Abstract
Recent progress in the field of human induced pluripotent stem cells (iPSCs) has led to the efficient production of human neuronal cell models for in vitro study. This has the potential to enable the understanding of live human cellular and network function which is otherwise not possible. However, a major challenge is the generation of reproducible neural networks together with the ability to interrogate and record at the single cell level. A promising aid is the use of biomaterial scaffolds that would enable the development and guidance of neuronal networks in physiologically relevant architectures and dimensionality. The optimal scaffold material would need to be precisely fabricated with submicron resolution, be optically transparent, and biocompatible. Two-photon polymerisation (2PP) enables precise microfabrication of three-dimensional structures. In this study, we report the identification of two biomaterials that support the growth and differentiation of human iPSC-derived neural progenitors into functional neuronal networks. Furthermore, these materials can be patterned to induce alignment of neuronal processes and enable the optical interrogation of individual cells. 2PP scaffolds with tailored topographies therefore provide an effective method of producing defined in vitro human neural networks for application in influencing neurite guidance and complex network activity.
Highlights
The last decade has seen the emergence of neuronal cultures derived from human induced pluripotent stem cells that has promise to be a technology that addresses many of the major issues currently facing the field
We identify two suitable materials for the fabrication of micropatterned surfaces using 2PP to guide human neuronal network development
Dental LT Clear (DClear), and a combination of PEG-DA and Irgacure 2959 were found to meet the required criteria of optical transparency with a low auto-fluorescence profile, amenability to 2PP at the micron scale and biocompatibility with cultured human neurons
Summary
Animal models have played a central role in elucidating key mechanisms of neurodevelopment and neurotransmission, but despite many attempts at “humanising” models by implementing disease-causing mutations from human studies, it is not always possible to replicate complex neurological disorders in animals.[4,5,6,7] To determine relevant functional human mechanisms and their disease related dysfunction it is becoming clear that patient-derived human cell models are required that allow functional interrogation of cell–cell interactions from single cells to the whole-network level.8–141792 | Lab Chip, 2020, 20, 1792–1806 PaperThe last decade has seen the emergence of neuronal cultures derived from human induced pluripotent stem cells (iPSCs) that has promise to be a technology that addresses many of the major issues currently facing the field. Developments with iPSC-derived neural cultures are advancing rapidly, a majority have so far focused on deriving two dimensional planar cultures featuring a single subtype of neural cells, and a limited number of functional endpoints.[15,16] In addition, a high variability of derived neuronal cell types and a lack of defined network architecture makes deriving conclusions from the analysis potentially problematic. Recent focus has moved to using organoids which exhibit an in vivo-like morphogenic development program that generates self-organising structures and a wide spectrum of neural cells.[17,18,19] Whilst organoids present an excellent emerging model for neurodevelopmental studies, robust cellular functional interrogation presents difficulties due to variable organoid size and structure.[20,21,22]
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