连翘脂素对脂多糖联合正常人血浆诱导的A549细胞炎症损伤的作用  

Effect of phillygenin on inflammatory response of A549 cells induced by lipopolysaccharide and normal human plasma

作　　者：郭静 张启云[2,4] 靳翔 薛焕焕 路青瑜 郭丽 孙黔云[2,4] 张立伟 GUO Jing;ZHANG Qi-yun;JIN Xiang;XUE Huan-huan;LU Qing-yu;GUO Li;SUN Qian-yun;ZHANG Li-wei(Mordern Research Center for Traditional Chinese Medicine,Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education,Institute of Molecular Science,Shanxi University,Taiyuan 030006,China;The Key Laboratory of Chemistry for Natural Products,Guizhou Province and Chinese Academy of Science,Guiyang 550014,China;KweiChow Moutai Hospital,Renhuai Guizhou 564500,China;State Key Laboratory of Function and Applications of Medicinal Plants,Guizhou Medical University,Guiyang 550014,China)

机构地区：[1]山西大学,分子科学研究所化学生物学与分子工程教育部重点实验室,中医药现代研究中心,山西太原030006 [2]贵州省中国科学院天然产物化学重点实验室,贵州贵阳550014 [3]贵州茅台医院,贵州仁怀564500 [4]贵州医科大学省部共建药用植物功效与利用国家重点实验室,贵州贵阳550014

出　　处：《中国药理学通报》2023年第3期503-511,共9页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No U1812403);山西省重点研发计划项目(No 201603D3114010);山西省科技攻关项目(No.2014ZD0402);贵州省科技计划及平台人才项目(黔科合平台人才[2019]5702号)。

摘　　要：目的通过脂多糖(lipopolysaccharide,LPS)联合正常人血浆(NHP)刺激A549细胞,探究连翘脂素(phillygenin,PHI)对A549细胞炎症损伤的干预作用及机制。方法采用1 mg·L^(-1)LPS联合5%NHP刺激A549细胞建立炎症损伤模型。MTT和Hoechst 33342染色检测PHI对细胞活力、数目、面积、形态和DNA含量的影响。采用ELISA、免疫荧光染色和实时荧光定量PCR检测炎症因子、NF-κB p65入核及mRNA表达;Western blot检测NF-κB和MAPK关键蛋白表达。结果LPS联合NHP共刺激组IL-6、IL-8含量及NF-κB p65核内转录活性明显上调;PHI(1、10、50和100μmol·L^(-1))可以剂量依赖性降低IL-6和IL-8表达、NF-κB p65核内转录活性及p65 mRNA表达,明显下调IκBα、p65及p38、JNK、ERK蛋白磷酸化水平,上调IκBα表达。结论LPS联合NHP可以成功诱导A549炎症损伤模型,PHI可抑制该炎症反应,其机制与抑制LPS-TLR4-NF-κB/MAPK信号通路的活化有关。Aim To investigate the effect of phillygenin(PHI)on lipopolysacchride(LPS)and normal human plasma(NHP)induced inflammatory injury on alveolar type II epithelial A549 cells and the related mechanism.Methods A549 cells were exposured to 1 mg·L^(-1)of LPS and 5%NHP to build the inflammatory injury model.The effects of PHI on cell viability,cell number,area,morphology,and DNA content of A549 cells were detected by thiazole blue colorimetric(MTT)and Hoechst 33342 staining.The A549 cells were pretreated with PHI for 2 h,and then exposed to 1 mg·L^(-1)of LPS and 5%NHP.The contents of inflammatory cytokines and NF-κB p65 were determined by ELISA and quantitative real-time PCR.The transfer of NF-κB p65 from cytoplasm to nucleus was detected by cellular immunofluorescence.The protein expressions of NF-κB and MAPK signaling pathways were detected by Western blot.Results The contents of IL-6 and IL-8 and the transfer of NF-κB p65 from cytoplasm to nucleus were significantly up-regulated in LPS combined with NHP co-stimulated group.PHI at 1,10,50,and 100μmol·L^(-1)decreased the mRNA and protein expressions of IL-6 and IL-8,inhibited the transfer of NF-κB p65 from cytoplasm to nucleus,reduced the NF-κB p65 mRNA level,down-regulated the phosphorylation levels of IκBα,NF-κB p65,p38,JNK,and ERK,and up-regulated the protein expression of IκBαin a dose-dependent manner.Conclusions LPS combined with NHP successfully induces inflammatory injury in alveolar type II epithelial A549 cells.PHI could improve the inflammatory response by inhibiting the activation of LPS-TLR4-NF-κB/MAPK signaling pathways.

关 键 词：脂多糖 正常人血浆 A549细胞 急性肺损伤 连翘脂素 CD14 NF-κB/MAPK信号通路 

分 类 号：R284.1[医药卫生—中药学] R364.5[医药卫生—中医学] R345.57R563.8R734.202.2