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 85days. 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.

Role of Human-Induced Pluripotent Stem Cell-Derived Spinal Cord Astrocytes in the Functional Maturation of Motor Neurons in a Multielectrode Array System

“…Previous MEA studies have focused on hiPSC‐derived cortical neurons, either alone or in coculture with primary rodent cortical astrocytes and more recently with hiPSC‐derived astrocytes . Despite an established body of evidence suggesting that astrocytes contribute to neuronal electrophysiological maturation by promoting synaptogenesis , MEA‐ and hiPSC‐based paradigms, still influenced by a traditionally neuron‐centered perspective, have not yet been used to model this unique glial contribution.…”

Literature review and appraisal on alternative neurotoxicity testing methods

“…This is an interesting approach, especially with regards to brain region-specific differentiation. Others have used the concept of producing NPC as an intermediate cell population with a distinct proliferation capacity to generate reliable terminal differentiated neural tissue (neuron-glia and neuron subtype proportions) in manageable time intervals (Li et al, 2011;Hofrichter et al, 2017). More work is needed here to understand if such methods can be used for reproducible generation of brain region-specific neural cultures with human cells.…”

Application of Human‐Induced Pluripotent Stem Cells (hiPSCs) to Study Synaptopathy of Neurodevelopmental Disorders

“…To fully mimic the natural architecture within the human brain and develop a method to generate human neurons on a large scale for clinical research, a threedimensional (3D) neuronal culture was developed from NPC or rosette aggregates (Mariani et al, 2012;Hogberg et al, 2013;Gouder et al, 2015;Hookway et al, 2016;Chandrasekaran et al, 2017;Hofrichter et al, 2017;Chen et al, 2018b) under the support of 3D scaffolds (Edgar et al, 2017;Gu et al, 2017;Hoveizi et al, 2017;Ishikawa et al, 2017;Lei and Schaffer, 2013;Leong et al, 2016;K. Li et al, 2017;Lu et al, 2012;Medda et al, 2016;Pellett et al, 2015;Uemura et al, 2011;Zhang et al, 2016).…”