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Axonal Reconnection with Polyethylene Glycol and Methylene Blue after Sciatic Nerve Transection Results in Rapid Behavioral Recovery
Cameron P. Keating, MBBS, BMedSc; Joshua M. Britt, BS; Jacqueline R. Kane, BS; Aleksej Zuzek, BS; Christopher S. Spaeth, BS; Jerry D. Fan, BS; Francisco Gonzalez-Lima, PhD; Timothy Schallert, PhD; George D. Bittner, PhD; Jonathan M. Winograd, MD
Plastic Surgery Research Laboratory, Mass. General Hospital, Boston, MA, USA
Objective
The objective was immediate restoration of axonal membrane continuity and distal motor and sensory innervation after peripheral nerve transection, using both the lipid membrane fusogen polyethylene glycol (PEG) and the antioxidant methylene blue (MB), enhancing functional recovery.
Methods
Animals were subjected to sciatic nerve transection (with the exception of sham) followed by: no repair; PEG alone; suture alone; suture+PEG; and suture+PEG/MB. Compound action potentials (CAPs) were measured by directly stimulating/recording from the Sprague Dawley rat sciatic nerve: before transection; after transection; and after repair as a physiological measure of axonal continuity. Ex vivo nerve dye diffusion studies were performed. Histological parameters of nerve regeneration including: nerve fiber counts; axon/fiber diameter; g ratio, and myelin thickness were measured 5 mm proximal and distal to the repair at 1,2 and 12 weeks. Weekly hindlimb behavioral assessments were undertaken for 12 weeks with two validated scoring systems: Sciatic Function Index (SFI – distal limb function); and Foot Fault testing (FF – proximal limb function).
Results:
Pre-injury sciatic nerve CAPs were similar in all groups (mean 7.6 mV) and post-transection CAPs were undetectable in all animals. 100% of the suture+PEG and suture+PEG/MB groups demonstrated return of CAPs post-repair (87% of baseline) compared with 0% in all other groups. Similarly, dye diffusion studies confirmed morphologically that dye traversed the lesion site through reconnected axons in all animals with detectable post-repair CAPs and none of the other groups. Histology data is currently being processed. Behavioral recovery was both more rapid and effective in the suture+PEG/MB group on SFI scoring with 40% recovery at 72 hours (vs 0% - suture alone) and 70% at 12 weeks (vs 20%; Fig. 1). Curve-fit parameters indicated statistically significant behavioral improvement (p<0.001). Similar more rapid and advanced recovery was demonstrated with FF scoring and there was high inter-experimenter reliability with this technique.
Conclusions:
Successful axonal reconnection was achieved in all of the suture+PEG and suture+PEG/MB groups and none of the controls after sciatic transection, providing compelling evidence of physiologic and morphologic restoration of axonal continuity. Functional outcomes in the suture+PEG/MB group demonstrated dramatic improvements in the speed and extent of motor and sensory recovery compared to standard of care suture alone on two behavioral assessments. Translation to human nerve repair could lead to a paradigm shift in the treatment of peripheral nerve injuries.
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