基于脑肠互动和脑靶向分布研究痛泻要方中防风的脾经引经作用及机制

Medicinal guiding can guide other traditional Chinese medicine to specified entrails or meridian and enhance the therapeutic effect of the prescription. The project supported by National Natural Science Foundation of China which confirms that the guiding action was correlated with tissue distribution. As a Chinese classical prescription, Tongxie Yaofang is commonly used to treat diarrhea due to spleen deficiency and liver hyperactivity. Saposhnikoviae Radix guides other drugs to the spleen meridian in TongxieYaofang. Our study indicates that Saposhnikoviae Radix can improve the effect of Tongxie Yaofang and the concentration of active ingredient in brain, the mechanism needs further research. It is known that the blood-brain barrier can affect the distribution of drugs in brain, and the central nervous system can regulate the intestinal function through brain-gut axis. On these grounds, we put forward the hypothesis that Saposhnikoviae Radix raises the distribution of other drugs in brain by the regulation of blood-brain barrier, and regulates the intestinal function through brain-gut axis to play guiding role. The design idea of this study is brain-gut axis, distribution of brain tissue, blood-brain barrier and the regulation of tight junction and transporters. We will establish the rat model with irritable bowel syndrome and blood-brain barrier model, with the method of Western Blot, UPLC-MS/MS, eletronic microscope observation and immunofluorescence, observe the effect of Saposhnikoviae Radix on the synergy action of TongxieYaofang and the tissue distribution of effective components, and further explore the effect of Saposhnikoviae Radix on blood-brain barrier permeability and the function and expression of transpoters. Our study will be helpful to reveal the meridine guiding effect of Saposhnikoviae Radix and explore the connotation of medicinal guiding and provide a new idea for the research of medicinal guide theory.

引经药能够引导药物到达特定的脏腑经络，增强方剂疗效。我们刚完成的国家自然科学基金证实：中药引经作用与药物靶向分布高度相关。痛泻要方主治脾虚肝旺之痛泻，方中防风为脾经引经之药。前期实验显示防风可提高全方药效，增加有效成分的脑分布，其机制有待深入研究。已知血脑屏障可影响药物脑分布，中枢神经可通过脑肠轴调节肠道功能。据此，我们推测：防风可能通过调控血脑屏障增加药物的脑组织分布，并经脑肠轴作用于肠道，从而表现脾经引经效应。根据脑肠轴→脑组织分布→血脑屏障→紧密连接和转运体调控的思路，本研究拟通过肠易激综合征大鼠和体外血脑屏障模型，采用Western Blot、UPLC-MS/MS、电镜观察、免疫荧光等手段，明确防风对痛泻要方的增效作用，考察防风对痛泻要方成分组织分布的影响，探讨防风对血脑屏障通透性、紧密连接和转运体的调控，揭示防风引经作用及机制，阐明防风脾经引经的科学内涵，为中药引经研究提供示范。

引经药能够引导药物到达特定的脏腑经络，增强方剂疗效，引经药是中药实现靶向给药的载体。防风在痛泻要方中具有脾经引经作用。本项目以中药引经与药物靶向分布具有相关性为突破点，中枢神经系统可通过脑肠轴调节肠道功能为主线，探讨防风是否调控血脑屏障增加药物的脑组织分布，并经脑肠轴作用于肠道，从而表现脾经引经效应。. 首先通过腹泻型肠易激综合征大鼠评价防风对痛泻要方的影响，结果显示防风可通过调节脑肠互动中脑肠肽、下丘脑-垂体-肾上腺轴及5-羟色胺信号系统增强痛泻要方的药效。在此基础上通过药代动力学及组织分布研究发现防风可增加肝、脾、脑和小肠中白术内酯I、芍药苷、橙皮苷的分布浓度，减少肺中白术内酯I和没食子酸的分布；通过计算组织与血浆中的药物浓度比发现，防风可显著提高白术内酯I、芍药苷、橙皮苷的脑/血浆比，降低白术内酯I、芍药苷的肺/血浆比和芍药苷的脾/血浆比，推测防风可能通过增加痛泻要方有效成分的脑分布发挥引经作用。采用大鼠原代脑微血管内皮细胞和星形胶质细胞联合培养建立体外血脑屏障模型，明确了防风及其成分香柑内酯可促进白术内酯Ⅰ、芍药苷和橙皮苷通过血脑屏障。进一步研究发现防风和香柑内酯可下调血脑屏障紧密连接Occludin、Claudin-5和ZO-1的mRNA及蛋白表达。药物转运实验显示P-gp、MRP1介导了芍药苷外排，橙皮苷则经P-gp、BCRP外排出血脑屏障，防风及香柑内酯可抑制P-gp、MRP1的功能及表达。以上表明防风可调控血脑屏障上紧密连接和转运体促进痛泻要方相关成分透过血脑屏障，进而通过脑肠互动增强痛泻要方的疗效。. 本项目以经典方剂痛泻要方为载体，基于脑肠互动和脑靶向分布探讨防风的脾经引经机制，阐明了防风引经作用的科学内涵，为中药引经理论研究提供了示范。