基于抗氧化反应元件的药物筛选模型的建立

Establishment of a drug screening model based on antioxidant response element

目的建立基于抗氧化反应元件(antioxident responseelement,ARE)和SEAP报告基因的细胞筛选模型,用此模型对金雀异黄素、芹菜苷配基、儿茶素、茨非醇4种化合物进行活性比较,观察上述化合物拮抗过氧化氢损伤的作用,寻找具有强抗氧化活性作用的黄酮类化合物。方法构建重组报告基因表达载体pARE-TAL-SEAP与对照载体分别转染HEK-293细胞,检测上述黄酮类化合物作用后对SEAP的活性和抗氧化作用的影响。结果在加入25μmol.L-1H2O2进行氧化损伤的HEK-293细胞中,加入100μmol.L-1的4种化合物进行比较时金雀异黄素的抗氧化作用最强,金雀异黄素诱导SEAP的最大表达水平较对照孔升高1.3倍;加入25μmol.L-14种化合物的抗氧化作用时茨非醇的抗氧化作用最强,茨非醇诱导SEAP的最大表达水平较对照孔升高1.5倍。结论利用此模型可以检测H2O2的氧化应激反应并可筛选到抗氧化活性强的黄酮类化合物。

Aim To establish a screening model based on antioxidant response element and SEAP reporter gene; To screen four compounds （Genistin, Apicidin, Catehin gallate and Kaempferol） for antioxidant activity; To search for flavonol compounds with high antioxidant activity. Methods A recombinant reporter vector pARE-TAL-SEAP was constructed, pARE-TAL-SEAP and control vector were transiently transfected into HEK-293 cells pretreated with 25 μmol ·L^-1 H2O2. SEAP activity was assayed after flavonol compounds treatment. Results In the 25 μmol · L^-1 H2O2 pretreated HEK-293 cell oxidative damage model, Genistin showed the highest antioxidant activity with SEAP induction 1.3 times higher than control in 100 μmol · L^-1 group; Kaempferol showed the highest antioxidant activity with SEAP induction 1.5 times higher than control in 25 μmol · L^-1 group. Conclusion This model can be used to detect H2O2 oxidative stress response and screen flavonol compounds with high antioxidant activity.