Satellite cells from dystrophic (Mdx) mice display accelerated differentiation in primary cultures and in isolated myofibers

Abstract

In the dystrophic (mdx) mouse, skeletal muscle undergoes cycles of degeneration and regeneration, and myogenic progenitors (satellite cells) show ongoing proliferation and differentiation at a time when counterpart cells in normal healthy muscle enter quiescence. However, it remains unclear whether this enhanced satellite cell activity is triggered solely by the muscle environment or is also governed by factors inherent in satellite cells. To obtain a better picture of myogenesis in dystrophic muscle, a direct cell-by-cell analysis was performed to compare satellite cell dynamics from mdx and normal (C57Bl/10) mice in two cell culture models. In one model, the kinetics of satellite cell differentiation was quantified in primary cell cultures from diaphragm and limb muscles by immunodetection of MyoD, myogenin, and MEF2. In mdx cell cultures, myogenin protein was expressed earlier than normal and was followed more rapidly by dual myogenin/MEF2A expression and myotube formation. In the second model, the dynamics of satellite cell myogenesis were investigated in cultured myofibers isolated from flexor digitorum brevis (FDB) muscle, which retain satellite cells in the native position. Consistent with primary cultures, satellite cells in mdx myofibers displayed earlier myogenin expression, as well as an enhanced number of myogenin-expressing satellite cells per myofiber compared to normal. The addition of fibroblast growth factor 2 (FGF2) led to an increase in the number of satellite cells expressing myogenin in normal and mdx myofibers. However, the extent of the FGF effect was more robust in mdx myofibers. Notably, many myonuclei in mdx myofibers were centralized, evidence of segmental regeneration; all central nuclei and many peripheral nuclei in mdx myofibers were positive for MEF2A. Results indicated that myogenic cells in dystrophic muscle display accelerated differentiation. Furthermore, the study demonstrated that FDB myofibers are an excellent model of the in vivo state of muscle, as they accurately represented the dystrophic phenotype. Developmental Dynamics 235:203–212, 2006.