Effects of Lycium barbarum Polysaccharides on Anti-inflammatory Activity and NF-κB Signaling Pathway Induced by LPS in BV2 Microglia
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摘要: 为探究枸杞多糖（Lycium barbarum polysaccharide，LBP）对脂多糖（lipopolysaccharide，LPS）诱导的BV2小胶质细胞的保护作用。本研究利用MTT法检测细胞活性，ELISA检测炎症因子的分泌，Western Blot检测蛋白表达情况。结果显示，单独LPS处理BV2小胶质细胞后，细胞的活性无显著变化而细胞上清液中的炎症因子PGE2、IL-1β、IL-6及TNF-α释放显著增多；将枸杞多糖梯度与LPS诱导的BV2小胶质细胞共孵育24 h后，有效地提升了LPS诱导的BV2小胶质细胞的活性，同时显著抑制了LPS激活的BV2小胶质细胞炎症因子的释放（P<0.05）；流式结果表明，LPS组小胶质细胞可产生大量ROS，小胶质细胞经不同浓度LBP处理12 h后，ROS含量显著降低（P<0.05）；Western Blot检测结果显示，与对照组相比，LPS组和LBP组中NF-κB信号通路相关蛋白表达及细胞核内p65蛋白表达水平均显著上调（P<0.05），且与LBP浓度成反比，而细胞质内p65蛋白表达显著降低（P<0.05），且随着LBP浓度增加而降低。本研究表明，枸杞多糖对LPS激活的BV2小胶质细胞有显著的保护作用，对于LPS激活的BV2小胶质细胞引起的炎症反应具有抑制作用，同时能抑制小胶质细胞中ROS的含量以发挥抗氧化应激作用，同时有效地抑制LPS刺激后细胞内p-TAK1、iκB、p-iκB及p-p65的表达。Abstract: To explore the protective effect of Lycium barbarum polysaccharide(LBP)on lipopolysaccharide(LPS)-induced BV2 microglia. In this study,MTT assay was used to detect cell viability,ELISA was used to detect the secretion of inflammatory factors,and Western Blot was used to detect protein expression. The results showed that the activity of BV2 microglia treated with LPS alone had no significant change,but the release of inflammatory factors PGE2,IL-1 β,IL-6 and TNF-α in the supernatant of BV2 microglia cells increased significantly. After LBP gradient was co-incubated with LPS-induced BV2 microglia for 24 h,the activity of LPS-induced BV2 microglia was effectively enhanced and the release of LPS-activated BV2 microglia inflammatory factors was significantly inhibited(P<0.05). The results of flow cytometry showed that microglia in LPS group could produce a large amount of ROS,and the content of ROS in microglia was significantly decreased after being treated with different concentrations of LBP for 12 h. The results of Western Blot showed that the protein expression of NF-κB signaling pathway and nucleusp 65 protein were significantly up-regulated in LPS group and LBP group,and were inversely proportional to LBP concentration,while p65 protein expression in cytoplasm of control group was significantly higher than that in LPS group and LBP group,and decreased with the increase of LBP concentration. This study showed that LBP had a significant protective effect on LPS activated BV2 microglia,inhibited the inflammatory response induced by LPS activated BV2 microglia,and inhibited the content of ROS in microglia to play an anti-oxidation stress effect,and effectively inhibited the expression of p-TAK1,iκB,p-iκB and p-p65 after LPS stimulation.
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