Rutka J T, Giblin J R, Apodaca G, DeArmond S J, Stern R, Rosenblum M L
Cancer Res. 1987 Jul 1;47(13):3515-22.
We devised a model system to study the effects of extracellular matrix proteins on the malignant phenotype of an anaplastic glioma cell line, U 343 MG-A. Well-characterized cultures derived from normal human leptomeninges were grown to confluence and maintained for 2 weeks. The leptomeningeal cells were then removed with base and detergent, leaving behind an extracellular matrix enriched in laminin, fibronectin, type I and IV collagen, and procollagen III. U 343 MG-A tumor cells planted on top of this normal extracellular matrix were profoundly growth inhibited compared with glioma cells grown on plastic alone. Glioma cells grown on the extracellular matrix developed multiple, slender processes and assumed a more differentiated astrocytic phenotype; immunostains for glial fibrillary acidic protein revealed a more extensive intracytoplasmic network of intensely staining filaments than in control glioma cells. When glioma cells grown on the extracellular matrix were analyzed by an enzyme-linked immunosorbent assay for glial fibrillary acidic protein, the amount of this intermediate filament per cell was increased 20-fold compared with glioma cells growing on plastic. The growth and differentiation of U 343 MG-A glioma cells in flasks coated with purified fibronectin or laminin was not significantly perturbed; however, glioma cell cultures grown in flasks coated with purified type I or IV collagen showed decreased cellular proliferation, stellate cell formation, and increased levels of glial fibrillary acidic protein per cell compared with glioma cells growing on plastic. Gelatin gel analysis showed that U 343 MG-A glioma cells growing on plastic secreted a 65,000-D metalloproteinase that was not secreted by glioma cells grown on the leptomeningeal extracellular matrix. We conclude that in this system, the extracellular matrix of a normal human leptomeningeal culture substantially inhibited the proliferation of and induced differentiation in an anaplastic glioma cell line. Our analysis of single components of the extracellular matrix suggests that these effects may be mediated in part by type I and IV collagen. The mechanism by which the leptomeningeal extracellular matrix inhibits glioma cell proliferation may be by diminishing tumor-associated protease secretion so that the degradation of extracellular matrix macromolecules in the tumor cell microenvironment is prevented and tumor cell migration becomes less likely.
我们设计了一个模型系统，以研究细胞外基质蛋白对间变性胶质瘤细胞系U 343 MG-A恶性表型的影响。将源自正常人软脑膜的特征明确的培养物培养至汇合状态并维持2周。然后用碱和去污剂去除软脑膜细胞，留下富含层粘连蛋白、纤连蛋白、I型和IV型胶原以及III型前胶原的细胞外基质。与仅在塑料上生长的胶质瘤细胞相比，接种在这种正常细胞外基质上的U 343 MG-A肿瘤细胞的生长受到显著抑制。在细胞外基质上生长的胶质瘤细胞形成多个细长的突起，并呈现出更分化的星形胶质细胞表型；胶质纤维酸性蛋白的免疫染色显示，与对照胶质瘤细胞相比，其胞质内有更广泛的强染色丝网络。当通过酶联免疫吸附测定法分析在细胞外基质上生长的胶质瘤细胞中的胶质纤维酸性蛋白时，每个细胞中这种中间丝的量比在塑料上生长的胶质瘤细胞增加了20倍。在涂有纯化纤连蛋白或层粘连蛋白的培养瓶中，U 343 MG-A胶质瘤细胞的生长和分化没有受到明显干扰；然而，与在塑料上生长的胶质瘤细胞相比，在涂有纯化I型或IV型胶原的培养瓶中生长的胶质瘤细胞培养物显示细胞增殖减少、星状细胞形成以及每个细胞中胶质纤维酸性蛋白水平增加。明胶凝胶分析表明，在塑料上生长的U 343 MG-A胶质瘤细胞分泌一种65000-D的金属蛋白酶，而在软脑膜细胞外基质上生长的胶质瘤细胞不分泌这种酶。我们得出结论，在这个系统中，正常人软脑膜培养物的细胞外基质显著抑制了间变性胶质瘤细胞系的增殖并诱导其分化。我们对细胞外基质单一成分的分析表明，这些作用可能部分由I型和IV型胶原介导。软脑膜细胞外基质抑制胶质瘤细胞增殖的机制可能是通过减少肿瘤相关蛋白酶的分泌，从而防止肿瘤细胞微环境中细胞外基质大分子的降解，降低肿瘤细胞迁移的可能性。
