Evidence for distinct fast and slow myogenic cell lineages in human foetal skeletal muscle

To analyse the myogenic cell lineages in human foetal skeletal muscle, muscle cell cultures were prepared from different foetal stages of development. The in vitro muscle cell phenotype was defined by staining the myotubes with antibodies to fast and slow skeletal muscle type myosin heavy chains using immunoperoxidase or double immunofluorescence procedures. The antibodies to fast skeletal muscle myosin heavy chains stained nearly all myotubes dark in cell cultures prepared from quadriceps muscles at 10-18 weeks of gestation. The antibodies to slow skeletal muscle myosin heavy chains, in contrast, stained only 10-40% of the myotubes very dark. The remaining myotubes were further subdivided into two populations, one of which was unstained while the other stained with variable intensity for slow myosin heavy chain. The slow myosin heavy chain staining was not influenced by the nature of the substratum used to culture these cells, although the growth of muscle cell cultures was greatly improved on matrigel-coated dishes. The presence of both slow and fast myosin heavy chains was detected even when myotubes were grown on uncoated petri dishes. The myotube diversity was further investigated by analysing the clonal populations of human foetal skeletal muscle cells in vitro. When cultured at clonal densities, two types of myogenic clones were identified by their differential staining with antibodies to slow myosin heavy chain. As was the case with the high density muscle cell cultures, virtually all myotubes in both groups of clones stained with antibodies to fast myosin heavy chains. Antibodies to slow myosin heavy chains stained nearly all myotubes dark in one group of myogenic clones, but only a subset of the myotubes stained dark for slow myosin heavy chain in the second group of clones. The proportion of slow myosin heavy chain positive myotubes in this group varied in different clones. The myogenic diversity was thus apparent in both high density and clonal human muscle cell cultures, and myogenic cells retained their ability to modify their muscle cell phenotype.