Interferon‐α (IFN‐α) inhibits granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) expression at the post‐transcriptional level in murine bone marrow stromal cells

Summary Recently it has been shown that IFN‐α inhibits expression of GM‐CSF in adherent cells of human long‐term bone marrow cultures (LTBMC) stimulated with interleukin‐1 (IL‐1), tumour necrosis factor‐α (TNF‐α) or endotoxin. The murine bone marrow stromal cell line +/+‐ 1.LDA11 was used to further define regulatory mechanisms of IFN‐α inhibition on GM‐CSF expression. This cell line originated from a murine Dexter type culture and exhibits a pre‐adipocytic phenotype. As in human LTBMC, we could demonstrate a inhibitory effect of IFN‐α co‐incubation on GM‐CSF activity in serum‐free supernatants of +/+‐1.LDA11 stromal cell cultures stimulated with IL‐1 or TNF‐α or the combination of IL‐1 plus TNF‐α. IFN‐α inhibitory effect on GM‐CSF expression was shown to be dose dependent with minimal response at 10U/ml and maximal inhibition at a dose of 500U/ml. Northern blot analysis confirmed these data at the mRNA level. Reprobing of Northern blots for interleukin‐6 (IL‐6) mRNA showed increased expression after IFN‐α incubation, demonstrating specific and differential regulatory effects of IFN‐α on cytokine production in bone marrow stromal cells. Inhibition of GM‐CSF mRNA by IFN‐α was time dependent, starting at about 90–120 min post‐treatment. Cycloheximide (CHX) incubation abolished the inhibitory effect of IFN‐α on GM‐CSF expression, suggesting the requirement of a labile protein. Reporter gene studies were used in order to evaluate the effect of IFN‐α incubation on GM‐CSF mRNA transcription in stromal cells. For this purpose, GM‐CSF promoter fragments were subcloned into a luciferase expression vector. Neither constitutive nor TNF‐α stimulated GM‐CSF transcription was inhibited by IFN‐α co‐incubation. On the other hand, actinomycin‐D chase experiments revealed a reduced GM‐CSF mRNA stability after IFN‐α incubation. The induction of a RNAase, possibly a 2–5A‐dependent RNAase, by IFN‐α may be a possible cause for the increased GM‐CSF mRNA decay. These results show a regulatory role for IFN‐α in the bone marrow microenvironment possibly involved in the myelosuppres‐sive effect of IFN‐α therapy or viral infections.