异体移植_____去细胞神经；_异体移植；

中国医药生物协会骨库分会主审化学去细胞异体神经修复神经缺损长度的实验研究

http://www.100md.com 《中国矫形外科杂志》 2006年第8期

异体移植,,去细胞神经；,异体移植；,再生,1材料与方法,2结果,3讨论,参考文献：

     摘要：[目的]研究神经长度是否影响化学去细胞神经的质量及化学去细胞异体神经修复犬神经缺损的适当长度范围。[方法]切取犬的坐骨神经全长，截取直径近似段长度12 cm，去除神经外膜的脂肪组织及血管，在50%Triton X100和50%脱氧胆酸钠中重复消化得到化学去细胞神经；部分神经用于组织学评价，将12 cm神经按顺序切割为6段，石蜡包埋制备切片，厚度5 μm。HE染色和Fastblue染色，观察去细胞程度、神经细胞外基质结构完整性、髓鞘染色强度，观察每段神经的差异。在体内实验中，用化学去细胞异体神经桥接犬的坐骨神经缺损，缺损长度分别为8、10 cm组，每组各有6条成年犬，随访时间12个月。观察动物肢体功能恢复、肌电图变化及再生神经组织学表现，包括HE染色、S100免疫组织化学染色、乙酰胆碱酯酶染色。[结果]去细胞神经全长各段在去细胞程度、结构完整性及残余髓鞘染色综合评价中无差异。体内移植实验结果显示8 cm去细胞异体神经移植组的犬在术后12个月踝关节可直立行走，而10 cm去细胞异体神经移植组犬12个月时踝关节不能直立行走。肌电图检查结果显示，两组动物的手术肢体均可记录到诱发的运动和感觉电位。肌电图波幅、时限及运动神经传导速度结果显示，8 cm移植组明显高于10 cm移植组(P<005)。组织学检查显示8 cm移植组再生的神经轴突通过移植段神经到达远侧的神经中，并与所支配的肌肉建立新的运动终板联系，坐骨神经支配的小腿三头肌中有大量新生的运动终板形成。在10 cm移植组有部分再生的神经纤维通过移植段神经进入远端的神经，小腿三头肌中新生的运动终板数量和大小明显低于8 cm移植组(P<005)。[结论]在化学去细胞神经的制备中，神经的长度对去细胞的质量无影响；化学去细胞异体神经修复神经缺损的长度若不超过8 cm可取得较满意的功能康复结果。

    关键词：去细胞神经；异体移植；再生

    Experimental study of repairable length of nerve defects with acellular nerve allografts∥SUN Mingxue, LU Shibi, TANG Jinshu, et al. Institute of Orthopedics, PLA General Hospital of Chinese, Beijing, 100853

    Abstract：[Objective]The purpose of this paper is to demonstrate whether the nerve length could affect the quality of acellular nerve and investigate the properly repairable distance of nerve defects with acellular nerve allografts. [Method]Fresh sciatic nerves were obtained from adult dogs and divided into 12 cm long segments. The nerve segments were decellularized via an improved chemical decelluarization treatment as following: Nerve segments were rinsed with cold sterile Ringer's solution and submergedin 5% Triton100 solution 12h ,and then soaked the nerve segments into 5% sodium deoxycholate for 12h. The treated nerve segments were washed in distilled water for 3h. This procedures were repeated once again. In vitro, the degrees of decellularization , demyelination and integrity of nerve fiber tubal of chemically extracted acellular nerves were observed with microscope and assessed by a score system. In vivo,the sciatic nerve of dogs on the right was exposed.In 8 cm grafted group (n=6),a 7 cm segment of sciatic nerve was removed from the midthigh level. In 10 cm grafted group (n=6) , a 9 cm segment of sciatic nerve was removed at the same level. The gaps were bridged with acellular nerve allografts by 8 cm and 10 cm long segments respectively. The followup period was 12 month postoperatively. Motor functional recovery of the right hind following allografting was examined by neurobehavioral, electrophysiological, histological and immunohistochemical assessment. [Result]There was no difference on the degrees of decellularization, demyelination and integrity of nerve fiber tubal among every fraction of the acellular nerve from the two ends to the central portion. In 8 cm grafted group , all survival dogs(n=5) were held upright with the affected hindlimb extended so that the body's weight was supported by the distal metatarsus and toes. In 10 cm grafted group, animals were failed to held upright with the affected hindlimb. Electrophysiological studies showed that elctromyographic activity was observed in both groups.After 12 month the conduction velocity was 321+51 m/s in 8 cm grafted animals and 183+6.0m/s in 10 cm grafted group. In normal animals, the conduction velocity was 1066+164 m/s. The conduction velocity in 10 cm grafted group was lower than 8 cm grafted(P<0.05) and normal nerve(P<001) significantly. Histological examination showed that axons sprouted from the proximal portion and reached the distal stump across the gap, but the myelinated axons was lower in 10 cm grafted animals than in 8 cm grafted. S100 staining indicated that Schwann cells presented in the central portion of both graft groups. In 8 cm grafted group, a great deal of Schwann cells arranged along the longitudinal directions. Motor endplates at the nervemuscle junction were observed in both grafted groups with acetylcholinesterase staining. The number and size of endplate in 10 cm grafted group was lower than 8 cm grafted(P<0.05) and normal nerve(P<0.01) significantly. [Conclusion]In the procedure of chemically extracted acellular nerve, the length of treated nerve does not affect the quality of acellular nerve. Eightcm gap may be the properly repairable extent of nerve defects with acellular nerve allografis. ......

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