目的 建立hAhR介導(dǎo)的CYP1A1體外誘導(dǎo)活性評價模型，應(yīng)用于體外快速篩選通過hAhR途徑介導(dǎo)的對CYP1A1具有激活能力的藥物。方法 利用雙熒光素酶報告基因系統(tǒng)，將CYP1A1啟動子序列插入報告基因質(zhì)粒上游，構(gòu)建pGL4.17-CYP1A1質(zhì)粒，并與含有hAhR編碼區(qū)序列的pcDNA3.1-hAhR表達(dá)質(zhì)粒瞬時共轉(zhuǎn)染HepG2細(xì)胞， pGL4.17-control、pcDNA3.1（+）分別為pGL4.17-CYP1A1、pcDNA3.1-hAhR的空載體，建立hAhR-CYP1A1報告基因模型；應(yīng)用AhR的完全激動劑TCDD（2 nmol/L）驗證該報告基因模型的可靠性，應(yīng)用該模型考察人參皂苷Rf、Rc、Re、Rg₁、Rb、Rd、Rh₂和Rg₃（20 μmol/L）通過hAhR途徑對CYP1A1的誘導(dǎo)作用。結(jié)果 經(jīng)驗證，報告基因模型構(gòu)建成功；人參皂苷Rf、Re、Rg₁和Rc對AhR具有激活作用，其中與對照組比較，人參皂苷Rg₁、Rc對AhR激活效應(yīng)顯著（P<0.05、0.01），人參皂苷Rb、Rd、Rh₂和Rg₃對AhR無激活作用。結(jié)論 本研究建立基于hAhR的CYP1A1體外誘導(dǎo)活性評價模型，可為具有潛在CYP1A1誘導(dǎo)作用的目標(biāo)化合物的篩選提供有效、快速的體外篩選手段，人參皂苷Rg₁、Rc對AhR-CYP1A1激活效應(yīng)顯著。

Objective The aim of this study is to establish and validate an in vitro model to screen potential drug inducers of CYP1A1 mediated via nuclear receptor AhR, which can be applied to evaluate the induction activity toward CYP1A1 by a large number of drugs rapidly and effectively.Methods Dual luciferase reporter gene system was performed. The promoter of CYP1A1 DNA was cloned into the firefly luciferase reporter gene vector, pGL4.17[luc2/Neo], in order to construct pGL4.17-1A1. Transiently cotransfecting the pGL4.17-1A1 reporter vector and hAhR expression plasmid pcDNA3.1-hAhR to HepG2 cells, PGL4.17-control and pcDNA3.1 (+) were empty vectors of pGL4.17-CYP1A1 and pcDNA3.1-hAhR respectively; And hAhR-mediated CYP1A1 reporter gene model was established. As the full agonist of AhR, TCDD was applied to ensure the reliability of hAhR-mediated CYP1A1 reporter gene model. The model was used to investigate the induction of CYP1A1 by hAhR pathway of ginsenoside Rf, Rc, Re, Rg₁, Rb, Rd, Rh₂ and Rg₃. Results Nuclear receptor hAhR-mediated CYP1A1 induction model has been established successfully. Ginsenoside Rf, Re, Rg₁ and Rc (20 μmol/L) could activate AhR. Compared with the control group, ginsenoside Rg₁ and Rc had significant activation effect on AhR (P<0.05, 0.01). Ginsenoside Rb, Rd, Rh₂ and Rg₃ had no activation on AhR.Conclusion The establishment of this induction model is an efficient and rapid in vitro way of investigation the drug inducers or inhibitors of CYP1A1.

國家重大科技專項資助項目（2015ZX09501004-003-002）