A Novel Model for Evaluating Nerve Regeneration in the Composite Tissue Transplant: The Murine Heterotopic Limb Transplant

Abstract

Purpose For individuals who have experienced debilitating upper extremity injury or amputation, hand transplantation holds the potential for drastic quality of life improvement. This potential depends on adequate nerve regeneration into the transplant and reanimation of graft musculature. In this study, we demonstrate the use of a murine heterotopic limb transplant model for evaluation of nerve regeneration in a composite tissue allograft (CTA). We also compare the effects of various immunosuppressive regimens on nerve regeneration in this model. Methods The study consisted of five groups of mice, all of which underwent heterotopic limb transplant with coaptation of the recipient and donor sciatic nerves. The groups received the following immunosuppressive regimens: group A (positive control)-syngeneic transplant, no immunosuppression; group B (negative control)-allogeneic transplant, no immunosuppression; group C-allogeneic transplant, FK-506+MR1; group D-allogeneic transplant, MR1+ CTLA4-Ig; group E-syngeneic transplant, FK-506 treatment with preloading. Results Group B animals showed signs of transplant rejection as early as 5 days postoperatively. Except for one mouse from group C and one mouse from group D, all other animals had viable transplants and nerve regeneration present in the donor sciatic nerve at the 3-week endpoint of the study. Conclusions To our knowledge, this represents the first report of the use of a mouse CTA model for evaluation of nerve regeneration. The mouse heterotopic limb transplant model will be a valuable tool for CTA research since it can be performed with more ease, and with less host morbidity and mortality than the mouse orthotopic model.

Robust Axonal Regeneration in a Mouse Vascularized Composite Allotransplant Model Undergoing Delayed Tissue Rejection

“…As it was previously determined that robust axonal regeneration proceeds with strong immunosuppression to prevent overall tissue rejection following VCA, 14 the current study used a sub-immunosuppressive dose of Tacrolimus (FK506) to induce selective and delayed rejection of the VCA. This sub-immunosuppressive drug regimen allowed us to consider the antigenicity of nerve and its regenerative capacity under these conditions.…”

“…The criteria used to diagnose rejection have been previously detailed. 14,15 Briefly, these signs include erythema, progressive edema, and exudation, followed by eschar formation, necrosis, mummification, and slough.…”

“…In the study, a negative control group (allogeneic transplant without FK506 treatment) was not used as our previous data demonstrated transplant rejection signs by 5 days postoperatively and resulting nerve tissue was completely rejected leaving essentially no tissue to analyze. 14 Subcutaneous delivery of Tacrolimus (FK506, LOT-ATH-114; LC Laboratories, Woburn, Massachusetts) was performed daily for immunosuppression from the time of surgery until the end point of the study at a dose of 2 mg/kg for the allogeneic (group 2) and semi-allogeneic (groups 3 and 4) mice. All mice in the study underwent serial live imaging of their sciatic nerve of the transplanted limb at 2, 4, 6, and 8 weeks post transplantation for monitoring axonal progression in groups 1 and 2 and SC migration in groups 3 and 4.…”

“…(Sharpoint, Reading, Pennsylvania). The anastomoses were performed nearly "underwater," with the vessels submerged in heparinized saline (50 U/mL), as previously described, 12,14 for better filling and visualization of the lumen. Following reperfusion, host and donor sciatic and saphenous nerves were coapted with epineurial neurorrhaphy using 11-0 nylon suture.…”

Mouse Models of Experimental Vascularized Composite Allotransplantation

“…The team's latest study with the heterotopic model was in 2011, in which the most prominent modification was the rejoining of recipient sciatic nerve with the limb allograft counterpart. Microscopic analysis of post-allotransplantation nerve regeneration was also performed [18].…”