title: Embryotoxicity assessment of developmental neurotoxicants using a neuronal endpoint in the embryonic stem cell test.
abstract: The embryonic stem cell test (EST) is a validated in vitro embryotoxicity test; however, as the inhibition of cardiac differentiation alone is used as a differentiation endpoint in the EST, it may not be a useful test to screen embryotoxic chemicals that affect the differentiation of noncardiac tissues. Previously, methylmercury (MeHg), cadmium and arsenic compounds, which are heavy metals that induce developmental neurotoxicity in vivo, were misclassified as nonembryotoxic with the EST. The aim of this study was to improve the EST to correctly screen such developmental neurotoxicants. We developed a neuronal endpoint (Tuj-1 ID₅₀) using flow cytometry analysis of Tuj-1-positive cells to screen developmental neurotoxicants (MeHg, valproic acid, sodium arsenate and sodium arsenite) correctly using an adherent monoculture differentiation method. Using Tuj-1 ID₅₀ in the EST instead of cardiac ID₅₀, all of the tested chemicals were classified as embryotoxic, while the negative controls were correctly classified as nonembryotoxic. To support the validity of Tuj-1 ID₅₀) , we compared the results from two experimenters who independently tested MeHg using our modified EST. An additional neuronal endpoint (MAP2 ID₅₀), obtained by analyzing the relative quantity of MAP2 mRNA, was used to classify the same chemicals. There were no significant differences in the three endpoint values of the two experimenters or in the classification results, except for isoniazid. In conclusion, our results indicate that Tuj-1 ID₅₀ can be used as a surrogate endpoint of the traditional EST to screen developmental neurotoxicants correctly and it can also be applied to other chemicals.
authors: Center for Drug Development Assistance, National Institute of Food and Drug Safety Evaluation, Korea Food and Drug Administration, Cheongwon-gun, Chungcheongbuk-do 363-951, South Korea.
authors: Dae Hyun
authors: DH
authors: Baek
authors: Tae Gyun
authors: TG
authors: Kim
authors: Hwa Kyung
authors: HK
authors: Lim
authors: Jin Wook
authors: JW
authors: Kang
authors: Su Kyoung
authors: SK
authors: Seong
authors: Seung Eun
authors: SE
authors: Choi
authors: So Yun
authors: SY
authors: Lim
authors: Sung Hee
authors: SH
authors: Park
authors: Bong-hyun
authors: BH
authors: Nam
authors: Eun Hee
authors: EH
authors: Kim
authors: Mun Sin
authors: MS
authors: Kim
authors: Kui Lea
authors: KL
authors: Park
mesh_terms: D000818:Animals; D001149:Arsenates / Q000633:toxicity; D018053:Arsenites / Q000633:toxicity; D041702:BALB 3T3 Cells; D002454:Cell Differentiation; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D053595:Embryonic Stem Cells / Q000187:drug effects*; D023381:Endpoint Determination; D007928:Lethal Dose 50; D008767:Methylmercury Compounds / Q000633:toxicity; D051379:Mice; D009474:Neurons / Q000187:drug effects*; D015203:Reproducibility of Results; D017670:Sodium Compounds / Q000633:toxicity; D018675:Toxicity Tests / Q000379:methods*; D014635:Valproic Acid / Q000633:toxicity
chemical_list: D001149:Arsenates; D018053:Arsenites; D008767:Methylmercury Compounds; D017670:Sodium Compounds; C017947:sodium arsenite; D014635:Valproic Acid; C009277:sodium arsenate