丁苯酞通过诱导自噬调控BV-2小胶质细胞炎症反应

Butylphthalide regulates the inflammatory response of BV-2 microglial cells by inducing autophagy

目的研究丁苯酞(NBP)对β-淀粉样蛋白(Aβ25-35)诱导的BV-2小胶质细胞炎症反应的影响及可能机制。方法培养BV-2小胶质细胞,Aβ25-35诱导构建阿尔茨海默病(AD)炎症反应细胞模型,将其分为空白对照组(Control组)、模型组(Model组)、丁苯酞组(NBP)和丁苯酞联合6-氨基-3-甲基嘌呤(3-MA)组(3-MA组)。采用CCK-8法检测NBP和Aβ25-35对BV-2小胶质细胞活力的影响,Western blotting检测各组细胞自噬蛋白LC3B-Ⅱ和p62的表达情况,实时PCR检测自噬基因ATG5、Beclin1和炎症指标白细胞介素(IL)-1β、IL-6 mRNA的表达水平,ELISA检测细胞上清液中IL-1β的分泌量。结果Aβ25-35浓度≥20μmol/L时对细胞生存率具有显著损伤作用,NBP浓度≤40μmol/L时细胞活力与未处理细胞基本无差异。Model组IL-1β、IL-6 mRNA表达量较Control组明显上调,细胞上清液中IL-1β的分泌量增加(P<0.05)。与Model组相比,NBP组IL-1β、IL-6 mRNA表达量显著降低,ATG5、Beclin1 mRNA和LC3B-Ⅱ蛋白表达上调,p62蛋白明显减少,细胞上清液中IL-1β含量降低(P<0.05)。与NBP组相比,3-MA组IL-1β、IL-6 mRNA表达增加,ATG5、Beclin1 mRNA和LC3B-Ⅱ蛋白表达降低,p62蛋白和细胞上清液中IL-1β的分泌量增加(P<0.05)。结论NBP可能通过诱导自噬减轻Aβ25-35构建的BV-2小胶质细胞炎症反应。

Objective To investigate the effect of butylphthalide(NBP)onβ-amyloid protein(Aβ)25-35-induced inflammatory response of BV-2 microglial cells and the underlying mechanism.Methods Related indicators were assessed by CCK-8 assay,Western blotting,real-time PCR,and ELISA.Results The expression levels of IL-1βand IL-6 in the model group were significantly upregulated compared with those in the control group(P<0.05).However,compared with the Model group,the NBP group showed significantly downregulated IL-6 and IL-1βmRNA and protein,and p62 protein levels,and significanlty upregulated ATG5 and Beclin1 mRNA,and LC3B-Ⅱprotein levels(P<0.05).These effects of NBP were significantly suppressed by the PI3-K/autophagy inhibitor 3-methyladenine(P<0.05).Conclusion NBP may reduces the Aβ25-35-induced inflammatory response of BV-2 microglial cells through autophagy pathway.