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Tissue Engineered Nerve Grafts Serve as a Living Scaffold to Facilitate Regeneration and Functional Recovery Following a 5 cm Nerve Lesion in Swine
D. Kacy Cullen, PhD1; Mindy I. Ezra, PhD1; Kevin D. Browne, BS1; John Dutton, BS1; Laura A. Struzyna, BSE1; Kritika S. Katiyar, BS1; Joseph P. Morand, BS1; John A. Wolf, PhD1; Harry C. Ledebur, PhD2 ; Douglas H. Smith, MD1
1Neurosurgery, University of Pennsylvania, Philadelphia, PA; 2Axonia Medical, Inc, Kalamazoo, MI
Surgical repair strategies following peripheral nerve injury (PNI) are currently inadequate, especially following major lesions involving the loss of segments several centimeters in length. We have developed tissue engineered nerve grafts (TENGs) that are lab-grown nervous tissue comprised of neurons spanned by long axonal tracts. Axon “stretch growth” - a natural axon growth mechanism that we replicate in custom mechanobioreactors - is used to rapidly generate long, aligned axonal tracts (5-10 cm in 14-21 days).
Three-dimensional TENGs are then created by embedding the living axonal tracts in an extracellular matrix. In rodent and porcine models of PNI, we previously found that TENGs serve as a “living scaffold” to accelerate and direct host axon regeneration and Schwann cell infiltration across short nerve gaps. In the current study, TENGs were used to bridge major 5 cm lesions in the deep peroneal nerve (~20 cm from distal muscle target) in comparison to the sural nerve autograft in minipigs. We found that TENG neurons/axons survived long-term (absent immunosuppression) and directly interacted with host axons and Schwann cells. In particular, we found that TENGs accelerated acute axonal regeneration across the 5 cm lesions based on direct axon-facilitated axon regeneration, demonstrating that this new mechanism of axon regeneration that was observed in 1 cm lesions was scaled to more challenging 5 cm lesions. Remarkably, this allowed a sub-population of host axons to cross the graft within 5 weeks (averaging 1.6 mm/day) - before host Schwann cells had infiltrated the graft. By 3 months following TENG repair, the bulk of host axons had crossed the graft and there was a reconstituted compound nerve action potential. Initial muscle reinnervation and evoked hoof twitch were achieved by 7 months following TENG repair, and at this time the repaired nerve contained a high density of myelinated host axons. Currently, the extent of functional recovery and regenerated nerve morphometry are being compared between TENGs and sural nerve autografts at 9-12 months post-repair.
These results demonstrate that our unique tissue engineering strategy accelerated regeneration based on axon-facilitated axon regeneration, which recapitulates an axonal pathfinding mechanism used during embryonic development and allowed for an initial bolus of axon regeneration to occur independent of host Schwann cells.
Overall, TENGs enabled rapid axon regeneration, target reinnervation and functional recovery when used to bridge a challenging 5.0 cm nerve lesion, demonstrating the efficacy of TENGs to facilitate nerve regeneration following major PNI.
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