Studies on uptake and effect of triclosan on production of inflammatory mediators in human gingival fibroblasts

Abstract: Triclosan (2,4,4'-trichloro-2'-hydroxy-diphenyl ether) is a lipid soluble anti-bacterial agent widely used in dentifrices, mouthrinses, soap and cosmetics. Although reports on the efficacy of triclosan on periodontal diseases are conflicting, it is documented in clinical studies that dentifrices containing triclosan reduce plaque accumulation and improve gingival health. Several clinical reports have suggested that the effect of triclosan on gingival inflammation might be related to its anti-inflammatory effect in addition to what can be accounted for the anti-bacterial effect of the agent. The present thesis is based on a series of experimental studies investigating the cellular uptake of triclosan and its effect on the production of inflammatory mediators using cultures of human gingival fibroblasts. The analyses were performed by Autoradiography, ELISA, EIA, RIA and Western Blot. mRNA expression was determined using in situ hybridization and reverse transcription polymerase chain reaction. Information about the intracellular distribution of triclosan is fundamental to understanding the mechanisms underlying its anti-inflammatory effects. Thus the initial Study (I) in this series was concerned with its uptake by gingival fibroblasts. It was found that triclosan was taken up rapidly and distributed in the nucleus and cytoplasm of gingival fibroblasts. The washing experiments showed that the agent remained associated with the nucleus even after repeated washing. Study II was conducted to investigate the effect of triclosan on the production of interleukin (IL)-1beta, IL-6 and prostaglandin E2 (PGE2). Triclosan reduced the production of cell associated IL-beta as well as the number of cells expressing IL-beta mRNA in gingival fibroblasts treated with tumor necrosis factor a (TNFalpha) Furthermore, triclosan reduced the TNFalpha induced PGE2 formation and the basal production of PGE 2 in control cells. On the contrary, triclosan did not affect the production of IL-6 induced by TNFalpha. In Study III, the effect of triclosan on the production of interferon-gamma (IFN-gamma) and the expression of major histocompatibility complex (MHC) class II antigen was studied. Treatment of gingival fibroblasts with triclosan reduced both IFN-gamma production as well as IFN-gamma-induced MHC class II expression. Based on the finding in Study II that triclosan reduces the production of PGE2 in gingival fibroblasts, Study IV explored the hypothesis that the agent may target the enzymes involved in the prostanoid biosynthetic cascade. Triclosan reduced the basal level as well as the stimulatory effect of TNFalpha on PGE2 biosynthesis, by reducing the mRNA and the protein expression of prostaglandin E synthase-1 (mPGES-1). Triclosan, however, did not affect the translocation of NF-kappaB or the expression of COX-2. The following conclusions are drawn from the results of these four studies: triclosan is taken up by gingival fibroblasts and remains associated with the nucleus. The agent attenuates the production of PGE2 by reducing mPGES-1 at the transcriptional and translational level. Triclosan reduces the formation of IL-1beta, IFN-gamma and MHC class II expression. The ability to inhibit the production of these important inflammatory mediators may have relevance for the clinically demonstrated anti-inflammatory effect of the agent.