嗅鞘细胞移植对脊髓损伤后损伤区髓鞘相关轴突生长抑制因子受体表达的影响

Effect of olfactory ensheathing cells transplantation on level of NgR in the injuried region after spinal cord injury

目的:观察嗅鞘细胞移植前后脊髓损伤区髓鞘相关轴突生长抑制因子受体(NgR)的变化,探讨嗅鞘细胞移植治疗脊髓损伤的有关机制。方法:实验于2006年9月至2007年5月在西安交通大学医学院环境与基因重点实验室完成。①实验动物:成年健康SD雄性大鼠40只,用随机数字表法分为正常组、模型组、嗅鞘细胞组和DF12对照组,每组10只。另取30只健康成年雄性SD大鼠作为嗅鞘细胞的来源。②实验方法:除正常组外,其余各组均建立全横切脊髓损伤模型。嗅鞘细胞组将原代培养12d的嗅鞘细胞悬液调整为(1×1011)个/L,在距损伤缘上下各1mm处分4点应用微量注射器注射,深度1.0mm,每处各注射1μl;DF12对照组同法每点注射等量DF12培养液;模型组、正常组不进行任何处理。③观察项目与方法:各组分别于移植后1、4、8周采用免疫组化技术动态检测脊髓损伤区NgR表达的变化。同时在移植后8周后行嗜银染色检测组织形态学变化。结果:①NgR表达的变化:正常组NgR表达的吸光度值明显低于其余3组(P<0.05)。嗅鞘细胞组于移植后1,4,8周脊髓损伤区NgR的表达均明显低于模型组和DF12对照组(P<0.05或P<0.01),而模型组和DF12对照组差异无统计学意义(P>0.05)。②组织形态学变化:嗅鞘细胞移植8周后除正常组外,其余各组均可见明显的神经纤维再生,但模型组与DF12对照组大部分纤维排列紊乱,再生纤维方向性较差;嗅鞘细胞组可见明显的新生轴突,且神经纤维跨越损伤部位修复脊髓损伤,无论在数量还是质量上均优于模型组及DF12对照组。结论:嗅鞘细胞移植可能通过降低脊髓损伤区NgR蛋白的表达,从而促进损伤脊髓的修复。

Objective：To observe the change of NgR in the region of spinal cord injury（SCI） before and after transplantation of olfactory ensheathing cells（OECs） and explore the relevant mechanism of OECs transplantation in treating SCI. Methods：The experiment was enforced from September 2006 to May 2007 in the Key Laboratory of Environment and Genes Related to Diseases of Ministry of Education of Xi＇an Jiaotong University. ①Forty adult SD rats were divided into normal group,model group,OECs group and DF12 control group with random digits table. Each group had 10 rats. In addition,30 healthy male SD rats were accepted as the material for cell culture. ②The rats in model group,OECs group and DF12 control group were cut off T10 spinal cord to establish SCI model. Then the primary culture OECs suspension[（1×1011）piece/ L] was injected into the region about 1 mm from the margin of injury （about 1.0 mm depth and every place with 1μl ） of OECs group and DF12 culture fluid was injected into DF12 control group in a same dose and method. The normal group and model group did not any disposal. ③In the 1st,4th and 8th weeks after injection,immunohistochemistry were used to evaluate the results of OECs transplantation. And the histological silver staining was applied to evaluate the effect of OECs transplantation in the 8th week after injection. Results：All 40 rats survived and were accepted for analysis. ①At the 1st,4th and 8th weeks after injection,immunohistochemistry and western blot results showed the level of NgR in normal group were obviously lower than other groups（P0.05）,and in OECs group was obviously lower than that of DF12 control and model group（P0.05 or P0.01）. There was no significant difference between model group and DF12 control group. ②At the 8th week after injection,histological silver staining showed that nerve fibre obviously regenerated in model group,OECs group and DF12 control group,but the regenerated fiber in model group and DF12 control group arranged in disorder compared with that of OECs group. Conclusion：OECs transplantation may enhance the repair of spinal cord injury by depressing the level of NgR.