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Surgical Angiogenesis To Nerve Allografts Improves Early Functional Recovery In A Rat Sciatic Nerve Defect Model
Tiam M. Saffari, MD, Femke Mathot, MD, Allen T. Bishop, Prof, MD, Alexander Y. Shin, Prof, MD.
Mayo Clinic, Rochester, MN, USA.

PURPOSE: It is widely accepted that functional results are poor when nerve grafts are transplanted in severely scarred tissue beds and that independent blood supply of nerve grafts could improve outcomes. Optimized processed allografts (OPA) have no vascular supply. Addition of vascularization to the OPA promotes immediate revascularization and minimizes the period of ischemia, diminishing fibrosis and central necrosis potentially enhancing functional recovery. The purpose of this study was to determine how addition of vascularization to nerve allografts would affect functional recovery.
METHODS: Sixty Lewis rats were divided into three groups of 20 animals each. Nerves harvested from Sprague Dawley rats were decellularized according to protocol (Hundepool et al, 2017), irradiated and stored until use as OPA. Unilateral 10 mm sciatic nerve defects were repaired with (i) OPA, (ii) OPA wrapped with a superficial inferior epigastric fascial (SIEF) flap to provide vascularization to the nerve bed, and (iii) autografts. Twelve and 16 weeks after surgery, nerve regeneration was assessed using functional, electrophysiologic, histomorphologic, and immunofluorescence analyses (N=10/time point). Ultrasound was used during the survival period to noninvasively evaluate muscle atrophy.
RESULTS: Rats in which a well-vascularized SIEF flap was provided to the nerve allograft showed significantly better isometric tetanic force (ITF) recovery at 12 weeks, compared to allograft alone. ITF recovery normalized between groups at 16 weeks. Ultrasound showed a trend toward less atrophy in SIEF rats, compared to allograft only, however, this was not significant. Electrophysiology showed superiority of autografts at both time points. Histomorphometric analysis revealed no differences between groups with regard to axon area, axon count and myelin area, however, the N-ratio was significantly inferior in allografts at 12 weeks compared the autograft and SIEF group. Using immunofluorescence, the expression of CD34, indicating vascularity, showed significantly improved levels in SIEF samples compared to allografts at 12 weeks, and highest expression at 16 weeks compared to all groups. Protein gene product 9.5 (PGP9.5), a pan-neuronal marker, did not reveal any differences between groups at 12 or 16 weeks.
CONCLUSION: This study suggests that addition of a well-vascularized fascial flap to the nerve allograft bed increases vascularity in the nerve allograft, subsequently improving early functional recovery, comparable to autografts.

Figure 1. Compound muscle action potential (CMAP, 1A) and ITF (1B) measurements for autograft, allograft and SIEF. Results are expressed as a percentage of the unoperated side and given as meanSEM.


Figure 2. Histological analysis of PGP 9.5 and CD34. Micrographs showing density in control nerve samples(A), autograft(B), allograft(C) and SIEF(D) at 12 weeks. Density of axons and vascularity (normalized to the combined area of tissues positively stained for any of PGP9.5, CD34 and DAPI) is shown in E and F, respectively. Scale bars are set at 100 Ám.


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