小续命汤治疗缺血性脑卒中的脑内整合药代特征研究及干预血脑屏障损伤的多靶点机制

Under the traditional Chinese medicine (TCM) theories, Xiao-xu-ming decoction (XXMD), the classic TCM prescription, has been with systematic pharmacodynamic studies granted by the National Natural Science Foundation in the prophase work stage. Now, it will be further studied on the pharmacokinetic profile in the current project by the weight mode of pharmacological action factor. The current project intends to put the focus on the systematic pharmacokinetic mode for TCM prescription study in brain, via researching on the pharmacokinetic profiles of active ingredient group of XXMD, evaluating the weight factor for pharmacological action, then, developing the new mathematics models to reveal the pharmacokinetic characteristics of the integrity composition of XXMD. During the studies, the gerbil pathological and physiological self-control brain regions model and the real-time microdialysis technique are applied in order to sample from brain regions dynamically and continuously in the periods of ischemia, reperfusion and recovery. Furthermore, the research puts the highlight on illuminating the protective effect and molecular mechanism of XXMD against ischemic brain injury at blood-brain barrier (BBB). Whether the anti-ischemia effect of XXMD presented by protecting the physical membrane barrier system and efflux transporter system of the BBB, and regulating the mitochondrial proteome in endothelial cell, we will investigate in-depth studies. Finally，By exploring the metabolism characteristics and the pharmacological action of the effective components, we try to clarify the substance foundation of prescription action in vivo. Via studying the effective components (including prototype drugs, metabolites and adduct products), we intend to elucidate the reasonable and scientific TCM compatibility theories in the aspects of pharmacokinetic characteristics and pharmacodynamic molecular mechanism of XXMD, and aim at achieving the purpose of providing new modes for the TCM prescription study and convincing scientific data to support the rich connotation of the TCM compatibility theories.

本课题以中药理论为指导，针对在自然基金资助下经过系统药效学研究的传统经典复方"小续命汤"，应用沙鼠自身双侧脑区的病理、生理对照模型，采用微透析技术实时、动态、连续监测动物给予小续命汤有效成分组后，在缺血期、再灌注期及恢复期双侧脑区各药物成分的经时变化过程，依据同步检测的药效特征，尝试以药效权重的新模式进行药代特征轮廓研究，实现药效权重模式下的小续命汤各物质成分整合药代特性评价。并进一步从血脑屏障的物理性膜屏障系统、外排转运体系统、内皮细胞线粒体蛋白质组系统阐释小续命汤入脑、入血、入血未入脑成分对缺血性血脑屏障损伤的保护作用及相关分子靶点机制，阐明其发挥药效的物质基础，探讨其脑靶向优势。进而针对效应成分（包括药物原型、代谢产物及加合产物）进行药材来源的归属，最终实现从药物代谢层面和药理分子机制层面共同揭示其组方的深层配伍机制，为诠释传统中医药配伍理论的丰富内涵提供有力的科学数据支持。

本课题关注经典复方“小续命汤”，应用沙鼠自身双侧脑区的病理、生理对照模型，采用微透析技术监测小续命汤有效成分组在缺血期、再灌注期及恢复期双侧脑区各药物成分的经时变化过程，依据同步检测的药效特征，实现药效权重模式下的小续命汤各物质成分整合药代特性评价。并进一步从血脑屏障的物理性膜屏障系统、外排转运体系统、内皮细胞线粒体蛋白质组系统阐释小续命汤对缺血性血脑屏障损伤的保护作用及相关分子靶点机制，阐释其发挥药效的物质基础。进而针对小续命汤效应成分进行药材来源的归属，最终实现从药物代谢层面和药理分子机制层面共同揭示其组方的深层配伍机制。.此外，基于UPLC/Q-TOF-MSE分析小续命汤有效成分组的化学成分, 共鉴定出42个化合物，主要包括黄酮类、生物碱类、三萜皂苷类，归属了各化合物的单味药来源，为其药效物质基础研究奠定了基础。.应用UPLC-Q-Exactive技术的蛋白质组学法研究发现，MCAO模型组和小续命汤组差异蛋白主要可归为参与细胞转化、生物调节、代谢过程、应激反应、细胞组分组成与合成、免疫系统过程，相关蛋白靶点及其涉及的生物体调控通路可能是小续命汤有效成分组对治疗局灶性脑缺血发挥作用的生物学基础。.小续命汤有效成分组对局灶性脑缺血再灌注大鼠损伤恢复早期即产生神经保护作用，可能与调节脑内氧化应激水平有关。其能够明显减轻脑缺血再灌注早期导致的脑组织能量代谢紊乱，改善脑线粒体结构和功能损伤，表明其对脑线粒体的保护作用可能是其发挥神经保护的作用机制之一。.课题组已按任务书中相关研究内容完成研究计划。并在国内外学术期刊上发表相关研究论文18篇。SCI论文2篇，EI论文1篇，国内核心期刊15篇，国际会议论文4篇，获得国际会议分组报告优胜奖1项。培养硕士研究生8人，其中毕业6人，在读2人。获得国家发明专利授权1项。.本课题组对中药配伍理论研究新模式进行了较为深入的探讨，有助于为同类复方的配伍研究提供思路借鉴和技术示范。并为诠释传统中医药配伍理论的丰富内涵做出有益尝试。.