Isolation, Culture, and Differentiation of Fibro/Adipogenic Progenitors (FAPs) from Skeletal Muscle.

Fibro/Adipogenic Progenitors (FAPs) are a multipotent progenitor population resident in skeletal muscle. During development and regeneration, FAPs provide trophic support to myogenic progenitors that is required for muscle fiber maturation and specification. FAPs also represent a major cellular source of fibrosis in degenerative disease states, highlighting them as a potential cellular target for anti-fibrotic muscle therapies. Effective and reproducible methods to isolate and culture highly purified FAP populations are therefore critical to further understand their biology. Here, we describe a fluorescent activated cell sorting (FACS) based protocol to isolate CD31-/CD45-/Integrin-α7-/Sca1+ FAPs from murine skeletal muscle including details of tissue collection and enzymatic muscle digestion. We also incorporate optimized methods of expanding and differentiated FAPs in vitro. Together, this protocol provides a complete workflow to study skeletal muscle derived FAPs and compliments downstream analytical, drug screening, and disease modeling applications.