Immunoelectron microscopic and freeze etch studies of muscle plasma membrane and extracelular matrix in normal and diseased skeletal myofiber

采用抗-异常糖聚糖抗体和抗美罗辛单克隆抗体对人体骨骼肌进行免疫金电镜观察。我们还打算使用抗α -糖酐异常抗体来进行研究。根据已发表的α -糖异常肽cDNA序列，合成了抗α -糖异常肽抗体（Nature 355: 696-702,1992）。然而，我们的α -糖醛酸异常抗体目前正在分析其特性。抗β -肌营养不良多糖在骨骼肌纤维肌层下的超微结构定位[J] .电子显微杂志43:327-331,1994；中华病理学杂志146:189-196,1995。人骨骼肌纤维中肌红蛋白的超微结构定位在肌质膜外和基底层成分内。此外，冷冻蚀刻复制品的基板电镜显示，短的细胞骨架垂直于肌质膜表面，并与基板的这种小纤维结构的网络形成相连接，基板中也含有较厚的胶原型原纤维。merosin的信号定位于人骨骼肌纤维复杂基底层的内侧。因此，与金标记的美罗红分子相关的细胞骨架元件被认为是与肌营养不良蛋白（dystrophin vir -dystroglycan）相关的α -肌营养不良蛋白（α - dystropcan）。

Immunogold electron microscopic investigation was carried out in human skeletal muscles by using anti beta-dystroglycan antibodies and anti merosin monoclonal antibody. We also intended to perform the study by using anti alpha-dystroglycan antibodies. Anti alpha-dystroglycan antibodies were generated against three synthetic peptides deduced on the published alpha-dystroglycan cDNA sequence (Nature 355 : 696-702,1992). However, our alpha-dystroglycan antibodies are now under analysis for their characterization. The anti beta-dystroglycan was ultrastructurally localized in the subsarcolemmal area of skeletal myofibers (J Electron Microsc 43 : 327-331,1994 ; Am J Pathol 146 : 189-196,1995). In contrast, the ultrastructural localization of merosin in human skeletal myofiber was present just outside of muscle plasma membrane and just inside area of basal lamina components. In addition, electron microscopy of basal lamina of freeze etched replica showed that the short cytoskeleton was standing perpendicular to the surface of muscle plasma membrane and it was connected to the network formation of this small fibrillar structures of basal lamina which also contained more thick collagen type IV fibrills. The signal of merosin was localized at the inside part of the complicated basal lamina of huma skeletal myofibers. The merosin was turned out to be the alpha-dystroglycan receptor (Sunada Y.et al.J Biol Chem 269 : 13729-13732,1994), so the cytoskeletal element associated with the gold labelled merosin molecules is thought to be the alpha-dystroglycan which is connected with dystrophin vir beta-dystroglycan.