A vitamin D₃ analog augmented interleukin-8 production by human monocytic cells in response to various microbe-related synthetic ligands, especially NOD2 agonistic muramyldipeptide

Active metabolite vitamin D₃, 1α,25-dihydroxyvitamin D₃, is a pleiotropic factor and exhibits various physiological functions, including immunomodulating activities. In this study, the possible regulation of innate immune responses of human monocytic cells by a vitamin D₃ analog was examined. Human monocytic THP-1 cells were pre-treated with OCT, vitamin D₃ analog, 1α,25-dihydroxy-22-oxavitamin D₃, followed by stimulation with various chemically synthesized Toll-like receptors (TLR) and NOD1 and NOD2 ligands. OCT-treated cells produced more IL-8 than non-treated cells upon stimulation with various chemically-synthesized ligands: TLR2-agonistic lipopeptide (FSL-1), TLR3-agonistic poly I:C, TLR4-agonistic lipid A (E. coli-type LA-15-PP), NOD1-agonistic FK565 and NOD2-agonistic muramyldipeptide (MDP). Among the ligands, MDP was the highest inducer of IL-8 production in OCT-treated THP-1 cells, and IL-8 production increased depending on the treatment time until 72 h. OCT up-regulated the expression of NOD2 in THP-1 cells, and OCT-treated cells exhibited higher activation of p38, JNK and ERK in the MAPK pathway, IκBα in the NF-κB pathway, and TAK1 upstream in response to MDP than non-treated cells. Analysis using siRNA against NOD2 and inhibitors of specific signal molecules indicated that the existence of NOD2 and activation of the above signaling molecules are required for enhanced production of IL-8 in OCT-treated THP-1 cells. These findings suggested that NOD2, NF-κB and MAPK pathways are involved in the activity of OCT to augment the response of human monocytic cells to MDP.