ReProGlo: a new stem cell-based reporter assay aimed to predict embryotoxic potential of drugs and chemicals.
A stem cell-based reporter assay was developed to detect drug-induced alterations in the canonical Wnt/beta-catenin signaling pathway, which is involved in the regulation of early embryonic development. The so-called ReProGlo assay allows simultaneous determination of cell viability and luciferase reporter activity in a high throughput 96-well microtiter format. A clone of mouse embryonic stem (mES) cells stably expressing the SuperTopFlash reporter was established. This allows Wnt pathway activity determinations in undifferentiated mES cells and their differentiated descendants. Several test chemicals were analyzed in the new assay system. Known embryotoxicants like retinoic acid or lithium chloride induced concentration-dependent increases in reporter activity. The potency of valproic acid and a series of structural analogs to activate the Wnt pathway correlated well with their reported teratogenic activity in the mouse. Cyclophosphamide was also active but only after metabolic activation by hepatocytes. The new test may help to predict embryotoxic potential of chemicals.
Uibel
Frederik
F
Institute of Experimental and Clinical Pharmacology and Toxicology, Department of Toxicology, University of Tübingen, Tübingen, Germany.
Mühleisen
Annette
A
Köhle
Christoph
C
Weimer
Marc
M
Stummann
Tina C
TC
Bremer
Susanne
S
Schwarz
Michael
M
D000826:Animal Testing Alternatives; D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D018920:Coculture Techniques; D004305:Dose-Response Relationship, Drug; D053595:Embryonic Stem Cells / Q000187:drug effects* / Q000378:metabolism; D017930:Genes, Reporter; D022781:Hepatocytes / Q000378:metabolism; D049409:Luciferases, Firefly / Q000235:genetics; D051379:Mice; D015203:Reproducibility of Results; D012098:Reproduction / Q000187:drug effects*; D013723:Teratogens / Q000737:chemistry / Q000633:toxicity*; D018675:Toxicity Tests / Q000379:methods* / Q000592:standards; D014162:Transfection; D051153:Wnt Proteins / Q000235:genetics*; D051176:beta Catenin / Q000235:genetics*
D013723:Teratogens; D051153:Wnt Proteins; D051176:beta Catenin; D049409:Luciferases, Firefly