佛波酯对LPS/IFN-γ诱导的巨噬细胞活性氧产生的影响

The influence of phorbol-1,2-myristate-1,3-acetate on the production of reactive oxygen species by LPS/IFN-γ inducted in RAW264.7 macrophages

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 study the influence of phorbol-1,2 myristate-1,3-acetate （PMA） on the production of reactive oxygen species（ROS） by LPS/IFN-γ inducted in RAW264.7 macrophages. Methods RAW264.7 macrophages were stimulated with 1 μg/mL I.PS and 100 U/mL IFN-γ at 0,3,6,9,12,18 h, respectively. The expression of inducible nitric oxide synthase （iNOS） were assayed by western blotting. The 200 ng/mL PMA was added at last concentration to detected the production of ROS by laser scanning confocal microscope. Results The expression of iNOS increased significantly after treatment by LPS/IFN-γ. The inducible RAW264.7 macrophages produced respiratory burst by stimulated with PMA. Conclusion PMA promote a great amount of reactive oxygen species（ROS） produced in the inducible RAW264.7 macrophages with LPS/IFN -γ.