Skeletal muscle cell populations. Separation and partial characterization of fibroblast-like cells from embryonic tissue using density centrifugation.
Cell suspensions from the breast muscles of 10-day old chicken embryos were separated into non-myogenic, fibroblast-like cell fractions and a mononucleated, myogenic cell fraction by Percoll density centrifugation. Isolated populations were characterized by their morphology in both mass cultures and individual macroscopic clones and by the immunocytochemical detection of skeletal muscle- and smooth muscle-specific proteins in individual cells. Cell populations were also characterized by their protein patterns using sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The less dense, non-myogenic cells comprised 16% of the cells. In culture they were predominantly flattened, stellate cells and gave rise to clones lacking myotubes. These fibroblast-like cells were negative for skeletal muscle myosin or muscle type creatine phosphokinase. Less than 0.1% of these cells demonstrated strong fluorescence when stained with anti-desmin or anti-smooth muscle specific actin. This observation suggested that the vast majority of these cells were not related to vascular smooth muscle cells. Also, over 99% of the non-myogenic cells did not display characteristic properties of endothelial cells. The denser myogenic cell fraction comprised over 80% of the cells and in clonal cultures gave rise to about 70% myogenic clones. An additional 30% of clones from this fraction were non-myogenic indicating heterogeneity in this population. We conclude that Percoll centrifugation can be employed for the isolation of myogenic and non-myogenic cell populations directly from the embryonic muscle. Moreover, this procedure allows the direct analysis of cell-specific proteins (e.g., by gel electrophoresis) without the need for cell culturing. The results thus obtained closely reflect the status of the cells in the intact muscle.
D000199:Actins / Q000032:analysis; D000818:Animals; D002453:Cell Cycle; D002469:Cell Separation / Q000379:methods*; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D002642:Chick Embryo; D003893:Desmin / Q000032:analysis; D005347:Fibroblasts / Q000166:cytology*; D007158:Immunologic Techniques; D009124:Muscle Proteins / Q000032:analysis; D009130:Muscle, Smooth / Q000166:cytology; D009132:Muscles / Q000166:cytology / Q000196:embryology*; D011205:Povidone; D012822:Silicon Dioxide
D000199:Actins; D003893:Desmin; D009124:Muscle Proteins; C016039:Percoll; D012822:Silicon Dioxide; D011205:Povidone