姜黄素衍生物通过Notch1/Hes1信号通路抑制RSV感染后人肺上皮细胞MUC5AC分泌的研究  

Curcumin derivatives inhibit secretion of MUC5AC in RSV-infected human pulmonary epithelial cells via Notch1/Hes1 signaling pathway

作　　者：叶文静 翁婷婷 王乐颖 李海燕 陈小芳 董琳 YE Wenjing;WENG Tingting;WANG Leying;LI Haiyan;CHEN Xiaofang;DONG Lin(Department of Pediatric Respiratory Diseases,the Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University,Wenzhou 325027,China)

机构地区：[1]温州医科大学附属第二医院、育英儿童医院儿童呼吸科,325027

出　　处：《浙江医学》2021年第15期1600-1604,1609,共6页Zhejiang Medical Journal

基　　金：重大新药创制科技重大专项(2020ZX09201002)。

摘　　要：目的探讨姜黄素衍生物通过Notch1/发状分裂相关增强子1(Notch1/Hes1)信号通路对呼吸道合胞病毒(RSV)感染后人肺上皮细胞BEAS-2b黏蛋白5AC(MUC5AC)分泌的影响及其作用机制。方法建立体外RSV感染细胞模型,随机分成6组:姜黄素衍生物B6组、γ分泌酶抑制剂(DAPT)+B6组、DAPT组、地塞米松(DXM)组、RSV组和细胞对照组。B6组在RSV感染造模0.5 h前予10μmol/L B6预处理,DAPT+B6组在B6预处理前先以20μmol/L DAPT预处理1 h。造模开始前1 h,DXM组加入0.01 mg/ml DXM。B6组、DAPT+B6组、DAPT组、DXM组及RSV组以100倍半数感染量(TCID50)浓度的RSV感染吸附2 h后加维持液培养12、24 h收获细胞及上清液进行检测。应用RT-PCR、Western blot法分别检测各组细胞Notch1、Hes1、MUC5AC mRNA与蛋白质的表达;ELISA法检测上清液中IL-25蛋白水平。结果与细胞对照组相比,B6组、DAPT+B6组、DAPT组、RSV组Notch1、Hes1、MUC5AC mRNA和蛋白表达水平及IL-25蛋白水平在12、24 h两个时间点均显著升高(均P<0.05),其中MUC5AC表达水平在24 h升高更显著。与RSV组相比,两个时间点其他组上述指标表达水平均有所降低(均P<0.05),以DXM组表达水平最低;而B6组、DAPT+B6组、DAPT组同一时间点上述指标表达水平相近。结论RSV感染人肺上皮细胞后,通过Notch1/Hes1信号通路促进IL-25的释放,并上调MUC5AC表达,导致气道黏液高分泌,而姜黄素衍生物对此具有抑制效应。Objective To explore the effect of curcumin derivatives on the secretion of Mucin-5AC(MUC5AC)in human pulmonary epithelial cells infected with respiratory syncytial virus(RSV)and its mechanism.Methods The BEAS-2b cells were randomly divided into 6 groups:curcumin derivatives B6 group,γsecretase inhibitor(DAPT)+B6 group,DAPT group,dexamethasone(DXM)group,RSV group and cell control group.The B6 group was pretreated with 10μmol/L B60.5 h before modelling.The DAPT+B6 group was pretreated with 20μmol/L DAPT for 1 h before the B6 pretreatment.The DXM group was pretreated with 0.01 mg/ml DXM 1h before modelling.B6 troup,DAPT+B6 group,DAPT group and RSV group were infected with 100×TCID50 RSV for 2 h,and then cultured for 12 h and 24 h,the cells and supernatants were harvested for detection.Expression of Notch1,Hes1 and MUC5AC mRNA and protein were detected by qRT-PCR and Western blot,respectively.ELISA was used to measure the protein level of IL-25 in the culture supernatant.Results Compared with cell control group,the mRNA and protein expression levels of Notch1,Hes1,MUC5AC and the protein levels of IL-25 in B6 group,DAPT+B6 group,DAPT group,and RSV group increased significantly at 12 h and 24 h(P<0.05),and the expression levels of MUC5AC increased more significantly at 24 h.While compared with RSV group,the expression levels of the above indicators in other groups decreased at both time points(P<0.05),and the expression levels of DXM group were the lowest.The expression levels in B6 group,DAPT+B6 group and DAPT group were similar at the same time point.Conclusion RSV infection can promote the release of IL-25 through Notch1/Hes1 signaling pathway,upregulating the expression of MUC5AC and leading to hypersecretion of airway mucus,and curcumin derivative(B6)has an inhibitory effect on this.

关 键 词：呼吸道合胞病毒 黏蛋白5AC Notch1/Hes1信号通路 白介素-25 姜黄素衍生物 

分 类 号：R285[医药卫生—中药学]