IL-6 production in human intestinal epithelial cells following stimulation with IL-1β is associated with activation of the transcription factor NF-κB

Recent studies suggest that interleukin-1β (IL-1β) stimulates interleukin-6 (IL-6) production in human intestinal epithelial cells, but the intracellular mechanisms of this response are not known. In other reports, the nuclear factor-κB (NF-κB) regulated IL-6 production in certain cell types. We tested the hypothesis that IL-6 production in the enterocyte is associated with activation of NF-κB. Caco-2 cells, a human intestinal epithelial cell line, were grown in tissue culture whereafter they were treated with IL-1β (0.5 ng/ml). Cells were preincubated with pyrrolidine dithiocarbamate (PDTC; 10-500 μM), tosyl-lys-chloromethylketone (TLCK; 10- 500 μM), or genistein (25-75 μM), all of which are known inhibitors of NF- κB. IL-6 levels in the culture media were measured after 24 hr by enzyme- linked immunosorbent assay (ELISA) and IL-6 messenger RNA (mRNA) levels were determined after 4 hr by competitive reverse-transcriptase polymerase chain reaction (RT-PCR). NF-κB activity was determined by electrophoretic gel mobility shift assay (EMSA). PDTC, TLCK, and genistein each inhibited IL- 1β-induced IL-6 production by the Caco-2 cells in a dose-dependent fashion. These responses were also associated with a decrease in IL-6 mRNA levels. There was no NF-κB activity in untreated cells, but the addition of IL-1β resulted in the activation of NF-κB as determined by EMSA. The results suggest that IL-1β-induced IL-6 production in the enterocyte is associated with activation of NF-κB. The inhibition of IL-6 production by the NF-κB inhibitors indicates that the IL-6 production is regulated by NF-κB, although further experiments are needed to test that hypothesis.