Schwann Cell Isolating,Culturing And The Study Of Transplanting On The Self-assembly Which Repairing Peripheral Nerve

Schwann cells (SCs), one of glia cells, forming peripheral nerve Myelin sheath, were the first described by Theodor Schwann in 1839. SCs which transplanted in peripheral nerve can survive , proliferate, migrates and differentiation, and they may secrete various nerve nutrition factors (NTFs) , cell adhesion molecule, and extracellular matrix, promoting and guiding the regeneration of the axis of peripheral nerve. SCs and functional recovering are the most important two factors of peripheral nerve regeneration. SCs is playing very important role in the process of peripheral nerve regeneration.The method of the paper study seek one kind of isolating of SCs , which can isolate and culture SCs from sciatic nerve of new-born Wister rat within short time, moreover the number reaches over 10~6/ml, the purity reaches above 90%, then they can clinic coming to satisfy the clinic require of repairing peripheral nerve. The certain quantity of SCs transplanted in self-assembling material PDLLA-CHS-CS of repaired peripheral nerve , study the biocompatibility of PDLLA-CHS-CS and the material influence the proliferation of SCs. Studying the impact of the material PDLLA-CHS-CS to peripheral nerve regeneration, used PDLLA-CHS-CS renovated Wister rat sciatic nerve.SCs were isolated by warm enzyme digestion, tissue culture, cold enzyme digestion of substep and cold enzyme digestion of combine. After the double 30min adhesion culture, comparison the total number, the number of live cells and the purity, we found the method of the cold enzyme digestion of combine can satisfy the necessary of clinical in vitro and in vivo from the number, the time and the purity, which need isolate Schwann cells from autologus nerve.Culture solution of SCs used DMEM/F12+10%FBS(fetus bovine serum ). Through Ara-C eliminated and suppressed the fibroblasts growth and the basic fibroblasts growth factor(bFGF) proliferated the SCs. the number of Schwann cells can reach 10~6/ml and the purify of Schwann cells can reach over 90% , which can satisfy the necessary of clinical. The growth curve of Schwann cells used the MTT assay and identified SCs by HE dyeing and anti-s100 immunohistochemistry of Horseradish peroxidase mark and anti-NGFRp75 of FITC fluorescence mark.PDLLA, CHS have good biocompatibility, CS is one of extracellular matrix, which can promote the peripheral nerve regeneration. Though the technique of static electricity self-assembling, PDLLA , CHS, CS fabricated multilayer compound material. It namely overcome PDLLA degradation slow shortcoming , have also have overcome CHS machinery intensity weakness , added a few CS can promote the SCs proliferation, should be one kind of ideal repairing peripheral nerve material. In vitro, such as: The MTT assay, laser scanning together focusing microscope and ESEM , observed SCs cell growth on the surface of PDLLA-CHS-CS compound material. The result showed PDLLA-CHS-CS and the SCs have good biocompatibility and good affinity, and the material promote the proliferation of SCs.In vivo, defect and loss peripheral nerve is restored PDLLA-CHS-CS material, discovering PDLLA-CHS-CS material being able to promote the defect and loss rat sciatic nerve regeneration, moreover NGF beneficial to regeneration of peripheral nerve.