脐血间充质干细胞诱导分化为类雪旺细胞修复大鼠坐骨神经损伤的实验研究  

AN EXPERIMENTAL STUDY ON REPAIR OF SCIATIC NERVE INJURY BY Schwann-LIKE CELLS DERIVED FROM UMBILICAL CORD BLOOD MESENCHYMAL STEM CELLS

作　　者：王玺[1] 王胜[2,3] 肖玉周[3,4] 

机构地区：[1]蚌埠市第三人民医院骨科,安徽蚌埠233000 [2]蚌埠医学院第二附属医院骨科 [3]组织移植安徽省重点实验室 [4]蚌埠医学院第一附属医院骨科

出　　处：《中国修复重建外科杂志》2015年第2期213-220,共8页Chinese Journal of Reparative and Reconstructive Surgery

摘　　要：目的评价将人脐血间充质干细胞（human umbilical cord blood mesenchymal stem cells,h UCBMSCs）来源的类雪旺细胞（Schwann cells,SCs）作为种子细胞,修复大鼠坐骨神经15 mm缺损的效果,为h UCBMSCs应用于临床治疗周围神经缺损提供实验依据。方法 SPF级3月龄雄性SD大鼠45只,体重200-250 g。取新生儿脐带血,淋巴细胞分离液复合高分子量羟乙基淀粉分离培养h UCBMSCs并鉴定;取第3代h UCBMSCs,采用改良化学诱导法联合细胞因子方法诱导分化培养类SCs并鉴定。取15只SD大鼠坐骨神经,采用液氮反复冻融振荡洗涤法制备去细胞神经基膜管作为支架材料;将密度1×10^7个/m L的类SCs细胞悬液多点注射至该支架内复合培养7 d构建组织工程神经。取SD大鼠30只制备长约15 mm坐骨神经缺损动物模型,根据缺损神经修复方法不同,实验分为A、B、C 3组（n=10）,A组采用组织工程神经缝合,B组采用未复合类SCs的去细胞神经基膜管缝合,C组采用自体坐骨神经原位缝合。术后行大体观察、坐骨神经功能指数（sciatic function index,SFI）测定、神经电生理功能检测、腓肠肌湿重测定、Masson染色评价神经修复情况。结果分离培养的h UCBMSCs高表达MSCs表面标志;诱导培养后类SCs经免疫细胞化学染色检测示神经胶质细胞标志物S100b、胶质纤维酸性蛋白、P75表达呈阳性。术后8周,大体观察示A组组织工程神经管壁无坏死及液化,周围轻度粘连,吻合处连续性较好;B组支架外观与A组相似;C组自体神经周围粘连较A、B组轻,吻合口光滑,无明显膨大,颜色与正常神经相似。各组大鼠术后SFI随时间延长呈逐渐降低趋势,C组SFI恢复优于A、B组,A组优于B组（P〈0.05）。术后各组大鼠远端吻合口处均可检测到神经复合动作电位,波幅及传导速度C组均优于A、B组,A组优于B组（P〈0.05）。术后各组大鼠实验侧小腿腓肠肌与健侧相比,均发生不同程度萎�Objective To evaluate the effect of using Schwann-like cells derived from human umbilical cord blood mesenchymal stem cells（h UCBMSCs） as the seed cells to repair large sciatic nerve defect in rats so as to provide the experimental evidence for clinical application of h UCBMSCs. Methods Fourty-five male Sprague Dawley（SD） rats in SPF grade, weighing 200-250 g, were selected. The h UCBMSCs were harvested and cultured from umbilical cord blood using lymphocyte separating and high molecular weight hydroxyethyl starch, and then was identified. The h UCBMSCs of 3rd generation were induced to Schwann-like cells, and then was identified by chemical derivatization combined with cytokine. The acellular nerve basal membrane conduit was prepared as scaffold material by the sciatic nerve of SD rats through repeated freezing, thawing, and washing. The tissue engineered nerve was prepared after 7 days of culturing Schwann-like cells（1×10^7 cells/m L） on the acellular nerve basal membrane conduit using the multi-point injection. The 15 mm sciatic nerve defect model was established in 30 male SD rats, which were randomly divided into 3 groups（10 rats each group）. Defect was repaired with tissue engineered nerve in group A, with acellular nerve basal membrane conduit in group B, and with autologous sciatic nerve in group C. The nerve repair was evaluated through general observation, sciatic function index（SFI）, nerve electrophysiology, weight of gastrocnemius muscle, and Masson staining after operation. Results The h UCBMSCs showed higher expression of surface markers of mesenchymal stem cells, and Schwann-like cells showed positive expression of glia cell specific markers such as S100 b, glial fibrillary acidic protein, and P75. At 8 weeks after operation, the acellular nerve basal membrane conduit had no necrosis and liquefaction, with mild adhesion, soft texture, and good continuity at nerve anastomosis site in group A; group B had similar appearance to group A; adhesion of group C was milder than that of

关 键 词：组织工程神经 人脐血间充质干细胞 类雪旺细胞 去细胞神经基膜管 神经修复 

分 类 号：R651.3[医药卫生—外科学]