腺相关病毒介导的脑源性神经营养因子对DBA／2J小鼠视网膜神经节细胞的保护作用  

Neuroprotection of adenoassociated vira-mediated brain-derived neurotrophic factor on retinal ganglion cells in DBA/2J mice with glaucoma

作　　者：黄楚开[1] 张铭志[1] 刘丽芳[1] 岑令平[1] 韩森森 

机构地区：[1]汕头大学·香港中文大学联合汕头国际眼科中心,515041

出　　处：《中华实验眼科杂志》2011年第4期314-318,共5页Chinese Journal Of Experimental Ophthalmology

基　　金：国家自然科学基金丽上项目（30672282）、2009年汕头市科技计划项目（E200900184）

摘　　要：背景神经营养因子的缺乏与青光眼的视神经损害密切相关。外源性神经营养因子的补充具有短暂的保护作用。腺相关病毒（AAV）介导的神经营养因子可在眼内长期表达，但是否对青光眼动物的视神经具有持久的保护作用有待研究。目的评估AAV介导的脑源性神经营养因子（BDNF）基因在DBA／2J小鼠眼内的表达及其对视网膜神经节细胞（RGCs）的保护作用。方法健康清洁级DBA／2J小鼠10只从4月龄起，每月使用Tonolab眼压计测量眼压。6月龄时左眼玻璃体腔内注射AAV介导的BDNF和绿色荧光蛋白（GFP）基因（AAV—BDNF—GFP）1μ1，右眼注射等量的生理盐水作为对照。注射后3个月心脏灌流后取出视网膜，荧光显微镜下观察GFP在视网膜中的表达，免疫组织化学法计算存活的RGCs数目并进行比较。结果DBA／2J小鼠4月龄时AAV—BDNF—GFP眼眼压平均为11．90mmHg，对照眼眼压为11．40mmHg。实验眼与对照眼眼压5月龄时均开始升高，8月龄时达到高峰。从4月龄到9月龄，实验组和对照组眼压比较差异无统计学意义（t=-1．78～0．61，P=0．11—0．90）。玻璃体腔注射AAV—BDNF—GFP3月龄后视网膜可以观察到GFP阳性细胞，转染率为46．33％±8．08％。AAV—BDNF—GFP组实验眼的平均RGCs的密度为（3168．13±1319．33）mm^2，对照眼为（2024．81±796．38）mm^2，差异有统计学意义（t=2．75，P=0．02）。结论AAV介导的BDNF对DBA／2J小鼠RGCs有保护作用。Background Deficiency of neurotrophic factor is associated with the damage of optic nerve in glaucoma. Reaserches showed that eetopically applied neurotrophic factor has a transient neuroprotective effect in glaucoma model, and the viral expression of adeno-associated neurotrophic factor may provide long-term supplementation of neurotrophie factor and neuroprotection in tissues. Objective The present study was to investigate the neuroproteetive effect of adeno-associated viral （AAV）-mediated brain-derived neurotrophic factor （BDNF） on retinal ganglion cells （RGCs） in DBA/2J mice with experimental glaucoma. Methods 10 clean DBA/2J mice were administered intravitreal injection with 1 mieroliter of AAV-BDNF-GFP in the left eyes at the age of 6 months,and the right eyes were injected with the same volume of saline solution as control. Intraoeular pressure （IOP） was measured with Tonolab in the mice every month. Retinas were obtained after 3 months for the investigation of GFP expression in RGCs using fluorescence microscopy. Immunohistoehemistry was performed by applying TUJ1 and Cy3 antibodies to identify surviving RGCs. Results The IOP of DBA/2J mice were 11.90 mmHg and 11.40 mmHg in the right eyes and left eyes, respectively, at 4 months. The IOP of DBA/2J mice began to rise at 5 months and reached its peak in 8 month-old mice. There was no statistically significant difference in IOPs betweenthe right eyes and the left eyes from 4 month- through 9 month-old mice（ t =- 1.78-0.61 ,P= O. 11-0.90）. Three months after intravitreal injection of AAV-BDNF-GFP, GFP was positively expressed in RGCs of retinas with the expression rate of 46.33%±8.08%. The over-expression of BDNF led to more RGCs survival than the control eyes （3168. 13±1319.33/mm2 vs 2024.81±796.38/mm2,t=2.75,P=0. 02）. Conclusion These data suggest that BDNF can exert a protective effect in DBA/2J glaucoma mice.

关 键 词：青光眼 眼压 脑源性神经营养因子 视网膜神经节细胞 

分 类 号：R77[医药卫生—眼科]