LBP inhibitory peptide reduces endotoxin-induced macrophage activation and mortality

Abstract

The aim of this study was to investigate whether P12, a lipopolysaccharide (LPS)-binding protein (LBP) inhibitory peptide could reduce LPS induced inflammation in vitro and in vivo. Human monocyte-like cell line (U937 cells) was grown in RPMI 1640 and stimulated with PMA in order to induce differentiation to the macrophage stage. A total of 70 Kunming mice (8-12 wk old) were used in our experiments. The effects of P12 on the binding of LPS to U937 cells and alveolar macrophages (AMs) were determined by flow cytometric analysis. Nuclear factor kappa B (NF-kappa B) translocation was evaluated with subunit P65 by Western blotting. The production of tumor necrosis factor-alpha (TNF-alpha), alanine transaminase (ALT), and nitric oxide (NO) as measured by ELISA, enzymatic activity assay, and enzymatic assay with nitrate reductase. Differences among groups were determined using one-way ANOVA test and Fisher exact test. U937 cells were treated with LPS, LBP, and indicated concentrations of P12. Mice were administered LPS intraperitoneally and P12 via the tail vein. P12 inhibited the binding of FITC-conjugated LPS (FITC-LPS) to U937 cells and AMs. NF-kappa B translocation and the production of TNF-alpha, ALT, and NO induced by LPS was also significantly suppressed by P12. Furthermore P12 protected mice from LPS-induced death. The results suggest that blockade of LBP at inflammation sites might attenuate LPS-induced circulatory shock. This results in a beneficial effect in a mouse model of endotoxemia.