Inhibition of growth and induction of differentiation in a malignant human glioma cell line by normal leptomeningeal extracellular matrix proteins

Abstract

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.