Suppression of lipopolysaccharide-induced cytokine production of gingival fibroblasts by a soybean, Kunitz trypsin inhibitor

Background: Human bikunin, a Kunitz-type trypsin inhibitor, inhibits inflammation by down-regulating the expression of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) in tumor cells and inflammatory cells. Objectives: We analyzed the effect of a soybean-derived Kunitz trypsin inhibitor (KTI) on TNF-alpha production in human gingival fibroblasts stimulated by lipopolysaccharide (LPS), an inflammatory inducer. Material and methods: Mitogen-activated protein kinase (MAPK) activation and cytokine levels were monitored using western blot and a specific enzyme-linked immunosorbent assay (ELISA). Results: Here, we show (i) a soybean KTI abrogates LPS-induced up-regulation of TNF-alpha mRNA and protein expression in a dose-dependent manner in gingival fibroblasts, (ii) KTI also blocks the induction of TNF-alpha target molecules interleukin-1 beta (IL-1 beta) and IL-6 proteins, (iii) inhibition by KTI of TNF-alpha induction correlates with the suppressive capacity of extracellular signal-regulated kinase 1/2 (ERK1/2) and p38 signaling pathways, implicating repressed ERK1/2 and p38 signalings in the inhibition, and (iv) pretreatment of cells with KTI blocked LPS-induced nuclear factor kappa B (NF kappa B) activation. Conclusion: Our results indicate that KTI inhibits LPS-induced up-regulation of cytokine expression possibly through suppression of ERK1/2 and p38 kinase-mediated NF kappa B activation. These findings may identify anti-inflammatory properties of KTI at the level of gingival fibroblasts and may be relevant to the use of KTI in modulating inflammation, including periodontal disease.