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A Novel Forelimb Orthotopic Allotransplantation Model in Mouse
Xianyu Zhou, M.D.1,2, Georg J. Furtmüller, M.D.1, ByoungChol Oh, D.V.M., Ph.D.1, Maria Chicco, M.B.B.S.1, Jun Yang, M.D., Ph.D.2, W.P. Andrew Lee, M.D.1, Gerald Brandacher, M.D.1.
1Vascularized Composite Allotransplantation Laboratory, Department of Plastic and Reconstructive Surgery, Johns Hopkins University School of Medicine, Baltimore, MD, USA, 2Department of Plastic and Reconstructive Surgery, The Ninth People's Hospital of Shanghai, Shanghai Jiaotong University School of Medicine, Shanghai, China.

A novel forelimb orthotopic allotransplantation model in mouse
Xianyu Zhou, M.D.1,2*, Georg J. Furtmüller, M.D.1*, ByoungChol Oh, D.V.M., Ph.D.1, Maria Chicco, M.B.B.S.1, Jun Yang, M.D., Ph.D.2, W.P. Andrew Lee, M.D.1,Gerald Brandacher, M.D.1
1Vascularized Composite Allotransplantation Laboratory, Department of Plastic and Reconstructive Surgery, Johns Hopkins University School of Medicine, Baltimore, MD, USA
2The Ninth People’s Hospital of Shanghai, Department of Plastic and Reconstructive Surgery, Shanghai Jiaotong University School of Medicine, Shanghai, China.
*These two authors contributed equally to this work
Purpose: In translational research related to Vascularized composite allotransplantation (VCA), mouse models provide a profoundly relevant means to successfully studying of crucial aspects related to the immunobiology of graft rejection and acceptance, neuronal reinnervation of sensory and motor function and associated cortical reintegration of lost extremity function. The model developed here is designed to address this critical void to allow for mouse orthotopic forelimb transplantation, a tool to studying both aspects of functional recovery and immunobiology related to VCA in a genetically versatile mouse model system.
Methods: Male Balb/c donors and C57BL/6 recipients (8-10 weeks) mice were used to perform syngeneic and allogeneic orthotopic forelimb transplantation. Co-stimulation blockaded (CTLA4-Ig & MR-1) was used for immunosuppression. A circumferential skin incision was made at the shoulder. The pectoralis major muscle insertion was released at the humerus and reflected. The vascular pedicle was dissected from the axilla to the cubital fossa and transected at the level of the proximal axillary artery. Median and ulnar nerves were identified and transected proximal to the cubital fossa. The radial nerve was identified and transected dorsally at the intersection of medial and distal third of the humerus. Both flexor and extensor muscle groups were cauterized and transected at mid humerus level. In the recipient animal shoulder mobility was preserved and vessels and nerves were transected proximal to the cubital fossa. The graft was secured by muscle approximation with 6-0 Polysorb and by using a 26G spinal needle as an intramedullary rod for osteosynthesis. A non-suture cuff technique was employed to anastomose donor axillary vessels with recipient brachial (mid-arm) vessels. Tension free primary end-to-end neurorrhaphy was performed in median, ulnar and radial nerves using 11-0 microsuture.
Results: Long term animal and graft survival in syngeneic transplant combinations demonstrated the feasibility of orthotopic mouse forelimb transplantation with only limited behavioral disabilities (>60 days; N=3).The non-suture cuff technique allows for successful supermicrovascular anastomosis of the donor axillary vessel with the recipient brachial
vessel. The feasibility of radial, median and ulnar nerve coaptation provides the basis for studying functional recovery as well as cortical reintegration of upper extremity function after VCA. Long term allograft survival (>4 months; N=3) underpins the applicability of orthotopic forelimb transplantation for studying the immunobiology of functional extremity reconstruction using VCA.
Conclusion: The described orthotopic forelimb transplantation model in the mouse is designed to add an additional research model for VCA by providing the option of using the entire range of genetically engineered mice for research related to transplant immunology, neuroregeneration, and functional recovery after upper extremity transplantation.


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