Elucidation of Molecular Mechanism of Muscle Regeneration of Dystrophin-dificient Duchenne Muscular Dystorophy

我们先前鉴定了α1-促突触蛋白敲除小鼠中的异常肌肉再生（J.Cell Biol 158：1097-1107，2002）。这些发现可能代表了在肌营养不良蛋白缺陷型Duchenne肌营养不良中发现的异常肌肉再生，因为肌营养不良蛋白缺陷型肌膜中也缺乏肌营养不良蛋白结合蛋白之一α1-突触体蛋白的表达。本研究试图进一步阐明肌肉再生的分子机制。本文重点研究了微重力条件下骨骼肌萎缩的尾部悬吊模型。非常有趣的是，这些萎缩的肌肉在地面上重新加载后迅速恢复。我们深入分析了该模型上肌营养不良蛋白-糖蛋白复合物（DGC）组分的表达。我们发现，神经元型一氧化氮合酶（nNOS）的表达减少，从肌膜在尾部悬吊。nNOS是通过DGC的α1-syntrophin锚定在肌膜上的功能分子之一。因此，我们研究了nNOS和α1-syntrophin基因敲除小鼠的悬尾过程。非常有趣的是，nNOS基因敲除小鼠在尾部悬吊过程中显示出肌肉萎缩的恢复，而α1-syntorophim基因敲除小鼠则没有。此外，nNOS基因敲除小鼠在再负荷过程中表现出肌肉再生困难。在肌肉再生过程中，卫星细胞的增殖是最重要的，正如在肌肉再生中所发现的那样。最后，我们检查了nNOS敲除小鼠的肌肉卫星细胞的增殖能力，并揭示了这些细胞的增殖能力受损。因此，我们得出结论，nNOS的表达是非常重要的，无论是在肌肉萎缩的尾巴悬挂和卫星细胞的增殖过程中肌肉重新加载后的地面上的生长。

We previously identified abnormal muscle regeneration in α1-syntrophin knock-out mice (J.Cell Biol 158:1097-1107, 2002). These findings may represent abnormal muscle regeneration found in dystrophin-deficient Duchenne muscular dystrophy, since expression of α1-syntorophin, one of dystrophin-binding proteins, was also absent from the dystrophin-deficient sarcolemma. In this study, we further tried to elucidate molecular mechanism of muscle regeneration. We paticulaly paid attention on tail suspension model, where skeletal muscle was suffered from muscle atroply due to micro-gravity. Very interestingly those atrophied muscle revealed prompt recovery after re-loading on the ground. We intensively analyzed expression of constituents of dystrophin-glycoprotein complex (DGC) on the model. We found a decrease of expression of neuronal Nitric Oxide synthase (nNOS) from the sarcolemma during the tail suspension. nNOS is one of functional molecules, which is anchored to the sarcolemma through α1-syntrophin of DGC. Therefore, we examined process of tail suspension of both nNOS and α1-syntrophin knock out mice. Very interestingly, nNOS knock-out mice revealed resistency in muscle atrophy during tail suspension process, but α1-syntorophim knock-out mice did not. Moreover, nNOS knock-out mice showed difficulty in muscle regrowth in re-loading process. In muscle regrowth process, proliferation of satellite cells were most important, as found in muscle regeneration. We finally examined proliferation capacity of muscle satellite cells of nNOS knock out mice and revealed impaired proliferation capacity of these cells. Therefore, we concluded the expression of nNOS is very important both in muscle atrophy during tail suspension and in proliferation of satellite cells during muscle re-growth after reloading on the ground.