Elafin在人支气管上皮细胞系16HBE胞质内对黏蛋白5AC生成的影响  

The effect of Elafin in 16HBE cytoplasm on the mucin5AC production

作　　者：魏华生[1] 尤列.皮尔曼 维克多.科罗索夫 周向东[1] 

机构地区：[1]重庆医科大学附属第二医院呼吸内科,重庆400010 [2]俄罗斯医学科学院远东呼吸生理与病理中心,布拉戈维申斯克675000

出　　处：《基础医学与临床》2011年第7期809-813,共5页Basic and Clinical Medicine

基　　金：国家自然科学基金(81070031);国家自然科学基金中俄合作项目(81011120108)

摘　　要：目的探讨Elafin在胞质内对表皮生长因子(EGF)诱导的黏蛋白(MUC)5AC生成的影响及其作用机制。方法体外培养人支气管上皮细胞系16HBE,将已构建好的pEGFP-N1-Elafin载体通过LipofectamineTM2000转染到16HBE细胞中,以转染空质粒载体的16HBE细胞作为对照,均给予外源性EGF作为刺激物共同孵育。以RT-PCR和ELISA分别检测细胞内黏蛋白(MUC)5AC mRNA水平和MUC5AC含量。间接免疫荧光细胞化学法结合激光共聚焦显微镜检测细胞核中核因子(NF)-κB的活性。免疫共沉淀法结合Western blot检测SUMO-1-p-IκBα含量。结果转染重组质粒的细胞中MUC5ACmRNA水平、MUC5AC的含量(0.36±0.09)以及NF-κB的活性显著低于转染空载体的对照组细胞(MUC5AC的含量0.55±0.07)(P<0.01),而SUMO-1-p-IκBα的含量则显著高于后者(P<0.01)。结论 Elafin能够通过促进p-IκBα的类泛素化阻止NF-κB的活化,从而下调MUC5AC的生成。Objective To investigate the effect of Elafin in cytoplasm on epidermal growth factor（EGF）-induced mucin（MUC）5AC production and the mechamism.Methods After cultivating 16HBE cells in vitro,the constructed pEGFP-N1-Elafin eukaryotic expression vector was transfected into the 16HBE cells by LipofectamineTM2000,and 16HBE cells transfected the vector without Elafin gene were classified as control group.The cells were then incubated with exogenous EGF.The mucin（MUC）5AC mRNA level was analyzed by RT-PCR,the MUC5AC protein production was assayed by ELISA,the NF-κB activity in nuclear was detected by immunofluorescence and laser scanning confocal microscope,the SUMO-1-p-IκBα production was assayed by co-immunoprecipitation（CO-IP） and Western blot.Results The MUC5AC mRNA level,the MUC5AC protein production（0.36±0.09） and the NF-κB activity of the experimental group were all significantly lower than those of the empty vector group（MUC5AC production of the later is 0.55±0.07）,but the SUMO-1-p-IκBα production was significantly higher than that of thelatter（P0.01）.Conclusion Elafin prevents NF-κB activation via its effect on p-IκBα sumoylation,thus reducing epidermal growth factor-induced mucin5AC production.

关 键 词：ELAFIN 核因子-ΚB 小泛素样修饰蛋白-1 类泛素化 黏蛋白5AC 

分 类 号：R562[医药卫生—呼吸系统]