香烟烟雾提取物通过蛋白激酶C-核因子相关因子2调节大鼠气道上皮细胞血红素加氧酶1表达

发布时间：2018-09-07 08:39

【摘要】：目的:探讨蛋白激酶C(PKC)-红系衍生的核因子相关因子2(Nrf2)对香烟烟雾提取物(CSE)诱导的大鼠气道上皮细胞血红素加氧酶1(HO-1)表达的影响。方法:通过CSE刺激雄性SD大鼠气道上皮细胞,使用PKC抑制剂RO318220和Nrf2 siRNA,将细胞分为对照组、CSE 3 h组、RO318220组、Nrf2 siRNA组和RO318220+Nrf2 siRNA组,用Western blotting法分别检测HO-1、Nrf2和p-PKC蛋白表达,免疫细胞化学法观察HO-1蛋白表达,逆转录-聚合酶链反应(RT-PCR)法检测HO-1 mRNA表达,免疫荧光法检测Nrf2蛋白定位,测定HO-1活性。结果:暴露CSE 3 h后,Nrf2蛋白主要表达在胞核,胞核蛋白表达增强,p-PKC蛋白、HO-1的mRNA和蛋白高表达,HO-1活性增强。预先给予RO318220,PKC蛋白、Nrf2胞浆和胞核蛋白、HO-1的mRNA和蛋白表达均明显减弱,HO-1活性显著降低。预先用siRNA沉默Nrf2,胞浆和胞核的Nrf2蛋白表达均减弱,HO-1活性、mRNA和蛋白水平明显降低。RO318220联合Nrf2 siRNA处理后,PKC蛋白、Nrf2胞浆和胞核蛋白、HO-1 mRNA和蛋白表达均明显降低,HO-1活性明显降低。结论:CSE通过PKC激活Nrf2,诱导Nrf2核转位,从而上调HO-1的表达水平。
[Abstract]:Aim: to investigate the effect of protein kinase C (PKC)-erythroid derived nuclear factor-related factor 2 (Nrf2) on the expression of heme oxygenase 1 (HO-1) in rat airway epithelial cells induced by cigarette smoke extract (CSE). Methods: the airway epithelial cells of male SD rats were stimulated by CSE. The cells were divided into two groups: control group (control group) for 3 h with PKC inhibitor RO318220 and Nrf2 siRNA,. The expression of HO-1,Nrf2 and p-PKC protein was detected by Western blotting method. The expression of HO-1 protein was detected by immunocytochemistry, the expression of HO-1 mRNA was detected by reverse transcription-polymerase chain reaction (RT-PCR), the localization of Nrf2 protein and the activity of HO-1 were detected by immunofluorescence. Results: after exposure to CSE for 3 h, the expression of nrf2 protein was mainly found in the nucleus, and the expression of nuclear protein enhanced the activity of mRNA and HO-1 of p-PKC protein. The expression of mRNA and protein of RO318220,PKC protein nrf2 in cytoplasm and nuclear protein was significantly decreased. The expression of Nrf2 protein in cytoplasm and nucleus of Nrf2, was inhibited by siRNA in advance. The activity and protein level of Nrf2, HO-1 were significantly decreased after treated with Nrf2 siRNA combined with .RO318220. The expression of mRNA and protein in the cytoplasm and nucleoprotein of nrf2 and nucleoprotein HHO-1 were significantly decreased after treatment with .RO318220, and the activity of HO-1 was significantly decreased. Conclusion Nrf2 nuclear translocation was induced by Nrf2, activated by PKC, and the expression of HO-1 was upregulated.
【作者单位】：湖南省人民医院呼吸内科;
【分类号】：R563.9

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