肠道菌群介导的内酯类组分C-位磺酸化代谢在穿心莲保肝利胆中的作用

Andrographis paniculata has exact hepatoprotective effect. Its active ingredients are lactones. The low blood concentration of lactones and its strong efficacy are "paradox". Gut microbiota metabolism is one of the important reasons for this paradox. In our previous studies we found that the bioavailability of lactones was extremely low, but lactones were greatly metabolized into C-sulfonated metabolites by gut microbiota in rats and healthy volunteers. The exposures of C-sulfonated metabolites in rat liver and its protective effects were significantly greater than that of the prototype, where hepatic FXR and bile acid transporters BSEP/MRP2/MRP3/MRP4 expression was up-regulated. Thus, we propose the scientific hypothesis: the active ingredients of Andrographis paniculata are converted into C- sulfonated metabolites by gut microbiota, and then activate hepatic FXR, up regulate BSEP/MRPs expression to protect liver. We aim to use pseudo germ-free/germ-free rats, 16SrDNA and proteomics technology to identify the strains and key enzymes involved in C-sulfonated metabolism, and then elucidate its metabolic mechanism. Secondly, we intend to elaborate the relationship of lactones-gut microbiota- hepatoprotective effect. Finally, we plan to use of FXR knockout mice and siRNA interference technology to clarify the mechanism of Andrographis paniculata hepatoprotective effect. The results will provide new theoretical basis for rational use of Andrographis paniculata.

穿心莲的功效“清热燥湿”，具有确切的保肝利胆作用，其有效组分内酯类极低的血药浓度与较强的药效作用形成“悖论”。从肠道菌群代谢的角度诠释此悖论已成为热点。我们前期研究发现虽然内酯类组分的生物利用度极低，但在大鼠和健康志愿者体内均大量转化为由肠道菌介导的C-位磺酸化代谢物，其大鼠肝脏暴露量和作用显著地大于原型，且导致肝脏FXR和胆汁酸转运体BSEP/MRP2/MRP3/MRP4表达上调。由此提出科学假说：穿心莲口服后内酯类组分经肠道菌转化为C-位磺酸化代谢物，进而激活肝脏FXR，上调BSEP/MRPs，发挥作用。本项目拟利用伪无菌/无菌鼠、16SrDNA和蛋白质组学技术，鉴定参与内酯类C-位磺酸化代谢的菌株及其关键酶，阐明其代谢的机理；明确内酯类组分-肠道菌群-保肝利胆的内在关联；最后运用FXR基因敲除鼠及siRNA干扰技术，阐明穿心莲保肝利胆的作用机制，为其临床合理应用提供新的理论依据。

穿心莲的功效“清热燥湿”，具有确切的保肝利胆作用，然而穿心莲的有效成分在肠道会发生强烈的C-位磺酸化代谢。本项目拟基于前期工作基础，研究肠道菌群介导的C-位磺酸化代谢的机理、肠道菌群介导的C-位磺酸化代谢在穿心莲保肝利胆中的重要作用和穿心莲如何发挥抗胆汁淤积作用，阐明有效成分-肠道菌群-保肝利胆的内在关联及穿心莲保肝利胆的作用机制，为其临床合理应用提供新的理论依据。我们利用伪无菌、无菌小鼠模型及结合其它生物学方法首次阐明肠道菌群介导的穿心莲C-位磺酸化代谢的机理，发现了一种全新的药物代谢途径：肠道菌群分泌的APS还原酶能够将机体的硫酸根还原为亚硫酸氢根，亚硫酸氢根进攻穿心莲内酯的C=C不饱和双键发生亲核加成反应从而形成其C-位磺酸化代谢产物；在伪无菌和APS还原酶抑制小鼠模型，我们进一步发现该代谢反应显著影响穿心莲的抗胆汁淤积作用；最后，我们利用FXR敲基因小鼠等阐明穿心莲通过激活肝脏FXR，上调胆汁酸转运体而发挥抗胆汁淤积作用。