从血脂调控JAK2/STAT3通路探索黄连吴茱萸苦辛化浊效应的关键机制

Hyperlipidemia is frequently occurring illnesses in clinical. JAK2/STAT3 Signal Pathway is closly connected with regulation of fat metabolism. In Traditional Chinese Medicine, it is thought that dampness and heat, phlgem turbid obstructed in the body and abnormal function of the spleen and stomach is the main pathogenesis of disruption of fat metabolism. Combination of bitter tasted Huang Lian(Coptis chinensis) for dispersion and pungent tasted Wu Zhuyu for purgation has the effect of transporting fat and blood stasis which has the function of lowering blood lipids, but there are few studies on them. To discuss the molecular signal mechanism of combination of bitter Huanglian for purgation and acrid Wu Zhuyu for dispersion to lowering blood lipids and interpret the scientific connotation of description compatibility theory, we put on the hypothesis that 'the most important mechanism of lowering blood lipids and transporting the turbid by the combination of Huang Lian and Wu Zhuyu is depended on the regulation of JAK2/STAT3 signal pathway' on the basis of related researches before, selecting the combination of Huang Lian and Wu Zhuyu as the observing target, studing the internal connection between the effect of lowering blood lipids by the combination of Huang Lian and Wu Zhuyu and the regulation of key target genes of JAK2/STAT3 signal pathway by molecular biology technique with models in vivo and in vitro on the basis of research work before. The research can not only express the molecular signal mechanism of lowering blood lipids by the combination of Huang Lian and Wu Zhuyu and is very important in widening the clinical application, it can also interpret the scientific connotation of prescription compatibility theory, providing thoughts and methods for deeper and systematical studies on the combination of components of classic descriptions of TCM.

高脂血症是当今临床高发病， JAK2/STAT3信号通路与脂代谢调控密切相关。中医认为湿热痰浊内蕴、升降失常是脂质代谢紊乱的重要病机。黄连、吴茱萸配伍苦辛通降，运化膏脂血浊，具有明确降脂作用，但二者配伍调节脂代谢的研究甚少。为探讨其配伍苦辛化浊以降脂的分子信号机制，阐释方剂配伍理论的科学内涵，在前期研究基础上，提出"黄连-吴茱萸辛开苦降配伍以降脂化浊的关键机制在于通过调控JAK2/STAT3信号通路达到降脂效应"假说，借助体内外两种模型，以黄连-吴茱萸配伍为对象，结合前期研究基础，运用分子生物学技术，研究黄连-吴茱萸配伍降血脂效应与JAK2/STAT3信号通路关键靶基因调控的内在联系。该研究不仅对揭示黄连-吴茱萸配伍降血脂的分子信号机制、拓宽其临床适用范围具有重要意义，还可诠释辛开苦降方剂配伍理论的科学内涵，为深入、系统开展中医经典小方的配伍机制研究提供思路与方法。

本项目以高脂模型大鼠为载体，以黄连吴茱萸1:1配伍为研究对象，从脂代谢的重要调控通路Leptin-AMPK和Leptin-JAK2-STAT3信号通路以及胆固醇吸收关键因子NPC1L1、ACAT2、ApoB48入手，探讨了黄连吴茱萸配伍对于JAK2/STAT3 信号通路及胆固醇吸收相关因子表达的影响，明确了黄连-吴茱萸配伍调控脂质代谢的分子机制。 .本研究以SD大鼠为实验对象，通过持续喂养高脂饲料4周建立高脂血症动物模型。造模成功后，分别给予小檗碱高剂量（72.8mg/kg）、小檗碱低剂量（36.4mg/kg）、吴茱萸碱高剂量（16.8mg/kg）、吴茱萸碱低剂量（8.4mg/kg）、小檗碱吴茱萸碱配伍高剂量（89.2mg/kg）、小檗碱吴茱萸碱配伍低剂量（44.6mg/kg）灌胃干预8周。研究发现，44.6mg/kg和89.2mg/kg配伍剂量组能显著降低高脂血症大鼠血清TC、LDL-C水平(P＜0.05)，且降脂作用优于同等剂量的小檗碱和吴茱萸碱。通过脂代谢重要通路Leptin-AMPK和Leptin-JAK2-STAT3中关键靶点表达检测发现，小檗碱吴茱萸碱配伍黄连吴茱萸辛开苦降配伍对Leptin-AMPK 和Leptin-JAK2-STAT3信号通路中各靶点的调节具有协同增效作用。.此外，本项目在前期研究基础上，升高小檗碱与吴茱萸碱配伍剂量，采用89.2mg/kg、178.4mg/kg配伍组干预高脂模型大鼠4周，研究小檗碱与吴茱萸碱配伍对高胆固醇血症大鼠小肠ACAT2 、ApoB48和NPC1L1表达的影响。结果表明，178.4mg/kg配伍组大鼠血清总胆固醇、β-谷固醇水平较模型对照组显著降低(P＜0.05)，89.2 mg/kg 配伍组极显著降低(P＜0.01)；小檗碱与吴茱萸碱配伍89.2mg/kg和178.4mg/kg剂量组可降低高脂模型大鼠小肠组织ACAT2、ApoB48 和NPC1L1 蛋白的表达(P＜0.05或P＜0.01)。.综上所述，本项目明确了黄连吴茱萸配伍对高脂血症模型动物脂质水平的调控研究，并从分子水平阐明黄连吴茱萸配伍降脂的作用机制与调控脂代谢重要通路Leptin-AMPK和Leptin-JAK2-STAT3以及降低小肠胆固醇吸收有关，对于揭示方剂辛开苦降配伍理论及中药降脂研究提供新思路。