Hydrogel-Based Bioprocess for Scalable Manufacturing of Human Pluripotent Stem Cell-Derived Neural Stem Cells

Neural stem cells derived from human pluripotent stem cells (hPSC-NSCs) are of great value for modeling diseases, developing drugs, and treating neurological disorders. However, manufacturing high-quantity and -quality hPSC-NSCs, especially for clinical applications, remains a challenge. Here, we report a chemically defined, high-yield, and scalable bioprocess for manufacturing hPSC-NSCs. hPSCs are expanded and differentiated into NSCs in microscale tubes made with alginate hydrogels. The tubes are used to isolate cells from the hydrodynamic stresses in the culture vessel and limit the radial diameter of the cell mass to less than 400 μm to ensure efficient mass transport during the culture. The hydrogel tubes provide uniform, reproducible, and cell-friendly microspaces and microenvironments for cells. With this new technology, we showed that hPSC-NSCs could be produced in 12 days with high viability (∼95%), high purity (>90%), and high yield (∼5 × 10⁸ cells/mL of microspace). The volumetric yield is about 250 times more than the current state-of-the-art. Whole transcriptome analysis and quantitative real-time polymerase chain reaction showed that hPSC-NSCs made by this process had a similar gene expression to hPSC-NSCs made by the conventional culture technology. The produced hPSC-NSCs could mature into both neurons and glial cells in vitro and in vivo. The process developed in this paper can be used to produce large numbers of hPSC-NSCs for various biomedical applications in the future.