【摘要】 目的研究IL-1β对健康人支气管上皮细胞(HBE)高迁移率族蛋白1(HMGB1)表达和释放的影响。方法培养HBE135-E6E7,四唑盐(MTT)法检测不同浓度IL-1β对支气管上皮细胞活力的影响,荧光定量PCR检测IL-1β刺激HBE后HMGB1mRNA表达,双抗体夹心ELISA法检测细胞上清液中HMGB1水平,蛋白免疫印迹方法检测IL-1β刺激HBE总蛋白、胞核、胞质HMGB1表达。免疫荧光观察IL-1β刺激后对HBE HMGB1的移位的影响。结果 0.1、1.0、10.0ng/mL IL-1β刺激HBE 24h对其活力无影响。IL-1β以浓度和时间依赖性方式刺激HBE的HMGB1mRNA水平升高,IL-1β增加HBE的HMGB1蛋白表达水平。在浓度依赖性实验,与对照组比较,1.0、10.0ng/mL IL-1β刺激HBE 24h后培养上清液HMGB1水平显著增加;在时间依赖性实验中,与对照组比较,10.0ng/mL IL-1β刺激HBE 12h、24h后细胞上清液中的HMGB1显著升高(P<0.05);10.0ng/mL IL-1β刺激HBE 24h后,HBE胞质蛋白明显增加,胞核蛋白减少。免疫荧光检测显示,HMGB1主要表达正常HBE的细胞核内,少量分布于细胞质,IL-1β(10.0ng/mL)刺激HBE 24h,HMGB1明显从HBE胞核向胞质转位。结论 IL-1β可以显著诱导支气管上皮细胞HMGB1的表达和主动释放,提示HMGB1可能参与了IL-1β介导慢性气道炎症过程。更多还原

【Abstract】 Objective To investigate the effect of IL-1βon the expression and release of high mobility group box 1(HMGB1)from normal human bronchiolar epithelial cell(HBE).Methods HBE135-E6E7 was developed and methyltetrazolium(MTT)assay was used to assess the viability of HBE cell line under different concentration of IL-1β.The mRNA expression of HMGB1 in HBE after stimulating with IL-1βwere determined by Real-time PCR;the level and location of HMGB1 in the cytoplasm,nucleus and culture medium of HBE after stimulating with IL-1βwere detected by Western blot and ELISA assay.The expression and translocation of HMGB1 of HBE after stimulating with IL-1β were detected by Immunofluorescence.Results 0.1,1.0,10.0ng/mL IL-1βdid not influence the cell viability of HBE;IL-1βincreased the mRNA expression of HMGB1 in HBE in does-and time-dependent manner and increased the protein expression of HMGB1 in HBE.In comparison with control group,the levels of HMGB1 in the culture medium significantly increased after stimulation with IL-1βat 1.0,10.0ng/mL for 24h(P<0.05)in the dose dependent experiments;and in time dependent experiments,10.0ng/mL IL-1βsignificantly increased HMGB1 level in the culture medium after stimulation for both 12 hand 24h(P<0.05).After stimulation with IL-1β(10.0ng/mL)for 24 h,the HMGB1 expression in cytoplasm significantly increased in plasmosin and decreased in nucleus.HMGB1 translocated from nuclei to cytoplasm after stimulation with IL-1β(10.0ng/mL)for 24 h.Conclusion IL-1βcould induce HMGB1 expression and release in human bronchial epithelial cells,which indicates that HMGB1 may involve in IL-1β-mediated chronic airway inflammation.更多还原