胰岛素样生长因子结合蛋白相关蛋白1通过细胞外调节蛋白激酶信号通路抑制视网膜新生血管的研究  

Inhibitory effect of insulin-like growth factor binding protein-related protein 1 on retinal angiogenesis via ERK signaling pathway

作　　者：张平[1] 王泓[1] 曹晖[1] 许迅[1] 孙涛[1] 

机构地区：[1]上海交通大学附属上海市第一人民医院眼科上海市眼底病重点实验室上海眼视觉与光医学工程技术研究中心,200080

出　　处：《中华眼科杂志》2017年第3期207-211,共5页Chinese Journal of Ophthalmology

基　　金：国家自然科学基金青年项目（81200701）;上海市卫生局青年科研项目（20114Y054）

摘　　要：目的 探讨抑血管新生因子胰岛素样生长因子结合蛋白相关蛋白1（IGFBP-rPl）在调控视网膜新生血管方面的作用及其相关分子机制.方法 实验研究.将120只7日龄C57BL/6J小鼠采用随机数字表法随机分为对照组（24只）、氧诱导视网膜病变（OIR）未干预组（24只）及不同浓度IGFBP-rP1OIR干预组（72只）.采用改良Smith法将96只小鼠建立OIR模型;12日龄OIR干预组小鼠再次随机予玻璃体腔内注射3种质量浓度重组小鼠IGFBP-rPI（50、100、200 μg/L）进行干预,每组24只;17日龄小鼠予高分子量异硫氰酸荧光素-右旋糖酐灌注后行视网膜铺片并测量各组小鼠视网膜新生血管相对面积;行石蜡切片HE染色计数突破视网膜内界膜的血管内皮细胞核数目;行免疫印迹法检测各组小鼠视网膜组织磷酸化ERK1/2（p-ERK1/2）、ERK1/2和VEGF的表达.结果 成功建立OIR小鼠模型.OIR未干预组小鼠与对照组小鼠相比,p-ERK1/2和VEGF表达均明显上调,差异均有统计学意义（t=100.068,P=0.000;t=6.526,P=0.003）;50、100、200 μg/L IGFBP-rP1干预组小鼠视网膜新生血管相对面积分别是OIR未干预组小鼠的（76.53±1.35）％、（34.27±2.18）％、（19.72±2.79）％,差异有统计学意义（F=1920,P=0.000）;50、100、200 μg/L IGFBP-rP1干预组小鼠视网膜新生血管内皮细胞核数分别为（11.62±2.54）、（6.07±2.47）、（3.25±1.50）个,差异有统计学意义（F=852.387,P=0.000）.同时干预组小鼠p-ERK1/2和VEGF表达明显下调（F=859.587,P=0.000;F=24.301,P=0.000）,且表现为IGFBP-rP1浓度依赖性.结论 IGFBP-rP1通过阻断ERK信号通路和下调VEGF的表达,抑制视网膜新生血管形成,可能成为未来新生血管性眼底病基因治疗的靶点.Objective To explore the inhibitory effect of insulin-like growth factor binding protein-related protein 1 （IGFBP-rP1）,a novel anti-angiogenic factor,on retinal angiogenesis and its underlying molecular mechanisms.Methods Experimental study.C57BL/6J mice were classified into three groups：control group （n=24）,oxygen-induced retinopathy （OIR） non-intervention group （n=24） and OIR intervention group （n=72）.The OIR mouse model was established using improved Smith＇s methods （n=96）.Twelve-day-old mice in the OIR intervention group were randomly assigned into three groups receiving intravitreal injection of recombinant mouse IGFBP-rP1 （50 μg/L,100 μg/L and 200 μg/L,respectively）.Five days later,the proliferative neovascular responses were estimated by quantifying the new vessel area relative to the total retinal area in flattening retinas stained by high molecular FITC-Dextran and counting the number of neovascular cell nuclei breaking through the internal limiting membrane （ILM） in cross-sections.Retinal phosphor-ERK1/2 （p-ERK1/2）,ERK1/2 and vascular endothelial growth factor （VEGF） protein expression was assessed by Western blot.Results In the fluorescence angiograms,irregular neovascularization and fluorescence leakage were observed in the OIR model.In the OIR non-intervention group,the expression of p-ERK1/2 and VEGF was significantly up-regulated in comparison with the control group （t=100.068,P=0.000.t=6.526,P=0.003）.The area ratios of new retinal vessels and the number of neovascular cell nuclei in mice receiving intravitreal injection of recombinant mouse IGFBP-rP1 both decreased significantly （F=1920,P=0.000.F=852.387,P=0.000）,following the down-regulation of retinal p-ERK1/2 protein expression （F=859.587,P=0.000） and VEGF protein expression （F=24.301,P=0.000） in a dose-dependent manner （P〈0.05）.There was no significant difference in ERK1/2 protein expression （P〉0.05）.Conclusions IGFBP-rP1 inhibits retinal angiogenesis by blocking ERK si

关 键 词：视网膜新生血管化 胰岛素样生长因子结合蛋白质类 MAP激酶信号系统 血管内皮生长因子A 

分 类 号：R774.1[医药卫生—眼科]