REGULATION OF INTERLEUKIN-8 GENE-EXPRESSION BY OXIDANT STRESS

Interleukin 8 (IL-8) is a recently described cytokine that functions a s a potent neutrophil chemoattractant and activator. We sought to exam ine the link between the generation of reactive oxygen intermediates ( ROI) and the regulation of IL-8 gene expression to specifically test t he hypothesis that ROI would induce production of IL-8 mRNA and protei n. In lipopolysaccharide-stimulated human whole blood, the OH radical scavenger dimethyl sulfoxide (Me2SO) dramatically inhibited (approxima tely 90%) IL-8 production, but had minimal effects on the production o f tumor necrosis factor, interleukin 1beta (IL-1), and IL-6. To determ ine whether NADPH-oxidase-generated free radicals were critical in the regulation of IL-8, studies were performed using blood from patients with chronic granulomatous disease. In both normal individuals and pat ients with chronic granulomatous disease, production of IL-8 could be initiated with lipopolysaccharide, phytohemagglutinin, or aggregated i mmune complexes, and this production could be inhibited by Me2SO (1% v /v). To examine if oxidant stress represents a ubiquitous mechanism fo r the induction of IL-8, experiments were performed in cultured cell l ines. In the human hepatoma cell line Hep-G2, Me2SO dose-dependently i nhibited tumor necrosis factor-stimulated IL-8 production, with a 74 /- 1% reduction observed at a Me2SO concentration of 1%. Direct exposu re to ROI demonstrated that H2O2 stimulated IL-8 production in a dose- dependent manner in Hep-G2 cells, A549 pulmonary type II epithelial ce lls, and human skin fibroblasts; this induction could be prevented by addition of catalase. The production of IL-8 appeared to be specific t o an oxidant stress since exposure of the cells to heat shock or chemi cal stress did not induce expression of IL-8. These studies demonstrat e that oxidant stress is an important regulator of IL-8 gene expressio n and support the hypothesis that low levels of ROI may serve to initi ate IL-8 production which then serves to recruit neutrophils to sites of inflammation.