Mechanisms of skeletal muscle regeneration and factors affecting it.

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The time courses of the expression and distribution of submembranous cytoskeletal proteins, dystrophin and vinculin, were investigated in regenerating fibers of the anterior latissimus dorsi muscle (ALD) in the chicken after cold injury. At 2-7 days after injury, serial transverse and longitudinal frozen sections were prepared from the ALD,stained with antibodies against dystrophin, vinculin and cardiac C-protein (CCP) and then incubated with FITC-labeled secondary antibody and rhodamine-labeled phalloidin. The specimens were subsequently observed under a confocal microscope. The antibody against CCP was used to detect regenerating fibers. CCP-positive regenerating fibers first appeared at 3 days after injury. At this time, dystrophin was faintly and nonuniformly distributed along the cell membrane of some regenerating fibers. Some of these dystrophin-positive fivers were also reactive to anti-vinculin. At 7 days after injury, regenerating fibers exhibited various degrees of anti-CCP staining reactivity, reflecting a variety of regeneration stages. In longitudinal sections, fibers intensely stained with anti-CCP were found to contain sparsely distributed thin actin fibrils (miofibrils) reactive with phalloidin. In en face views of such fibers, dystrophin was faintly and uniformly distributed along the cell membrane. Fibers positive with anti-CCP at moderate levels exhibited a striated pattern of dystrophin to some extent along the sarcolemma in addition to the faint and diffuse distribution. In fibers weakly labeled with anti-CCP,well-developed myofibrils filled the cytoplasm, and dystrophin was found to exhibit distinct striations and/or a lattice-like pattern as those in intact fibers. These results suggest that dystrophin is expressed earlier than vinculin during regeneration of skeletal muscle fibers and that the distribution of dystrophin along the sarcolemma changes from the uniform to the striated, and to the lattice-like pattern as the regeneration advances.