摘要
目的探讨血管紧张素Ⅱ对大鼠胰星状细胞(PSC)核因子κB(NF-κB)信号转导通路活化的影响。方法采用电泳迁移率改变分析(electrophoretic mobility shift assay,EMSA)检测细胞NF- κB的DNA结合活性,Western印迹方法检测NF-κB抑制蛋白(IκBs)降解。单核细胞趋化蛋白-1 (MCP-1)基因和蛋白表达分别采用Northern印迹和ELISA方法检测。结果血管紧张素Ⅱ可诱导大鼠PSC内NF-κB发生双相活化,活化高峰分别出现在15 min和6 h后。NF-κB活化伴有IκBα和IκBβ联合降解。抗氧化剂吡咯烷二硫基甲酸盐(PDTC)、二碘基苯(DPI)和N-乙酰丝氨酸可明显抑制血管紧张素Ⅱ诱导的NF-κB活化,提示氧化应激反应在NF-κB活化中发挥了重要作用。NF-κB抑制剂 MG132和PDTC可显著下调血管紧张素诱导的MCP-1基因表达。结论血管紧张素Ⅱ可通过氧化应激机制活化PSC内NF-κB信号通路,诱导MCP-1表达,从而参与胰腺炎症和纤维化的发生。
Objective To investigate the impact of angiotension Ⅱ （Ang Ⅱ） on the activation of nuclear factor-kappaB （NF-κB） signaling transduction pathway in cultured rat pancreatic stellate cells （PSCs）. Methods Growth-arrested rat PSCs were incubated for indicted time intervals with increasing concentrations of Ang Ⅱ. The DNA binding activity of NF-κB was determined using electrophoretic mobility shift assay （EMSA）. Western blot analysis was used to examine the degradation of inhibitory proteins of NF-κB （IκBs）. Expression of monocyte chemoattractant protein-1 （MCP-1） mRNA and protein were assessed by Northern blot and enzyme-linked immunosorbent assay （ELISA）, respectively. Results Treatment of cells with Ang Ⅱ led to a biphasic activation of NF-κB, which peaked at 15 min and 6 h later, and it was correlated with differential degradation of IκBa and IκBβ. Ang Ⅱ-induced NF- κB activation was greatly inhibited by antioxidants pyrrolidine dithiocarbamate （PDTC）, diphenylene iodonium, and N-acetylcysteine, suggesting the involvement of oxidative stress in this process. In PSCs, Ang Ⅱ induced a dose-dependent increase in the expression of MCP-1 and this effect was markedly inhibited by blocking NF-κB activation with MG132 and PDTC, indicating that Ang Ⅱ -induced MCP-1 gene expression was NF-κB-dependent. Conclusion The present results suggest that Ang Ⅱ , through an oxidative stress-dependent mechanism, may activate a functional NF-κB signaling pathway in PSCs, through which it may participate in pancreatic inflammation and fibrosis.
出处
《中华消化杂志》
CAS
CSCD
北大核心
2006年第4期243-246,共4页
Chinese Journal of Digestion
