基于“代谢轮廓-菌群调节-药效靶点”模式研究生脉饮“益心复脉”的作用机制

Shengmai Yin is a prescription for nourishing heart and restoring pulse. The spleen, stomach and heart are connected by meridians, five elements and functions. Previously, our group found that Shengmai Yin polysaccharide could improve intestinal damage and intestinal flora Shengmai Yin, a classic formula to boost the heart and restore the pulse, has been widely used in the treatment of such cardiac diseases as myocardial ischemia with significant therapeutic effects. However, its mechanism of action has been unclear yet. Traditional Chinese medicine (TCM) has provided important insights into such diseases, including ‘supplementing the spleen to nourish the heart’ and ‘the heart is paired with the small intestine’; while in recent years, gut microbiota has been proved to be a new and effective medicine target for the treatment of cardiac diseases. The applicant's previous study found that Shengmai Yin and its polysaccharide could significantly improve the diversity of gut microbiota. Hereby, the applicant proposes the hypothesis that the heart-nourishing and pulse-restoring Shengmai Yin can activate the cardiac target via regulating the gut microbiota and affect the gastrointestinal absorption of small-molecule effective components so as to treat myocardial ischemia. In this project, the 16S rRNA high-throughput sequencing technology will be used to perform a comparative study on gut microbiota diversity in myocardial ischemia-model rats; the in vivo intestinal absorption model will be established to investigate the effect of Shengmai Yin on the absorption kinetics features of effective component groupings; the metabonomics approaches will be applied to screen the commonly differentially-expressed metabolites of the gut microbiota involved in the plasma and urine of the rats, as well as related targets and metabolic pathways; and the Spearman correlation analysis will be adopted to create the ‘Metabolic Profile-Microbial Regulation-Pharmacodynamic Target’ Model of Shengmai Yin. The results of this project will reveal the mechanism of Shengmai Yin's ‘boosting the heart and restoring the pulse’ action via regulating the gut microbiota for the treatment of myocardial ischemia and further provide a new idea to illuminate the ‘therapeutic effect’ of TCM classic formula for clinical application.

生脉饮为益心复脉的经典方剂，临床上被广泛用于治疗心肌缺血等心系疾病，疗效确切，但其作用机制亟待阐明。“以脾养心”、“心合小肠”是中医对心系疾病发病机制的重要论述；近年研究亦证实中药可通过调控肠道菌群治疗心系疾病。申请者前期研究发现生脉饮及其多糖能显著改善肠道菌群的多样性。鉴此，提出“生脉饮可通过调节肠道菌群激活心脏靶点并影响小分子有效组分的胃肠吸收发挥益心复脉的作用治疗心肌缺血”的假说。采用16S rRNA高通量测序技术，研究心肌缺血模型大鼠肠道菌群多样性；建立在体肠吸收模型，考察生脉饮对有效组分群吸收动力学特征的影响；运用代谢组学方法，筛选大鼠血浆和尿液中肠道菌群参与的共有的差异性代谢产物及相关靶点和代谢通路，采用spearman相关性分析，建立生脉饮“代谢轮廓-菌群调节-药效靶点”模型，以揭示生脉饮调节肠道菌群以“益心复脉”治疗心肌缺血的作用机制，为阐明经典名方临床疗效提供一种新思路。

中医的灵魂是整体观念和辨证论治，而中药汤剂是最能体现中医特色的经典剂型，然而“物质基础”和“作用机制”是研究中药汤剂的两个核心问题。生脉饮具有益气复脉、养阴生津之功效，为益心复脉之经方，但其作用机制有待进一步深入探索。为深入探讨传统汤剂的制剂内涵以及大分子和小分子活性成分在汤剂中的作用，从化学成分、药效表达、网络药理、在体吸收等多维度，研究了生脉饮改善心肌缺血的药效物质基础和作用机制。本项目采用UPLC-Q-Orbitrap HRMS技术，较全面表征了生脉饮中的主要化学成分；建立垂体后叶素所致心肌缺血大鼠模型，采用HE染色、酶联免疫等方法分析了相关生物学指标，研究结果提示生脉饮预防脂质过氧化损伤、降低细胞损伤；采用高通量测序技术，运用生物信息学分析心肌缺血大鼠的肠道菌群差异性；采用双变量spearman分析生物学指标和标识菌群的相关性；采用MicroPITA分析代表性样本，发现生脉饮可能通过升高拟杆菌属的丰度，参与调节心肌损伤的水平，发挥“益心复脉”的作用；采用大鼠在体肠灌流模型，从病证模型角度研究生脉饮主要化学成分的吸收特点，从生物学指标-肠道微生物层面研究大分子多糖参与生脉饮发挥疗效的特点，同时对生脉饮中经典药对人参-五味子中有效成分人参皂苷以及五味子木脂素进行组分配伍研究，揭示了生脉饮及含人参-五味子药对复方的组方原理以及配伍规律。本项目的开展研究了生脉饮中化学成分-肠道菌群-药效表达发挥作用的主要途径，揭示了生脉饮主要通过调节免疫炎症反应、氧化应激等通路以发挥疗效，有利于进一步推动生脉饮的临床应用和制剂开发，为阐明中药传统汤剂的科学内涵提供一种研究思路。