Tumor Necrosis Factor‐Induced Proliferation of Astrocytes from Mature Brain Is Associated with Down‐Regulation of Glial Fibrillary Acidic Protein mRNA

Previous results from this laboratory have shown that tumor necrosis factor (TNF) is mitogenic for bovine astrocytes in chemically defined (CO) medium. The maximum mitogenic response was detected with 200 U/ml at 48 h. We have now extended these studies to assess the effect of TNF on message levels for the intermediate filament proteins glial fibrillary acidic protein (GFAP) and vimentin. The results have shown that, whereas TNF had only a slight effect on vimentin mRNA, TNF induced a marked decrease to 4.3 ± 2.0% of controls in GFAP mRNA which was both time and dose dependent. The lowest effective dose was 50 U/ml and the maximal effective dose was 200 U/ml. Kinetic analysis of this response demonstrated that a marked decrease in GFAP mRNA was present at 12 h and continued to decrease through 72 h. To determine the reversibility of the TNF effect, astrocyte cultures were exposed to 200 U/ml TNF for varying periods of tee and then cultured in fresh CD medium. A 1‐h pulse with TNF was sufficient to reduce GFAP mRNA levels when measured 24 h later. However, cultures incubated with 200 U/ml TNF for 48 h followed by incubation in CD medium without TNF for 7 days showed that GFAP mRNA levels had returned to 60% of the control values. Nuclear runoff assays showed that the effect of TNF on GFAP mRNA was at the posttranscriptional level. Polyacrylamide gel electrophoretic analysis of astrocyte cytoskeletal proteins demonstrated that GFAP levels were reduced after a 5‐day incubation with 200 U/ml TNF whereas protein levels of vimentin and actin were not significantly changed. Western blots confirmed that GFAP levels were reduced to 36% of the control values. Thus the effect of TNF on GFAP mRNA expression was not due to a generalized effect on intermediate filament metabolism.