Encapsulated Neural Stem Cell Neuronal Differentiation in Fluorinated Methacrylamide Chitosan Hydrogels


Neural stem/progenitor cells (NSPCs) are able to differentiate into the primary cell types (neurons, oligodendrocytes and astrocytes) of the adult nervous system. This attractive property of NSPCs offers a potential solution for neural regeneration. 3D implantable scaffolds should mimic the microstructure and dynamic properties found in vivo, enabling the natural exchange of oxygen, nutrients, and growth factors for cell survival and differentiation. We have previously reported a new class of materials consisting of perfluorocarbons (PFCs) conjugated to methacrylamide chitosan (MAC), which possess the ability to repeatedly take-up and release oxygen at beneficial levels for favorable cell metabolism and proliferation. In this study, the neuronal differentiation responses of NSPCs to fluorinated methacrylamide chitosan (MACF) hydrogels were studied for 8 days. Two treatments, with oxygen reloading or without oxygen reloading, were performed during culture. Oxygen concentration distributions within cell-seeded MACF hydrogels were found to have higher concentrations of oxygen at the edge of the hydrogels and less severe drops in O2 gradient as compared with MAC hydrogel controls. Total cell number was enhanced in MACF hydrogels as the number of conjugated fluorines via PFC substitution increased. Additionally, all MACF hydrogels supported significantly more cells than MAC controls (p < 0.001). At day 8, MACF hydrogels displayed significantly greater neuronal differentiation than MAC controls (p = 0.001), and among MACF groups methacrylamide chitosan with 15 fluorines per addition (MAC(Ali15)F) demonstrated the best ability to promote NSPC differentiation.

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We are grateful for funding from the University of Akron that supported this work. The authors would also like to thank Dr. Rebecca Willits for allowing us to perform rheological measurements in her laboratory as well as assistance with interpreting mechanical and swelling results.

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Correspondence to Nic D. Leipzig.

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Associate Editor Michael S. Detamore oversaw the review of this article.

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Supplementary Fig. S1 Representative images of scaffolds with no oxygen resupplementation obtained by confocal microscope at day 8 at the center of scaffolds, each picture represents a stack of 50 images with a spacing of 200 μm. MAC(Ali15)F shows the highest cell number indicated by nuclear Hoechst 33342 stain. Corresponding 3D reconstruction movies generated from these images are presented as supplemental material (Movies 5–8). Scale bar equals 100 μm (PDF 736 kb)

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Li, H., Wijekoon, A. & Leipzig, N.D. Encapsulated Neural Stem Cell Neuronal Differentiation in Fluorinated Methacrylamide Chitosan Hydrogels. Ann Biomed Eng 42, 1456–1469 (2014) doi:10.1007/s10439-013-0925-0

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  • Chitosan
  • Oxygen
  • Hydrogel
  • Perfluorocarbons
  • Neural stem cells
  • Neuronal differentiation