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

The transcription factor, nuclear factor kappaB (NF-kappaB), is activated by various stimuli including cytokines, radiation, viruses and oxidative stress. Here we show that, although induction with H2O2 gives rise to NF-kappaB 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 la 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-kappaB 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-kappaB inhibitors clasto-lactacystin-beta -lactone and E-64D (L-3-trans-ethoxy-carbonyloxirane-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.