Plastic Surgery Research Council
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PSRC 60th Annual Meeting
Program and Abstracts

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Notch Signaling In Senescent Schwann Cells: A Target For Improving Nerve Regeneration
Gwendolyn M. Hoben, MD, PhD, Xueping Ee, MD, Lauren Prange, BS, Daniel Hunter, RA, Amy M. Moore, MD, Sheila Stewart, PhD, Susan E. Mackinnon, MD, Matthew D. Wood, PhD.
Washinton University School of Medicine, St.Louis, MO, USA.

PURPOSE: Nerve gaps repaired with long nerve grafts (autografts or allografts) are associated with poor axonal regeneration and the accumulation of senescent Schwann cells (SenSCs). Senescent cells are characterized by altered gene and protein expression, and we recently found that SenSCs upregulate expression of Notch ligands (Jag1 and Dll4). Based on these results, we hypothesize that increased Notch signaling due to SenSCs reduces axonal regeneration and inhibition of Notch signaling will increase axonal outgrowth.

METHODS: Schwann cells (SCs) derived from rat sciatic nerves were cultured and treated with aphidicolin (damages DNA) to induce senescence. In vivo, cultured SenSCs or normal SCs (control) were implanted in a rat sciatic nerve transection using a 5mm conduit. After 4 weeks, axonal regeneration inside and across the conduit was assessed using histomorphometry. The L4 and L5 dorsal root ganglia (DRG) from the afflicted nerve were explanted and examined with immunohistochemistry for Notch receptor activation. A known inhibitor of Notch receptor activation, DAPT, was used in cell culture to establish a causal relationship between Notch activation in neurons and reduced axonal growth in the presence of SenSCs. Rat DRG neurons were co-cultured with SCs or SenSCs and exposed to DAPT supplementation and maximal neurite extension was measured over time.

RESULTS: In vivo, axonal regeneration was decreased in the presence of SenSCs. Myelinated axon numbers and regenerative quality were reduced in the SenSC conduits compared to SC conduits. Concomitant with decreased axonal growth, DRG neurons in the rats with implanted SenSC conduits had increased activation of Notch receptors. In the presence of cultured SenSCs, DRG neurite extension was decreased 28-40% compared to neurons co-cultured with healthy SCs. With the addition of the Notch activation inhibitor, DAPT, neurite extension was only reduced 13-21% compared to neurons co-cultured with healthy SCs representing an up to 50% rescue of neurite extension.

CONCLUSION: These data suggest SenSCs activate Notch signaling in neurons via increased expression of Notch ligands: this signalling ultimately results in decreased axonal extension. Neurite extension was partially rescued by treating neurons with a Notch receptor activation inhibitor. This work implicates Notch signalling as an important mechanism by which Schwann cell senescence reduces axonal regeneration. Thus, inhibition of Notch signaling may improve nerve regeneration in SenSC-rich environments, such as long nerve grafts.


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