Decreased IL-6 secretion by fibroblasts following repeated doses of TNFα or IL-1α: Post-transcriptional gene regulation

Abstract

Tumor necrosis factor-α (TNFα) and interleukin-1α (IL-1α) are pluripotent cytokines mediating the host response to sepsis, injury, and cancer. Animals can be protected from the lethal effects of TNFα by repeated administration of sublethal doses, but the mechanism of this effect is not known. Human foreskin fibroblasts (FS4 cells), which rapidly elaborate interleukin-6 (IL-6) when stimulated with TNFα or IL-1α, were grown in culture as confluent monolayers and their secretion of IL-6 was quantitated using the murine B9-hybridoma bioassay against an external reference of human recombinant IL-6 (Genetics Institute). When FS4 cells were incubated with human recombinant TNFα (50 ng/ml; Cetus) or recombinant IL-1α (30 pg/ml; Genzyme) a rapid increase in IL-6 production was measured over control, rising to IL-6 levels of 71.7 ± 5.9 units/ml with TNFα and 54.0 ± 1.2 units/ml with IL-1α after 7.5 hr incubation. FS4 cells which were exposed to cytokine, rinsed, and then reexposed to cytokine 24 hr later produced significantly less IL-6 [38.1 ± 2.8 units/ml with second exposure to TNFα ( P < 0.05), and 18.3 ± 1.9 units/ml with second exposure to IL-1α ( P < 0.01)]. Successive daily exposure to TNFα or IL-1α caused a further stepwise diminution of IL-6 secretion. When RNA was extracted from FS4 cells incubated daily with TNFα or IL-1α for 1 to 3 days, Northern blot analysis revealed no apparent change in IL-6 mRNA gene expression following repeated exposure to TNFα or to IL-1α. We conclude that fibroblasts become refractory to the stimulatory effects of TNFα and IL-1α after a single exposure, and that repetitive exposure can cause a further decrease in IL-6 production which probably occurs by post-transcriptional regulation. These observations may have applicability to clinical conditions in which toxicity or untoward effects are mediated through the secretion of cytokines.