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Updated: Aug 29, 2026
Human Pluripotent Stem Cell Based Developmental Toxicity Assays for Chemical Safety Screening and Systems Biology Data Generation
Published on: June 17, 2015
Molecular multiple endpoint embryonic stem cell test--a possible approach to test for the teratogenic potential of
N I zur Nieden1, G Kempka, H J Ahr
1Research Toxicology, Bayer AG, Aprather Weg, D-42096 Wuppertal, Germany. nicole.zur.nieden@gmx.de
Abstract:
The embryonic stem cell test (EST) examines the cytotoxicity of chemical compounds on embryonic stem (ES) cells and 3T3.A31 fibroblasts. Additionally, the EST measures the ability of ES cells to differentiate into contracting cardiomyocytes following drug exposure. In this study, we introduce new endpoints to obtain a molecular multiple endpoint EST (mme-EST), enabling the identification of potential chemical effects on osteogenic, chondrogenic and neural differentiation in addition to the traditional endpoint of cardiomyocyte differentiation. Six compounds in three classes with known teratogenic in vivo potential were assayed with the mme-EST in a pilot study: penicillin G (non-teratogenic), 5-fluorouracil and retinoic acid (strongly teratogenic), diphenylhydantoin, valproic acid and thalidomide (moderately teratogenic). While the traditional EST measures a morphological endpoint, we included molecular markers of differentiation as endpoints. With the mme-EST, every compound could be classified correctly according to its known teratogenic potential in vivo. Penicillin G, 5-fluorouracil and diphenylhydantoin inhibited differentiation of all endpoints equally. Interestingly, valproic acid showed the strongest inhibition of neural differentiation, while thalidomide specifically inhibited osteogenic development. Retinoic acid, on the other hand, supported neural but inhibited chondrogenic and osteogenic differentiation concentration-dependently. Valproic acid and thalidomide, classified incorrectly with the established EST model, were classified correctly with the mme-EST according to their effects on specific endpoints. This pilot study indicates that the predictive value of the EST may be enhanced by including further differentiation endpoints.
Insights
The molecular multiple endpoint EST (mme-EST) improves teratogenic potential prediction by assessing chemical effects on multiple cell differentiation pathways. This enhanced assay correctly classified compounds, unlike the traditional EST.
Area of Science:
- Developmental toxicology
- Stem cell biology
- Chemical safety assessment
Background:
- The traditional embryonic stem cell test (EST) assesses chemical cytotoxicity and cardiomyocyte differentiation.
- Existing methods lack comprehensive evaluation of chemical impacts on diverse differentiation pathways.
- Accurate prediction of in vivo teratogenic potential remains a challenge.
Purpose of the Study:
- To introduce and validate a molecular multiple endpoint EST (mme-EST) for enhanced teratogenicity prediction.
- To assess the mme-EST's ability to identify chemical effects on osteogenic, chondrogenic, and neural differentiation.
- To compare mme-EST performance against the traditional EST using known teratogenic compounds.
Main Methods:
- Assayed six compounds with known in vivo teratogenic potential using the mme-EST.
- Included molecular markers for osteogenic, chondrogenic, neural, and cardiomyocyte differentiation.
- Compared mme-EST classifications with established teratogenic classifications.
Main Results:
- The mme-EST correctly classified all tested compounds according to their known in vivo teratogenic potential.
- Valproic acid and thalidomide, misclassified by the traditional EST, were accurately classified by the mme-EST.
- Specific differentiation pathways were differentially affected by compounds (e.g., valproic acid on neural, thalidomide on osteogenic).
Conclusions:
- The mme-EST significantly enhances the predictive value of the EST for chemical teratogenicity.
- Incorporating multiple molecular differentiation endpoints provides a more nuanced understanding of chemical toxicity.
- The mme-EST offers a more accurate and comprehensive tool for developmental toxicity screening.
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