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Skeletal muscle regeneration after injury involves various processes, such as infiltration by inflammatory cells, the proliferation of satellite cells and fusion to myotubes. The c-ski nuclear protein has been implicated in the control of cell proliferation and/or terminal differentiation in the growth of skeletal muscle. However, there have been no reports concerning the involution of c-ski in the regeneration of injured skeletal muscle in mammals. A possible role for c-ski in the proliferation of myogenic cells in rat skeletal muscle during regeneration has been investigated with the assistance of in vitro experiments with L6 skeletal muscle cells. The expression levels of c-ski mRNA in regenerating tissues increased to approximately threefold that of intact tissues at 2 days after injury and decreased to normal levels at 2 weeks after injury. Many mononuclear cells among the Ski-positive cells expressed desmin and proliferating cell nuclear antigen, indicating that Ski-producing cells include the proliferating myogenic cells. The proliferation of L6 cells was significantly retarded by expression of the antisense ski gene. The results of the present study reveal that the c-ski gene plays an important role in the proliferation of myogenic cells in the regeneration of injured skeletal muscle.

Abstract. Growth/Differentiation Factor 8, or GDF-8, is a member of the TGF-β family and is expressed specifically in myogenic precursor cells of the myotome during development as well as in adult axial and paraxial muscles of the mouse. It acts as a negative regulator of skeletal muscle mass, and is known as myostatin. As a part of our studies to elucidate mechanisms underlying the growth of skeletal muscle mass due to physical training in mammals including horses and humans, we isolated and cloned myostatin cDNA from equine (Thoroughbred) skeletal muscles by an RT-PCR method. The base sequence of the entire coding region of equine myostatin cDNA (DDBJ accession no. AB033541) exhibited a high degree of homology to those of other species so far reported. In the deduced amino acid sequence, 4 amino acid positions were unique to the equine myostatin, i. e., Val 15 instead of Met, Leu 18 instead of Val or Ala, Ala 201 instead of Thr, Arg 244 instead of Gly or Glu in other species. Western blot analysis of the equine skeletal muscles using anti-myostatin antibody, yielded 3 immunoreactive bands, i. e., 26, 45 and 50 kDa. The band at approximately 26 kDa probably represents mature myostatin and another at approximately 50 kDa corresponds in size to the unprocessed precursor myostatin proteins reported in the literature. Western blot analysis of equine sera of 2 independent breeds, Thoroughbred and Kiso-uma, revealed the possible presence of precursor myostatin proteins at relatively high levels. The molecular identity and the biological significance of serum myostatin immunoreactivities remain to be clarified.