Article

Skeletal muscle satellite cells: Background and methods for isolation and analysis in a primary culture system

Department of Biological Structure, School of Medicine, University of Washington, Seattle, WA, USA.
Methods in molecular biology (Clifton, N.J.) (Impact Factor: 1.29). 01/2012; 798:21-52. DOI: 10.1007/978-1-61779-343-1_2
Source: PubMed

ABSTRACT Repair of adult skeletal muscle depends on satellite cells, myogenic stem cells located between the basal lamina and the plasmalemma of the myofiber. Standardized protocols for the isolation and culture of satellite cells are key tools for understanding cell autonomous and extrinsic factors that regulate their performance. Knowledge gained from such studies can contribute important insights to developing strategies for the improvement of muscle repair following trauma and in muscle wasting disorders. This chapter provides an introduction to satellite cell biology and further describes the basic protocol used in our laboratory to isolate and culture satellite cells from adult skeletal muscle. The cell culture conditions detailed herein support proliferation and differentiation of satellite cell progeny and the development of reserve cells, which are thought to reflect the in vivo self-renewal ability of satellite cells. Additionally, this chapter describes our standard immunostaining protocol that allows the characterization of satellite cell progeny by the temporal expression of characteristic transcription factors and structural proteins associated with different stages of myogenic progression. Although emphasis is given here to the isolation and characterization of satellite cells from mouse hindlimb muscles, the protocols are suitable for other muscle types (such as diaphragm and extraocular muscles) and for muscles from other species, including chicken and rat. Altogether, the basic protocols described are straightforward and facilitate the study of diverse aspects of skeletal muscle stem cells.

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    • "This is done, for example, by seeding a heterogeneous cell suspension on a common un-functionalized glass or plastic substrates. Under these conditions some cell types adhere to the bare substrate while others remain in suspension for a longer period of time (see for Danoviz and Yablonka-Reuveni, 2012). The selective cells-adhesivity reflects differences in the nature and rates of ECM secretion and CAM expression. "
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    • "Upon injury, SatCs are activated and migrate to the wound, proliferate, differentiate, and form new myofibers or repair damaged ones [18]. SatCs are located between the basal lamina and the plasma membrane [17], [19], and express the transcription factor Pax7 [20], [21]. A distinct gene expression profile characterizes the SatC progeny [22], [23]. "
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