摘要
目的 观察重组腺相关病毒（rAAV）-多聚嘧啶序列结合蛋白相关剪接因子（PSF）对氧诱导视网膜新生血管的抑制作用.方法 7日龄C57BL/6J小鼠18只,随机分为正常组、rAAV-PSF注射组、rAAV注射组,各组均6只.蛋白免疫印迹法（Western blot）检测各组小鼠视网膜组织中PSF的表达.7日龄C57BL/6J小鼠48只随机分为正常组、氧诱导视网膜病变（OIR）模型组、OIR rAAV-PSF治疗组、OIR rAAV治疗组,各组均为12只.除正常组外,其余各组小鼠与哺乳母鼠共同置于氧浓度为（75±2）％的氧箱内饲养5d后转移至正常环境中饲养5d,建立OIR模型.12日龄时,OIR rAAV-PSF治疗组小鼠玻璃体腔注射病毒滴度为5×10^13pfu/ml的rAAV-PSF重组载体2μI; OIR rAAV治疗组小鼠玻璃体腔注射相同病毒滴度的单纯rAAV载体2μl.OIR模型组小鼠出氧箱后不作任何处理.石蜡切片苏木精-伊红染色并计数突破视网膜内界膜的血管内皮细胞核;Western blot检测视网膜中VEGF蛋白表达含量.结果 正常组小鼠视网膜PSF蛋白表达与rAAV-PSF注射组比较,差异有统计学意义（F=16.05,P=0.001）,与rAAV注射组比较,差异无统计学意义（F=16.05,P=0.890）.正常组、OIR模型组、OIR rAAV-PSF治疗组、OIR rAAV治疗组突破内界膜的视网膜新生血管内皮细胞核数比较,正常组与OIR模型组差异有统计学意义（F=101.00,P=0.007）;rAAV-PSF治疗组与OIR模型组差异有统计学意义（F=101.00,P=0.002）;OIR rAAV治疗组与OIR模型组差异无统计学意义（F=101.00,P=0.550）.各组视网膜VEGF蛋白表达比较,正常组与OIR模型组差异有统计学意义（F=13.20,P=0.005）,OIR模型组与OIR rAAV-PSF治疗组差异有统计学意义（F=13.20,P=0.001）;OIR模型组与OIR rAAV组差异无统计学意义（F=13.20,P=0.071）.结论 rAAV-PSF玻璃体腔注射可以抑制OIR小鼠视网膜VEGF表达,从而抑制其新生血管生成.
Objective To observe the inhibitory effect of intraocular injection of recombinant adenoassociated virus-polypyrimidine tract-binding protein-associated splicing factor （rAAV-PSF） in oxygen induced retinopathy mice model.Methods Eighteen C57BL/6J mice were divided into 3 groups randomly,including normal group,rAAV-PSF injection group,rAAV injection group.Western blot analysis was applied to detect the transfection expression level of PSF.The other 48 C57BL/6J mice were randomly divided into 4 groups：normal group,ischemia-induced retinopathy （OIR） group,rAAV-PSF treated OIR group,rAAV treated OIR group,12 mice in each group.Placed all mice （excepted the mice in control group） in cages of （75±2） % oxygen concentration environment for 5 days then moved to a normal environment for 5 days to induced the OIR model.At the 12th day the OIR rAAV-PSF treated OIR group was intravitreal injected with 2 μl 5×10^13 pfu/ml rAAV-PSF and the rAAV treated OIR group was treated with intravitreal injection of 2 μl 5 ×10^13 pfu/ml rAAV.The mice in OIR group were left intact after moved out of oxygen cages.Five days after injection,the eyeballs were harvested and retinal sections were stained to count the nuclear of retinal endothelium cells.Western blot analysis was applied to detectthe protein level of vascular endothelial growth factor （VEGF） in retina.Results There was a significant difference of the expression PSF between normal group and rAAV-PSF treated group （F=16.05,P=0.001）.There was no significant difference of the PSF expression between normal group and rAAV treated group（F=16.05,P=0.890）.There was a significant difference of the number of retinal endothelium cells nuclear between normal group and OIR group （F=101.00,P=0.007）.There is a significant difference of the number of retinal endothelium cells nuclear between rAAV-PSF treated group and OIR group （F=101.00,P=0v002）.There was no significant difference of the number of retinal endothelium cells nuclear between OIR rAAV treate
出处
《中华眼底病杂志》
CAS
CSCD
北大核心
2014年第5期504-508,共5页
Chinese Journal of Ocular Fundus Diseases
基金
国家自然科学基金
