RESUMO
The application of alternative methods in developmental and reproductive toxicology is challenging in view of the complexity of mechanisms involved. A battery of complementary test systems may provide a better prediction of developmental and reproductive toxicity than single assays. We tested twelve compounds with varying mechanisms of toxic action in an assay battery including 24 CALUX transcriptional activation assays, mouse cardiac embryonic stem cell test, ReProGlo assay, zebrafish embryotoxicity assay, and two CYP17 and two CYP19 activity assays. The battery correctly detected 11/12 compounds tested, with one false negative occurring, which could be explained by the absence of the specific mechanism of action of this compound in the battery. Toxicokinetic modeling revealed that toxic concentrations were in the range expected from in vivo reproductive toxicity data. This study illustrates added value of combining assays that contain complementary biological processes and mechanisms, increasing predictive value of the battery over individual assays.
Assuntos
Alternativas aos Testes com Animais , Teratogênicos/toxicidade , Testes de Toxicidade/métodos , Animais , Aromatase/metabolismo , Bioensaio , Linhagem Celular , Células Cultivadas , Embrião não Mamífero/efeitos dos fármacos , Células-Tronco Embrionárias/efeitos dos fármacos , Humanos , Camundongos , Ratos , Receptores de Esteroides/metabolismo , Reprodutibilidade dos Testes , Reprodução , Esteroide 17-alfa-Hidroxilase/metabolismo , Peixe-ZebraRESUMO
Alternative assays are highly desirable to reduce the extensive experimental animal use in developmental toxicity testing. In the present study, we developed an improved test system for assessing neurodevelopmental toxicity using differentiating embryonic stem cells. We advanced previously established methods by merging, modifying and abbreviating the original 20-day protocol into a more efficient 13-day neural differentiation protocol. Using morphological observation, immunocytochemistry, gene expression and flow cytometry, it was shown predominantly multiple lineages of neuroectodermal cells were formed in our protocol and to a lower extent, endodermal and mesodermal differentiated cell types. This abbreviated protocol should lead to an advanced screening method using morphology in combination with selected differentiation markers aimed at predicting neurodevelopmental toxicity. Finally, the assay was shown to express differential sensitivity to a model developmental neurotoxicant, methyl mercury.
