茵陈术附汤抗肝内胆汁淤积的物质基础和分子机制研究：从药动学和调控胆汁酸转运体/代谢酶-炎症通路解析

Intrahepatic cholestasis (IHC) is common in liver diseases, and there is a shortage of effective treatment. Cholestasis belongs to the category of "traditional Chinese medicine (TCM) jaundice". Yinchenzhufu Decoction (YCZFD) has been a representative TCM prescription for the treatment of jaundice. In our previous studies in mice models, results showed that YCZFD exhibited an obvious protective effect against IHC, and alleviated jaundice, liver inflammation and hepatic damage. YCZFD up-regulated the mRNA expression of CYP2b10, UGT1a1, BSEP, MRP2, and upstream nuclear receptor FXR, while down-regulated the mRNA expression of TNF-alpha and IL-1beta. Accordingly, we propose a hypothesis that YCZFD could suppress IHC by regulating the expression and function of bile acid transporters, metabolizing enzymes and inflammatory factors. To test this hypothesis, the effects of YCZFD on liver function, histopathology, and pharmacokinetic-pharmacodynamic analysis will be investigated to determine the effective composition of YCZFD in acute and chronic mice IHC models. Furtherly, the effect of YCZFD on serum metabonomics, bile acid profiles, bile acid transporters and metabolizing enzymes, as well as on related nuclear receptor and inflammatory factor will be investigated in animal models and cell models. This research would help elucidate the effective constituents and molecular mechanism of YCZFD for anti-IHC, alleviation of jaundice and hepatoprotective effect, and provide a basis for further research and development of YCZFD in anti-IHC treatment.

肝内胆汁淤积(IHC)广泛存在于慢性肝病，其治疗药物不足。胆汁淤积属中医“黄疸”范畴，茵陈术附汤(YCZFD)是中医治疗黄疸的代表方剂。我们前期动物实验表明YCZFD可改善小鼠IHC，减轻黄疸、肝脏炎症和肝损伤，上调Cyp2b10、UGT1a1、BSEP、MRP2及上游FXR的mRNA表达，下调TNF-alpha和IL-1 beta的mRNA表达。据此，我们提出YCZFD通过调控转运体/代谢酶-炎症通路，产生抗IHC和退黄保肝作用的假说。本项目采用急性和慢性小鼠IHC模型，研究YCZFD对肝功能及病理学的影响，通过体内药动学-药效学的联合分析确定药效成分；并进一步采用整体动物模型和多种细胞模型，研究YCZFD对代谢组学、胆汁酸谱、胆汁酸转运体/代谢酶及其调控因子、炎症因子的影响，阐明YCZFD抗IHC和退黄保肝的效应成分与分子机制，为YCZFD治疗IHC的进一步研发提供基础。

肝内胆汁淤积(IHC)广泛存在于慢性肝病，其治疗药物不足，中医在临床实践中形成了多个治疗IHC（中医属“黄疸”范畴）的经典方剂，具有独特优势,茵陈术附汤(YCZFD)是中医治疗IHC的代表方剂。本项目采用小鼠急性和慢性IHC模型和多种细胞模型，研究YCZFD对肝功能及病理学、代谢组学、胆汁酸谱、胆汁酸转运体/代谢酶及其调控因子、肠道屏障、肝脏炎症通路和胆管损伤等的影响，并进行体内药动学的联合分析，阐明YCZFD抗IHC的效应成分与分子机制。结果：①YCZFD对ANIT诱导的胆汁淤积性肝损伤具有明显的保护作用，其作用机制与调节胆汁酸转运体和代谢酶表达，改善胆汁酸紊乱有关，并与抑制肝脏炎症以及改善胆管损伤有关。②YCZFD具有抗DDC诱导的胆汁淤积性肝损伤作用，其与诱导胆汁酸代谢酶表达、促进胆汁酸代谢、降低胆汁酸在体内淤积有关，还与修复胆汁淤积小鼠肠道屏障、减少内毒素移位、抑制TLR4/Myd88/NF-κB炎症通路介导的肝脏炎症反应有关。③药动学研究获得了YCZFD在动物体内多成分的药动学规律，阐明了潜在的药效成分；用HepG2细胞、三明治培养原代肝细胞及胆汁外排受阻模型、FXR和NF-B双荧光素酶报告基因模型、TLR4基因诱导过表达细胞模型和RAW264.7细胞等多种细胞模型研究了这些成分抗胆汁淤积性肝损伤的作用机制，阐明滨蒿内酯、肉桂酸、芒柄花素、芒柄花苷、白术内酯III、绿原酸、甘草次酸、甘草苷、甘草素、苯甲酰乌头原碱、苯甲酰新乌头原碱、苯甲酰次乌头原碱分别通过提高胆汁酸外排转运体/代谢酶表达和活性、促进胆汁酸外排、激动FXR、抑制NF-κB转录和抑制TLR4和炎症因子表达等不同机制改善胆汁淤积性肝损伤，为体内发挥药效的成分。研究结果阐明了YCZFD抗IHC的机制和物质基础，对于经典方剂的传承及创新具有重要的意义，为YCZFD治疗IHC的进一步新药研发提供了基础。