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Updated: May 28, 2026
Profiling Individual Human Embryonic Stem Cells by Quantitative RT-PCR
Published on: May 29, 2014
Transcriptomic concentration-response evaluation of valproic acid, cyproconazole, and hexaconazole in the neural
Peter T Theunissen1, Joshua F Robinson, Jeroen L A Pennings
1Laboratory for Health Protection Research, National Institute for Public Health and the Environment (RIVM), 3720 BA Bilthoven, The Netherlands. peter.theunissen@rivm.nl
Abstract:
Alternative developmental toxicity assays are urgently needed to reduce animal use in regulatory developmental toxicology. We previously designed an in vitro murine neural embryonic stem cell test (ESTn) as a model for neurodevelopmental toxicity testing (Theunissen et al., 2010). Toxicogenomic approaches have been suggested for incorporation into the ESTn to further increase predictivity and to provide mechanistic insights. Therefore, in this study, using a transcriptomic approach, we investigated the concentration-dependent effects of three known (neuro) developmental toxicants, two triazoles, cyproconazole (CYP) and hexaconazole (HEX), and the anticonvulsant valproic acid (VPA). Compound effects on gene expression during neural differentiation and corresponding regulated gene ontology (GO) terms were identified after 24 h of exposure in relation to morphological changes on day 11 of culture. Concentration-dependent responses on individual gene expression and on biological processes were determined for each compound, providing information on mechanism and concentration-response characteristics. All compounds caused enrichment of the embryonic development process. CYP and VPA but not HEX significantly enriched the neuron development process. Furthermore, specific responses for triazole compounds and VPA were observed within the GO-term sterol metabolic process. The incorporation of transcriptomics in the ESTn was shown to enable detection of effects, which precede morphological changes and provide a more sensitive measure of concentration-dependent effects as compared with classical morphological assessments. Furthermore, mechanistic insight can be instrumental in the extrapolation of effects in the ESTn to human hazard assessment.
Insights
This study enhances the embryonic stem cell test (ESTn) for developmental toxicity by incorporating transcriptomics. This approach provides earlier and more sensitive detection of chemical effects on neural development than traditional methods.
Area of Science:
- Developmental Toxicology
- Stem Cell Assays
- Toxicogenomics
Background:
- Regulatory developmental toxicology urgently requires alternatives to animal testing.
- The in vitro murine neural embryonic stem cell test (ESTn) was previously developed for neurodevelopmental toxicity.
- Integrating toxicogenomic approaches can improve ESTn predictivity and mechanistic understanding.
Purpose of the Study:
- To investigate the concentration-dependent effects of known developmental toxicants on gene expression during neural differentiation using transcriptomics.
- To assess the utility of incorporating transcriptomics into the ESTn for enhanced neurodevelopmental toxicity testing.
- To gain mechanistic insights into the effects of cyproconazole, hexaconazole, and valproic acid.
Main Methods:
- Utilized a transcriptomic approach to analyze gene expression changes in murine neural embryonic stem cells after exposure to cyproconazole, hexaconazole, and valproic acid.
- Assessed concentration-dependent effects on gene expression and gene ontology (GO) terms after 24 hours of exposure.
- Correlated transcriptomic findings with morphological changes observed on day 11 of culture.
Main Results:
- All tested compounds (cyproconazole, hexaconazole, valproic acid) enriched the embryonic development process.
- Cyproconazole and valproic acid significantly enriched the neuron development process, while hexaconazole did not.
- Triazole compounds and valproic acid showed specific responses within the sterol metabolic process GO term.
- Transcriptomics detected effects preceding morphological changes, offering a more sensitive measure of concentration-dependent toxicity compared to morphological assessments.
Conclusions:
- Incorporating transcriptomics into the ESTn enhances its sensitivity and provides mechanistic insights for developmental toxicity assessment.
- This approach allows for earlier detection of chemical-induced effects on neural development.
- Mechanistic data from transcriptomics can aid in extrapolating in vitro findings to human hazard assessment.
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