低镁对脂多糖诱导巨噬细胞表达和释放高迁移率族蛋白1的影响

Effect of low magnesium on the expression and release of high mobility group box-1 in macrophages induced by lipopolysaccharide

目的探讨低镁对脂多糖(LPS)诱导小鼠巨噬细胞RAW264.7表达和释放高迁移率族蛋白1(HMGB1)的促进作用。方法传代培养的小鼠RAW264.7细胞分为4组,接种于6孔板,每组为3孔,4组分别为含1mmol/L硫酸镁的RPMI1640培养基组(C1组)、含0.1mmol/L硫酸镁的RPMI1640培养基组(C0.1组)、含1mmol/L硫酸镁的RPMI1640培养基加500ng/mLLPS的刺激组(L1组)和含0.1mmol/L硫酸镁的RPMI1640培养基加500ng/mLLPS的刺激组(L0.1组)。孵育24h后,收集细胞及细胞培养上清液,用反转录-聚合酶链反应和酶联免疫吸附试验分别检测各组细胞内HMGB1mRNA表达水平和细胞培养上清液中HMGB1蛋白含量的变化。结果 C0.1组与C1组间HMGB1mRNA及蛋白表达的差异均无统计学意义(P值均>0.05),L1组和L0.1组HMGB1mRNA及蛋白的表达均显著高于C0.1组和C1组(P值均<0.05),L0.1组又显著高于L1组(P值均<0.05)。结论低镁促进LPS诱导的RAW264.7细胞表达和释放HMGB1,可能对脓毒症患者的预后有一定损害作用。

Objective To explore the effect of low magnesium on the expression and release of the high mobility group box-1 （HMGB1） in mouse macrophage RAW264.7 induced by lipopolysaccharide （LPS）. Methods RAW264.7 cells were incubated in 6-well tissue culture plates with RPMI1640 medium and divided into 4 groups, each with three holes： C1 group containing 1 mmol/L magnesium sulphate; CO. 1 group containing 0. 1 mmol/L magnesium sulphate; L1 group containing 1 mmol/L magnesium sulphate ＋ 500 ng/mL LPS; and L0. 1 group containing 0. 1 mmol/L magnesium sulphate ＋ 500 ng/mL LPS. The HMGB1 expression and releasing were detected 24 h after incubation by reverse transcription-polymerase chain reaction （RToPCR） and enzyme linked immunosorbent assay （ELISA）, respectively. Results There were no significant differences in the HMGB1 mRNA and protein expression between C1 group and C0. 1 group （both P~〉 0. 05）. The HMGB1 mRNA and protein expression in L1 group and L0.1 group were significantly higher than those in C1 group and CO. 1 group （both P〈0.05）. And the two indices in L0. 1 group were significantly higher than that in L1 group （all P〈0. 05）. Conclusion Low magnesium promotes the expression and release of HMGB1 in LPS-induced RAW264.7 cells, which may play a negative role in the prognosis of sepsis. （Shanghai Med J, 2012, 35= 302-305）