【作者】 陈军；

【导师】 沈华； 蔡燕娴； 章开衡； 贾万新；

【作者基本信息】 上海交通大学 ， 外科学（整形外科）（专业学位）， 2017， 硕士

【摘要】 目的:观察检测PLGA神经导管的生物相容性,探讨内置自体脂肪和神经的PLGA神经导管修复周围神经缺损的可行性。材料和方法:显微镜下从SD大鼠分离获取坐骨神经和腹股沟皮下脂肪,贴壁培养,分离提纯鉴定雪旺细胞、成纤维细胞和脂肪干细胞;将分离提纯鉴定的细胞与PLGA微丝共培养,钙黄绿素染色后观察细胞在PLGA微丝上的粘附和伸展,将材料的浸提液与细胞共培养,CCK8法检测浸提液对细胞的毒性。24只SD大鼠平均分成4组,于无菌条件下切断右侧的坐骨神经,制成10mm长的坐骨神经缺损模型,PLGA组采用PLGA神经导管连接缺损神经,PLGASN组由内置自体神经碎屑的PLGA神经导管连接,PLGA-SN-Fat组由内置自体脂肪与神经碎屑的PLGA神经导管连接,Autograft组采用自体神经移植进行修复。12周后通过对再生神经的一般观察、HE染色、Masson染色、免疫荧光S100染色以及甲苯胺蓝染色评价神经的再生情况;通过肌电图检测、腓肠肌的HE染色及Masson染色评价腓肠肌的神经再支配情况。结果:从SD大鼠成功地分离提纯获得了高纯度的雪旺细胞、成纤维细胞和脂肪干细胞,三种细胞可以很好地在PLGA微丝上粘附和伸展,且PLGA神经导管浸提液对这三种细胞没有毒性作用。一般观察、肌电图检测以及组织学染色观察分析均提示内置自体脂肪和神经的PLGA神经导管可以比较有效地修复大鼠坐骨神经缺损。结论:内置自体脂肪和神经的PLGA神经导管具有支持,引导和促进周围神经再生的作用,有利于周围神经缺损的修复,为周围神经的修复提供了一种新的方法。更多还原

【Abstract】 Objective:The aim of our research is to study the biocompatibility of the PLGA nerve conduit and to evaluate whether the combination of autologous fat and minced nerve tissue with poly-lactic-co-glycolic-acid(PLGA)scaffolds can improve peripheral nerve regeneration more effectively.Methods:The sciatic nerves and subcutaneous inguinal fat were isolated from SD rats under microscope,then adherent culture of adipose tissue and nerve tissue was performed.Proliferation and purification of Schwann cells,fibroblasts and adipose derived stem cells(ADSCs)were conducted after the isolation of the sciatic nerves and subcutaneous inguinal fat from the newborn Sprague-Dawley rats.Then Schwann cells,fibroblasts and ADSCs were cultured with PLGA fibre yarn to detect their ability to attach to and extend on the biomaterial using Calcein staining.CCK8 assay was performed to test the cytotoxicity of the extracts from the PLGA nerve conduit.Following the establishment of the 10 mm sciatic nerve defect models,the twenty four Sprague-Dawley rats were divided equally and randomly into four groups,they are group PLGA,PLGA-SN,PLGA-SN-Fat and Autograft.The 10 mm sciatic nerve defects in the four groups were rehabilitated with empty PLGA nerve scaffolds,PLGA nerve scaffolds with autologous minced nerve tissue,PLGA nerve scaffolds combined with autologous fat and autologous minced nerve tissue,autograft respectively.After 12 weeks,detection and analysis involved general observation,HE staining,Masson staining,immunofluorescence S100 and toluidine blue staining of the regenerated nerves to assess the nerve regeneration.Electromyogram,HE and Masson staining of the gastrocnemius muscle were conducted to evaluate the targeted muscle reinnervation.Results:We achieved highly purified Schwann cells,fibroblasts and ADSCs and our results demonstrated that Schwann cells,fibroblasts and ADSCs were able to adhere to and extend on fibre yarn avtively.CCK8 assay results showed that the extracts from the PLGA nerve conduit had no cytotoxicity to Schwann cells,fibroblasts and ADSCs.Besides,general observation,detection and analysis of electromyogram and histomorphometric indicate that the PLGA scaffolds containing autologous fat and autologous minced nerve tissue can rehabilitate rats’ sciatic nerve defects effectively.Conclusions: PLGA nerve conduits containing autologous fat and autologous minced nerve tissue can support,guide and enhance peripheral nerve regeneration,which makes contributions to the repair of peripheral nerve defects,providing a new method for peripheral nerve rehabilitation.更多还原