The clinical use of pluripotent stem cell (PSC)-derived neural cells requires an efficient differentiation process for mass production in a bioreactor. Toward this goal, neural differentiation of murine embryonic stem cells (ESCs) in three-dimensional (3D) polyethylene terephthalate microfibrous matrices was investigated in this study. To streamline the process and provide a platform for process integration, the neural differentiation of ESCs was induced with astrocyte-conditioned medium without the formation of embryoid bodies, starting from undifferentiated ESC aggregates expanded in a suspension bioreactor. The 3D neural differentiation was able to generate a complex neural network in the matrices. When compared to 2D differentiation, 3D differentiation in microfibrous matrices resulted in a higher percentage of nestin-positive cells (68% vs. 54%) and upregulated gene expressions of nestin, Nurr1, and tyrosine hydroxylase. High purity of neural differentiation in 3D microfibrous matrix was also demonstrated in a spinner bioreactor with 74% nestin + cells. This study demonstrated the feasibility of a scalable process based on 3D differentiation in microfibrous matrices for the production of ESC-derived neural cells. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:1013–1022, 2013
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