咖啡酸苯乙酯对抗脂多糖诱导的小胶质细胞炎症反应的作用及其机制的研究

Inhibitory Effects of Caffeic Acid Phenethyl Ester on Inflammatory Responses of Microglial Cells(BV-2)Induced by Lipopolysaccharide

目的探究咖啡酸苯乙酯对脂多糖诱导的小胶质细胞BV-2神经炎症反应的抑制作用及其潜在的作用机制。方法采用脂多糖(1μg·mL-1)诱导小胶质细胞BV-2活化,制作炎症反应模型,利用四甲基偶氮唑蓝法检测细胞存活率,并用酶联免疫法测定咖啡酸苯乙酯对炎症因子一氧化氮、白细胞介素-1β和白细胞介素-6合成释放的影响。然后用蛋白印迹法研究了咖啡酸苯乙酯对炎症相关蛋白(诱导型一氧化氮合酶、环氧化酶-2、IκB以及P-IκB)表达的作用。此外,进一步研究了咖啡酸苯乙酯对炎症转录因子NF-κB核转位的作用。结果咖啡酸苯乙酯与脂多糖(1μg·mL-1)同时作用于BV-2小胶质细胞,分别于温箱中孵育24、8 h收集上清液,检测一氧化氮、白细胞介素-1β、白细胞介素-6的释放量。结果显示,脂多糖(1μg·mL-1)可以造成细胞损伤,增加炎症因子释放,咖啡酸苯乙酯则可以降低脂多糖诱导的小胶质细胞对一氧化氮、白细胞介素-1β和白细胞介素-6的释放,并可抑制诱导型一氧化氮合酶、环氧化酶-2、P-IκB、P-NF-κB蛋白的表达,还可以抑制NF-κB的核转位。结论上述研究说明,咖啡酸苯乙酯(5,10,20μmol·L-1)可以通过抑制炎性因子的表达而减少对小胶质细胞的损伤,其抗炎活性可能是通过抑制NF-κB信号通路介导的炎症反应实现的。

OBJECTIVE To investigate the inhibitory effects of caffeic acid phenethyl ester （CAPE） on LPS-induced inflammatory responses in BV-2 microglial cells and its mechanism. METHODS The model was build by BV-2 microglial cells induced by LPS （μg·mL^-1 ） , which was used to inspect the cell survivability by means of MTT, investigate the effect of CAPE on the release of inflammatory factor NO, IL-1β, IL-6 by ELISA, and the expression of inflammation-related protein （iNOS, COX-2, IκB, P-IKB, NF- κB, P-NF-κB） were analyzed by used of Western Blot. Finally, the regulatory effects of CAPE on inflammatory transcription factor NF- κB activation were inspected. RESULTS CAPE and LPS affected on BV-2 cells, which are collected after being incubated at 37 for 24 or 8 h, then test the release of NO, IL-1 β, IL-6. As is shown, LPS （ 1 μg·mL^-1 ） can injury the cells,and increase inflammatory cytokines. CAPE （5, 10, 20 μg·mL^-1 ） can decrease these inflammatory cytokines, down-regulate iNOS, COX-2, IKB, P-IKB expression and restrain NF-κB nuclear transloeation. CONCLUSION The anti-inflammatory activity of CAPE may be achieved through inhibiting NF-κB signaling pathway.