基于"有故无殒"理论的莪术神经发育毒性评价模式的研究

The theory of YOU-GU-WU-YUN, which is criterion about safety of medications during pregnancy, originated from Huangdi Neijing. The present research on toxicity of traditional Chinese medicine based on normal animal have restricted rational dose of traditional Chinese medicine in clinical practice, and security of traditional Chinese medicine also is questioned in the world. Clinical practice of traditional Chinese medicine demonstrated that drastic traditional Chinese medicine applied in pregnant women with indication exhibit therapeutic effect, and not appear developmental neurotoxicity. Therefore, we propose the working hypothesis that blood stasis syndrome is foundation for different expression of developmental neurotoxicity of removing blood stasis and resolving static blood Chinese medicine and its therapentic effect. To test this hypothesis, this project use animal model of blood stasis syndrome as the study object, and Rhizoma Curcumae, which be forbidden to take for pregnant women, as tool traditional Chinese medicine, and the modern scientific method such as toxicology, pharmacodynamics, pharmacokinetics and molecular biology will be used in this project. Developmental neurotoxicity of Rhizoma Curcumae will be evaluated through the observation on the effect of Rhizoma Curcumae on the physiology and reproduction of pregnant rats and neural development of fetal rats, and further to elucidate the underlying molecular mechanisms. This project will evidence the theory of toxicity-attenuating based on syndrome differentiation of traditional Chinese medicine, and has the important theory and the practical significance for establishing research system of developmental neurotoxicity of traditional Chinese medicine.

"有故无殒"理论源自于《黄帝内经》，是中医对妊娠期用药安全的权衡准则。目前基于正常动物的中药毒性研究限制了临床用药的合理剂量，也使中药安全性在国际上受到质疑。中医临床实践表明，峻烈药物使用于患有适应证候的妊娠妇女，表现出治疗作用而并未出现神经发育毒性。因此，我们提出"血瘀证是破血化瘀类中药神经发育毒性与母体治疗作用选择性表达的基础"这一科学假说。为了验证该假说，本项目采用毒理学、药效学、药物动力学、分子生物学等现代科学研究方法和手段，以血瘀证大鼠模型为研究对象，选用适用于此证的孕妇禁服中药莪术为工具药，通过观察其对妊娠大鼠生理生殖状态及仔鼠神经发育的影响，对莪术神经发育毒性进行整体评价并探讨其分子机理。科学地佐证"有故无殒"的中药辩证减毒理论，对构建符合中医理论和临床实际的中药神经发育毒性研究体系具有重要的理论和现实意义。

“有故无殒”理论源自于《黄帝内经》，是中医对妊娠期用药安全的权衡准则，目前基于正常动物的中药毒性研究限制了中药的临床用药。本资助项目以正常孕大鼠和血瘀证孕大鼠为研究对象，探讨了血瘀证在莪术神经发育毒性中的作用及其机制，本项目按计划完成了莪术活血化瘀作用、生殖毒性、神经发育毒性、莪术体内代谢等研究内容。我们采用肾上腺素联合冰浴成功的复制大鼠血瘀证模型，莪术灌胃对正常大鼠和血瘀证模型孕大鼠的体重、摄食量和窝均产仔数等指标的影响具有显著性差异，对正常大鼠的生殖发育毒性更明显。妊娠期莪术灌胃后，正常大鼠和血瘀证孕鼠的仔鼠体重、肛殖距、负趋地性、断崖回避和学习记忆等指标具有显著性差异，莪术对正常大鼠的生殖发育和神经行为发育的毒性更明显。莪术具有降低血瘀证模型大鼠血液流变学、血栓素B2、6-酮-前列腺素F1α、可溶性血管细胞粘附分子、可溶性细胞间粘附分子-1和血小板聚集率至正常孕大鼠水平的作用，而对正常大鼠这些指标的抑制作用不明显。妊娠期莪术灌胃后，正常孕大鼠子代大脑神经细胞粘附分子（neural cell adhesion molecule, NCAM）、α2,8-多聚唾液酸转移酶（α2, 8-sialyl transferase, ST8Sia）-II、ST8Sia -IV、Fyn和粘着斑激酶（focal adhesion kinase, FAK）蛋白和mRNA表达显著降低，而血瘀证大鼠子代大脑NCAM、ST8Sia II、ST8Sia IV、Fyn和FAK蛋白和mRNA水平较正常对照组子代大鼠无显著性降低。抑制子代大鼠脑组织粘附分子表达可能是莪术子代神经发育毒性的机制之一。莪术灌胃后，在正常大鼠和血瘀证大鼠脑组织和血液中的莪术醇浓度有显著性差异，在正常大鼠脑组织中莪术醇含量显著增加。莪术醇是莪术神经发育的毒性成分，通过减少神经元活性、抑制神经突触生长和神经元氧化应激而诱导神经发育毒性，NCAM-Fyn-FAK信号通路可能涉及莪术醇介导的神经突触生长抑制。本资助项目构建符合中医理论和临床实际的中药神经发育毒性研究体系，佐证了"有故无殒"的中药辩证减毒理论，有助于提高中药治疗水平及扩大应用范围。本项目研究结果发表论文12篇，其中SCI收录论文4篇。