Since cysteinyl-leukotrienes are major protagonists in the pathophysiology of human asthma and since neutrophils are involved in the more severe form of asthma, we studied the potential for leukotriene (LT)D₄ to induce synthesis of the chemokine IL-8 through activation of the CysLT1 receptor. We found LTD₄ to induce IL-8 gene expression in monocytic THP-1 cells and human dendritic cells with complete abrogation by selective CysLT1 antagonists. HEK293 cells stably transfected with CysLT1 were used to better study the transcriptional regulation of the IL-8 promoter. Stimulation of the cells with graded concentrations of LTD₄ resulted in a time- and concentration-dependent induction of IL-8 transcription and protein synthesis. Use of IL-8 promoter mutants with substitutions in their NF-B, AP-1 and NF-IL-6 binding elements revealed a requirement for NF-B and AP-1, but not NF-IL-6, in LTD₄-induced activation of the IL-8 promoter. Overexpression of dominant negative IB inhibited the IL-8 transactivation induced by LTD₄. NF-B DNA binding activity was induced by LTD₄ as determined by electrophoretic mobility shift assays and could be supershifted by antibodies against p50 and p65. Supershift assays following LTD₄ stimulation also indicated the formation of a c-Jun/c-Fos complex. Moreover, our results demonstrated that LTD₄ upregulates the expression of c-fos and c-jun at the mRNA level. Our data show for the first time that LTD₄, via the CysLT1 receptor, can transcriptionally activate IL-8 production, with involvement of the transcription factors p50, p65, Fos and Jun. These findings provide mechanistic and potentially therapeutic elements for modulation of the inflammatory component of asthma.