Combination of multiple neural crest migration assays to identify environmental toxicants from a proof-of-concept chemical library

  • Johanna Nyffeler
  • Xenia Dolde
  • Alice Krebs
  • Kevin Pinto-Gil
  • Manuel Pastor
  • Mamta Behl
  • Tanja Waldmann
  • Marcel Leist
In vitro systems

Abstract

Many in vitro tests have been developed to screen for potential neurotoxicity. However, only few cell function-based tests have been used for comparative screening, and thus experience is scarce on how to confirm and evaluate screening hits. We addressed these questions for the neural crest cell migration test (cMINC). After an initial screen, a hit follow-up strategy was devised. A library of 75 compounds plus internal controls (NTP80-list), assembled by the National Toxicology Program of the USA (NTP) was used. It contained some known classes of (developmental) neurotoxic compounds. The primary screen yielded 23 confirmed hits, which comprised ten flame retardants, seven pesticides and six drug-like compounds. Comparison of concentration–response curves for migration and viability showed that all hits were specific. The extent to which migration was inhibited was 25–90%, and two organochlorine pesticides (DDT, heptachlor) were most efficient. In the second part of this study, (1) the cMINC assay was repeated under conditions that prevent proliferation; (2) a transwell migration assay was used as a different type of migration assay; (3) cells were traced to assess cell speed. Some toxicants had largely varying effects between assays, but each hit was confirmed in at least one additional test. This comparative study allows an estimate on how confidently the primary hits from a cell function-based screen can be considered as toxicants disturbing a key neurodevelopmental process. Testing of the NTP80-list in more assays will be highly interesting to assemble a test battery and to build prediction models for developmental toxicity.

Keywords

Cell migration Cell tracking Cytotoxicity High content imaging Developmental toxicity Human stem cells 

Abbreviations

AOP

Adverse outcome pathways

AraC

Cytosine arabinoside

cMINC

Circular MINC

CytoD

Cytochalasin D

DMSO

Dimethyl sulfoxide

DNT

Developmental neurotoxicity

EC

Effective concentration

EC90V

EC90 of viability

EC75M

EC75 of migration

EGCG

Epigallocatechin gallate

EGF

Epidermal growth factor

FBS

Foetal bovine serum

FGF

Fibroblast growth factor

FR

Flame retardant

GRIND2

Grid-independent descriptors

hESC

Human embryonic stem cell

logP

Octanol–water distribution coefficient

MeHgCl

Methylmercury(II) chloride

MINC

Migration of neural crest cell

MW

Molecular weight

NCC

Neural crest cell

NHBA

Number of hydrogen bond acceptors

NOAELV

No observed adverse effect level for viability

NRB

Number of rotatable bonds

NTP

National Toxicology Program

PAH

Polycyclic aromatic hydrocarbon

PBDE

Polybrominated diphenyl ether

PBS

Phosphate buffered saline

PCA

Principal component analysis

QSAR

Quantitative structure-activity relationship

ROI

Region of interest

TB-BPA

Tetrabromobisphenol A

TPSA

Polar surface

Supplementary material

204_2017_1977_MOESM1_ESM.xls (852 kb)
Supplementary material 1 (XLS 852 kb)
204_2017_1977_MOESM2_ESM.pdf (1.1 mb)
Supplementary material 2 (PDF 1135 kb)

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Copyright information

© Springer-Verlag Berlin Heidelberg 2017

Authors and Affiliations

  • Johanna Nyffeler
    • 1
    • 2
  • Xenia Dolde
    • 1
    • 3
  • Alice Krebs
    • 1
    • 3
  • Kevin Pinto-Gil
    • 4
  • Manuel Pastor
    • 4
  • Mamta Behl
    • 5
  • Tanja Waldmann
    • 1
  • Marcel Leist
    • 1
    • 2
    • 3
  1. 1.In Vitro Toxicology and Biomedicine, Department inaugurated by the Doerenkamp-Zbinden FoundationUniversity of KonstanzKonstanzGermany
  2. 2.Research Training Group RTG1331KonstanzGermany
  3. 3.Konstanz Research School Chemical Biology (KoRS-CB)KonstanzGermany
  4. 4.Research Programme on Biomedical Informatics (GRIB), Department of Experimental and Health SciencesUniversitat Pompeu FabraBarcelonaSpain
  5. 5.Division of National Toxicology ProgramNational Institute of Environmental Health SciencesResearch Triangle ParkUSA

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