黄芪甲苷对A549细胞胸腺基质淋巴细胞生成素表达的影响

Effect of astragaloside Ⅳ on the expression of thymic stromal lymphopoietin in A549 cells

目的观察黄芪甲苷对胸腺基质淋巴细胞生成素(TSLP)基因表达的影响,并探讨此作用是否与核因子-κB(NF-κB)有关。方法肿瘤坏死因子-α(TNF-α)+白介素-4(IL-4)刺激A549细胞4 h、10 h和18 h后,使用实时荧光定量PCR(RT-PCR)方法观察A549细胞TSLP mRNA表达的变化;同时使用RT-PCR方法观察黄芪甲苷以及NF-κB抑制剂PDTC对TSLP mRNA表达的影响,使用免疫荧光观察NF-κB P65核转录的变化,并用western blot方法检测IκBα表达的变化。结果 TNF-α+IL-4刺激A549细胞后TSLP mRNA表达明显增加,4 h时增加最明显,10 h逐渐下降,18 h后基本降到正常;PDTC能够显著抑制TSLP mRNA表达,而黄芪甲苷抑制作用不明显;黄芪甲苷能够显著抑制A549细胞NF-κB P65的核转录,升高胞浆IκBα的蛋白水平。结论气道上皮细胞A549细胞中NF-κB对TSLP基因具有重要的调控作用,黄芪甲苷能够升高胞浆IκB-α的蛋白水平,抑制NF-κB的核转录,但黄芪甲苷对TSLP的基因表达没有抑制作用。

Objective To investigate the effect of astragaloside Ⅳ on the expression of TSLP mRNA in A549 cells,and to explore whether nuclear factor κB（ NF-κB） is involved. Methods A549 cells were cultured with TNF-α and IL-4,and the expression of TSLP mRNA at 4 h,10 h and 18 h was detected by real time-PCR. The effects of astragaloside Ⅳand PDTC（ NF-κB inhibitor） on the expression of TSLP mRNA were also quantified by real time-PCR. The cytoplasm protein expression of IκBα was evaluated by western blot,and the nuclear translocation of NF-κB P65 was detected by immunofluorescence assays. Results Compared with the control group,the expression of TSLP mRNA was significantly increased in A549 cells stimulated with TNF-α and IL-4 at 4 h,and decreased at 10 h,almost reduced to the baseline at 18 h. PDTC greatly inhibited the level of TSLP mRNA. Astragaloside Ⅳ had little inhibitory effects on the expression of TSLP mRNA. However,Astragaloside Ⅳ suppressed the nuclear translocation of NF-κB P65,and enhanced the protein level of IκBα in the cytoplasm. Conclusions NF-κB pathway may play an important role in regulating TNF-α and IL-4 stimulating TSLP mRNA expression. AstragalosideⅣ may suppress the nuclear translocation of NF-κB P65,enhancing the protein level of IκB-α in the cytoplasm. But,Astragaloside Ⅳ has little inhibitory effects on the expression of TSLP mRNA.