阿司匹林对Poly-IC诱导小胶质细胞激活的影响及其机制

Effect of aspirin on microglia activation induced by Poly-IC and its regulatory mechanism

目的研究阿司匹林对Poly-IC诱导小胶质细胞激活的影响及其机制。方法通过体外培养小鼠小胶质细胞BV2细胞株,建立Poly-IC刺激诱导小胶质细胞免疫激活模型。本研究分为对照组(不做处理)、模型组(Poly-IC 10μg/ml)、高剂量阿司匹林组(1 mmol/L阿司匹林)、低剂量阿司匹林组(0.1 mmol/L阿司匹林)、高剂量阿司匹林预处理组(Poly-IC 10μg/ml+1 mmol/L阿司匹林)、低剂量阿司匹林预处理组(Poly-IC 10μg/ml+0.1 mmol/L阿司匹林)。应用细胞免疫荧光法检测小胶质细胞吞噬能力﹑活性氧﹑Iba1蛋白表达,RT-qPCR法检测各组小胶质细胞炎症因子IL-1β﹑IL-6﹑IL-10﹑TNF-α﹑COX-2的mRNA表达。结果与对照组相比较,模型组中小胶质细胞形态发生改变,吞噬能力增强,活性氧产生增加,Iba1蛋白表达下降。且模型组中IL-1β(20.55±1.92)﹑IL-6(63.98±7.83)﹑TNF-α(16.84±3.19)﹑COX-2(6.78±0.42)的mRNA表达较对照组IL-1β(1.01±0.14)﹑IL-6(0.95±0.17)﹑TNF-α(1.22±0.38)﹑COX-2(0.87±0.11)显著增加(t=26.14,10.22,17.06,37.07;均P<0.01)。经阿司匹林预处理的小胶质细胞吞噬能力较模型组减弱,活性氧产生较模型组降低,且Iba1蛋白表达较模型组有一定恢复。高剂量阿司匹林预处理组促炎因子IL-1β(9.95±0.52)﹑IL-6(39.64±6.89)﹑TNF-α(1.57±0.42)﹑COX-2(2.47±0.14)的mRNA表达较模型组明显降低(t=14.18,3.69,16.68,27.03;均P<0.01)。结论阿司匹林对Poly-IC诱导小胶质细胞激活有抑制作用,其机制可能与阿司匹林抑制胶质细胞炎性因子的表达有关。

Objective To study whether aspirin has inhibitory effect on microglia activation induced by Poly-IC and its mechanism.Methods Microglia cell line BV2 were cultured in vitro to establish a Poly-IC stimulation-induced microglia cell immune activation model.The experiment groups were divided into control group(no treatment),model group(Poly-IC 10μg/ml),high dose aspirin group(1 mmol/L aspirin),low dose aspirin group(0.1 mmol/L aspirin),high dose aspirin pretreatment group(Poly-IC 10μg/ml+1 mmol/L aspirin)and low dose aspirin pretreatment group(Poly-IC 10μg/ml+0.1 mmol/L aspirin).The phagocytosis ability of microglia cells,reactive oxygen species(ROS)and Iba1 protein expression were detected by using immunofluorescence method.The expression of the inflammatory cytokines Il-1β,Il-6,Il-10,TNF-αand cox-2 mRNA in microglia cells were detected by real-time quantitative PCR(RT-qPCR).Results Compared with the control group,the morphology of microglia cells in model group changed significantly,and the phagocytosis ability and production of reactive oxygen species(ROS)increased.At the meantime,the expression of Iba1 protein was strongly decreased.In the model group,The mRNA expressions of IL-1β(20.55±1.92),IL-6(63.98±7.83),TNF-α(16.84±3.19),COX-2(6.78±0.42)were higher than IL-1β(1.01±0.14),IL-6(0.95±0.17),TNF-α(1.22±0.38),COX-2(0.87±0.11)in the control group.(Il-1β(t=26.14),Il-6(t=10.22),TNF-α(t=17.06)and COX-2(t=37.07),all P<0.01).In the aspirin pretreatment group,the phagocytic ability of microglia cells was inhibited compared with the model group,and the production of reactive oxygen species(ROS)reduced.The expression of Iba1 protein was also partly recovered.Meanwhile,the effect of the high aspirin dose pretreatment group on pro-inflammatory factors IL-1β(9.95±0.52),IL-6(39.64±6.89),TNF-α(1.57±0.42),COX-2(2.47±0.14)were lower than those in the model group significantly.(IL-1β:t=14.18,IL-6:t=3.69,TNF-α:t=16.68,COX-2:t=27.03,all P<0.01).Conclusion Aspirin has an inhibitory effect on microglial activation induced by Poly-IC,which may be related with inhibiting the expression of inflammatory factors.