Proinflammatory cytokines induce liver and activation-regulated chemokine/macrophage inflammatory protein-3 alpha/CCL20 in mucosal epithelial cells through NK-kappa B

Liver and activation-regulated chemokine (LARC)/CCL20 is expressed by surfa ce-lining epithelial and epidermal cells, and is likely to link innate and acquired immunity by attracting immature dendritic cells, effector memory T cells and B cells via CCR6. Here we examined the mechanism of LARC express ion in epithelial-type cells. Either IL-1 beta or tumor necrosis factor (TN F)-alpha strongly induced LARC mRNA in intestinal cell lines Caco-2 and T84 , while both were effective on HEK 293T cells. Induction of LARC was also d emonstrated in the intestinal epithelium of BALB/c mice upon treatment with IL-1 alpha or TNF-alpha. Transient transfection assays using murine LARC p romoter-reporter constructs identified a region essential for IL-1 beta, or TNF-alpha -induced promoter activation in Caco-2 and 293T cells. Using sit e-directed mutagenesis, we demonstrated that an NF-kappaB site located betw een -96 and -87 bp upstream from the transcriptional start site was both ne cessary and sufficient for IL-1 beta- or TNF-alpha -induced promoter activa tion in Caco-2 and 293T cells. Electrophoretic mobility shift assays demons trated that p50/p65 heterodimer and p65 homodimer of NF-kappaB bound to thi s site in 293T cells upon treatment with IL-1 beta and TNF-alpha, and p50/p 65 heterodimer bound to this site in Caco-2 cells upon treatment with IL-1 beta. Co-expression of constitutively active p65 strongly activated the pro moter construct carrying the intact NF-kappaB site in 293T and Caco-2 cells . Collectively, LARC expression in intestinal epithelial-type cells is indu ced by proinflammatory cytokines such as IL-1 and TNF-alpha primarily throu gh activation of NF-kappaB.