Archives of Toxicology

, Volume 91, Issue 4, pp 2017–2028 | Cite as

Omnisphero: a high-content image analysis (HCA) approach for phenotypic developmental neurotoxicity (DNT) screenings of organoid neurosphere cultures in vitro

  • Martin R. Schmuck
  • Thomas Temme
  • Katharina Dach
  • Denise de Boer
  • Marta Barenys
  • Farina Bendt
  • Axel Mosig
  • Ellen Fritsche
Protocols

Abstract

Current developmental neurotoxicity (DNT) testing in animals faces major limitations, such as high cost and time demands as well as uncertainties in their methodology, evaluation and regulation. Therefore, the use of human-based 3D in vitro systems in combination with high-content image analysis (HCA) might contribute to DNT testing with lower costs, increased throughput and enhanced predictivity for human hazard identification. Human neural progenitor cells (hNPCs) grown as 3D neurospheres mimic basic processes of brain development including hNPC migration and differentiation and are therefore useful for DNT hazard identification. HCA of migrated neurospheres creates new challenges for automated evaluations because it encompasses variable cell densities, inconsistent z-layers and heterogeneous cell populations. We tackle those challenges with our Omnisphero software, which assesses multiple endpoints of the ‘Neurosphere Assay.’ For neuronal identification, Omnisphero reaches a true positive rate (TPR) of 83.8 % and a false discovery rate (FDR) of 11.4 %, thus being comparable to the interindividual difference among two researchers (TPR = 94.3, FDR = 11.0 %) and largely improving the results obtained by an existing HCA approach, whose TPR does not exceed 50 % at a FDR above 50 %. The high FDR of existing methods results in incorrect measurements of neuronal morphological features accompanied by an overestimation of compound effects. Omnisphero additionally includes novel algorithms to assess ‘neurosphere-specific’ endpoints like radial migration and neuronal density distribution within the migration area. Furthermore, a user-assisted parameter optimization procedure makes Omnisphero accessible to non-expert end users.

Keywords

Developmental neurotoxicity (DNT) Neurospheres High-content image analysis (HCA) Neuronal identification Neurite outgrowth Migration 

Supplementary material

204_2016_1852_MOESM1_ESM.pdf (1.7 mb)
Supplementary material 1 (PDF 1732 kb)

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Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Martin R. Schmuck
    • 1
  • Thomas Temme
    • 2
  • Katharina Dach
    • 1
  • Denise de Boer
    • 1
  • Marta Barenys
    • 1
  • Farina Bendt
    • 1
  • Axel Mosig
    • 2
  • Ellen Fritsche
    • 1
  1. 1.IUF - Leibniz Research Institute for Environmental MedicineDüsseldorfGermany
  2. 2.Department of Biophysics, ND04/596Ruhr-University BochumBochumGermany

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