金合欢素通过调节Sirt1介导的AMPK/Nrf2信号通路改善肺炎链球菌感染引起的肺泡上皮细胞损伤  

Acacetin ameliorates alveolar epithelial cell damage caused by Streptococcus pneumoniae infection by regulating Sirt1-mediated AMPK/Nrf2 signaling pathway

作　　者：范彩霞 张宗林[2] 伏瑶 姜红[2] FAN Caixia;ZHANG Zonglin;FU Yao;JIANG Hong(Clinical Pharmacology Research Center,Linyi People's Hospital,Linyi 276000,China;Department of Pharmacy,Linyi People's Hospital,Linyi 276000,China;Central Lab,Linyi People's Hospital,Linyi 276000,China)

机构地区：[1]临沂市人民医院临床药理研究中心,临沂276000 [2]临沂市人民医院药学部,临沂276000 [3]临沂市人民医院中心实验室,临沂276000

出　　处：《中国免疫学杂志》2024年第3期513-518,共6页Chinese Journal of Immunology

摘　　要：目的:探讨金合欢素通过调节沉默信息调节因子相关酶1(Sirt1)介导的5'-磷酸腺苷激活的蛋白激酶(AMPK)/核因子E2相关因子2(Nrf2)信号通路对肺炎链球菌(SP)感染引起的肺泡上皮细胞损伤的影响。方法:SP感染体外培养的肺泡上皮细胞A549建立细胞损伤模型,采用终浓度分别为0、5、25、50、100、150、200μmol/L的金合欢素处理,CCK-8检测各处理组细胞活力并筛选金合欢素最佳作用浓度。体外培养的A549细胞随机分为5组:对照组、模型组、金合欢素(150μmol/L)组、EX527(Sirt1抑制剂,40μmol/L)组、金合欢素(150μmol/L)+EX527组(40μmol/L),对照组不进行处理,其余各组以SP感染建立细胞损伤模型,150μmol/L金合欢素和40μmol/L EX527分别处理,CCK-8、流式细胞术分别检测各组细胞活力与凋亡率;试剂盒检测各组细胞活性氧(ROS)、超氧化物歧化酶(SOD)、丙二醛(MDA)、乳酸脱氢酶(LDH)水平及IL-10、IL-1β、TNF-α水平;免疫印迹法检测各组细胞增殖相关蛋白Ki-67、增殖细胞核抗原(PCNA)、凋亡相关蛋白caspase-9、Bax表达、Sirt1与AMPK/Nrf2信号通路蛋白p-AMPK/AMPK、Nrf2表达。结果:与对照组比较,模型组A549细胞活力、SOD、IL-10水平、p-AMPK/AMPK、Sirt1、Nrf2、Ki-67与PCNA蛋白表达降低(P<0.05),凋亡率、MDA、LDH、ROS水平、IL-1β、TNF-α水平升高(P<0.05)。与模型组、金合欢素+EX527组分别比较,金合欢素组A549细胞活力、SOD、IL-10水平、p-AMPK/AMPK、Sirt1、Nrf2、Ki-67与PCNA蛋白表达均升高(P<0.05),凋亡率、MDA、LDH、ROS、IL-1β、TNF-α水平均降低(P<0.05);EX527组A549细胞活力、SOD、IL-10水平、p-AMPK/AMPK、Sirt1、Nrf2、Ki-67与PCNA蛋白表达均降低(P<0.05),凋亡率、MDA、LDH、ROS、IL-1β、TNF-α水平均升高(P<0.05)。结论:金合欢素可通过上调Sirt1表达激活AMPK/Nrf2信号,进而促进抗炎因子分泌,减少ROS和促炎因子产生,减轻炎症与氧化应激,最终缓解神经元损伤。Objective:To investigate effect of acacetin on alveolar epithelial cell damage caused by Streptococcus pneumoniae(SP)infection by regulating sirtuin 1(Sirt1)-mediated 5'-AMP activated protein kinase(AMPK)/nuclear factor erythroid-2 related factor 2(Nrf2)signaling pathway.Methods:Alveolar epithelial cells A549 cultured in vitro were infected with SP to establish a cell damage model.After treatment with acacetin at final concentrations of 0,5,25,50,100,150,200μmol/L,CCK-8 was performed to detect cell viability of each treatment group and optimal concentration of acacetin was screened.A549 cells cultured in vitro were ran-domly separated into five groups:control group,model group,acacetin(150μmol/L)group,EX527(Sirt1 inhibitor,40μmol/L)group,acacetin(150μmol/L)+EX527(40μmol/L)group,control group was not treated,other groups were infected with SP to establish a cell damage model,and then treated with 150μmol/L acacetin and 40μmol/L EX527,CCK-8 and flow cytometry were performed to measure cell viability and apoptosis rate in each group;kits were performed to measure levels of reactive oxygen species(ROS),superoxide dismutase(SOD),malondialdehyde(MDA),lactate dehydrogenase(LDH)and IL-10,IL-1β,TNF-αlevels of cells in each group;Western blot was performed to measure proliferation-related proteins Ki-67,proliferating cell nuclear antigen(PCNA),apoptosis-related proteins caspase-9,Bax,Sirt1 and AMPK/Nrf2 signaling pathway proteins p-AMPK/AMPK,Nrf2 expres-sions of cells in each group.Results:Model group had decreased A549 cell viability,SOD and IL-10 levels,p-AMPK/AMPK,Sirt1,Nrf2,Ki-67 and PCNA protein expressions(P<0.05),and increased apoptosis rate,MDA,LDH,ROS,IL-1βand TNF-αlevels than control group(P<0.05).Compared with model group and acacetin+EX527 group,acacetin group had increased A549 cell viability,SOD and IL-10 levels,p-AMPK/AMPK,Sirt1,Nrf2,Ki-67 and PCNA protein expressions(P<0.05),and decreased apoptosis rate,MDA,LDH,ROS,IL-1βand TNF-αlevels(P<0.05);EX527 group had decreased A549 cell viability,SOD a

关 键 词：金合欢素 SIRT1 AMPK/Nrf2信号 SP 肺泡上皮细胞 损伤 

分 类 号：R378.14[医药卫生—病原生物学]