基于抗缺氧和氧化应激损伤的古方“四妙勇安汤”防治缺血性心脏病药效物质与作用机制研究

The famous ancient prescription Simiao Yong’an Decoction based on stasis-dispelling and toxin-resolving efficacy was widely and effectively used for the treatment of ischemic heart disease (IHD) in clinic, but the active ingredients and mechanisms have not yet been clarified. According to the important pathologic process of IHD, hypoxia and oxidative stress induced injury in cardiomyocytes were used as the screening and evaluation indexes. Our preliminary study found Simiao Yong’an Decoction offered protection against oxidative and hypoxia stress-induced myocardial cell damage. On this basis, the active ingredients and mechanisms of Simiao Yong’an Decoction for prevention and treatment of IHD will be investigated in this project. The active fractions will be determined through multi-level activity evaluation of the cardioprotective effects. A systematic chemical investigation of active fractions will be fulfilled through modern chromatographic and spectral techniques under the guidance of HPLC fingerprint spectrum-effect relationship. And then, the corresponding evaluations of the ingredients isolated from the active fractions will be carried out. The regulation function of MAPK/Pim-2 of the potential active compounds will be futher investigated. Finally, SAR relationship between trace elements by UPLC-TOF-MS and Pim-2 will be discussed, in order to reveal the scientific content of the famous ancient prescription Simiao Yong’an Decoction, a synergistic action against IHD by multiple active ingredients.

著名中药复方四妙勇安汤遵循“祛瘀”和“化毒”的治疗要点，古方新用，广泛用于缺血性心脏病的临床防治，疗效明显，但其相关药效物质基础及作用机制研究尚属空白。本项目从缺血性心脏病缺氧、氧化应激诱导心肌损伤的重要病理基础出发，选择相应活性筛选体系对四妙勇安汤进行药效评价，实验结果证实了四妙勇安汤抗心肌细胞氧化，提高缺血心肌耐缺氧的药效。在此基础上本项目首先通过多水平活性评价确定四妙勇安汤心肌保护的活性部位，并建立活性部位的谱-效相关性指纹图谱，以此为指导对活性部位进行系统的化学成分研究；对分离得到的化学成分采用相应活性测试体系进行活性评价，并进一步探讨潜在活性化合物预处理心肌细胞后对重要通路MAPK/Pim-2的共同调控作用；最后结合CDOCKER考察UPLC-TOF-MS检识的微量成分与Pim-2蛋白的构效关系，从而阐明“四妙勇安汤”药效物质群共同作用防治缺血性心脏病的科学内涵。

著名中药复方四妙勇安汤遵循“祛瘀”和“化毒”的治疗要点，古方新用，广泛用于心血管和内分泌疾病的临床防治，疗效明显，但其相关药效物质基础及作用机制研究尚属空白。本项目首先从动物和细胞两个层面，发现四妙勇安汤可以改善ISO诱发的心力衰竭大鼠模型的心脏功能，进一步通过RNA-seq数据分析和分子生物学实验，明确了四妙勇安汤抗心肌纤维化的分子机制主要是由AMPK激活引起的，从而抑制了其下游Akt/mTOR和TGF-β/SMAD3途径。接下来建立了四妙勇安汤活性部位的特征图谱，以此为指导对活性部位进行系统的化学成分研究，共分离得到32个化合物。通过UPLC-HR-ESI-MS技术，对主要的26个色谱峰进行了准确指认，并准确归属了其药材来源，同时采用该方法发现了大鼠血浆中的16个具有潜在活性的入血原型成分。最后构建和分析了四妙勇安汤入血成分—心力衰竭症靶点网络，发现入血成分与氧化还原调节信号通路、MAPK信号通路、细胞增殖紧密相关；入血成分的活性检测发现Angoroside C在ISO诱导的心力衰竭大鼠模型中改善了心脏功能降低的症状，3,5-dicaffeoylquinic acid表现出与阳性药地高辛相似的心肌保护作用，故而完成了四妙勇安汤入血原型成分心肌保护作用的初步机制研究工作。