Methods: One and 4 week periods of UWS cold preservation treatment of nerve allografts were compared to untreated allograft and isograft controls. To assess the functional and histomorphometric effects, Lewis recipient rats were randomized to 4 groups of 8. After each animal received the appropriate 2 cm long graft (ACI donor for allografts), walking track analyses was performed to assess function and nerve tissue was harvested at 9 weeks for histomorphometric analysis. To assess the immunologic response, C57 recipient mice were randomized into 4 groups of 8 and received the appropriate inlay sciatic nerve graft (Balb/c donor for allografts). At 10 days, nerve tissue, the spleen and blood were harvested. Nerve tissue was examined for degree of cellular infiltration. The cellular immune response was quantified using the enzyme-linked immunosorbent spot assay (Elispot) and the humoral response was evaluated by measuring for donor specific antibodies using flow cytometry.

Results: In the rat model, functional analysis did not show significant differences between groups. Preliminary histomorphometric data showed that the isograft group had robust nerve regeneration, the allograft group had poor regeneration and the cold preserved groups had intermediate regeneration: the 4 week group showed a trend toward better regeneration compared to the 1 week cold preserved group. In the mouse model, the Elispot assay showed a shift from a predominantly interferon-ã producing T cell response to an interleukin-4 producing response with increased duration of cold preservation. At the 10 day endpoint, there were no detectable IgM or IgG alloantibodies by flow cytometry. Examination of the nerve tissue is pending.

Conclusion: Work in the rat model confirmed that cold preservation of nerve allograft tissue does not preclude subsequent nerve regeneration. The immunologic effect, as observed in the mouse model, showed that increased duration of UWS cold preservation shifted the cellular immune response from an T helper cell type 1 (pro-inflammatory) to a type 2 (tolerance inducing) response. Therefore, cold preservation of nerve allograft tissue in UWS diminished immunogenicity while maintaining nerve regeneration and could serve as a means to produce acellular, non-antigenic graft material for use in peripheral nerve reconstruction.