van Kuppevelt T H, Benders A A, Versteeg E M, Veerkamp J H
Department of Biochemistry, University of Nijmegen, The Netherlands.
Exp Cell Res. 1992 Jun;200(2):306-15. doi: 10.1016/0014-4827(92)90177-a.
Basement membrane (BM) components were studied on human muscle and skeletal muscle cells cultured on different media by immunofluorescence and electron microscopy. Their topographical relation with acetylcholine receptors was investigated. Myotubes cultured on a combination of the serum substitute Ultroser G and brain extract show a continuous layer of heparan sulfate proteoglycans (HSPGs), laminin, and type IV collagen. In contrast, myotubes cultured on serum-containing media are associated with granular depositions of HSPG and laminin and only with wisps of type IV collagen. Omission of brain extract or substitution by chicken embryo extract results in an intermediate staining pattern. For all types of cultures, fibronectin is localized in and around mononuclear cells, but hardly associated with myotubes. A codistribution between clusters of acetylcholine receptors and HSPG and laminin and Vicia villosa B4 lectin-positive material exists only in Ultroser G/brain extract-based myotubes like in muscle in vivo. No clustering is observed in serum-based myotubes. Electron microscopy reveals that the former myotubes are surrounded by a continuous BM consisting of a lamina lucida, lamina densa, and lamina fibroreticularis. Proteoglycans are present on the external site of the lamina densa and associated in a regular fashion with collagen fibrils. In conclusion, BMs associated with myotubes cultured on Ultroser G/brain extract resemble in many ways the in vivo situation, including synaptic specializations. Cultured myotubes may serve as a model system for studies on the structure and function of human muscular (synaptic) BM under normal and pathological conditions.
通过免疫荧光和电子显微镜技术，对在不同培养基上培养的人肌肉和骨骼肌细胞的基底膜（BM）成分进行了研究。研究了它们与乙酰胆碱受体的拓扑关系。在血清替代物Ultroser G和脑提取物的组合培养基上培养的肌管显示出连续的硫酸乙酰肝素蛋白聚糖（HSPG）、层粘连蛋白和IV型胶原蛋白层。相比之下，在含血清培养基上培养的肌管与HSPG和层粘连蛋白的颗粒状沉积物相关，仅与IV型胶原蛋白的细丝相关。省略脑提取物或用鸡胚提取物替代会导致中间染色模式。对于所有类型的培养物，纤连蛋白定位于单核细胞内和周围，但几乎与肌管无关。乙酰胆碱受体簇与HSPG、层粘连蛋白和野豌豆B4凝集素阳性物质之间的共分布仅存在于基于Ultroser G/脑提取物的肌管中，就像在体内肌肉中一样。在基于血清的肌管中未观察到聚集。电子显微镜显示，前者的肌管被由透明层、致密层和纤维网状层组成的连续BM包围。蛋白聚糖存在于致密层的外部，并以规则的方式与胶原纤维相关联。总之，与在Ultroser G/脑提取物上培养的肌管相关的BM在许多方面类似于体内情况，包括突触特化。培养的肌管可作为研究正常和病理条件下人类肌肉（突触）BM的结构和功能的模型系统。
