基于肠道菌群介导的五味子醋制增强收敛止泻效应机制研究

The processing of traditional Chinese medicine (TCM) is the traditional pharmaceutical technology with the most independent intellectual property rights.The processing of detoxification and synergism is the characteristic of clinical practice of TCM. However, the theory of Chinese medicine processing which guides the rational and effective application of Chinese herbal medicine is still lacking enough modern scientific evidence, becoming a bottleneck restricting the development of TCM modernization. The occurrence of disease is related to intestinal flora imbalance, and it is now a hot topic in the whole theory of TCM. Schisandrae Chinensis Fructus (SCF) has the effect of convergence, liver preservation and so on. SCF after processed with vinegar can enhance the convergence antidiarrheal effect, but the mechanism of its synergistic effect has not yet been studied. In view of the above problems, the project is based on the previous research, taking the intestinal flora - convergent antidiarrheal as the breakthrough point, modern techniques such as metagenome are applied to processing mechanism. To identify the effector substances through network pharmacology, screening out the target protein, combined with metabolomics pathway analysis, to evaluate the correlation between effective substances, target proteins, metabolites, pathways and the convergence antidiarrheal effect. This project combines macro system of TCM and modern microtechnology, to clarify the hypothesis that SCF after processed with vinegar exerts therapeutic effect possibly through regulating metabolism and metabolic transformation of effector substances and intestinal flora, regulating intestinal microenvironment, target protein and metabolic pathway. From multi-component, multi-target, multi-channel system interpretation of Chinese medicine theory, it provides a scientific basis for clinical application of SCF, and also provides a demonstration for the modern research of Chinese herbal pieces.

中药炮制是最具自主知识产权的传统制药技术，炮制解毒增效为中医临床用药特色，但指导其合理用药的炮制理论尚缺乏现代科学论据，成为制约中药现代化的瓶颈。疾病发生与肠道菌群失调密切相关，目前为中医药整体理论研究热点。五味子具收敛、保肝等功效，醋制增强收敛止泻作用，但其增效机制未见研究。针对上述问题，项目在前期研究基础上，以肠道菌群-收敛止泻为切入点，将宏基因组等现代技术引入炮制机制研究。通过网络药理学辨识效应物质群，分析关联靶点，结合代谢组学通路分析，评价效应物质、靶蛋白、代谢组分及通路与收敛止泻的关联性。将中医宏观体系与现代微观技术有机结合，阐明五味子醋制是“通过效应物质与肠道菌群的相互调节与代谢转化，多元调控机体肠道微环境、靶蛋白及代谢通路等，导致五味子醋制效应差异变化”假说的科学内涵。从多成分、多靶点、多途径系统诠释中药炮制理论，为五味子临床应用提供科学依据，也为中药饮片现代研究提供示范。

五味子具收敛固涩等功效，醋制可增强其涩肠止泻作用。项目在前期研究基础上，以肠道菌群-收敛止泻为切入点，将16S rRNA测序等现代技术引入炮制机制研究。通过网络药理学辨识效应物质群，分析关联靶点，结合代谢组学通路分析，评价效应物质、靶蛋白、代谢组分及通路与收敛止泻的关联性。从多成分、多靶点、多途径系统诠释中药炮制理论，为五味子临床应用提供科学依据，也为中药饮片现代研究提供示范。.以DSS成功诱导建立了小鼠溃疡性结肠炎模型，结合16S rRNA基因测序、基于UPLC-Q-TOF/MS技术的代谢组学及多元统计分析对肠道菌群的结构组成和多样性进行分析，发现生、醋五味子多糖及提取物的摄入能显著缓解和改善肠道菌群失调，减轻肠道炎症疾病的症状显著调节SCFAs的含量（代谢水平），且醋五味子调节作用更显著；并鉴定得PE、LysoPE、PC、LysoPC、PG 等32个组间差异性潜在生物标记物，生、醋五味子主要调控UC小鼠的酪氨酸代谢、精氨酸生物合成、丙氨酸、天冬氨酸和谷氨酸等氨基酸代谢通路以及谷胱甘肽代谢通路，且醋五味子调控作用更显著。Western blot分析发现，生、醋五味子对结肠TLR4/NF-κB/NLRP3通路及紧密连接蛋白调控作用显著，且醋五味子调控趋势更佳，结合生、醋五味子治疗UC生物标志物和入血成分相关性分析，筛选出当归酰戈米辛H、五味子醇甲、五味子乙素、五味子醇乙、Diisobutylphthalate、戈米辛S、苯甲酰戈米辛H为五味子醋制增强涩肠止泻的潜在效应物质。.综上所述，生、醋五味子通过调节肠道菌群的结构组成和多样性、影响短链脂肪酸代谢以及炎症因子的表达水平、调控TLR4/NF-κB/NLRP3信号通路、紧密连接蛋白表达，调节下游酪氨酸代谢等氨基酸代谢通路，改善UC肠道炎症反应，发挥涩肠止泻功效，且醋五味子效果更佳。此外，当归酰戈米辛H等化合物可能是生、醋五味子涩肠止泻效应物质。为五味子饮片临床合理应用提供科学依据，也为中药炮制传统理论的现代研究提供示范。