摘要
目的探讨银杏酮酯(GBE50)通过抑制小胶质细胞的炎症反应对神经元突触损伤的影响。方法①采用格里斯(Griess)试剂检测不同浓度GBE50(25~400 mg·L^(-1))干预对经脂多糖(LPS)刺激后BV2小胶质细胞培养液中亚硝酸盐含量的影响。②不同浓度GBE50(25~400 mg·L^(-1))干预经LPS刺激的BV2小胶质细胞,收集上清液作为条件培养基(CM)处理PC12细胞,采用四甲基偶氮唑盐比色(MTT)法检测BV2小胶质细胞和PC12细胞活力。③将对数生长的BV2和PC12细胞以2×106个/孔铺板于6孔板中,分为空白对照组(Ctrl组,仅培养液培养),对照加药组(Ctrl+GBE50组,GBE50干预浓度为200 mg·L^(-1)),模型组(LPS组,LPS刺激浓度为100μg·L^(-1)),模型加药组(LPS+GBE50组,GBE50预处理1 h后,加入LPS),各组进行相应干预。采用Western blot、实时荧光定量逆转录聚合酶链式反应(RT-qPCR)法检测CM处理的PC12细胞N-甲基-D-天门冬氨酸受体(NMDAR)亚基蛋白及基因表达量。结果①Greiss法结果显示,GBE50干预可降低LPS刺激BV2细胞产生的一氧化氮(NO)水平(P<0.05)。②MTT法结果显示,GBE50对BV2细胞活力无影响;LPS刺激后的CM处理PC12细胞活力显著下降,该现象在GBE50干预的CM中有所改善,GBE50浓度在200 mg·L^(-1)时细胞活力已无明显下降(P<0.05)。③Western blot结果显示,与LPS组比较,LPS+GBE50组中NMDAR亚基NR1、NR2A和NR2B蛋白表达升高(P<0.05);RT-qPCR结果显示,与LPS组比较,LPS+GBE50组中NMDAR亚基基因表达量均增加(P<0.05)。结论GBE50可通过抑制激活小胶质细胞的炎症反应,减少NMDAR的损伤,对神经元突触起到保护作用。
Objective To investigate the effect of Ginkgo biloba extract 50(GBE50)on synaptic damage of neurons by inhibiting the inflammatory response of microglia.Methods①Griess reagent was used to detect the effect of different concentrations of GBE50(25-400 mg·L^(-1))on the content of nitrite in the culture medium of BV2 microglia after LPS stimulation.②LPS stimulated BV2 microglia were treated with different concentrations of GBE50(25-400 mg·L^(-1)),and the supernatant was collected as conditioned medium(CM)to treat PC12 cells.MTT assay was used to detect the viability of BV2 microglia and PC12 cells.③Logarithmic BV2 and PC12 cells were laid in a 6-well plate with 2×106 cells/well plates and divided into blank control group(Ctrl group,cultured only in culture medium),control plus drug group(Ctrl+GBE50 group,GBE50 intervention concentration was 200 mg·L^(-1)),model group(LPS group,LPS stimulation concentration was 100μg·L^(-1)),model treatment group(LPS+GBE50 group,GBE50 pretreatment for 1 h,adding LPS),and each group was intervened accordingly.The protein and gene expression of N-methyl-D-aspartate receptor(NMDAR)subunit in PC12 cells treated with CM were detected by Western blot and real-time fluorescence quantitative reverse transcriptase polymerase chain reaction(RT-qPCR).Results①The results of Greiss method showed that GBE50 intervention reduced the level of nitric oxide(NO)produced by BV2 cells stimulated by LPS(P<0.05).②The results of MTT assay showed that GBE50 had no effect on the viability of BV2 cells,but the viability of PC12 cells treated with CM stimulated by LPS decreased significantly,which was improved in CM treated with GBE50,and there was no significant decrease in cell viability when the concentration of GBE50 was 200 mg·L^(-1)(P<0.05).③Western blot results showed that compared with the LPS group,the protein expression of NMDAR subunits NR1,NR2A and NR2B in the LPS+GBE50 group was increased(P<0.05).RT-qPCR results showed that compared with the LPS group,the NMDAR subunit gene expres
作者
热爱拉·阿合力江
刘新娟
杨光
童雨婷
高外毛
胡凯帆
潘洁
高崎
邢越阳
王星禹
赵妍
王健
徐颖
AHELIJIANG Reaila;LIU Xinjuan;YANG Guang;TONG Yuting;GAO Waimao;HU Kaifan;PAN Jie;GAO Qi;XING Yueyang;WANG Xingyu;ZHAO Yan;WANG Jian;XU Ying(School of Rehabilitation,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China;School of Integrative Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China;SPH Xing Ling Sci.&Tech.Pharmaceutical Co.,Ltd,Shanghai 201712,ChinaAbstract)
出处
《上海中医药杂志》
CSCD
2024年第6期82-87,100,共7页
Shanghai Journal of Traditional Chinese Medicine
基金
国家自然科学基金面上项目(82174003)
上海市教委产教融合项目(2022310031002310)。
关键词
神经退行性疾病
银杏酮酯
小胶质细胞
神经元
炎症反应
中药研究
作用机制
neurodegenerative diseases
Ginkgo biloba extract 50
microglia
neurons
inflammatory response
traditional Chinese herbal medicine research
mechanism of action
