Quantitative increase of neuroglia-specific GFA protein in rat C-6 glioma cells in vitro

Abstract

AMONG neural cell lines the C-6 glioma, a cell line cloned from an N-methylnitrosourea (MNU)-induced experimental glioma in rat, has generated considerable interest as a model system for the study of neuroglia¹. The clone was originally selected on the basis of a high production of the protein S-100, a protein believed to be characteristic of glial cells in vivo². Since its introduction, the line has been further characterised with regard to a number of other biochemical features³. Among these is the recent demonstration of the glial fibrillary acidic (GFA) protein by immunofluorescent staining of C-6 cells maintained in certain in vitro conditions⁴. The GFA protein, isolated from multiple sclerosis plaques and areas of sub-ependymal gliosis in the central nervous system, has been shown to be a specific marker for fibrillary astrocytes, is believed to be a component of the 80–90 Å diameter glial filaments^5–8, and has not been reported in non-glial neural cell lines. It is, therefore, a useful biochemical marker for the differentiation of a type of mature astrocyte. In previous work with the C-6 line, GFA protein was found to be present by immunofluorescence in only an occasional C-6 cell maintained in monolayer or suspension cultures, and to be markedly increased in many C-6 cells when they were maintained for 12–14 d in an organ culture system using Gelfoam matrices⁴. Such a system is known to favour structural differentiation in vitro⁹. No increase of filaments was observed, however, in any of the in vitro systems used, suggesting that the synthesis of GFA protein could proceed without the formation of intracytoplasmic glial filaments⁴. We compared the quantitative levels of GFA protein in C-6 and other cell types in various culture conditions, using a “two-site” immunoradiometric assay recently developed in this laboratory¹⁰.