基于雌激素受体调节Nrf2-ARE通路的黄芩中抗氧化成分的筛选

目的建立基于Nrf2-ARE通路的报告基因筛选模型,筛选黄芩中基于雌激素受体发挥抗氧化作用的活性成分。方法 ARE荧光素酶报告质粒p GL4. 37和海肾荧光素酶报告质粒pRL-TK共同转染293T细胞。将黄芩中汉黄芩素、黄芩素、黄芩苷等3个主要活性成分和(或)雌激素受体(ER)特异性抑制剂加入Nrf2-ARE双荧光素酶报告基因系统,检测其是否通过ER影响Nrf2-ARE通路,发挥抗氧化作用。将筛选出的成分和(或) ER抑制剂、Nrf2-ARE通路抑制剂加入Ha Ca T细胞中,验证是否通过ER影响Nrf2-ARE通路发挥抗氧化作用。结果黄芩苷(100μmol·L^-1)可明显激活293T细胞中的Nrf2-ARE通路,诱导表达倍数为空白组的(1. 56±0. 01)倍(P <0. 01)。预给予ER抑制剂后,诱导表达倍数下降至(1. 02±0. 23)倍,抗氧化作用消失。分别预给予ER抑制剂、Nrf2-ARE通路抑制剂后,黄芩苷干预UVB损伤的Ha Ca T细胞中的ROS值明显上升,SOD值明显下降。结论黄芩苷可以通过ER影响Nrf2-ARE通路,发挥抗氧化作用。

Screening of antioxidant components in Scutellariae Radix through mediating Nrf2-ARE signaling pathway based on estrogen receptor

Aim To establish ARE dual-luciferase reporter assay system and used it to identify the antioxidant substance of Scutellaria baicalensis Georgi. Methods 293T cells were transiently co-transfected with ARE luciferase reporter plasmid PGL 4.37 and sea kidney luciferase reporter plasmid PRL-TK. Three major active ingredients of Scutellaria baicalensis Georgi such as scutellarin, baicalein, baicalin and/or estrogen receptor(ER) specific inhibitor were added to Nrf2-ARE luciferase reporter assay system to detect whether they exerted antioxidant effect through the estrogen receptor affecting the Nrf2-ARE signaling pathway. Results Baicalin(100 μmol·L^-1 ) could obvi- ously activate Nrf2-ARE pathway in 293T cells, and the induced expression was(1.56±0.01) times that of blank group( P <0.01).After pre-administration of ER specific inhibitor, the induced expression decreased to(1.02±0.23) times, and the antioxidant effect disappeared. After pre-administration of ER and Nrf2-ARE pathway specific inhibitor respectively, ROS in HaCaT cells injured by UVB significantly increased and and SOD was markedly down-regulated by baicalin. Conclusion Baicalin plays antioxidant activity through mediating Nrf2-ARE signaling pathway based on estrogen receptor.