基于肠道菌群-胆汁酸轴研究黄连-吴茱萸调控脂代谢作用机制

FXR/FGF15/CYP7A1 Signal Pathway which is modulated by the gut microbiota is closly connected with regulation of fat metabolism. In Traditional Chinese Medicine, it is thought that blocking of the spleen qi, dampness and phlegm turbid obstructed in the body is the main pathogenesis of disruption of fat metabolism. Compatibility of bitter tasted Huang Lian(Coptis chinensis) for dispersion and pungent tasted Wu Zhuyu(Fructus evodiae) for purgation has the effect of transporting fat and blood stasis, restoring the function of the spleen qi, which has the function of lowering blood lipids, but there are a very few studies on them. Our preliminary study found that mechanisms the down-regulating blood lipid effect of Huanglian- Wuzhuyu compatibility had connection with the inhibiting of cholesterol absorptions. Therefore, we proposed the scientific hypothesis that the mechanism of lowering blood lipids by the compatibility of Huang Lian and Wu Zhuyu is depended on the regulation of gut microbiota to activate FXR/FGF15/CYP7A1 signal pathway which impacts on the synthesis of bile acid. .As a starting point that the key target of gut microbiota and FXR/FGF15/CYP7A1 pathways mediated by gut microbiota, using metagenomic sequencing, liquid chromatography–tandem mass spectrometry (LCMS/MS), qPCR, Western blot and other detection techniques, we will research the effects of compatibility of Huang Lian and Wu Zhuyu on the HLP mice models of FXR、FGF15、FGFR4、CYP7A1 mRNA and protein expressions, which have a close relationship with synthesis of bile acid. The aim of the research is to explore the mechanism on FXR/FGF15/CYP7A1 pathway regulated by compatibility of Huang Lian and Wu Zhuyu, explicit the biological effects of compatibility of Huang Lian and Wu Zhuyu on HLP, and clarify the scientific connotation of acrid opening and bitter downbearing compatibility theory, provide the ideas for HLP in prevention research with TCM.

肠道菌群调控下的胆汁酸合成通路FXR/FGF15/CYP7A1与高脂血症（HLP）的发生密切相关。中医认为脾气壅遏，痰湿阻滞是高脂血症的重要病机。黄连、吴茱萸配伍辛开苦降，具有明确降脂作用，但二者配伍降脂机制尚不明确。前期研究发现，黄连吴茱萸配伍降脂作用与抑制胆固醇吸收有关，而胆汁酸是胆固醇的重要代谢途径，因此提出“黄连-吴茱萸配伍通过调控肠道菌群而激活FXR/FGF15/CYP7A1通路促进胆汁酸合成以达到降脂效应”假说，采用宏基因组测序、高效液相色谱-质谱联用、qPCR等方法，通过黄连-吴茱萸配伍对HLP小鼠肠道菌群结构及胆汁酸合成途径关键因子FXR、FGF15、FGFR4、CYP7A1表达的研究，探索其对FXR/FGF15/CYP7A1通路的调控机制，明确其干预HLP的生物学效应，阐明辛开苦降配伍理论的科学内涵，为HLP中医药防治研究提供思路。

本项目以高脂饮食诱导的高脂血症模型动物为载体，以黄连吴茱萸1:1配伍为研究对象，从肠道菌群以及胆汁酸合成信号通路FXR/FGF15/CYP7A1为切入点，采用宏基因组学以及代谢组学技术，探讨了黄连-吴茱萸1:1配伍对高脂血症模型动物肠道菌群、胆汁酸代谢、以及胆固醇合成关键通路FXR/FGF15/CYP7A1的影响，明确了黄连-吴茱萸1:1配伍调控高脂模型动物肠道微环境的分子机制。.本研究分别以SD大鼠和C57BL/6 小鼠作为实验对象，通过持续喂养高脂饲料建立高脂血症动物模型，给予黄连吴茱萸1:1配伍水煎液高中低剂量，以及等量小檗碱、吴茱萸碱、小檗碱吴茱萸碱不同剂量进行干预。研究结果发现，黄连吴茱萸等比配伍对于实验性高脂模型大鼠与小鼠均具有明确的降脂效应，主要体现在降低血脂水平、改善肝脏功能、调节炎症水平，以及调控血清胆汁酸水平等方面。其次，黄连吴茱萸等比配伍可增加高脂模型大鼠肠道菌群多样性，减少有害菌群表达、增加有益菌及肠道屏障修复菌群表达；相比两者单用，其等比配伍更能恢复高脂模型大鼠肠道菌群的多样性。另外，通过对高脂模型动物粪便胆汁酸代谢谱表达研究发现，黄连吴茱萸等比配伍可以上调高脂模型小鼠粪便中胆汁酸 CA、GCA、UDCA、7-KDCA 的表达，下调 12-KLCA、ApoCA、IsoLCA 的表达。此外，黄连吴茱萸等比配伍还可通过抑制 FXR/FGF15/FGFR4通路，上调 CYP7A1 的表达，促进胆汁酸的合成，影响胆汁酸代谢发挥调脂作用。.综上所述，本项目明确了黄连吴茱萸等比配伍对高脂血症模型动物血脂以及炎症水平的调控作用，初步探明了黄连吴茱萸等比配伍调控高脂模型动物肠道菌群及胆汁酸代谢的作用机制，并从分子水平阐释了黄连吴茱萸配伍降脂作用机制与调控胆汁酸代谢重要通路FXR/FGF15/CYP7A1有关，对于揭示方剂配伍理论及中药降脂抗炎研究提供新思路。