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目的研究佛波酯(PMA)对脂多糖/γ-干扰素(LPS/IFN-γ)诱导的巨噬细胞RAW264.7活性氧产生的影响。方法于实验前0、3、6、9、12、18h,用1μg/mL的LPS和100 U/mL IFN-γ诱导处于对数生长期的RAW264.7细胞,采用蛋白质印迹技术检测细胞中一氧化氮合酶(iNOS)的表达;对经LPS/IFN-γ诱导12h的RAW264.7细胞加入终浓度为200ng/mL的PMA,应用激光共聚焦显微技术观察RAW264.7细胞中活性氧的产生。结果 LPS/IFN-γ可使RAW264.7细胞的诱导型iNOS表达增加,同时PMA可刺激经LPS/IFN-γ诱导后的RAW264.7细胞产生呼吸爆发,细胞的荧光强度较未刺激组增强(P<0.01)。结论 PMA可促使经LPS/IFN-γ诱导的RAW264.7细胞产生大量活性氧。
Objective To investigate the effect of phorbol ester (PMA) on reactive oxygen species production by macrophages induced by LPS / IFN-γ. Methods RAW264.7 cells in logarithmic growth phase were induced with 1μg / mL LPS and 100 U / mL IFN-γ at 0, 3, 6, 9, 12 and 18 h before the experiment. Nitric oxide synthase (iNOS) was detected in RAW264.7 cells. The RAW264.7 cells induced by LPS / IFN-γ were treated with PMA at a final concentration of 200ng / mL. Laser scanning confocal microscopy was used to observe the expression of reactive oxygen species produce. Results LPS / IFN-γ could induce iNOS expression in RAW264.7 cells, while PMA could stimulate the respiratory burst of RAW264.7 cells induced by LPS / IFN-γ, and the fluorescence intensity of cells increased compared with that of unstimulated group P <0.01). Conclusion PMA can induce large amount of reactive oxygen species (ROS) in RAW264.7 cells induced by LPS / IFN-γ.