Comparative performance analysis of human iPSC-derived and primary neural progenitor cells (NPC) grown as neurospheres in vitro

Abstract

Developmental neurotoxicity (DNT) testing performed in rats is resource-intensive (costs, time, animals) and bears the issue of species extrapolation. Thus, reliable alternative human-based approaches are needed for predicting neurodevelopmental toxicity. Human induced pluripotent stem cells (hiPSCs) represent a basis for an alternative method possibly being part of an alternative DNT testing strategy. Here, we compared two hiPSC neural induction protocols resulting in 3D neurospheres: one using noggin and one cultivating cells in neural induction medium (NIM protocol). Performance of Nestin + /SOX2 + hiPSC-derived neural progenitor cells (NPCs) was compared to primary human NPCs. Generally, primary hNPCs first differentiate into Nestin + and/or GFAP + radial glia-like cells, while the hiPSC-derived NPCs (hiPSC-NPC) first differentiate into βIII-Tubulin + neurons suggesting an earlier developmental stage of hiPSC-NPC. In the ‘Neurosphere Assay’, NIM generated hiPSC-NPC produced neurons with higher performance than with the noggin protocol. After long-term differentiation, hiPSC-NPC form neuronal networks, which become electrically active on microelectrode arrays after 85 days. Finally, methylmercury chloride inhibits hiPSC-NPC and hNPC migration with similar potencies. hiPSC-NPCs-derived neurospheres seem to be useful for DNT evaluation representing early neural development in vitro . More system characterization by compound testing is needed to gain higher confidence in this method. • Establishment of the ‘Neurosphere Assay’ with human iPSC-derived neurospheres • Comparison of hiPSC-derived neurospheres with primary fetal neurospheres • iPSC-derived neurospheres show electrical activity on microelectrode arrays (MEAs). • Adverse effects of the developmentally neurotoxic compound MeHgCl on cell migration