Hantaï D, Festoff B W
Exp Neurol. 1987 Jan;95(1):44-55. doi: 10.1016/0014-4886(87)90005-7.
The turnover of basement membrane macromolecules in injured skeletal muscle has not been studied in contrast to other biologic systems undergoing remodeling. Plasminogen activators and other neutral proteases that are able to degrade these basement membrane macromolecules are secreted by cultured muscle cells. We sought to determine if locally released plasminogen activators could act on basement membrane components. Such degradation might be implicated in the disadhesion of nerve from muscle after motor nerve denervation. To test this hypothesis, we first undertook a study of the sensitivity of muscle extracellular matrix antigens following in vitro exposure to various proteases on frozen muscle sections. Fibronectin was found to be most sensitive, followed by type IV collagen and laminin. Of serine proteases, trypsin was the most active but was not selective, digesting matrix and sarcoplasmic components alike in less than 30 min. Purified urokinase was inactive unless plasminogen (also inactive alone) was previously added to tissue sections, at which time only matrix antigens were digested. Little if any observable degradation of sarcoplasmic proteins took place under these conditions. Using a highly sensitive and selective assay, we found that plasminogen activators were present in muscle tissue and increased 8- to 10-fold after 10 days of denervation. Using an extract of denervated muscle in the presence of plasminogen, we observed degradation of matrix antigens. No degradation was observed with control muscle extract. We next evaluated the degradation of these antigens in denervated muscle during a temporal study. The results, analyzed by quantitative image analysis, indicates that with increasing time after denervation a marked decrease of fibronectin and type IV collagen, followed by laminin occurred but, again, only in the present of plasminogen. These results indicate a selective sensitivity of basement membrane antigens of muscle and a role for plasminogen activators in the degradation of these adhesive basement membranes macromolecules after denervation.
与其他正在经历重塑的生物系统不同，受损骨骼肌中基底膜大分子的更新尚未得到研究。能够降解这些基底膜大分子的纤溶酶原激活剂和其他中性蛋白酶由培养的肌肉细胞分泌。我们试图确定局部释放的纤溶酶原激活剂是否能作用于基底膜成分。这种降解可能与运动神经去神经支配后神经与肌肉的分离有关。为了验证这一假设，我们首先对冷冻肌肉切片体外暴露于各种蛋白酶后肌肉细胞外基质抗原的敏感性进行了研究。发现纤连蛋白最敏感，其次是IV型胶原和层粘连蛋白。在丝氨酸蛋白酶中，胰蛋白酶活性最高，但没有选择性，在不到30分钟内就能消化基质和肌浆成分。纯化的尿激酶没有活性，除非事先将纤溶酶原（单独也无活性）添加到组织切片中，此时只有基质抗原被消化。在这些条件下，肌浆蛋白几乎没有可观察到的降解。使用一种高度敏感和选择性的检测方法，我们发现肌肉组织中存在纤溶酶原激活剂，去神经支配10天后增加了8至10倍。在纤溶酶原存在的情况下，使用去神经支配肌肉的提取物，我们观察到基质抗原的降解。对照肌肉提取物未观察到降解。接下来，我们在一项时间研究中评估了去神经支配肌肉中这些抗原的降解情况。通过定量图像分析分析结果表明，去神经支配后随着时间的增加，纤连蛋白和IV型胶原显著减少，随后层粘连蛋白也减少，但同样仅在纤溶酶原存在的情况下发生。这些结果表明肌肉基底膜抗原具有选择性敏感性，并且纤溶酶原激活剂在去神经支配后这些粘附性基底膜大分子的降解中起作用。
