Femke Mathot

Chapter 5 80 ABSTRACT Introduction Nerve regeneration involves multiple processes which enhance blood supply that can be promoted by growth factors. Currently, tools are lacking to visualize the vascularization patterns in transplanted nerves in vivo. The purpose of this study was to describe three-dimensional visualization of the vascular system in the rat sciatic nerve and to quantify angiogenesis of nerve reconstruction. Materials and Methods In 12 Lewis rats (weighing 250 – 300 grams), 10 mm sciatic nerve gaps were repaired with ipsilateral reversed autologous nerve grafts. At 12 and 16 weeks of sacrifice, Microfil® contrast compound was injected in the aorta. Nerve autografts (N=12) and contralateral untreated nerves (N=12) were harvested and cleared while preserving the vasculature. The amount of vascularization was measured by quantifying the vascular surface area using conventional photography (two dimensional) and the vascular volume was calculated with micro-computed tomography (three dimensional). For each measurement, a vessel/nerve area ratio was calculated and expressed in percentages (vessel%). Results The vascular volume measured 3.53 ± 0.43% in autografts and 4.83 ± 0.45% vessels in controls at 12 weeks and 4.95 ± 0.44% and 6.19 ± 0.29% vessels at 16 weeks, respectively. The vascular surface area measured 25.04 ± 2.77 % in autografts and 26.87 ± 2.13% vessels in controls at 12 weeks, and 28.11 ± 3.47% and 33.71 ± 2.60% vessels at 16 weeks respectively. The correlation between both methods was statistically significant (P=0.049). Conclusions Both methods are considered to successfully reflect the degree of vascularization. Application of this technique could be used to visualize and objectively quantify angiogenesis of the transplanted nerve graft. Moreover, this simple method is easily reproducible and could be extrapolated to any other desired target organ ex vivo in small animals to investigate the vascular network.