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研究姜黄素对Ⅱ相酶谷胱甘肽转移酶(GST)及NADP(H)醌氧化还原酶(NQO)活性的影响及其诱导机制。用光谱法检测细胞GST酶和NQO酶的活性,以及还原型谷胱甘肽(GSH)的含量;利用蛋白印迹法检测核转录因子Nrf2在胞浆与胞核的分布;采用凝胶电泳迁移率分析法(EMSA)检测Nrf2与Ⅱ相酶基因抗氧化反应序列(ARE)结合活性。不同浓度的姜黄素(10～30μmol.L-1)刺激结肠腺癌HT-29细胞后,能显著诱导GST酶及NQO酶活性的增加,同时能迅速提高细胞内GSH的含量;蛋白印迹和凝胶电泳迁移率结果显示,姜黄素诱导细胞核内转录因子Nrf2积聚,Nrf2-ARE的结合活性增加。姜黄素诱导的Ⅱ相酶GST酶及NQO酶活性增加与促进转录因子Nrf2由胞浆向胞核发生转位分布和增强Nrf2-ARE结合活性有关。
The effects of curcumin on the activity of phase II enzymes glutathione transferase (GST) and NADP(H) quinone oxidoreductase (NQO) and their induction mechanism were studied. The activity of GST enzyme and NQO enzyme and the content of reduced glutathione (GSH) were detected by spectroscopy; the distribution of nuclear transcription factor Nrf2 in cytoplasm and nucleus was detected by Western blotting; the electrophoretic mobility was measured by gel electrophoresis. The analysis method (EMSA) was used to detect the anti-oxidation reaction sequence (ARE) binding activity of Nrf2 and II phase enzyme genes. Different concentration of curcumin (10-30μmol.L-1) stimulated colon adenocarcinoma HT-29 cells, can significantly increase the activity of GST enzyme and NQO enzyme, and can rapidly increase the intracellular GSH content; Western blotting and coagulation The result of gel electrophoresis mobility showed that curcumin induced the accumulation of Nrf2, a nuclear transcription factor, and the binding activity of Nrf2-ARE increased. The increase of curcumin-induced phase II enzyme GST enzyme and NQO enzyme activity is related to the promotion of translocation of transcription factor Nrf2 from cytoplasm to nucleus and enhancement of Nrf2-ARE binding activity.