光化学法建立视网膜分支静脉阻塞大鼠模型及相关研究

摘要

目的:观察光化学法构建视网膜分支静脉阻塞(branch retinal vein occlusion, BRVO)大鼠模型的自然病程和不良事件.方法:选取30只SD(Sprague Dawley)大鼠尾静脉注射孟加拉红1min后,532nm激光(80mW,100μm,100ms)于视盘颞侧视网膜静脉二级分叉处进行单光点光凝50点建立BRVO模型.分别于1、3、5、7、10、14和21d检测全视野视网膜电图(electroretinogram,ERG)、荧光素眼底血管造影(fundus fluorescein angiography, FFA)和相干光断层扫描(optical coherence tomography,OCT).于1、5和21d各时间点随机处死2只大鼠行HE病理和血管内皮生长因子-a(vascular endothelial growth factor-a,VEGF-a)免疫组化染色.结果:光凝后,3只大鼠死亡,3只严重出血导致视网膜大部分脱离,1只出现视网膜凹陷,1只白内障.FFA和眼底(荧光)彩照发现BRVO大鼠模型造模成功率为73%(22/30),1d时近端变粗,远端变细,3~7d光凝血管完全再通.ERG示光凝1d后暗适应3.0反应b波降低至正常眼的0.694±0.042倍,5 ~7d下降至最低点约为正常眼0.487 ± 0.064倍,之后开始上升,21d 上升至初始值0.708±0.0465倍.OCT和HE病理切片分别于在体和离体水平发现第1d视网膜节细胞层和外核层水肿,3 ~5d水肿消失且激光光凝点视网膜附近250μm外核层开始凋亡变薄,到21d外核层变薄只剩3~4层细胞.免疫组化发现激光光凝部位VEGF-a第1d表达水平大于光凝前,第5d光凝处VEGF-a表达量无明显差异,21d光凝处VEGF-a表达略低于光凝前.结论:激光光凝制作 BRVO 模型是一种切实可行的方法,其疾病的演变和发展可以部分模拟人体BRVO的进程.同时由于造模成功率较低以及激光光凝相关并发症较多,在实际使用中有待进一步改进方法.%AIM:To investigate the natural course and adverse event of branch retinal vein occlusion (BRVO) rat model induced by laser photochemical method. METHODS: Thirty SD (Sprague Dawley) rats were administrated Bangladesh via tail vein. Then 532nm laser (80mW, 100μ m and 100ms) was performed on retinal vein secondary bifurcation of bitamporal optic disk for 50 spots. Electroretinogram (ERG), fundus fluorescein angiography ( FFA), optical coherence tomography (OCT) and fundus (fluorescein) photograph were applied on 1, 3, 5, 7, 10, 14 and 21d after BRVO model constructed. Two rats were sacrificed, respectively, on 1, 5 and 21d after photocoagulation to carry on HE (Hematoxylin - Eosin stain) and VEGF - α (vascular endothelial growth factor - α) immumohistochemical staining. RESULTS: There were three rats died, three rats with severe retinal detachment for excessive bleeding,one rat with retinal sunken, and one rat with cataract. FFA and fundus ( fluorescein) photograph showed that the successful BRVO rat model was 73% (22/30). It was found that the near-end photocoagulation vein became coarse, far - end became diminution on 1d and the photocoagulation vein total recanalization was on 3-7d. ERG showed the amplification of b wave (dark -adaptation 3.0 response) decreased to 0.694士0.042 of control eyes and on 5-7d decreased to rock bottom about 0.487士0.064 of control eyes. Then it increased Aii the time to 0.708士0.0465 of control eyes on 21d. OCT and HE staining found that retinal ganglion cells and outer nuclear layer became edema on 1d in vivo and in vitro.It was observed that the thickness of retina on photocoagulation vein (0μ m or 250μ m) decreased from 5d and there were 3-4 layer cells in ONL on 21d. The expression of VEGF-α at injured site were significantly more than control eyes on 1d and there were no significant difference on 5d;But the expression of VEGF-α were slightly less than control eyes on 21d. CONCLUSION: Photochemical method was a feasible method to establish BRVO rat model. The evolution and development of the BRVO model could partly mimic human BRVO phenomenon. At the same time, it should be improved to increase the successful model rate.