First steps in establishing a developmental toxicity test method based on human embryonic stem cells
First steps in establishing a developmental toxicity test method based on human embryonic stem cells
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DOI：
10.1016/j.tiv.2007.07.013
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发表时间：
2008-02-01
影响因子：
3.2
通讯作者：
Bremer, Susanne
中科院分区：
文献类型：
--
作者：
Adler, Sarah;Pellizzer, Cristian;Bremer, Susanne
胚胎干细胞的使用是目前评估体外发育毒性的最有前途的方法。此外，使用人类胚胎干细胞(HES)的可能性将增加消费者和患者的安全性，因为可以避免由于物种间差异而对物质进行错误分类。一种基于小鼠胚胎干细胞的验证测试-胚胎干细胞测试(EST)，包括以下终点：成纤维细胞和胚胎干细胞的IC50值以及抑制MES细胞向心肌细胞的分化。作为其成功验证研究的后续，我们建立了基于HES细胞和人成纤维细胞的细胞毒性试验，使用两种发育毒物：5-氟尿嘧啶(5-FU)和全反式维甲酸(RA)。将结果与EST的历史数据进行了比较。对5-FU，不同细胞系间无显著差异。然而，对于RA，两种测试系统对成纤维细胞的IC50值都高于对干细胞的IC50值，这是众所周知的发育毒物的影响。此外，还测试了几个标记基因作为可能的毒理学终点的可靠性和相关性。在OCT-4的早期分化过程中，hTERT和Dusp6的结果最为可靠。短臂和GATA-4被发现最适合于监测心脏分化。晚期心脏标记基因TNNT2直到第18天才表现出显著的结果。因此，这些标记基因最有可能作为发育毒性试验的终点。(C)2007爱思唯尔有限公司。保留所有权利。
The use of embryonic stem cells is currently the most promising approach to assess developmental toxicity in vitro. In addition, the possibility of using human embryonic stem (hES) cells will increase safety of consumers and patients as false classification of substances due to inter-species variations can be avoided.One validated test based on murine embryonic stem cells, the embryonic stem cell test (EST), consists of following endpoints: IC50 values of fibroblasts and embryonic stem cells as well as the inhibition of differentiation of mES cells into cardiomyocytes. As a follow up of its successful validation study we established a cytotoxicity assay based on hES cells and human fibroblasts employing two developmental toxicants: 5-fluorouracil (5-FU) and all-trans retinoic acid (RA). The results were compared to historical data from the EST. For 5-FU, no significant differences were obtained between the different cell lines. However, for RA, both test systems produced higher IC50 values for the fibroblasts than for the stem cells, which is a well-known effect of developmental toxicants. Moreover, the reliability and relevance of several marker genes as possible toxicological endpoints were tested. During early differentiation Oct-4, hTert and Dusp6 showed the most reliable results. Brachyury and GATA-4 were found to be best suited to monitor cardiac differentiation. The late cardiac marker gene TNNT2 demonstrated significant results until day 18. Therefore, these marker genes have the highest potential to serve as endpoints for a developmental toxicity test. (C) 2007 Elsevier Ltd. All rights reserved.