LPS致敏巨噬细胞产生O_2~及其信号机制

LPS induced macrophage priming effect on O_2~ production and its signal mechanisms

目的 :探讨LPS诱导的巨噬细胞 (MΦ)致敏及其信号机制。方法 :巨噬细胞样细胞株RAW 2 6 4 .7以10 μg/LLPS预处理 1h后洗去 ,再以 1mg/LPMA、1μmmol/LfMLP、10 0 μg/LLPS、1g/LZyms和 15mg/LMDP攻击 1h ,检测上清中超氧阴离子 (O 2 )的产生 ,并用荧光显微成像系统检测细胞内游离钙离子浓度 ( [Ca2 +]i) ,探讨 [Ca2 +]i与LPS致敏MΦ产生O 2 的关系。结果 :LPS预处理后以上述刺激剂攻击 1h均能不同程度致敏O 2 的产生 ,且致敏程度在一定范围内与LPS剂量和作用时间相关 ;细胞内静息态 [Ca2 +]i也呈LPS剂量和时间依赖性升高 ,均显著相关。且LPS预处理后用PMA攻击 ,细胞内瞬时 [Ca2 +]i升高幅度明显高于LPS未处理组。用A2 3187使细胞内 [Ca2 +]i升高可代替LPS致敏O 2 的产生 ,而预先用BAPTA和EGTA降低细胞内 [Ca2 +]i则可阻断LPS致敏O 2 的作用。结论 :LPS可致敏MΦ产生O 2 ;LPS预处理后细胞内静态 [Ca2 +]i的升高是LPS诱导O 2 致敏的重要原因。

AIM: To investigate the effect of lipopolysaccharide (LPS) priming on macrophage(MΦ). METHODS: Macrophage cell line RAW264.7 were pretreated with or without LPS for 1 h, then challenged with PMA, or LPS, muramyl dipeptide(MDP), Zymosan, formyl-methionyl-leucyl-phenylalanine(FMLP) for 1 h . O  2 production in supernatants and intracellular free calcium([Ca 2+ ]i ) were measured, and changes in [Ca 2+ ]i and LPS induced O  2 production were compared. RESULTS: LPS pretreatment significantly increased O  2 production in RAW264.7 cells challenged with the stimuli, and in a certain extent, both O  2 production and increase of resting intracellular [Ca 2+ ]i were dose- and time-dependent on LPS pretreatment.Furthermore,the peak [Ca 2+ ]i was significantly higher in LPS pretreated groups than that of LPS unpretreated groups when challenged with PMA. Pretreatment with Ca 2+ inophore A23187 mimicked the LPS priming effects on O  2 production, but pretreatment with Ca 2+ chelator BAPTA and EGTA blocked this priming effect. CONCLUSION: LPS induced MΦ priming effect on O  2 production is dependent on elevation of resting intracellular [Ca 2+ ]i .