全反式维甲酸干预鼠胚神经干细胞联合胶质细胞源性神经营养因子及硫酸软骨素酶ABC移植修复大鼠脊髓损伤的研究  

TRANSPLANTATION OF NEURAL STEM CELLS INDUCED BY ALL-TRANSRETINOIC ACID COMBINED WITH GLIAL CELL LINE DERIVED NEUROTROPHIC FACTOR AND CHONDROITINASE ABC FOR REPAIRING SPINAL CORD INJURY OF RATS

作　　者：廖烨晖 钟德君[1] 康敏[2] 姚帅辉[1] 张毅[1] 余永涛[1] 

机构地区：[1]泸州医学院附属医院脊柱外科,四川泸州646000 [2]泸州医学院附属医院消化内科,四川泸州646000

出　　处：《中国修复重建外科杂志》2015年第8期1009-1015,共7页Chinese Journal of Reparative and Reconstructive Surgery

基　　金：四川省卫生厅科研资助项目(2008-527-08 0178);泸州市重点科技计划资助项目(2008-23-5)~~

摘　　要：目的探讨全反式维甲酸(all-trans-retinoic acid,ATRA)干预培养的鼠胚神经干细胞(neural stem cells,NSCs)联合胶质细胞源性神经营养因子(glial cell line derived neur otrophic factor,GDNF)、硫酸软骨素酶ABC(chondroitinase ABC,Ch ABC)移植治疗大鼠脊髓损伤的效果。方法取健康成年雌性SD大鼠60只,体重200~250 g,随机分为5组(n=12),分别为假手术组(A组)、损伤对照组(B组)、NSCs+GDNF移植组(C组)、NSCs+Ch ABC移植组(D组)、NSCs+GDNF+Ch ABC移植组(E组)。B^E组于T10平面横断脊髓建立脊髓全横断损伤模型,A组仅显露硬脊膜但不损伤脊髓。于术后第8天将Brd U标记的ATRA干预培养的鼠胚NSCs移植至C、D、E组大鼠,第8~14天C^E组每天对应给予10μL GDNF、10μL Ch A BC,A、B组给予等量生理盐水。术后观察大鼠一般情况;于术前1 d、术后7 d及移植后1、2、5、8周采用BBB评分评估大鼠双后肢运动功能,采用体感诱发电位(somatosensory evoked potentials,SEP)评估神经传导功能。移植8周处死各组大鼠,取移植节段脊髓行HE染色和免疫荧光染色观察。结果造模术后共5只大鼠死亡,均补充。造模术后各时间点,B^E组大鼠BBB评分均较A组降低,SEP潜伏期均较A组延长,差异有统计学意义(P<0.05);造模术后7 d、移植后1周时,B^E组组间BBB评分、SEP潜伏期比较,差异无统计学意义(P>0.05);移植2、5、8周,C^E组大鼠双后肢功能逐步恢复,各时间点BBB评分均高于B组,SEP潜伏期均短于B组,差异有统计学意义(P<0.05);移植5、8周,E组BBB评分高于C、D组,SEP潜伏期短于C、D组,差异有统计学意义(P<0.05)。HE染色示,A组灰、白质分界清楚,细胞排列规则;B组损伤区血管形态欠完整,细胞排列紊乱,可见囊腔及胶质瘢痕形成;C^E组细胞增生明显,坏死囊腔较B组小。免疫组织荧光染色示,A、B组未见明显Brd U标记阳性细胞;C、D、E组Brd U阳性细胞胞体呈橘红色,E组阳性细胞多于C、D组(P<0.05)。C^E组神�Objective To observe the effect of transplantation of neural stem cells（NSCs） induced by alltrans-retinoic acid（ATRA） combined with glial cell line derived neurotrophic factor（GDNF） and chondroitinase ABC（Ch ABC） on the neurological functional recovery of injured spinal cord in Sprague Dawley（SD） rats. Methods Sixty adult SD female rats, weighing 200-250 g, were randomly divided into 5 groups（n=12）： sham operation group（group A）, SCI model group（group B）, NSCs＋GDNF treatment group（group C）, NSCs＋Ch ABC treatment group（group D）, and NSCs＋GDNF＋Ch ABC treatment group（group E）. T10 segmental transversal injury model of the spinal cord was established except group A. NSCs induced by ATRA and marked with Brd U were injected into the site of injury at 8 days after operation in groups C-E. Groups C-E were treated with GDNF, Ch ABC, and GDNF＋Ch ABC respectively at 8-14 days after operation; and group A and B were treated with the same amount of saline solution. Basso Beattie Bresnahan（BBB） score and somatosensory evoked potentials（SEP） test were used to study the functional improvement at 1 day before remodeling, 7 days after remodeling, and at 1, 2, 5, and 8 weeks after transplantation. Immunofluorescence staining and HE staining were performed to observe the cells survival and differentiation in the spinal cord. Results Five mouse died but another rats were added. At each time point after modeling, BBB score of groups B, C, D, and E was significantly lower than that of group A, and SEP latent period was significantly longer than that of group A（P0.05）, but no difference was found among groups B, C, D, and E at 7 days after remodeling and 1 week after transplantation（P0.05）. BBB score of groups C, D, and E was significantly higher than that of group B, and SEP latent period was significantly shorter than that of group B at 2, 5, and 8 weeks after transplantation（P0.05）; group E had higher BBB score and shorter SEP latent period than groups C and

关 键 词：神经干细胞 胶质细胞源性神经营养因子 硫酸软骨素酶ABC 脊髓损伤 大鼠 

分 类 号：R651.2[医药卫生—外科学]