药对“黄芪+白术”抗炎作用配伍增效的药动学分子机制及与药物转运体SLC22As的相关性研究

A pair of two herbs is a common medication form in Traditional Chinese medicine (TCM) practice. Evidences have proved that pharmacokinetic interactions of active components in herbs should be one of mechanisms which lead to synergism effects of herb pair prescription. In this study, a herb pair which is composed of Huangqi and Baizhu is chosen as a tool herb pair, and the synergism effects of Huangqi-Baizhu herb pair have been confirmed in earlier test. In next steps, fingerprints of Huangqi-Baizhu decoction, serum samples and target tissue samples will be set up, and “pharmacokinetic component group” will been found in order to investigate pharmacokinetic synergism mechanisms of Huangqi-Baizhu herb pair. HepG2.2.15、MDCK-hOCTN cells model will be used which can express SLC22As transporters in vitro study in order to screen substrates of SLC22As and to study pharmacokinetic interactions between Huangqi-Baizhu herb pair and drug transportors SLC22As. With modern pharmacokinetic technologies such as RT-PCR, gene transfection and RNAi, some researches will be done to explore the effects of drugs on the protein level and mRNA level of the transporters. Some researches will be done to explore regulation effects of herb pair on SLC22As via the nuclear receptor HNF4α signal pathway. The concept of “pharmacokinetic component group "is proposed in this study to clarify the correlation ship between synergistic actions of active components in the herbs and drug transporters, and to establish a method to investigate the relationships among compatibility program, synergistic effects and pharmacokinetic interactions for TCM prescriptions.

药对是中药方剂的主干和药效基础。药对配伍产生的有效成分间药动学相互作用，是其增效或减毒的机制之一。课题在前期已证实经典中药药对“黄芪+白术”配伍增效与其药动学相互作用有关的基础上，进一步优化药材煎剂、含药血清、靶组织样本“指纹图谱”，确定其中与药动学密切相关的“药动学成分群”，并以其为标志物，研究药对配伍增效的药动学机制。研究采用表达SLC22As转运体的HepG2.2.15、MDCK-hOCTN细胞模型及慢支大鼠模型，筛选药物跨膜转运的转运体和底物，药物对转运体活性、表达的干预作用以及与配伍增效的关系；应用定量RT-PCR、基因转染、RNAi等技术研究药物对转运体蛋白水平、mRNA水平的影响；考察药物经由核受体HNF4α信号通路对转运体的调控作用。本研究提出了“药动学成分群”的概念，旨在为阐释芪术药对配伍增效的药动学机制提供一种新方法，也可望为同类中药方剂配伍机制的研究提供一个新思路。

研究遵循方剂配伍特点，以多成分的药动学相互作用为切入点，进行方剂配伍增效的物质基础和机制研究。项目建立和完善了“脂多糖合并烟熏提取物刺激的A549细胞模型-烟熏合并气管滴入脂多糖诱导的大鼠慢性肺炎症模型”等体内外联合模型研究体系，筛选了方剂配伍增效的物质基础“药动学成分群”，并利用联合模型，进行以下研究：.1.考察慢性炎症状态下，SLC22A转运体亚族中OCTN1/2、OATP2B1、OAT1/3和ABC转运体MRP1、MRP2在模型动物、细胞中功能和蛋白表达的变化情况，并对分子机制进行了探讨。结果发现，慢性炎症状态下在模型大鼠和模型A549细胞中，①支气管扩张药沙丁胺醇等的转运体OCTN1/2蛋白、mRNA表达及摄取功能显著降低。而抑制剂PDTC、激动剂HSP70及沉默、过表达p56实验显示，下调作用是通过NF-κB信号通路进行调控的。②炎症相关因子白三烯、前列腺素E2等转运体OATP2B1炎症状态下蛋白、mRNA表达及摄取功能均上调。③烟草致癌物外排转运体MRP1、MRP2炎症状态下蛋白、mRNA表达均下调。机制研究表明，Wnt3a/β-catenin信号通路参与其调节过程，而MRP2的下调则是通过FXR信号通路来调控的。.2.考察了玉屏风、芪术药对及单味药对炎症状态下转运体活性和表达的干预作用。结果发现，玉屏风、黄芪等可明显降低模型A549细胞的炎症因子，尤以玉屏风的降低效果最显著。玉屏风可升高OCTN1/2的表达，且与黄芪、白术和防风组相比，玉屏风组OCTN1/2的表达最高。提示玉屏风能通过提高转运体OCTN1/2的表达及功能，促进药物跨肺泡上皮细胞的转运吸收，产生药效协同。.研究为阐释炎症状态下药物药动学相互作用的分子机制提供信息；同时，也阐释了方剂配伍增效与成分群药动学相互作用之间的联系及其分子机制，论证了组方的合理性，廓清疾病-转运体-药物三者关系。这对于揭示中药作用的物质基础、阐释复杂的中药方剂配伍增效分子机制，具有重要意义；也能为方剂质量控制、制剂改进、临床合理用药、发现慢性炎症性疾病的治疗新靶点，提供了更多信息。