Perineurial Window is Critical for Experimental Reverse End-to-Side Nerve Transfer

The depth of connective tissue window in the side of a recipient nerve in reverse end-to-side transfers (RETS) remains controversial.To test whether the depth of connective tissue disruption influences the efficiency of donor axonal regeneration in the context of RETS.Sprague-Dawley rats (n = 24) were assigned to 1 of the 3 groups for obturator nerve to motor femoral nerve RETS: group 1, without epineurium opening; group 2, with epineurium only opening; and group 3, with epineurium and perineurium opening. Triple retrograde labeling was used to assess the number of motor neurons that had regenerated into the recipient motor femoral branch. Thy1-GFP rats (n = 8) were also used to visualize the regeneration pathways in the nerve transfer networks at 2- and 8-week time point using light sheet fluorescence microscopy.The number of retrogradely labeled motor neurons that had regenerated distally toward the target muscle was significantly higher in group 3 than that in groups 1 and 2. Immunohistochemistry validated the degree of connective tissue disruption among the 3 groups, and optical tissue clearing methods demonstrated donor axons traveling outside the fascicles in groups 1 and 2 but mostly within the fascicles in group 3.Creating a perineurial window in the side of recipient nerves provides the best chances of robust donor axonal regeneration across the RETS repair site. This finding aids nerve surgeons by confirming that a deep window should be undertaken when doing a RETS procedure.