| Great progress was made on the study of injury and rehabilitating of peripheral nerve by tissue engineering in the recent years. The use of an artificial nerve conduit containing viable Schwann cells (SCs) is one of the most promising approaches to repair nerve injuries. SCs is the key ganglia cells which derived from pre - neurologic cell. Predegenerated nerves, so - called Wallerian degeneration, which occurs after nerve injuries, contain infiltrating macrophages and fibroblasts. Many investigators demonstrated that the presence of Schwann cells in the regenerating milieu benefits nerve regeneration. The Schwann cell -coated rolled grafts provided a modestly enhanced regenerative milieu, as evidenced by improved functional recovery. This is attributable to the known neuro-trophic properties of Schwann cells, both soluble and surface proteins augment the axonal migration process. In this study, we have developed a novel technique to isolate and purify SCs from neonatal rat peripheral nerves for an artificial nerve conduit, which is shown to have an effect on SC proliferation and survival. The combination of this technique and the altered medium promoted the migration and proliferation of SCs selectively by utilizing the supporting cells of SCs instead of discarding them by changing the culture dishes and media. The objective was to place adherent Schwann cells in artificial nerve grafts and to assess regeneration through the Schwann cell — laden grafts compared with that through acellular grafts and autografts. Schwann cells were isolated from neonatal SD rats. Using an artificial nerve conduit containing viable Schwann cells(SCs) is a more promising method for repairing peripheral nerve injuries than the use of a conduit alone tissue engineering offers some advantages over al-lograft nerve transplantation, it uses biodegradable materials and autologous cells that do not cause antigenic reactions, and it has already been applied to clinical fields. Thus, obtaining a large number of viable SCs in a short period ( which is necessary for the application of tissue engineering) is necessary for a clinical use. The nerve grafts are divided into allogeneic nerve grafts and heterogenous nerve grafts those have their benefits and disbenefits in bridging nerve gaps respectively . In addition, amount of cultured and highly purified Schwann cells play a key role as a seed cell in rehabilitating of peripheral nerve injury because of their secreting numerous neurotrophic factors. Presently, the autologous nerve grafting is an important method in reconstructive surgery of peripheral nerve lesions. However, there are still unresolved problems. The availability of auto-grafts is limited and the harvesting of autografts result in donor site morbidity such as sensory defects, scarring and neuroma formation. So the study of ideal graft is an important subject of peripheral nerve surgery. These results highlight the role of Schwann cells in nerve regeneration. Regenerative results approaching autograft levels in the Schwann cell laden group suggest that this methodology may ultimately be useful in clinical nerve repair.MATERIALS AND METHODSSchwann cells were isolated from SD rats and enzymatically digested with collagenase and dispase in vitro. Expression was monitored by immunohisto-chemical staining of the S -100 protein for Schwann cells and marked by Ho-echst33342. We made natural biodegradable nerve graft which could be cocul-tured with SCs. Morphological change can be observed by transmission electron microscope (TEM) , scanning electron microscope ( SEM ) and fluorescent microscope. Rats were randomly divided into three groups; autografts, acellular grafts, or Schwann cell - laden grafts. After injury and implantation, the animals were tested at intervals for return of function in 13 - weeks or 18 - weeks. Axon measurements were obtained in blinded fashion for fiber number, density, andaverage diameter as well as percentage of neural tissue in the total cross section by TEM and SEM. All groups were designed to investigate the motor functional recovery and NCV\ AMP N AREA of regenerated nerve after resconstruction by histochemical methods and electrophysiologic test. The data were expresses as mean ± SD and analyzed by SPSS10. 0 statistical package after taking a photograph under the microscope. Statistical comparisons were made using t - test or F - test required for statistical significance.RESULTS1. SCs performed mononuclear with two polar after culturing 24hr, then a-mount of regular SCs displayed activity and migration with a fairly stable orientation after 48 hr. The volumn of SCs becomes bigger according to the time changed. We can observe the morphological proformance with standard fluorescent microscope, owing to SCs nuclear staining labeled by Hoechst 33342. The expression of S - 100 of SCs was examines exceeding 90% by ABC immunohis-tochemical method.2. The axons and Schwann cell sheath disappear in the cross section of AR-SN. In the longitudinal section, typical long basal membrane consuit can be observed. The morphological change is also detective in toluidine blue stained specimen, while the group of transplanting SCs can observe the amount of mono-nuclear with two polar. The feature of cultures SCs shows shoulder by shoulder , head to head.3. After ARSN with cultured SCs bridged the gap of rat sciatic nerve gap, no anastomotic disruptions and no fractured conduits were observed. On gross inspection, all specimens contained neural regenerates. Based on the histomor-phometric data presented in Table. There was no significant difference in number of the regenerated nerve fibers, the thickness and the acre of regenerated my-elinated nerve between the experimental group and autograft group( P > 0.05). Whereas, there was significant difference between ARSN with SCs and ARSN(P< 0. 05 ) There was no significant difference between experimental group and control group in terms of the examined parameters of regeneration by electro-...
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