Mechanisms of skeletal muscle regeneration and factors affecting it.

研究了冷损伤后鸡背阔肌前肌再生纤维膜下细胞骨架蛋白dystrophin和vinculin的表达和分布的时程。在损伤后2-7天，从ALD制备连续的横向和纵向冷冻切片，用抗肌营养不良蛋白、粘着斑蛋白和心脏C蛋白（CCP）的抗体染色，然后与FITC标记的第二抗体和罗丹明标记的鬼笔环肽孵育。随后在共聚焦显微镜下观察标本。抗CCP抗体用于检测再生纤维。CCP阳性再生纤维在损伤后3天首次出现。此时，肌营养不良蛋白沿某些再生纤维的细胞膜呈微弱且不均匀的沿着分布。这些抗肌萎缩蛋白阳性的fivers中的一些也对抗黏着斑蛋白反应。损伤后7天，再生纤维表现出不同程度的抗CCP染色反应，反映了不同的再生阶段。在纵向切片中，发现用抗CCP强烈染色的纤维含有稀疏分布的与鬼笔环肽反应的细肌动蛋白原纤维（微原纤维）。从正面观察，肌营养不良蛋白沿细胞膜沿着均匀分布。中等水平抗CCP抗体阳性的纤维除了微弱和弥漫分布外，还表现出肌营养不良蛋白在一定程度上沿着肌膜的条纹状分布。在弱标记的纤维与抗CCP，发达的肌原纤维充满了细胞质，肌营养不良蛋白被发现表现出不同的条纹和/或格子样图案的那些在完整的纤维。这些结果表明，肌营养不良蛋白的表达早于纽蛋白在骨骼肌纤维的再生过程中，肌营养不良蛋白沿肌膜的分布变化从均匀的条纹状，沿着和网格状图案的再生进展。

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.