色素上皮衍生因子对氧诱导视网膜神经节细胞凋亡及Bcl-2表达的影响  

Effects of PEDF on apoptosis of retinal ganglion cells and expression of Bcl-2 in retina under hypoxia

作　　者：王亚娜[1] 高莎[1] 沈玺[1] 

机构地区：[1]上海交通大学医学院附属瑞金医院眼科,上海市200025

出　　处：《眼科新进展》2014年第6期501-505,共5页Recent Advances in Ophthalmology

基　　金：国家自然科学基金资助(编号:81170860);上海市科委自然基金(编号:112R1422000)~~

摘　　要：目的探讨色素上皮衍生因子(pigment epithelium-derived factor,PEDF)对氧诱导缺血性视网膜病变(oxygen-induced ischemic retinopathy,OIR)小鼠视网膜神经节细胞(retinal ganglion cell,RGC)凋亡的抑制作用及其可能的机制。方法选择7 d龄C57BL/6J小鼠140只,随机分为正常对照组、OIR模型组、PBS治疗对照组和PEDF药物治疗组。选择右眼为实验眼,将除正常对照组外的所有小鼠建立OIR模型。PEDF药物治疗组分别于小鼠12 d龄和14 d龄给予两次玻璃体内注射PEDF(2μg·μL-1)各1μL,PBS治疗对照组玻璃体内注射等量的PBS(10 mmol·L-1,pH 7.4)。于17 d龄处死小鼠,各组随机抽取5只,摘除右眼球,采用TUNEL法检测视网膜细胞凋亡情况以及免疫荧光染色法检测Bcl-2蛋白在小鼠视网膜中的表达情况;其余小鼠过量麻醉处死后取出视网膜,采用Western blot检测Bcl-2蛋白以及Real-time RT-PCR检测bcl-2 mRNA的表达。结果视网膜细胞的凋亡主要在RGC层,OIR模型组小鼠RGC层细胞凋亡率为(55.044±14.538)%,明显高于正常对照组的(3.317±1.657)%,差异有统计学意义(P<0.01);PBS治疗对照组细胞凋亡率为(55.852±4.490)%,明显高于PEDF药物治疗组的(5.897±2.079)%,差异有统计学意义(P<0.01);正常对照组与PEDF药物治疗组细胞凋亡率比较,差异无统计学意义(P>0.05)。免疫荧光显示,OIR模型组与PBS治疗对照组Bcl-2蛋白阳性染色较正常对照组明显减少,PEDF药物治疗组阳性染色较PBS治疗对照组增多。Western blot以及RT-PCR结果显示,OIR模型组Bcl-2蛋白和mRNA的表达量显著下降,分别为0.386 9±0.071 3和0.694 2±0.235 2,与正常对照组的0.778 7±0.097 4和1.000 0比较,差异均有统计学意义(均为P<0.05);PEDF药物治疗组Bcl-2蛋白和mRNA的表达量分别为0.610 1±0.109 5和0.818 3±0.308 0,与PBS治疗对照组的0.284 8±0.153 6和0.481 0±0.100 8比较,差异均有统计学意义(均为P<0.05);PEDF药物治疗组Bcl-2蛋白和mRNA的表达量较正常对照�Objective To study the effects of pigment epithelium derived factor （PEDF） on retinal ganglion cell（RGC） apoptosis and expression of Bcl-2 in mice with oxygen-induced ischemic retinopathy（OIR） and its possible mechanism. Methods A total of 140 postnatal day（ P）7 C57BL/5 mice were randomly divided into normal control group, OIR model group, PBS treatment group and PEDF treatment group, 35 mice in each group. OIR mice model were established in right eye of each mouse except normal control group. At P12 and P14 ,respectively,mice in PEDF treatment group were intravitreal injected of recombinant human PEDF（ 1 μL,2 μg · μL^-1） ,while mice in PBS treatment control group were injected the same volume of PBS（ 1 μL, 10 mmol · L^-1, pHT. 4）. All mice were euthanized at P17,5 mice in each group were randomly chosen and eyes were isolated, the retinal cell apoptosis was assessed bJ TUNEL staining and the expression of Bcl-2 protein in retina was detected by immunofluorescence. All other mice were euthanized and retinal specimens were prepared for Bcl-2 protein and mRNA analysis by Western blot and RT-PCR. Results Retinal cell apoptosis was mainly localized in RGC layer. The apoptosis rate of RGC was （55.044 ± 14. 538 ）% in the OIR model group and（3. 317 ±1. 557）% in the normal control group,there was significant difference （ P 〈 0.01 ） ; Compared with PEDF treatment group （ 5. 897 ±2.079 ） % , the apoptosis rate of RGC in PBS treatment group（55. 852 ± 4.490）% was significantly higher,there was significant difference（P 〈0.01 ） ;There was no statistical difference in apoptosis rate of RGC between normal control group and PEDF treatment group （P 〉 0. 05 ）. Immunofluorescence assay showed Bcl-2 protein expression in OIR model group and PBS treatment group were lower than that in normal control group, while PEDF treatment group was higher than PBS treatment group. The specific expression of Bcl-2 protein and mRNA were 0.386 9 ± 0.071 3 and 0.694 2 ± 0.235 2 in

关 键 词：氧诱导缺血性视网膜病变 色素上皮衍生因子 视网膜神经节细胞 Bcl-2 细胞凋亡 

分 类 号：R775[医药卫生—眼科]