AccScience Publishing / IJB / Online First / DOI: 10.36922/ijb.5973
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RESEARCH ARTICLE

Differentiation of iPSC-derived neural progenitors into motor neurons in 3D-printed bioscaffolds

Yilin Han1 ,  Marianne King2 ,  Hege Brincker Fjerdingstad3 ,  Maurizio Gullo4 ,  Lukas Zeger1 ,  Roland Kádár5 ,  Patrik Ivert1 ,  Joel C. Glover3,6 ,  Laura Ferraiuolo2 ,  Mimoun Azzouz2 ,  Elena N. Kozlova1
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1 Department of Immunology, Genetics and Pathology, Faculty of Medicine, Uppsala University, Uppsala, Sweden
2 Department of Neuroscience, University of Sheffield, Sheffield, United Kingdom
3 Norwegian Center for Stem Cell Research, Department of Immunology and Transfusion Medicine, Oslo University Hospital, Oslo, Norway
4 Institute for Medical Engineering & Medical Informatics, University of Applied Sciences and Arts Northwestern Switzerland, Muttenz, Switzerland
5 Department of Industrial and Materials Science, Chalmers University of Technology, Gothenburg, Sweden
6 Laboratory of Neural Development and Optical Recording (NDEVOR), Department of Molecular Medicine, Institute of Basic Medical Sciences, University of Oslo, Oslo, Norway
IJB, 5973 https://doi.org/10.36922/ijb.5973
Received: 14 November 2024 | Accepted: 27 February 2025 | Published online: 27 February 2025
© 2025 by the Author(s). This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution 4.0 International License ( https://creativecommons.org/licenses/by/4.0/ )
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Abstract

Translational medicine for neurodegenerative diseases can advance through the use of in vitro models incorporating human neural cells derived from patient-specific induced pluripotent stem cells (iPSCs). Previously, we investigated whether motor neuron (MN) progenitors derived from human iPSCs could differentiate from MNs within three-dimensional (3D)-printed scaffolds. While extensive neurite arborization was observed on the scaffold surface, no neurite outgrowth occurred within the scaffold interior. Here we showed the extensive growth of the neurites from iPSC-derived neural progenitors, imbedded into the gelatin scaffolds during 30 days of experimental time. We presented a bioink formulation that softens the scaffold while preserving its 3D structure, thereby facilitating neurite outgrowth throughout the scaffold. MN differentiation, evidenced by extensive neurite arborization and the expression of choline acetyltransferase (ChAT), was verified in 3D images deep within the scaffold structure. Notably, the degree of MN differentiation appeared to depend on two factors as follows: the delivery of MN differentiation factors via mesoporous silica particles (MSPs) embedded in the bioink and the method used to generate MN progenitors prior to 3D printing. In this paper, we provide a detailed protocol for 3D-printing human iPSC-derived MN progenitors, enabling their differentiation and survival within gelatin scaffolds. This protocol could be expanded to incorporate additional cell types, allowing the creation of more complex and standardized 3D neural tissues. Such advancements could facilitate investigations into the pathophysiology of motor neuron diseases and the development of new therapeutic strategies.

Graphical abstract
Keywords
3D bioscaffold
Differentiation
Gelatin
Induced pluripotent stem cells
Mesoporous silica
Motor neuron
Funding
E.K. was supported by the Swedish Research Council (Dnr 2019-02240) to E.K within the frame of EU Joint Programme of Neurodegenerative Diseases, and by the Swedish Natonal Space Agency (Dnr 2021-0005, Dnr 2024-00211). M.A., L.F., and M.K. were funded by MRC grant#MR/V000470/1.
Conflict of interest
The authors declare no conflict of interest.
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International Journal of Bioprinting, Electronic ISSN: 2424-8002 Print ISSN: 2424-7723, Published by AccScience Publishing
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