| Objective: Repairing of large gap defect of peripheral nerve injury is a big challenge for clinical therapy. Repairing with nerve autograft is the fundamental and ideal method for the peripheral nerve defect. But there isn't enough nerve autograft for the repairing of the large gap defect of peripheral nerve injury or nerve plexus injury. this makes the therapy difficult. There is a large resource of nerve allograft, but the immunological rejection limits the clinical application of nerve allografts. More and more scholars took tissue engineering into account to prepare the nerve graft. Recently they found that the "chemical extraction"method could remove the schwann cells and myelin sheaths in peripheral nerve effectively relieving the antigenicity of nerve allografts. So, this developed a new method to make substitute of nerve autograft. In this experiment, Triton X-100 was used to extract the schwann cells and myelin sheaths of peripheral nerve for obtaining the inartificial nerve extracellular matrix and preparing tissue-engineered nerve graft. This would provide experimental basis for the repairing of large gap defect of peripheral nerve. Methods: 60 pieces of sciatic nerve in 10 mm length were taken from 30 rabbits, and were randomly divided into 6 groups(A,B,C,D,E,F).Group A,B,C,D,E were treated with Triton X–100.Group F was control group. Before the extraction, sheared the adipose tissue and part of the epineurium with the operating microscope. Then they were dipped in 4oC distilled water immediately, Soaked for 6hs, distilled water made the cells and myelin sheaths expansion and disruption, in order that Triton X-100 can easily infiltrated. Put the nerves into 3% Triton X-100 solution, treated by Triton X-100 for 12hs.After this, put the nerves into 4oC distilled water again, washing for 6hs to make the complex of Triton X-100 and membran protein out from the nerve completely. Repeated the operation as before and changed the distilled water and Triton X-100 solution every time.5 groups of nerve were treated for 12hs,24hs,48hs,96hs,1week respectively. 2 pieces of nerve in 2 mm length were taken from the middle part of each piece of nerve. One was fixed with 10% formaldehyde and normaly embedded with paraffin, longitudinally sliced up, HE,PTAH,Alcian blue and laminin immunohistochemical staining were performed and investigated by light microscope. The laminin immunohistochemical stained sections were performed with image acquisition and analyzed with multicolor pathological image analysis system. Measure the laminin antibody reaction part of each section. All dates were analyzed by SPSS. The other piece of nerve was fixed with 2.5% glutaral, and observed with scanning electron microscope. Results: All the sections were observed by lightmicroscope. Schwann cells in fresh nerve distributed uniformly, myelin sheaths and basement membrane lined up in order. Schwann cells and myelin sheaths in peripheral nerve decreased gradually along with the elongation of the treated time. The delimitation between basement membrane and myelin sheath in treated nerve was not clear. Schwann cells and myelin sheaths disappeared when the nerves were treated for 96hs. Between the membranes was the basement membrane tube. When the nerves were treated for 1 week, the basement membrane was also clear, but the tube structure composed by collagen fibers was abnormity, part of the tube ruptured. Observed with scanning electron microscope, axons and myelin sheaths of the nerve decreased gradually along with treated time elongation. Axons and myelin sheathes disappeared when the nerves were treated for 96hs.The basal laminin tubes composed by collagen fibers were remained, collagen fibers maintained their former position, form and structure. When the nerves were treated for 1week, the basal lamin tubes structure was similar with the nerves treated for 96hs, but the structure was disordered a little, and the basal lamin tubes was abnormal. All the Laminin immunohistochemical stained sections were performed with image acquisition and analysis. The average gray degrees of the laminin antibody reaction part of each group were as follow: group A,140.1±3.41;group B,142.1±3.14;group C,142.1±3.14;group D,140.4±4.03;group E,141.7±2.62;group F,142.7±7.24. Dates were analyzed with SPSS, There wasn't...
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