Grouping of polychlorinated biphenyls according to inhibition of neural crest cell migration

Abstract

Introduction: The neural crest arises during early fetal development at the borders of the neural plate. Neural crest cells (NCCs) migrate in a well-defined manner in different directions and give rise to peripheral neurons, melanocytes, cranial bones and other tissues. These processes can be disturbed by developmental toxicants. Aims: 1. Establish a medium throughput human-cell based system to test for interference of toxicants with NCC migration. 2. Apply this test system to test several polychlorinated biphenyls (PCBs). Methods: NCCs were differentiated from human embryonic stem cells. To generate a defined migration target area, stoppers were inserted in culture dishes before cells were seeded. After removal of the stoppers, cells migrated inwards. Migration and cell viability were quantified after 48 h by high-content image analysis. Results: Time-lapse microscopy revealed that the cells migrate preferentially during 24 to 48 h after stopper removal. Therefore, toxicants were added only during that time period. Inhibitors of actin and microtubule function (cytochalasin D, taxol) inhibited NCC migration in a concentration-dependent manner without compromising cell viability. General cytotoxic compounds (etoposide, Triton X-100) and proliferation inhibitors (aphidicolin, cytosine arabinoside) were used to define assay specificity. Environmentally relevant PCBs (i.e. PCB138, 153, 170 and 180) reduced NCC migration very effectively at non-cytotoxic concentrations. Different PCB congeners could be assigned to three classes based on their efficacy; di-ortho-substituted PCBs were more effective than mono-ortho-substituted ones. Non-ortho-substituted, coplanar, PCBs did not inhibit NCC migration at concentrations ≤ 10 mM. Follow-up experiments revealed that PCBs inhibit NCC migration via a mechanism independent of the aryl hydrocarbon receptor (AhR) and ryanodine receptor (RyR). Conclusions: The new human-cell based assay allows medium throughput screening and QSAR studies of environmental toxicants. Mechanisms related to PCB effects will be further explored. This research was supported by the graduate school Research Training Group (RTG) 1331 funded by the German Research Foundation (DFG).