[Development of methodology for isolation and culture of mesenchymal stem cells derived from mouse skeletal muscle]

Abstract

To address the question whether there exists mesenchymal stem cells in adult mouse skeletal muscle, mononuclear cells from muscle were obtained by digestion and density gradient centrifugation and plated in alpha-MEM/F12 medium containing 10% fetal bovine serum. Cell biological properties including morphology, cytochemistry, growth pattern and phenotypes as well were evaluated. Likewise, the osteogenesis of cultured cells was also observed. The results showed that adherent cells homogenous in shape proliferated quickly in the culture system. The phenotypes of the cells were unique, which were positive for CD29 and Sca-1, and negative for CD34 and CD45. Cytochemistry evaluation showed that they were homogeneously positive for acid alpha-naphthl acetate esterase (ANAE) and acid phosphatase (ACP), and negative for alkaline phosphatase (ALP) and that, around 5% of them were positive for glycogen (periodic acid-Schiff reaction, PAS). Cells became ALP-positive after the induction by ascorbic acid, beta-phosphoglycerol and dexamethasone. It is concluded that mesenchymal stem cells exist in murine skeletal muscle and compose the complex heterogenous population of stem cells in muscle.