GRANULOCYTE-MACROPHAGE COLONY-STIMULATING FACTOR INCREASES 5-LIPOXYGENASE GENE-TRANSCRIPTION AND PROTEIN EXPRESSION IN HUMAN NEUTROPHILS

The complex process of neutrophil activation and accumulation is orche strated by a cascade of cytokines and bioactive lipids produced at the site of inflammation. Neutrophils are a rich source of the potent inf lammatory lipid leukotriene B-4(LTB(4)). Granulocyte-macrophage colony -stimulating factor (GM-CSF) can activate neutrophils for an exponenti al increase in LTB(4) production in response to a number of subsequent stimuli. in this report, we examined the temporal regulation, by GM-C SF, of the gene and protein expression of 5-lipoxygenase (5-LOX), a ke y enzyme of the LTB(4) production pathway. Human neutrophils were expo sed to 10 ng/mL of GM-CSF for various periods of time and 5-LOX mRNA w as measured by Northern blot analysis. We observed no change in 5-LOX mRNA at early time points (0.25 to 3 hours); however, by 18 hours we o bserved a significant augmentation of 5-LOX-specific message (4.3- +/- 1.7-fold increase; n = 6). Nuclear transcription assays indicated tha t the rate of 5-LOX gene transcription was augmented threefold in neut rophils incubated with GM-CSF, whereas the half-life of the message wa s not markedly changed. Parallel experiments lndicated that the levels of 5-LOX protein were also increased by GM-CSF. The augmentation was observed within 30 minutes after stimulation and was maximal (5.23 +/- 2.6; n = 4)at Is hours. Incubation of GM-CSF-stlmulated neutrophils w ith protein synthesis inhibitors resulted in a time-dependent impairme nt of their ability to produce LTB(4), with no inhibition seen during the first hours, a 75% decrease seen by 12 hours, and greater than 95% inhibition seen at 18 hours. Collectively, our data imply that GM-CSF can regulate LTB(4) production by two distinct mechanisms: a short-te rm increase that snot related to increased 5-LOX mRNA expression and i s independent of protein synthesis, and a sustained increase in LTB(4) production that is associated with the transcriptional activation of the 5-LOX gene, increase in 5-LOX mRNA levels, and dependence on prote in synthesis. Such transcriptional modulation of 5-LOX enzyme expressi on may provide new approaches for therapeutic intervention in protract ed inflammatory conditions. (C) 1995 by The American Society of Hemato logy.