醋甘遂泻水逐饮效应物质基础及"有故无殒"作用机制研究

As a diuresis holy medicine of TCM clinical, kansui Radix, the toxic Chinese materia medica have to be stir-baked with vinegar for oral administration. However, the existing research results not yet been able to fully answer the scientific question, namely, the material basis of expelling retained fluid by purgation and the action mechanism of “YOU GU WU YUN” of kansui stir-baked with vinegar. Based on the completed national natural science foundation object which has basically clarified the material basis and mechanism of toxicity reduction of kansui after processing with vinegar, the application guidelines of the toxic drug of TCM, and the clinical practice of expelling retained fluid by purgation by kansui stir-baked with vinegar through oral administration in TCM, We think that, compared with physiological state, there are higher therapeutic index and wider safety range under pathological state for kansui stir-baked with vinegar. So, there is “YOU GU WU YUN” in a certain range of dose, and play the effect of expelling retained fluid by purgation while ensuring drug safety in clinical Thus, the effect indexes which coincide with edema distention and fullness and hydrothorax and ascites in TCM were selected to tract the expelling retained fluid by purgation active substance of kansui stir-baked with vinegar by bio-guided method. To reveal the intrinsic nature of “YOU GU WU YUN” of kansui stir-baked with vinegar by studying the differences and characteristics of efficiency - poison - metabolism - metabonomics under physiological and pathological state of kansui stir-baked with vinegar from the different levels of whole, organs, cells and molecules. Provide scientific basis for clinical application safely and effectively of kansui, and exploration of ideas and methods for the basic research of scientific application of toxic Chinese materia medica.

有毒中药甘遂为中医临床"下水之圣药"，内服需醋炙。但现有研究结果尚未能全面解答醋甘遂泻水逐饮的效应物质基础及"有故无殒"作用机制等科学问题。基于前期所完成的国家自然基金课题已基本阐明甘遂醋炙减毒物质基础及作用机理，及中医传统有毒药应用"有故无殒"指导思想和中医临床采用醋甘遂内服泻下逐水的实践，我们认为：醋甘遂对于病理状态有比正常生理状态高的治疗指数和宽的安全范围，在一定剂量范围内表现为"有故无殒"，从而在临床上发挥泻水逐饮功效的同时又保证了用药安全。因此本研究选择符合醋甘遂中医辨证用药原则的水肿胀满、胸腹积水效应指标，应用生物导向分离技术追踪醋甘遂泻水逐饮功效的活性物质；从整体、组织、细胞及分子等不同水平研究醋甘遂在生理与病理状态下效-毒-代谢-代谢组学等各方面的差异及特点，以期揭示醋甘遂"有故无殒"的内在本质，为甘遂临床安全有效应用提供科学依据，为有毒中药科学应用基础研究提供思路与方法等方面的探索。

本项目基于中医传统有毒药应用“有故无殒”指导思想和中医临床采用醋甘遂内服泻下逐水的实践，复制与甘遂中医临床应用相适应的癌性腹水模型，建立甘遂泻水逐饮活性评价体系，采用适宜方法与技术开展甘遂在生理与病理状态下效-毒-代谢组学等研究，明确了醋甘遂泻水逐饮效应的物质基础，从实验水平揭示了醋甘遂“有故无殒”应用的科学内涵，为甘遂临床安全有效应用提供了科学依据，为有毒中药科学应用基础研究提供了良好示范，产生了较好的社会效益。.1.通过对给予癌性腹水模型动物生/醋甘遂及其提取物后大鼠腹围/体重比、尿量、尿中离子、尿液pH、腹水量、肾素-血管紧张素-醛固酮系统、抗利尿激素、胃肠AQP8表达及腹水中免疫抗炎因子含量等的研究表明，生、醋甘遂均可表现出显著的利尿消腹水作用，其作用机制可能与调节RAAS系统和免疫抗炎有关。对给予生/醋甘遂各样品的正常/模型动物肝、胃、肠组织病理形态学、肝功能、氧化损伤、炎性损伤和细胞凋亡等指标的测定结果表明，生、醋甘遂各样品均可致正常大鼠一定程度的组织损伤，而对模型动物则可显著缓解模型造成的组织损伤，使其毒性降低，即表现为“有故无殒”。.2.在明确乙酸乙酯部位为醋甘遂泻水逐饮活性部位基础上，以项目组分离得到的12个萜类成分为对照，采用UPLC/Q-TOF-MS法和UPLC-TQ-MS法对醋甘遂A、B、C、D各成分群中化学成分进行定性定量分析，共鉴定出31个化合物，其中成分群B主要含巨大戟烷型二萜类及三萜类成分，总量为90.1μg/mg；成分群C主要含假白榄烷型二萜类，总量为16.6μg/mg；成分群A仅含少量种类巨大戟烷型二萜类成分，不含三萜类成分，其可测定萜类成分总量为6.57μg/mg；成分群D基本不含所测二萜类和三萜类成分。进一步明确B、C成分群为醋甘遂泻水逐饮活性成群。.3.尿液和血液代谢组学研究结果表明，醋甘遂及其成分群B、C泻水逐饮效应发挥可能与调节脂质代谢、花生四烯酸代谢、胆汁酸代谢和氨基酸代谢异常有关，通过抑制炎症因子释放和肿瘤细胞生长，增强机体免疫功能和抗氧化能力，减轻炎症反应、氧化损伤和血管内皮细胞损伤，并调节肾素释放，对肾素-血管紧张素-醛固酮系统产生影响，纠正体内水盐代谢紊乱。该结果与效－毒研究结果相印证。.项目实施期间发表论文19篇，其中SCI收录8篇；获发明专利授权1项并获转让经费50万元；培养研究生6名，其中博士研究生1名。