炎性因子IL-1β对星形胶质细胞抗氧化应激能力的影响

Interleukin-1β Alters Anti-oxidative Regulation of Primary Cultured Astrocytes

目的:本研究应用原代培养星形胶质细胞,探讨炎症因子白细胞介素-1β(IL-1β)对星形胶质细胞抗氧化应激能力的影响。方法:在原代星形胶质细胞培养中分别加入1 ng/ml IL-1β,流式细胞仪检测不同作用时间(0、24、48、72 h)细胞线粒体膜电位改变,激光共聚焦显微镜观察活性氧(ROS)生成,分光光度法检测细胞内谷胱甘肽(GSH)含量,RT-PCR、Western Blot检测星形胶质细胞谷胱甘肽过氧化物酶(GSH-Px)的表达变化。结果:1 ng/ml IL-1β处理原代培养星形胶质细胞24~72 h,细胞线粒体膜电位无明显改变,细胞未出现明显损伤。IL-1β孵育24 h,细胞内ROS生成减少,GSH生成增加;48 h细胞内ROS生成增加,GSH较24 h组减低(P<0.05);72 h,原代培养星形胶质细胞内ROS生成明显增加,GSH较0 h组降低(P<0.05)。1 ng/ml IL-1β处理原代培养星形胶质细胞24 h,细胞内GSH-Px mRNA及蛋白水平较0 h组升高(P<0.05);但仅是一过性反应,IL-1β孵育72 h后,原代培养星形胶质细胞内GSH-Px mRNA及蛋白水平均较0 h组降低(P<0.05)。结论:持续炎性因子刺激下,星形胶质细胞抗氧化应激能力呈现动态改变,并且细胞内抗氧化物参与此变化过程。

Objective： To elucidate whether intracellular redox regulation could be altered in astrocytes responding to pro-inflammatory cytokine interleukin-1β （IL-1 β）. Methods：Primary cultured astroeytes were treated with 1 ng/ml IL-1β for 0, 24, 48 and 72 h. The effects of IL-1β on intracellular reactive oxide species （ROS） production, mitochondrial membrane potential （△ψm） changes, intracellular glutathione （GSH） content and protein as well as mRNA expressions of glntathione peroxidase （GSH-Px） were assayed by using laser confocal scanning, flow cytometry, Western blot and RT- PCR. Results：No significant A^ktm changes were observed in primary cultured astrocytes after 1 ng/ml IL-113 incubation for 24, 48 and 72h. A time-dependent production of ROS and GSH were observed. After incubated with IL-1 [3 for 24 h, decreased ROS production and increased GSH content intracellular （ P 〈 0.05 ） were detected. At 48 h, the ROS and GSH were decreased, and minimum ROS and GSH concentration were measured after 72 h incubation compared with 0 h group （ P 〈 0.05 ）. After treatment with IL-1 [3 for 24 h, down-regulation of GSH-Px mRNA and protein levels were ob- served, but returned to basal level at 48 h. Significant decreased mRNA and protein expressions of GSH-Px were found at 72 h compared with 0 h group （ P 〈 0.05 ）. Conclusion ： With sustained inflammatory factor＇s stimulation, redox regulation capacity of astrocyes changed dynamically. And intracellular anti-oxidative contents involved in this process.