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

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Enhancement of Nerve Regeneration After Delayed Repair with Controlled Release of Neurotrophic Factors
Kasra Tajdaran, MASc1, M. Cecilia Alvarez-Veronesi, MASc1, Matthew D. Wood, PhD2, Molly S. Shoichet, PhD1, Tessa Gordon, PhD1, Gregory H. Borschel, MD, FACS, FAAP1.
1University of Toronto, Toronto, ON, Canada, 2Washington University, St. Louis, MO, USA.

PURPOSE:
Peripheral nerve injuries frequently lead to functional and sensory dysfunction. Delays in surgical repair following nerve transection reduce axonal regeneration, which is in part, due to reduced neurotrophic support. Previously, local application of exogenous neurotrophic factors, including glial cell line derived neurotrophic factor (GDNF) was shown to have a positive effect on axon regeneration after delayed repair. Here we investigated a novel delivery system for GDNF following delayed repair using drug-loaded poly(lactic-co-glycolic acid) (PLGA) microspheres (MS) embedded in a fibrin gel, which serves to localize the microspheres around the nerve injury site and allows sustained GDNF release.
METHODS:
The characteristics of the drug delivery system including its biocompatibility and GDNF release profile were studied with cell toxicity assays and ELISA. The toxicity assay was performed by incubating PC-12 cells with the medium released from PLGA microspheres in vitro. . A delayed nerve repair model was used in which the rat common peroneal (CP) nerve was transected and regeneration was prevented by ligation for 2 months. Thereafter, the CP nerve stumps were coapted. The 3 experimental groups were implanted with GDNF-containing microspheres, free GDNF or empty microspheres in fibrin gel at the repair site. In a positive control group, the CP nerve was coapted immediately after transection. Four weeks after repair, the numbers of neurons that regenerated their axons were determined using retrograde labeling 15mm distal to the repair site. To analyze axonal regeneration through the nerve and identify possible axon coil formation, neurofilament (NF) immunohistochemistry analysis was performed on longitudinal sections of the CP nerves at the suture site, and at the proximal and distal parts of the distal stump (Fig.1).
RESULTS:
Using ELISA, GDNF release from microspheres was detected over 14 days in vitro. PC-12 cells incubated in vitro with released GDNF samples had similar viability to control cells cultured with normal media demonstrating that the drug delivery system was not toxic. Consistent with this, the numbers of motor and sensory neurons that regenerated their axons in vivo was the same as when there was no microspheres treament (Fig.2). The numbers of regenerating neurons increased when GDNF was administered in fibrin gel, but this increas was not significant. In contrast, GDNF administered within microspheres increased the numbers significantly, almost to the numbers after immediate nerve repair (Fig.2). Both qualitative and quantitative analysis of axon profiles in longitudinal sections showed the longitudinal outgrowth of axons with no evidence of any coiling (Fig.1).
CONCLUSION:
Addressing the deleterious effects of delayed repair holds promise for removing one of the major barriers to improve nerve regeneration. GDNF microspheres incorporated into fibrin gels improve axon regeneration after delayed nerve repair and offer a potential treatment of chronic peripheral nerve injuries.



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