Sasaki H, Kishiye T, Fujioka A, Shinoda K, Nagano M
Department of Anatomy, Kinki University School of Medicine, Osaka, Japan.
Cell Struct Funct. 1996 Apr;21(2):133-41. doi: 10.1247/csf.21.133.
Orthogonal aggregates of small intramembrane particles, termed "assemblies," are concentrated especially in the cell membranes of astrocytic processes that form the glia limitans at the outer surface of the brain and the perivascular sheath surrounding the parenchymal blood vessels. As an initial step to clarifying the totally unknown biochemical nature of this intramembrane structure, we have devised a culture system which enhances the differentiation of assemblies in secondary cultures of astrocytes derived from neonatal mouse neopallium. Since assemblies are most concentrated in the plasma membranes attaching to the basement membrane, we expected that extracellular matrix molecules constituting the basement membrane would be suitable candidates for our aim. We report here that a mixture of type IV collagen, laminin, and fibronectin, major components of the extracellular matrix, has the potency to increase assembly density in cultured astrocytes. We also report that, in freeze-fracture electron microscopy of cultured cells, one can satisfactorily preserve membrane structure and reliably obtain large replicas by inoculating cells on aluminium foil and peeling it from the cells in a freeze-fracture apparatus.
被称为“聚集体”的小膜内颗粒的正交聚集体尤其集中在星形胶质细胞突起的细胞膜中，这些突起在脑外表面形成胶质界膜以及围绕实质血管的血管周鞘。作为阐明这种膜内结构完全未知的生化性质的第一步，我们设计了一种培养系统，该系统可增强源自新生小鼠新皮质的星形胶质细胞二次培养物中聚集体的分化。由于聚集体最集中在附着于基底膜的质膜中，我们预计构成基底膜的细胞外基质分子将是实现我们目标的合适候选物。我们在此报告，细胞外基质的主要成分IV型胶原、层粘连蛋白和纤连蛋白的混合物具有增加培养星形胶质细胞中聚集体密度的能力。我们还报告，在培养细胞的冷冻断裂电子显微镜观察中，通过将细胞接种在铝箔上并在冷冻断裂装置中从细胞上剥离铝箔，可以令人满意地保存膜结构并可靠地获得大的复制品。
