Introduction: Hand and leg transplantation (Tx) in military and civilian populations is increasingly needed to improve the quality of life of individuals. Optimizing nerve regeneration is a key to successful outcomes. The objective of this study was to investigate whether Mesenchymal Stem Cells (MSC) derived from bone marrow can improve nerve regeneration and functional outcome in a limb transplant model.

Methods: Orthotopic syngeneic right hind-limb transplants were performed in Lewis (RT1.A^l) rats. The donor and recipient femoral artery/vein were anastamosed by Vascular Cuff method, and the sciatic nerve (epineurium) and muscles were approximated by suturing. The femur was stabilized with a stainless steel pin and bone cement. Animals received MSC (5 x10⁶; passage ≤7) or vehicle (saline control) locally around nerve, bone and vascular anastamoses sites, and 5×10⁶ MSC intravenously on the day of surgery. Intravenous MSC or vehicle injections were repeated every week for four weeks. Walking track and cutaneous pain reaction tests were performed at 1-2 week intervals post-Tx.

Results: MSC isolated from Lewis rat bone marrow and expanded ex vivo were CD29⁺, CD90⁺, CD34^–, CD31^–, CD45^low, MHC Class I⁺, Class II^–, CD80^low, and CD86^–. MSC were confirmed for pluripotency based on their differentiation potential in to adipocytes, osteocytes and chondrocytes in ex vivo cultures under specific conditions. At four weeks post-Tx, no animal bore complete weight on the transplanted limb, and we were not able to obtain clear foot prints to calculate Sciatic Function Index (SFI). However, 2 out of 5 vehicle treated animals and 1 out of 2 MSC treated animals showed slight foot prints by 2 weeks, which became more recognizable by 4 weeks. Also, 3 out of 5 control animals were Grade 1-2 on a scale of 0-3 (0=No Function; 3= Normal function) for peroneal, tibial and / or sural nerve sensory function at 4-6 weeks. In our pilot study (n=5) without MSC or vehicle therapy the sensory nerve function was Grade 1.53±0.69 (peroneal), Grade 1.79±0.29 (tibial) and Grade 1.39±0.55 (sural) at ∼8 weeks. Laser doppler analysis revealed normal vascularization in transplanted limbs. Limb transplant survival was 100%. The study is ongoing and awaiting data on long-term effects of MSC on neurogenesis, angiogenesis, myogenesis and limb function.

Conclusion: The limb transplant procedure was highly successful and MSC therapy appears to promote nerve functional recovery in our limb transplant model.

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