硫酸软骨素蛋白多糖酶结合骨髓基质干细胞修复亚急性大鼠脊髓挤压伤的实验研究  

Repair of subacute spinal cord crush injury by bone marrow stromal cell transplantation and chondroitinase ABC microinjection in adult rats

作　　者：张纯[1] 姚聪[2] 贺西京[1] 李浩鹏[1] 

机构地区：[1]西安交通大学医学院第二附属医院骨二科,陕西西安710004 [2]西安交通大学医学院第二附属医院肿瘤外科,陕西西安710004

出　　处：《南方医科大学学报》2010年第9期2030-2035,共6页Journal of Southern Medical University

基　　金：教育部博士点基金(20050698043);吴阶平医学基金

摘　　要：目的分析大鼠亚急性脊髓损伤(SCI)后联合应用骨髓基质干细胞移植和硫酸软骨素酶ABC的可行性,并探讨其相互作用。方法取新生SD大鼠的股骨和胫骨分离培养MSCs,调整细胞浓度为2.0×105/cm2备用。取体质量220～240g健康成年雄性SD大鼠24只制备SCI动物模型,并随机分为4组,A组对照组,B、C、D组每组6只,分别于SCI损伤区注射5μLMSCs;5μL硫酸软骨素酶(ChABC)和ChABC+MSCs。采用BBB评分法评价伤后1、2、3、7、14d大鼠双后肢的运动功能。伤后14d取材行HE染色并计算损伤区最大横径和坏死总面积;行GFAP-CS56、GFAP-GAP-43和GFAP-NF160免疫荧光染色评价神经纤维束的再生情况。结果损伤后算14天BBB评分结果示,A组与B、C、D组比较差异均有统计学意义(P<0.05),B、D组及C、D组间比较差异均有统计学意义(P<0.05),B、C组间无显著性差异。HE染色显示14天时各组均有空洞形成,伤后坏死总面积比较,B、C、D组与A组比较均有显著性差异(P<0.05),B、C、D三组之间D组与前两组比较存在显著性差异(P<0.05)。免疫组织化学染色示,A组GFAP反应最强,损伤区空洞明显可见且范围大于其他三组,D组介于B、C组之间;B、C和D组的GAP-43表达增多,且以D组为最;D组损伤区内的神经纤维通过明显多于其他三组。结论硫酸软骨素酶ABC联合骨髓基质干细胞移植对脊髓损伤有一定修复作用。Objective To investigate the possibility of repairing spinal cord injury by bone marrow stromal cell（MSC） transplantation and microinjection of chondroitinase ABC（ChABC） in adult rats.Methods MSCs isolated from the femoral and tibial bones of new-born Wistar rats were cultured and the cell density was adjusted to 1×105/ μl before transplantation.SD rats with T8 spinal cord crush injury were divided into 4 groups,namely group A（control） and groups B,C and D with injections of the MSCs,ChABC and MSCs＋ChABC at the injury site,respectively.At 0 h,1 day,2 days,3 days,1 week,and 2 weeks after the injury,the BBB score system was used to evaluate the motion function.At 14 days after the injury,the maximal transverse diameter and actual area of necrosis were evaluated on HE stained sections.GFAP-CS56,GFAP-GAP43 and GFAP-NF160 immunofluorescence double labeling staining were carried out to evaluate the regeneration of the nerve fibers.Results At the 14th day after the injury,BBB scores showed significant differences between group A and groups B,C and D（P〈0.05）,between the groups B and D and between groups C and D,butnot between groups B and C.HE staining showed cavity formation at the injury site in each group,with significant differences between group A and groups B,C and D and also between the latter3 groups.Immunofluorescence staining revealed more intense expression of GFAP in group A than in the other groups with apparent cavity formation at the lesion site,which was only moderate in groups B and C.The expression of GAP-43 was more intense in group D than in groups B and C.The expression of NF-160 was more intense in group D than in the other3 groups.Conclusion The strategy of MSC transplantation combined with ChABC can be effective forrepairing spinal cord injury in adult rats.

关 键 词：硫酸软骨素蛋白多糖酶 骨髓基质干细胞 脊髓损伤 

分 类 号：R651.2[医药卫生—外科学]