Early nerve regeneration on the crushing injury model and the autogenous nerve graft model of rat sciatic nerve

Park, Jong Chul; Ahn, Kang Min; Lee, Jee Ho; Lee, Jong Ho

Abstract

The number of Schwann cell in the early nerve regeneration on the crushing injury model and the autogenous nerve graft model was analysed. And the change pattern of myelination and axon growth after nerve injury on each nerve injury model was investigated. And the beta-Galactosidase activity on each nerve injury model after the injection of Adenoviral vector with LacZ gene was to be confirmed. For the crushing injury model, the injury was made by specially designed hemostat (Width: 3mm) at 10mm distal to sciatic notch for 10seconds(N=18). For autogenous nerve graft model, the 7mm segmental injury was made at same area and then microsuture was done immediately after injury(N=18). Three rats were sacrificed by cardiac perfusion at post injury 1, 3, 5, 7, 14, 21days and the sciatic nerves were harvested. Immunohistochemical staining for S-100 protein was made for detection of Schwann cell. For the observation of light and ultramicroscopic change, toluidine blue staining and TEM image were obtained(N=14). 3microliter Adenoviral vector with LacZ gene(3x10(11) PFU/ml) was injected after injury(N=6), and sacrificed at post injury 7, 14, 28days, then X-gal stain was made. For control group, we used the sciatic nerve of undamaged 3 rats(N=3). On the crushing injury model, the total number of positive cell for S-100 protein was significantly increased after injury and highest at post op 7days(p<0.05). The intensity of positive staining for S-100 protein was decreased immediately after crushing injury and restored over the time course. In the TEM image, myelination was detected at post op 7days. On the autogenous nerve graft model, the total number of positive cell was gradually increased and highest at post op 14days, and then the total number of positive cell was significantly decreased(p<0.05). The intensity of positive staining was gradually increased by the post op 14days. In the TEM image, myelination was detected at post injury 14days. On the each injury model, X-gal staining was positive around the area of adenovirus injection, and the intensity of X-gal staining was decreased over the time course. By the result mentioned above, preliminary study for comparing the transfection efficacy between immediate injection and delayed injection of adenoviral vector with neurotrophine gene at the time of highest Schwann cell number was made. As well, the control data of peripheral nerve regeneration on our nerve injury model was prepared for later experiment of adenovirus injection treated with neurotrophine gene for the peripheral nerve regeneration.