脂氧素对脂多糖诱导巨噬细胞炎症相关因子的影响及机制

Effects and mechanism of lipoxin on LPS-induced inflammatory mediators in RAW264.7 macrophages

目的:研究脂氧素(lipoxin A4)对脂多糖(lipopo-lysaccharide,LPS)诱导的小鼠巨噬细胞株RAW264.7中相关炎症因子如肿瘤坏死因子(TNF-α)、环加氧酶-2(COX-2)、前列腺素E2(PGE2)、血红素氧化酶-1(HO-1)和白介素-l0(IL-10)的影响,并进一步探讨其内在分子机制.方法:以1mg/LLPS刺激体外培养的RAW264.7细胞作为炎症模型,分别用脂氧素A4或脂氧素A4和ZnPPIX干预LPS刺激的RAW264.7细胞24 h后,收集培养上清并提取细胞总蛋白,酶联免疫法测定上清中TNF-α,IL-l0和PGE2浓度,免疫印迹法检测细胞COX-2和HO-l的蛋白表达量,分光光度计分析HO-1的活性.结果:1 mg/L LPS明显诱导TNF-α,COX-2和PGE2的生成,也适度增加IL-10及HO-l的产生;脂氧素A4可抑制LPS诱导的TNF-α(P<0.01vsLPS组),COX-2和PGE2(P<0.01vsLPS组)的生成,却进一步增加IL-10(P<0.01vsLPS组)和HO-1表达量和活性(P<0.01vsLPS组);ZnPPIX可减弱脂氧素A4对LPS诱导TNF-α(P<0.05vsLPS+脂氧素A4组),COX-2和PGE2(P<0.05vsLPS+脂氧素A4组)的生成抑制作用,同时也下调IL-10的生成量.结论:脂氧素A4可抑制LPS诱导的巨噬细胞中致炎介质TNF-α,COX-2及PGE2的生成,同时也促进IL-10的产生,这种效应在一定程度上是通过上调HO-1表达和活性实现.

AIM： To investigate the effects of lipoxin A4 on lipopolysaccharide （LPS）-induced inflammatory mediators in RAW264.7 macrophages and its underlying molecular mechanism. METHODS： RAW264.7 cells were stimulated with 1 mg/L LPS to induce inflammatory response and then lipoxin A4 at 100 ng/mL was administrated. ZnPPIX was added to RAW264. 7 cells by eotreatment of LPS and lipoxin An. The concentrations of TNF-α, IL-10 and PGE2 in the cell supernatants were measured by ELISA. COX-2 and HO-1 proteins were detected by Western blotting and HO-1 activity was analyzed by spectrophotometer. RESULTS： Compared with that in LPS group, lipoxin A4 significantly inhibited LPS-induced COX-2 protein expression and levels of TNF-α and PGE2 in RAW264.7 cells （ both, P 〈0.01） and this effect was accompanied by a parallel increase in HO-1 and IL-10（P〈0.01）. Treatment of ZnPPIX partially abolished the suppressive effects of lipoxin A4 on COX-2 protein expression and the levels of TNF-α and PGE2 in RAW264.7 cells induced by LPS, compared with those in LPS ＋ Lipoxin A4 group （ both, P 〈0.05）. CONCLUSION： Lipoxin A4 inhibits LPS-induced proinflammatory mediators, including COX-2, PGE2 and TNF-α,possibly by mediation through HO-1 pathway.