hBDNF-GFP基因转染神经干细胞对视神经损伤大鼠视网膜BDNF表达的影响  

Effects of hBDNF-GFP gene-transfected neural stem cell transplantation on BDNF expression in the retina of rats following optic nerve crush injury

作　　者：刘勇[1] 陈二涛[1] 冯东福[1] 潘栋超[1] 汪洋[2] 

机构地区：[1]上海交通大学医学院附属第三人民医院神经外科,上海交通大学医学院创伤医学研究所,201900上海 [2]复旦大学上海医学院解剖与组织胚胎学系

出　　处：《中华急诊医学杂志》2009年第8期841-845,共5页Chinese Journal of Emergency Medicine

基　　金：上海市教委自然科学基金重点项目（04BB23,07ZZ42）

摘　　要：目的探讨移植hBDNF-GFP基因转染神经干细胞后对视神经损伤大鼠视网膜BDNF表达的影响。方法①78只SD大鼠随机分为正常对照组（n=6）、视神经夹伤组（n=72）。视神经夹伤组动物行右侧视神经夹伤后随机平均分为三个亚组：分别于玻璃体腔内注射0．01mol／LPBS（PBS组），GFP基因转染神经干细胞（GFP-NSCs组）和hBDNF-GFP基因转染神经干细胞（hBDNF-GFP-NSCs组）。各组动物单纯暴露左侧视神经作为假损伤组。分别于移植后3d，7d，14d和28d处死动物取视网膜，ELISA法检测各组视网膜中BDNF含量。②另取6只视神经损伤大鼠移植hBDNF-GmNscs，分别于2周、4周和8周各处死2只大鼠，取眼球做冰冻切片观察移植细胞存活、迁移情况。③35只SD大鼠随机分为4组：正常对照组（n=5）、右眼视神经损伤NSCs治疗组（n=10）、GFP-NSCs治疗组（n：10）及hBDNF-GFP-NSCs治疗组（n=10）。④各组于术后4周和8周分别处死5只大鼠，Western-blot法检测视网膜组织中BDNF表达。结果①ELISA结果显示，正常对照组BDNF含量与假损伤组间比较差异无统计学意义（P〉0．05），移植手术后第3天时，三个损伤亚组视网膜匀浆上清中hBDNF含量明显增加（与假损伤组比较P〈0．05），而三组间差异无统计学意义（P〉0．05）；7d时，GFP-NSCs组与假损伤组比较差异具有统计学意义（P〈0．05），而PBS组、hBDNF-GFP-NSCs组与假损伤组比较差异无统计学意义（P〉0．05），PBS组与GFP-NSCs组间及hBDNF-GFP-NSCs组与GFP-NSCs组间差异具有统计学意义（P〈0．05）；移植第14天和28天时，PBS组和GFP-NSCs组中视网膜BDNF含量降低，而hBDNF-GFP-NSCs组中BDNF含量与其他三组比较明显升高，差异有统计学意义（P〈0．05）。②冰冻切片示移植后，hRDNF-GFP-Nscs可在宿主视网膜内存活，并逐渐迁移至视网膜各层。③Western-blot检测显示，各组内第4周和第8周BDNF表达�Objective To investigate the effects of human Rrain-derived neurotrophic factor-gamma fetoprotein （hBDNF-GFP） gene-transfected neural stem cell （NSC） transplantation on BDNF expressions in the retina of rats after optic nerve （ON） crush injury. Method ①Seventy-eight Sprague-Dawley （SD） rats were randomly assigned into a control group （ n =6） and ON crush group （ n = 72）. In the ON crush group, the right ON was crushed while the left NO was exposed as sham injury. Rats in the ON crush group were divided into three subgroups：PBS group （intravitreous injection of 0.01 mol/L phosphate buffered solution）; GFP group （intravitreous transplantation of GFP gene-transfected NSCs） ; and hBDNF-GFP group （intravitreous transplantation of hBDNF-GFP gene-transfected NSCs）. Rats were sacrificed 3, 7, 14 and 28 days after transplantation, and BDNF expressions in retinal homogenates was detected by using enzyme-linked imrnanosorbent assay （ELISA）.②The hBDNF- GFP-NSCs were transplanted intravitreous into six rats after ON crush injury. Following this, two rats were sacririced 2, 4 and 8 weeks after transplantation. The survival and location of NSCs in host retina were observed by frozen section analysis.③Adult SD rats were randomly divided into four groups： control group （n = 5）; NSC group （NSC transplantation, n = 10）; GFP-NSC group （GFP-NSC transplantation, n = 10）; and hBDNF-GFP- NSC group （hBDNF-GFP-NSC transplantation, n = 10）. Four and eight weeks after transplantation, five rats from every group were sacrificed. Western blot analysis was used to determine retinal BDNF expression. Results ① There was no significant difference in BDNF expression between the control group arid shanvinjury groups （ P 〉 0.05）. Three days after NSC transplantation, BDNF expression increased significantly in the three injured subgroups compared with the sham-injury group, （ P 〈 0.05）, whereas no significant inter-group differences in BDNF expressions among three injured

关 键 词：脑源性神经营养因子 干细胞 转染 视网膜 

分 类 号：R774.6[医药卫生—眼科] R741[医药卫生—临床医学]