Evaluation of a human neurite growth assay as specific screen for developmental neurotoxicants
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In vitro systems
Evaluation of a human neurite growth assay as specific screen for developmental neurotoxicants Anne K. Krug · Nina V. Balmer · Florian Matt · Felix Schönenberger · Dorit Merhof · Marcel Leist
Received: 14 February 2013 / Accepted: 2 May 2013 © Springer-Verlag Berlin Heidelberg 2013
Abstract Organ-specific in vitro toxicity assays are often highly sensitive, but they lack specificity. We evaluated here examples of assay features that can affect test specificity, and some general procedures are suggested on how positive hits in complex biological assays may be defined. Differentiating human LUHMES cells were used as potential model for developmental neurotoxicity testing. Forty candidate toxicants were screened, and several hits were obtained and confirmed. Although the cells had a definitive neuronal phenotype, the use of a general cell death endpoint in these cultures did not allow specific identification of neurotoxicants. As alternative approach, neurite growth was measured as an organ-specific functional endpoint. We found that neurite extension of developing LUHMES was specifically inhibited by diverse compounds such as colchicine, vincristine, narciclasine, rotenone, cycloheximide, or diquat. These compounds reduced neurite growth at concentrations that did not compromise cell viability, and neurite growth was affected more potently than the integrity of Electronic supplementary material The online version of this article (doi:10.1007/s00204-013-1072-y) contains supplementary material, which is available to authorized users. A. K. Krug (*) · N. V. Balmer · F. Matt · M. Leist Doerenkamp‑Zbinden Chair for In Vitro Toxicology and Biomedicine, University of Konstanz, Universitätsstr. 10, Box 657, 78457 Constance, Germany e-mail: anne.krug@uni‑konstanz.de F. Schönenberger · D. Merhof Interdisciplinary Center for Interactive Data Analysis, Modellingand Visual Exploration (INCIDE), University of Konstanz, Constance, Germany F. Schönenberger Bioimaging Center (BIC), University of Konstanz, Constance, Germany
developed neurites of mature neurons. A ratio of the EC50 values of neurite growth inhibition and cell death of >4 provided a robust classifier for compounds associated with a developmental neurotoxic hazard. Screening of unspecific toxicants in the test system always yielded ratios
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