An effective strategy for sustained neurotrophic factor (NTF) delivery to sites of peripheral nerve injury (PNI) would accelerate healing and enhance functional recovery, addressing the major clinical challenges associated with the current standard of care. In this study, scaffold-free cell sheets were generated using human dental pulp stem/progenitor cells, that endogenously express high levels of NTFs, for use as bioactive NTF delivery systems. Additionally, the effect of fibroblast growth factor 2 (FGF2) on NTF expression by dental pulp cell (DPC) sheets was evaluated. In vitro analysis confirmed that DPC sheets express high levels of NTF messenger RNA (mRNA) and proteins, and the addition of FGF2 to DPC sheet culture increased total NTF production by significantly increasing the cellularity of sheets. Furthermore, the DPC sheet secretome stimulated neurite formation and extension in cultured neuronal cells, and these functional effects were further enhanced when DPC sheets were cultured with FGF2. These neuritogenic results were reversed by NTF inhibition substantiating that DPC sheets have a positive effect on neuronal cell activity through the production of NTFs. Further evaluation of DPC sheets in a rat facial nerve crush injury model in vivo established that in comparison with untreated controls, nerves treated with DPC sheets had greater axon regeneration through the injury site and superior functional recovery as quantitatively assessed by compound muscle action potential measurements. This study demonstrates the use of DPC sheets as vehicles for NTF delivery that could augment the current methods for treating PNIs to accelerate regeneration and enhance the functional outcome.

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