Urinary trypsin inhibitor attenuates lipopolysaccharide-induced neutrophil activation

Abstract

BackgroundUrinary trypsin inhibitor (UTI), which is speculated to have anti-inflammatory effects, is one of serine protease inhibitors found in human urine and blood. The present study was conducted to clarify the effect of urinary trypsin inhibitor (UTI) on human neutrophil activation and its intracellular signaling mechanism in vitro.MethodsTo assess the possible interactions between UTI and lipopolysaccharide (LPS) in neutrophil activation, neutrophils from human blood were incubated with varying concentrations of UTI (1, 10, 100, 1,000 and 10,000 U/ml) plus LPS (100 ng/ml) or LPS alone in 24-well plates (5 × 106 cells/well). We measured protein levels for interleukin (IL)-6 and tumor necrosis factor-alpha (TNF-α) using enzyme-linked immunosorbent assay (ELISA) kits after 4 hours of incubation period. To elucidate the intracellular signaling pathway, we also measured the levels of phosphorylation of p38, ERK1/2 and JNK via Western blot analysis. Moreover, the nuclear levels of nuclear factor-kappa B (NF-κB) were determined with electrophoretic mobility shift assays (EMSA).ResultsUTI decreased the expression of inflammatory cytokines, including TNF-α and IL-6, and activation of intracellular signaling pathways, such as JNK, but not P38, ERK1/2 and nuclear translocation of NF-κB.ConclusionsUTI can attenuate LPS-induced neutrophil responses and may partially contribute to the treatment of neutrophil-mediated inflammatory diseases.