INTERLEUKIN-1-BETA INDUCTION OF TUMOR-NECROSIS-FACTOR-ALPHA GENE-EXPRESSION IN HUMAN ASTROGLIOMA CELLS

Cells that produce tumor necrosis factor-alpha (TNF-alpha) require the presence of signaling molecules since this cytokine is not normally expressed in a constitutive manner. It has been demonstrated that glial cells can produce TNF-alpha; however, the specific inducing molecules and their mechanisms(s) of action have not been clearly defined. In this study, we examined the effect of human recombinant interleukin-1-beta (IL-1-beta) on the expression of TNF-alpha by CH235-MG human malignant glioma cells. CH235-MG cells do not constitutively express TNF-alpha mRNA or protein; however, upon stimulation with IL-1-beta, these cells synthesize and secrete biologically active TNF-alpha IL-1-beta induces the expression of a 1.9 kb TNF-alpha mRNA species. Kinetic analysis demonstrated optimum TNF-alpha mRNA expression after a 4 h exposure to IL-1-beta, and peak TNF-alpha protein production at 18 h. Cycloheximide (CHX), an inhibitor of protein synthesis, markedly increased expression of TNF-alpha mRNA in IL-1-beta stimulated CH235-MG cells, indicating that de novo protein synthesis is not required for astroglioma TNF-alpha gene expression. Nuclear run-off analysis demonstrates that IL-1-beta causes transcriptional activation of the TNF-alpha gene, and CHX enhances IL-1-beta-induced TNF-alpha transcription. Studies of TNF-alpha mRNA stability using actinomycin D show that IL-1-beta-induced TNF-alpha mRNA has a half-life of approximately 30 min, and CHX increases the half-life of IL-1-beta-induced TNF-alpha mRNA to approximately 210 min. These results indicate that IL-1-beta, a cytokine present in the central nervous system during some pathological disease states, is a potent inducer of TNF-alpha in human malignant glioma cells.