摘要
目的探讨槟榔碱对脂多糖(LPS)诱导小鼠小胶质细胞BV2炎症反应的改善作用及机制。方法取对数生长期的BV2细胞分为空白组和LPS(0.01、0.1、1、10、20μg/mL)组,CCK-8法检测细胞活力,分光光度法检测一氧化氮(NO)含量。另取细胞分为空白组、模型组、槟榔碱(10、20、40μmol/L)组。CCK-8法检测细胞活力;分光光度法检测NO含量;酶联免疫吸附法(ELISA)检测上清中肿瘤坏死因子-α(TNF-α)、白细胞介素6(IL-6)、白细胞介素1β(IL-1β)和抗炎因子白细胞介素10(IL-10)水平;实时荧光定量PCR(qPCR)法检测细胞中TNF-α、IL-6、IL-1β、一氧化氮合酶(iNOS)mRNA表达表达;Western blotting法检测Toll样受体4(TLR4)、p-p65、p65、环氧化酶2(COX2)、iNOS、胞内磷脂酰肌醇激酶(PI3K)、蛋白激酶B(Akt)、p-Akt蛋白表达水平。结果0.01~20μg/mL LPS诱导对BV2小胶质细胞的细胞活力无显著影响,1μg/mL LPS诱导显著上调了细胞中NO含量;槟榔碱在10~40μmol/L内对细胞活力无显著影响,且均可缓解LPS诱导的BV2小胶质细胞损伤。与模型组相比,槟榔碱组可降低细胞内NO含量,下调炎症因子TNF-α、IL-6、IL-1βmRNA相对表达量及其血清水平;抑制TLR4、p-P65、iNOS、COX2、PI3K、p-Akt蛋白表达量。结论槟榔碱对LPS诱导的BV2小胶质细胞炎症模型具有显著改善作用,其作用机制可能与抑制NO生成、下调iNOS表达、调控TLR4/核因子-κB(NF-κB)、PI3K/Akt信号通路有关。
Objective To investigate the effects of arecoline on lipopolysaccharide(LPS)induced BV2 inflammatory response in mouse microglia and its mechanisms.Method BV2 cells were divided into blank group and LPS group(0.01,0.1,1,10,20μg/mL).The cell viability was detected by CCK-8 method,and the content of NO was detected by spectrophotometry.Cells were divided into blank group,model group and arecoline group(10,20,40μmol/L).Cell viability was detected by CCK-8 method.NO content was detected by spectrophotometry.The levels of TNF-α,IL-6,IL-1β,and anti-inflammatory factor IL-10 in supernatant were detected by enzyme-linked immunosorbent assay(ELISA).Real-time fluorescence quantitative PCR(qPCR)was used to detect the mRNA expression of TNF-α,IL-6,IL-1β,iNOS in cells.The expression levels of TLR4,p-P65,P65,COX2,iNOS,PI3K,Akt,and p-Akt were detected by Western blotting.Result In the concentration range(0.01—20μg/mL),LPS induction had NO significant effect on the cell viability of BV2 microglia,and 1μg/mL LPS induction significantly increased the content of NO in the cells.Arecaline had no significant effect on cell viability in the range of 10 to 40μmol/L,and could alleviate LPS-induced damage of BV2 microglia.Compared with the model group,arecoline group could decrease the intracellular NO content,down-regulate the mRNA expression of inflammatory factors TNF-α,IL-6,IL-1βand their serum levels.The expression levels of TLR4,p-P65,iNOS,COX2,PI3K and p-Akt were inhibited.Conclusion Arecarecine can significantly improve the LPS-induced inflammation model of BV2 microglia,and its mechanism may be related to inhibiting NO production,down-regulating the expression of iNOS,and regulating the TLR4/NF-κB and PI3K/Akt signaling pathways.
作者
孙源
王丹阳
孙晶
范蓓
宋洪波
刘新民
卢聪
王凤忠
SUN Yuan;WANG Danyang;SUN Jing;FAN Bei;SONG Hongbo;LIU Xinmin;LU Cong;WANGFengzhong(Institute of Food Science and Technology,Chinese Academy of Agricultural Sciences,Beijing 100193,China;College of food science,Fujian Agriculture and Forestry University,Fuzhou 350002,China;Institute of New Drug Technology,Ningbo University,Ningbo 315000,China)
出处
《现代药物与临床》
CAS
2024年第3期541-548,共8页
Drugs & Clinic
基金
国家重点研发计划(2022YFD1600303)
中国农业科学院农产品加工研究所创新工程(CAAS-ASTIP-2023-IFST)
三亚中国农业科学院国家南繁研究院“南繁专项”(YYLH05)
三亚中国农业科学院国家南繁研究院“南繁专项”(ZDXM2302)。
