灯盏细辛和赤芍配伍组方对小鼠部分肝细胞色素P450亚型活性的影响及其机制分析

目的:探讨灯盏细辛和赤芍配伍组方(辛芍组方)对小鼠细胞色素P450酶(CYP450s)主要亚型CYP1a2,CYP2d22,CYP2e1和CYP3a11代谢活性的影响以及相关机制。方法:小鼠分为5组,分别为正常组、苯巴比妥组(70 mg·kg-1)和辛芍组方低、中、高剂量组(0.31,0.62,1.25 g·kg-1)。给药结束后,取各组小鼠肝脏制备微粒体,考察CYP1a2,CYP2d22,CYP2e1和CYP3a11的代谢活性;并提取肝组织总RNA和蛋白,实时荧光定量聚合酶链式反应法(Real-time PCR)和蛋白免疫印迹法(Western blot)考察辛芍组方对CYP1a2,CYP2d22,CYP2e1,CYP3a11和孕烷X受体(PXR)mRNA及蛋白表达的影响;利用报告基因法考察辛芍组方对PXR的激活作用。结果:与正常组比较,辛芍组方可诱导CYP3a11酶活性,上调CYP3a11,PXR mRNA和蛋白水平,并具有PXR激活作用(P0.05,P0.01);与模型组比较,辛芍中、高剂量组方能抑制CYP1a2酶活性(P0.05),但对CYP1a2的mRNA水平和蛋白水平无明显影响;而辛芍组方对CYP2e1,CYP2d22的酶活性,mRNA水平和蛋白水平无明显影响。结论:辛芍组方具有CYP3a11活性诱导作用,该作用很可能与辛芍组方上调PXR表达并激活PXR从而促进CYP3a11表达有关;而其CYP1a2活性抑制作用机制与CYP1a2的表达调控无关,需要进一步研究。

Effect and Mechanism of Xinshao Formula on Activities of Cytochrome P450s in Mice Liver

Objective: To investigate the effects of erigerontis herba and paeoniae radix rubra formula (Xinshao formula) on the metabolic activity of main subtypes (CYP1a2, CYP2d22, CYP2e1 and CYP3a11) of cytochrome P450s (CYP450) in mice liver and explore its mechanism. Method: Mice were randomly divided into normal group, phenobarbital group (70 mg·kg^-1), and low-, middle-, high-dose Xinshao formula groups (0.31, 0.62, 1.25 g crude drug·kg^-1). After administration, the livers of mice were used to prepare microsomes to determine the metabolic activities of CYP1a2, CYP2d22, CYP2e1 and CYP3a11.The mRNA and protein expression levels of these four CYP450 enzymes and pregnane X receptor (PXR) in liver tissue of mice were evaluated by quantitative real-time polymerase chain reaction (Real-time PCR) and Western blot. The effect of Xinshao formula on the activation of PXR was estimated by reporter gene assay. Result: As compared with the normal group, Xinshao formula could induce the enzyme activity of CYP3a11, up-regulate the mRNA and protein expression levels of CYP3a11 and PXR, and activate PXR (P<0.05,P<0.01). As compared with the model group, middle and high dose Xinshao formula could inhibit CYP1a2 activity (P<0.05), but had no significant effect on the mRNA and protein expression levels of CYP1a2; while Xinshao formula had no significant effects on the activities and the expressions of CYP2e1 and CYP2d22. Conclusion: Xinshao formula could induce enzyme activity of CYP3a11, and the mechanism may be associated with up-regulating PXR expression, activating PXR and promoting CYP3a11 expression. Xinshao formula also could decrease the CYP1a2 activity, but it was not related to regulation of CYP1a2 expression, and the mechanism needs a further study.