Importance of post-transcriptional regulation of chemokine genes by oxidative stress

The transcription factor, nuclear factor κB (NF-κB), is activated by various stimuli including cytokines, radiation, viruses and oxidative stress. Here we show that, although induction with H₂O₂ gives rise to NF-κB nuclear translocation in both lymphocyte (CEM) and monocyte (U937) cells, it leads only to the production of mRNA species encoding interleukin-8 (IL-8) and macrophage inflammatory protein 1α in U937 cells. Under similar conditions these mRNA species are not observed in CEM cells. With the use of a transient transfection assay of U937 cells transfected with reporter constructs of the IL-8 promoter and subsequently treated with H₂O₂, we show that (1) IL-8-promoter-driven transcription is stimulated in both U937 and CEM cells and (2) the NF-κB site is crucial for activation because its deletion abolishes activation by H₂O₂. The production of IL-8 mRNA in U937 cells is inhibited by the NF-κB inhibitors clasto-lactacystin-β-lactone and E-64D (l-3-trans-ethoxycarbonyloxirane-2-carbonyl-l-leucine-3-methyl amide) but requires protein synthesis de novo. Moreover, inhibition of the p38 mitogen-activated protein kinase also decreases the IL-8 mRNA up-regulation mediated by H₂O₂. Taken together, these results show the importance of post-transcriptional events controlled by a p38-dependent pathway in the production of IL-8 mRNA in U937. The much lower activation of p38 in CEM cells in response to H₂O₂ could explain the lack of stabilization of IL-8 mRNA in these cells.