应用人肝细胞研究人参皂苷Rb1和Rg1、丹参素钠、冰片对CYP450药物代谢酶1A2、2B6和3A4的体外诱导作用

Study on induction of CYP450 enzyme 1A2、2B6 and 3A4 by ginsenoside Rb,Rg,danshensu sodium and borneol with human hepatocytes in vitro

目的使用原代培养的人肝细胞研究人参皂苷Rb1和Rg1、丹参素钠、冰片对细胞色素P450酶(CYP450)的诱导作用。方法分别将3批次的冷冻原代人肝贴壁细胞进行接种培养,使用人参皂苷Rb1和Rg1、丹参素钠、冰片(30μmol/L)对CYP1A2、CYP2B6和CYP3A4进行诱导,实时荧光定量PCR(qRT-PCR)法测定CYP450酶mRNA表达水平,评价4种单体成分对P450酶的诱导作用。阳性对照为20μmol/L利福平(CYP3A4诱导剂)、50μmol/L奥美拉唑钠(CYP1A2诱导剂)和1 mmol/L苯巴比妥钠(CYP2B6诱导剂);阴性对照组为10μmol/L红霉素。冰片(30μmol/L)与利福平联合给药,观察冰片对利福平的CYP1A2、CYP2B6和CYP3A4诱导作用的影响。结果3批次人肝细胞的诱导结果显示,阳性诱导剂奥美拉唑、苯巴比妥、利福平分别显著诱导CYP1A2、CYP2B6、CYP3A4的表达,阴性对照红霉素对CYP1A2、CYP2B6及CYP3A4的mRNA表达水平未见明显影响,人参皂苷Rb1和Rg1、丹参素钠、冰片在30μmol/L浓度给药时,对3批次人肝细胞的CYP1A2、CYP2B6和CYP3A4 mRNA表达水平未见明显影响;30μmol/L冰片与利福平联合给药,与利福平单独给药比较,3批次肝细胞的CYP2B6、CYP3A4 mRNA表达水平均减少,对CYP3A4的mRNA表达水平影响尤为显著。结论人参皂苷Rb1和Rg1、丹参素钠、冰片在30μmol/L浓度给药时,对3批次人肝细胞的CYP1A2、CYP2B6和CYP3A4均没有诱导作用。冰片与利福平联合用药时,可以阻断利福平对CYP2B6和CYP3A4的诱导作用,对CYP3A4的影响尤为显著。

Objective In this study,human primary cultured hepatocytes were used to study the induction of P450 enzyme by ginsenoside Rb1,Rg1,Danshensu Sodium and Borneol.Methods The CYP1 A2,CYP2 B6 and CYP3 A4 were induced by different inducers,and the real-time RT-PCR method was used to determine the mRNA expression level.The results of CYP450 mRNA expression level was used to evaluate the induction of P450 enzyme.The positive control group was 20μmol/L rifampicin(CYP3 A4 inducer),50μmol/L omeprazole sodium(CYP1 A2 inducer)and 1 mmol/L phenobarbital sodium(CYP2 B6 inducer);the negative control group was 10μmol/L erythromycin.Borneol(30μmol/L)combined with rifampicin was used to observe the effect of Borneol on the induction of CYP1 A2,CYP2 B6 and CYP3 A4 of rifampicin.Results The results of three donors of induction showed that the expression of CYP1 A2,CYP2 B6 and CYP3 A4 was significantly induced by omeprazole,phenobarbital sodium and rifampicin respectively,and the corresponding mRNA expression levels were significantly increased,all>4;negative control erythromycin which levels of CYP1 A2,CYP2 B6 and CYP3 A4 mRNA were not significantly change.The expression levels of CYP1 A2,CYP2 B6 and CYP3 A4 mRNA were not significantly affected by ginsenoside Rb1,Rg1,danshensu sodium and borneol at the concentrations of 30μmol/L.Comparedwithrifampinalone,when borneol(30μmol/L)was combined with rifampicin(20μmol/L),the expression of CYP2 B6 and CYP3 A4 mRNA in the three batches of hepatocytes decreased,and the effect on the expression of CYP3 A4 mRNA was particularly significant.No significant effect of CYP1 A2 mRNA expression levels was seen.Conclusion The expression levels of CYP1 A2,CYP2 B6 and CYP3 A4 mRNA were not significantly affected by Rb1,Rg1,danshensu sodium and borneol at the concentrations of 30μmol/L.When borneol(30μmol/L)was combined with rifampicin(20μmol/L),there was no effect on CYP1 A2 in the three batches of hepatocytes,but the induction effect of rifampicin on CYP2 B6 and CYP3 A4 in three batches of hepatocytes were reduced while the decrease of CYP3 A4 is particularly evident.