复合雪旺细胞的神经组织工程材料联合脉冲电磁场促进大鼠坐骨神经缺损的再生  

Schwann cells loaded neural tissue engineering scaffold combined pulsed electromagnetic field in vivo promote sciatic nerve regeneration in rats

作　　者：刘鐘阳[1] 刘靓[1] 黄景辉[1] 黄亮亮[1] 朱澍[1] 孙振[1] 权鑫[1] 杨亚锋[1] 马腾[1] 罗卓荆[1] 

机构地区：[1]第四军医大学西京医院骨科,西安710032

出　　处：《中华骨科杂志》2016年第8期465-478,共14页Chinese Journal of Orthopaedics

基　　金：国家自然科学基金面上项目（30973052）;国家自然科学基金青年项目（81201389）

摘　　要：目的观察在适宜脉冲电磁场（pulsed magnetic field，PMF）刺激下，复合雪旺细胞（Schwann cells，SCs）的神经导管对大鼠坐骨神经12mm缺损的修复效果。方法将SD雄性大鼠随机分为5组：自体神经组（Autograft group）、神经导管组（Scaffold group）、神经导管联合脉冲电磁场组（Scaffold＋PMF group）、复合雪旺细胞的神经导管组（Scaffold＋SCs group）和复合雪旺细胞的神经导管联合脉冲电磁场组（Scaffold＋SCs＋PMF group）。采用Ⅰ型胶原与壳聚糖在冷凝条件下制备胶原-壳聚糖神经导管，将雪旺细胞与神经导管复合构建组织工程神经材料，移植修复大鼠12mm坐骨神经缺损，术后给予每日4h的适宜脉冲电磁场（50Hz，2mT）刺激。移植术后第3、7和14天观察雪旺细胞存活情况，术后4、8和12周分别行功能学和形态学检测其修复疗效，并于术后1和3周进行神经营养因子RT-PCR检测。结果术后第3、7和14天，移植入体内的雪旺细胞活性好，Scaffold＋SCs＋PMF组术后第3、7和14天雪旺细胞活细胞百分比分别为85．28％＋2．47％，88．41％±1．39％，92．73％＋2．36％，明显高于Scaffold＋SCs组。术后12周，透射电镜观察证实Scaffold＋SCs＋PMF组有大量再生神经纤维成功长人神经导管内，其再生轴突总面积、有髓纤维数量、有髓纤维平均直径和有髓纤维髓鞘化程度与Autograft组接近，优于其他组。术后12周，腓肠肌靶位HE染色结果显示Scaffold＋SCs＋PMF组肌纤维平均面积为84．95％±3．92％，与Autograft组相近（86．52％±4．38％），优于其他组。荧光金逆行示踪结果显示，术后4、8和12周Scaffold＋SCs＋PMF组中标记阳性的运动神经元数目和感觉神经元数目与Autograft组相近，均优于其他组。神经电生理检测结果显示术后4、8和12周Seaffold＋SCs＋PMF组复合肌肉动作电位的波幅、潜伏期和神经传导速度与Autograft组相�Objective Under proper pulsed electromagnetic field （PMF, 50 Hz, 2 roT, 4 h） exposure, to observe the regenerative effects of Schwann cells （SCs） loaded nerve scaffold bridging 12 mm sciatic nerve defects in rats. Methods SD male rats were divided randomly into five groups as follows： Autograft group, Scaffold group, Seaffold＋PMF group, Scaffold＋SCs group, and Scaffold＋SCs＋PMF group. Then, the eollagen-chitosan nerve scaffolds were fabricated by I type collagen and chitosan in condensation, and SCs were loaded into the nerve scaffold to form neural tissue engineering material. Then, the 12 mm sciatic nerve defects of rats were bridged using the material. After surgery, the rats were exposed under PMF 4 hours every day. Three, seven and fourteen days after surgery, the survival of SCs in nerve scaffold was tested. Four, eight and twelve weeks after surgery, the neurologic function recovery and morphometrie analysis were performed. Meanwhile, RT-PCR test of neurotrophic factors was analyzed one and three weeks after operation. Results Three, 7, and 14 days after surgery, SCs transplanted into scaffolds were in good condition, the number of Live-labeled SCs in nerve scaffold under PMF were 85.28%±2.47%, 88.41%±1.39%, and 92.73%± 2.36%, which were significantly higher than that without PMF, respectively. Twelve weeks after surgery, Scaffold＋SCs＋PMF group was observed massive axons regenerated into scaffolds by TEM, and the total area of regenerated axons, the total number of myelinated axons, the mean diameter of the myelinated axons, and the degree of myelination （G-ratio） were in the similar range between Scaffold＋SCs＋PMF group and Autograft group, which were significantly higher than those in the remaining groups. Twelve weeks after surgery, the morphological analysis of gastrocnemius muscles showed that the percentage of muscle fiber area in the Scaffold＋ SCs＋PMF group was 84.95%±3.92%, which was in a similar range of Autograft group （86.52%±4.38%）, and was significantly

关 键 词：坐骨神经 许旺细胞 组织工程 电磁场 

分 类 号：R745.4[医药卫生—神经病学与精神病学] R318.08[医药卫生—临床医学]