应用胚胎干细胞试验(EST)检测妊娠期常用中药的胚胎安全性

Chinese Herbal Medicines (CHMs) have been widely used to relieve many symptoms and to treat complications during pregnancy. Systematic evaluation on the embryotoxicity is still lacking. Until now, there is no universal accepted evidence on how safe the CHMs are being used during pregnancy and if there are any adverse effects of the medicines to embryo-fetal development. There are more than 300 CHMs currently used for pregnancy in clinical practice. Our embryotoxicity laboratory has screened the embryotoxic potentials of 20 most frequently used CHMs in pregnant mice. Although the reproductive toxicity to both mothers and fetuses identified, in vivo test methods are very expensive, laborious and time consuming to determine the safety of all CHMs . Embryonic Stem cell Test (EST), as an alternative in vitro embryotoxicity test method has been well established and widely validated for chemicals, natural supplements and pharmaceuticals, but not yet for CHMs. In collaboration with Federal Institute for Risk Assessment in Germany, the founder of EST, we has primarily evaluated and validated the usefulness of EST in assessing the classification of CHMs for developmental toxicity. In this study, 130 well recorded CHMs which have been currently used during pregnancy will be screened for the embryotoxicity. We will determine the embryotoxic potentials of the study CHMs by the established EST protocol, comparing the biological and molecular endpoint measures obtained from the concentration response curves in cytotoxicity and differentiation. Secondly we will define the precise classification of embryotoxic potentials of all test CHMs based on the prediction models developed by linear analysis of discriminance. There is an urgent need in testing the safety of CHMs for local and international health care service and also commercial marketing. This study will provide rapid and reproducible alternative approach for large-scale validating and screening programme for CHMs. Our study will provide the scientific evidence on the safety of CHMs and promote the application and spread in western countries.

中药常用于治疗妊娠疾病及并发症，然而中药对妊娠期母体及胚胎胎儿发育的安全性，仍然缺乏系统和科学的评估。我们通过在体动物实验已初步证实部分妊娠期常用中药的潜在胚胎毒性。胚胎干细胞试验（EST），作为一种快速和精确的体外替代实验方法，已在科学界广泛应用于检验化学物质和药品的胚胎毒性。与创建EST的德国研究所的前期合作已证实EST可以用于评估中药的胚胎安全性和分类。基于此，本研究以临床上常用于妊娠期的130种中药为对象，通过比较添加单味中药的培养液与正常培养液培养得到的胚胎干细胞，分析其细胞毒性和分化的浓度毒性曲线，从而测定该种中药的胚胎安全性和毒性分类。该研究将完成对妊娠期常用中药的大规模筛查，在实验方法学上扩大EST应用范围、补充数据库中天然药物的信息，在临床应用上为中药的安全性提供科学的证明，并促进其在国际市场上的传播和发展。

研究开展时间表简述：第一年主要是进行中草药质控和检测，学习德国合作方的EST技术，掌握技术难点和关键点。第二年主要是在本课题组胚胎毒理实验室建模及初步开展部分中草药的胚胎安全评估。第三年主要是进一步进行中草药的胚胎安全评估及实现实验的重复性。.研究成果总结:.1）所有待测中药的进一步筛选.2）可进行胚胎安全性评估的中药共60种.无胚胎毒性的中药：42种 （与在体动物实验安全评估结果98%吻合）.有胚胎毒性的中药：18种 （与在体动物实验安全评估结果100%吻合）.弱胚胎毒性的中药：15种 （与在体动物实验安全评估结果87%吻合）.强胚胎毒性的中药：3种 （与在体动物实验安全评估结果100%吻合）.3）初步结论：EST技术应用于中药的胚胎安全评估可以准确地判断胚胎毒性的有和无，但在弱毒和强毒的分类上仍需更多数据研究以支持其准确度和精确度。这个结论为中药的安全方面的实验研究提供新方法、为其临床应用提供科学依据。同时项目的研究结果亦验证EST方法同样适用于中药的胚胎安全性筛查，并为EST 数据库增添中药的信息，从而补充和完善了EST 的应用方向和前景。.i）完成全部待测中草药的安全评估报告，大多数的胚胎毒性EST测试结果与文献中的in vivo报告一致；.ii）于6次国际性会议上发表研究进展与部分相关成果；.iii）已与德国合作方就本课题项目的实验结果开展讨论、分析数据并已经完成后续科研项目的计划书、尚待修改完善并递交2017年国自然项目申请。.iv）已成功建立新的合作伙伴，计划在本课题基础上应用新的技术方法检测中草药的胚胎安全性，希望为中草药在妊娠期疾病的应用领域增添新技术新方向。.v）发表:7篇会议论文被收录于国际性会议，由课题组负责人和相关成员作演讲及海报报告。4本专业书籍的章节已发表于NOVA和InTech，6篇学术论文已发表于SCI期刊，另有2篇学术论文已递交SCI期刊、等待接收和修改意见。.vi）人才培养：本课题负责人在本课题基础上申报新的科研项目，并获得2016年钱江人才D类计划择优资助，并作为”中药安全性评价学术带头人和重点培养人才”全职引进新的高校工作。课题负责人亦获得数个学术职务。本课题组及合作方、双方共同培养一名博士后已出站，并分别培养硕士生2名、已毕业，博士生1名、已毕业，和博士生1名、在读。