X Wu1, G Qian, Y Zhao, D Xu. 1. Institute of Respiratory Disease, Xinqiao Hospital, Third Military Medical University, Chongqing, 400037, China.
Abstract
OBJECTIVE AND DESIGN: 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. MATERIAL AND METHODS: 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. TREATMENT: U937 cells were treated with LPS, LBP, and indicated concentrations of P12. Mice were administered LPS intraperitoneally and P12 via the tail vein. RESULTS: 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. CONCLUSIONS: 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.
OBJECTIVE AND DESIGN: 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. MATERIAL AND METHODS: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. TREATMENT: U937 cells were treated with LPS, LBP, and indicated concentrations of P12. Mice were administered LPS intraperitoneally and P12 via the tail vein. RESULTS: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. CONCLUSIONS: 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.