Introduction: Schwann cells reorganize after nerve transection and provide a permissive environment for nerve regeneration. The present study was performed to determine if transplanted Schwann cells identified by GFP fluorescence and fluorescence in situ hybridization (FISH) for Y chromosome can integrate and participate in axonal regeneration, remyelination and node of Ranvier formation.

Methods: Schwann cells were prepared from dissociated sciatic nerves of GFP transgenic male rodents. Sciatic nerves of female rats were exposed and crushed for 20 sec with fine forceps to transect all axons. Immediately after nerve crush of a GFP-Schwann cell suspension was injected just distal to the crush site through a glass pipette attached to a Hamiliton syringe. 2-3 weeks later the nerves were removed and dissociated on glass slides. The engrafted cells were identified by GFP fluorescence and FISH for Y chromosome. Immunostaining for Na+ channel (Nav 1.6) and paranode (Caspr) was used to define nodes of Ranvier on regenerated axons.

Results: GFP+ cells distributed throughout the crush lesion and longitudinally distal to the nerve transection. GFP was observed only in the cytoplasmic compartments of the Schwann cells which included the cell body and the outer mesaxon. The nuclei of the GFP+ cells had Y chromosomes further indicating the donor origin of the myelin-forming cells. The GFP cells aligned longitudinally and surrounded the regenerated axons. Nodes of Ranvier between GFP+ cells were observed as breaks in green fluorescence and by intense sodium channel (Nav 1.6) staining on the regenerated axons.

Conclusion: The transplantation of identified Schwann cells into an acute axotomy model results in axonal regeneration and remyelination by the transplanted cells. The donor myelin-forming cells could be identified by GFP fluorescence and by FISH for Y chromosome. The transplanted cells were able to survive and form myelin with mature nodes of Ranvier on the regenerated axons. These results indicate that engrafted Schwann cells into injured peripheral nerve can integrate and participate in neural repair.