Scaffold-Free Nerve Conduits Engineered using Dental Pulp Stem Cells

Current facial nerve treatments result in poor clinical outcomes due to slow and inefficient axon regeneration and aberrant reinnervation. Bioactive scaffold-free nerve conduits engineered using neurotrophic dental pulp stem/progenitor cells (DPSCs) and their aligned extracellular matrix (ECM) offer a promising alternative that addresses these clinical challenges. DPSCs intrinsically express high levels of neurotrophic factors, growth factors that induce axon regeneration, and an aligned ECM supplies guidance cues to direct extending axons. To form such conduits, human DPSCs were cultured on a substrate with linear microgrooves, stimulating the cells to align and deposit a linearly oriented ECM. The subsequent aligned DPSC sheets contained levels of neurotrophic factors previously established sufficient to induce axon regeneration and stimulated oriented neurite outgrowth when directly co-cultured with neuronal cells. Moreover, when DPSCs in aligned cell sheets were differentiated into glial Schwann cells, chemical induction cues from the differentiation media and mechanotransductive cues from the oriented ECM synergistically enhanced differentiation. The resulting DPSC-derived glial cells resembled repair Schwann cells, secreting higher levels of neurotrophic factors and remodeling their ECM to be more conducive to nerve regeneration, and promoted neuritogenesis similar to the undifferentiated DPSC sheets. Furthermore, scaffold-free nerve conduits were engineered by rolling aligned DPSC sheets into robust cylindrical tissues that maintained their organized ECM and NTF expression. These aligned conduits were used to bridge critical sized segmental nerve defects in the buccal branch of rat facial nerves. After 12 weeks, treatment with the DPSC conduits induced regeneration of myelinated axons and functional recovery similar or better than treatment with the clinical standard of care. Therefore, scaffold-free DPSC nerve conduits are capable of promoting axon regeneration, through a continuous supply of neurotrophic factors, and directing axon extension, through an endogenous aligned ECM, offering a promising therapy for facial nerve repair