缺氧对人视网膜色素上皮细胞促血管生成素-1和促血管生成素-2表达的影响  

Effect of hypoxia on the expression of angiopoietin-1 and angiopoietin-2 in human retinal pigment epithelial cells

作　　者：雷润佳[1] 胡丹[1] 张鹏[1] 孙丽娟[1] 白倩[1] 闵捷[1] 

机构地区：[1]第四军医大学西京医院眼科,全军眼科研究所,陕西省西安市710032

出　　处：《眼科新进展》2013年第5期435-438,共4页Recent Advances in Ophthalmology

基　　金：陕西省科学技术研究发展计划(编号:2011K1402-01);西京医院学科助推计划(编号:XJZT10M16)~~

摘　　要：目的探讨缺氧环境下人视网膜色素上皮(retinal pigment epithelial,RPE)细胞中促血管生成素-1(angiopoietin-1,Ang-1)和促血管生成素-2(angiopoietin-2,Ang-2)表达变化及其可能的机制。方法将人RPE细胞培养在厌氧箱中,以建立缺氧模型。按缺氧时间0h、1h、4h、12h和24h分为5组,观察Ang-1、Ang-2与缺氧诱导因子-1α(hypoxia inducible factor-1α,HIF-1α)表达的变化;另取4h作为观察点,按处理因素分为缺氧对照组和HIF-1α抑制剂3-(5’-hydroxymethyl-2’-furyl)-1-benzyl indazole(YC-1)处理组,YC-1浓度分别为1μmol·L-1、10μmol·L-1和100μmol·L-1,观察抑制HIF-1α后对Ang-1和Ang-2表达的影响。应用免疫荧光染色法和Westernblot分别检测Ang-1、Ang-2与HIF-1α蛋白的表达。结果常氧条件下人RPE细胞内可见Ang-2蛋白表达,未见Ang-1及HIF-1α蛋白表达;缺氧条件下Ang-1和Ang-2分别与HIF-1α共表达于RPE细胞胞质中。HIF-1α与Ang-1在缺氧早期呈上升趋势,其中HIF-1α于缺氧后4h表达最高(0.451±0.022,与0h相比t=5.401,P<0.05),24h后有所下降;Ang-1于缺氧后24h表达最高(1.342±0.059,与0h相比t=20.100,P<0.01);Ang-2表达几乎不变(1.542±0.129、1.621±0.131、1.574±0.146、1.624±0.027,与0h相比t=0.044、0.673、0.244、0.700,均为P>0.05)。浓度为1μmol·L-1、10μmol·L-1和100μmol·L-1的YC-1均可有效抑制HIF-1α蛋白的表达(t=21.583、27.154、27.431,均为P<0.01)。HIF-1α蛋白表达下调后,Ang-1蛋白表达也随之受到抑制(t=9.492、15.487、17.942,均为P<0.05),而Ang-2蛋白表达无明显变化(t=0.859、0.178、1.565,均为P>0.05)。结论缺氧条件下人RPE细胞表达Ang-1上调,并与HIF-1α表达具有时相关系,Ang-2无明显变化;抑制HIF-1α表达后,Ang-1表达随之降低。提示在脉络膜新生血管生成过程中Ang-1的表达可能受HIF-1α选择性调控。Objective To investigate the effect of hypoxia on the expression of angiopoietin-1 （Ang-1） and angiopoietin -2（Ang-2） in human retinal pigment epithelial （RPE） cells and explore the mechanism. Methods Human RPE cells were cultured in anaerobic environment to establish the hypoxia model. All the cells were divided into 5 groups by their exposure time to the anaerobic enviroument：0 hour, 1 hour,4 hours, 12 hours and 24 hours, expression changes of Ang-1 ,Ang-2 and hypoxia-inducible fac- tor-1α（HIF-1α） in all groups were observed. Then the cells whose exposure time was 4 hours were divided into the hypoxia control group and the YC-1 [ 3-（5 ＇-hydroxymethyl- 2＇-furyl）-l-benzyl indazole,inducer of HIF-1 ] groups, according to the treatment factor,the concentrations of YC-1 were 1 μmol · L^-1 ,10 μmol · L^-1 and 100 μmol · L^-1, respectively. The expression change of Ang-1 ,Ang-2 in the YC-1 groups were observed. The expression of Ang-1 ,Ang-2 and HIF-1 a were respectively detected by immunofluorescence and Western blot. Results Under normoxia condition, expression of Ang-2 was observed, no Ang-1 or HIF-1α was observed. Under hypoxia condition, eoexpression of Ang-1 and HIF-1α and coexpression of Ang-2 and HIF-1α were observed in endoplasm of RPE cells. At the early stage of hypoxia,the expression of Ang-1 and HIF-1α tended to increase,the expression of HIF-1α peaked 4 hours after exposure（0.451 ± 0. 022 ）, which was significantly lower than that under normoxia condition （ t = 5. 401, P 〈 0.05 ） ,and decreased 24 hours after exposure. The expression of Ang-1 peaked 24 hours after exposure （ 1. 342 ± 0.059 ）, which was significantly lower than that under normoxia condition（ t = 20. 100 ,P 〈 0.01 ）. The expression of Ang-2 almost stay unchanged（ 1. 542±0. 129,1. 621±0. 131,1. 574±0. 146,1. 624±0.027） ,the difference between which and those under normoxia condition （ t = 0.044,0. 673,0. 244,0. 700） were of no statistical difference （all P 〉 0.05 �

关 键 词：脉络膜新生血管 促血管生成素-1 促血管生成素-2 缺氧诱导因子-1Α 视网膜色素上皮 眼 

分 类 号：R774.1[医药卫生—眼科]