Proliferation Differentiation Stem cells Aging 

References

  1. 1.
    Sherwood RI, Christensen JL, Conboy IM, Conboy MJ, Rando TA, Weissman IL et al (2004) Isolation of adult mouse myogenic progenitors: functional heterogeneity of cells within and engrafting skeletal muscle. Cell 119:543–554CrossRefGoogle Scholar
  2. 2.
    Cerletti M, Jurga S, Witczak CA, Hirshman MF, Shadrach JL, Goodyear LJ et al (2008) Highly efficient, functional engraftment of skeletal muscle stem cells in dystrophic muscles. Cell 134:37–47CrossRefGoogle Scholar
  3. 3.
    Conboy MJ, Cerletti M, Wagers AJ, Conboy IM (2010) Immuno-analysis and FACS sorting of adult muscle fiber-associated stem/precursor cells. Methods Mol Biol 621:165–173CrossRefGoogle Scholar
  4. 4.
    Bareja A, Holt JA, Luo G, Chang C, Lin J, Hinken AC et al (2014) Human and mouse skeletal muscle stem cells: convergent and divergent mechanisms of myogenesis. PLoS One 9:e90398CrossRefGoogle Scholar
  5. 5.
    Castiglioni A, Hettmer S, Lynes MD, Rao TN, Tchessalova D, Sinha I et al (2014) Isolation of progenitors that exhibit myogenic/osteogenic bipotency in vitro by fluorescence-activated cell sorting from human fetal muscle. Stem Cell Rep 2:92–106CrossRefGoogle Scholar
  6. 6.
    Charville GW, Cheung TH, Yoo B, Santos PJ, Lee GK, Shrager JB et al (2015) Ex vivo expansion and in vivo self-renewal of human muscle stem cells. Stem Cell Rep 5:621–632CrossRefGoogle Scholar
  7. 7.
    Liu L, Cheung TH, Charville GW, Rando TA (2015) Isolation of skeletal muscle stem cells by fluorescence-activated cell sorting. Nat Protoc 10:1612–1624CrossRefGoogle Scholar
  8. 8.
    Sincennes MC, Wang YX, Rudnicki MA (2017) Primary mouse myoblast purification using magnetic cell separation. Methods Mol Biol 1556:41–50CrossRefGoogle Scholar
  9. 9.
    Mauro A (1961) Satellite cell of skeletal muscle fibers. J Biophys Biochem Cytol 9:493–495CrossRefGoogle Scholar
  10. 10.
    Billin AN, Bantscheff M, Drewes G, Ghidelli-Disse S, Holt JA, Kramer HF et al (2016) Discovery of novel small molecules that activate satellite cell proliferation and enhance repair of damaged muscle. ACS Chem Biol 11:518–529CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.GlaxoSmithKline, Muscle Metabolism Discovery Performance UnitKing of PrussiaUSA
  2. 2.Gilead Sciences, Inc.Foster CityUSA

Personalised recommendations