人血管内皮细胞生长抑制因子基因转移抑制角膜新生血管的实验研究  

Inhibition of corneal neovascularization by vascular endothelia growth inhibitor gene

作　　者：王冰[1] 王传富[3] 王利华[2] 

机构地区：[1]山东大学附属省立医院眼科中心,济南250021 [2]山东大学附属省立医院神经外科,济南250021 [3]青岛大学医学院附属医院眼科

出　　处：《中华眼科杂志》2008年第10期929-933,共5页Chinese Journal of Ophthalmology

基　　金：山东省卫生厅青年基金资助项目（2007QZ017）

摘　　要：目的探讨以阳离子脂质体介导重组人pcDNA4-血管内皮细胞生长抑制因子（VEGI）基因转移对角膜新生血管的抑制作用。方法实验研究。40只健康成年新西兰大白兔（40只眼），采用随机数字表法分为4组：A组（10只兔）为阳离子脂质体-重组人pcDNA4-VEGI基因转染组；B组（10只兔）为空白载体转染组；C组（10只兔）为阳离子脂质体转染组；D组（10只兔）为空白对照组。自行构建真核表达的重组人血管内皮细胞生长抑制因子（VEGI）基因，并检测重组基因的正确性；角膜缝线法制作兔角膜新生血管模型，用结膜下注射的方法将阳离子脂质体包裹的VEGI重组质粒（pcDNA4-VEGI）转染入兔角膜，裂隙灯显微镜下观察记录各组兔角膜新生血管长出的时间、角膜新生血管的长度和数量，并分别于基因转染后1、3、7、14及21d以免疫组织化学方法观察VEGI基因表达情况，观察其对角膜新生血管的抑制作用。采用SPSS10．0单因素方差分析行数据统计。结果计算机自动测序证实成功构建真核表达的重组人VEGI基因；动物实验中基因转染组角膜新生血管平均出现时间为6．3d，对照组分别为3．1、3．2、3．2d不等，差异有统计学意义（F=39．838，P=0．00）；基因转染后第3天，转染组实验兔未出现角膜新生血管，对照组已有部分兔眼出现角膜新生血管；转染后第7天，转染组实验兔的角膜新生血管纤细，累及钟点数局限于缝线周围，对照组兔的角膜新生血管最长为2．9mm，血管密集；转染后第14天，转染组兔角膜新生血管最长达4．0mm，对照组新生血管最长达6．4mm，累及钟点数为3．2个；各时间段基因转染组角膜新生血管长度、平均面积与对照组相比，差异均有统计学意义。A组角膜新生血管长度与B组相比，第7、14、21天q值分别为17．386、20．944、8．892，P值均〈0．01；与C组相比，Objective To evaluate the effect of Effectene^TM lipofectine mediated plasmids encoding human pcDNA4-vascular endothelia growth inhibitor （pcDNA4-VEGI） gene on corneal neovascularization （CNV）. Methods It was an experimental study. Forty healthy New Zealand albino rabbits （40 eyes ） were divided into 4 teams according to the random digits table： team A （ 10 rabbits ） was the team which transfected by lipofectine mediated plasmids encoding human pcDNA4-vascular endothelia growth inhibitor （ pcDNA4-VEGI） gene; team B （ 10 rabbits ） was the team transfected by empty vector; team C （ 10 rabbits ） was the team transfected by lipofectine; team D （10 rabbits）was the empty control group which was transfected by saline. （1）Human VEGI gene fragment was connected with expressional vector plasmid pcDNA4 to construct the pcDNAa-VEGI gene expression vector. Computer automatic sequence analysis was used to identify the gene. （2） New Zealand albino rabbits were sutured by 5-0 silk in the superior cornea to establish the animal model and were transfected by pcDNA4-VEGI gene mediated by Effectene^TM lipofectine transfection. Length and area of CNV were measured by slit lamp every day after transfection, immunohistocbemistry was used to detected the expression of VEGI protein in cornea at the time 3, 7, 14 and 21 d. Results （1） Computer automatic sequence analysis confirmed the correct recombination of pcDNA4-VEGI gene. （2）Average occurrence of CNV in the pcDNA4-VEGI gene transfected team was 6. 3 days while that in the control teams were 3. 1,3.2,3.2 days （F =39. 838,P 〈0. 01 ）. Average length and area of CNV were also different in the VEGI team and the control teams（q between team A and B on average length of CNV was 17. 386, 20. 944 and 8. 892 on 7, 14 and 21 d; q between team A and C on average length of CNV was 19.488,19.795 and 7.483 on 7, 14 and 21 d; q between team A and C on average length ofCNV wasl9. 583,20.413 and 8.941 on 7, 14 and 21 d,and P were all 〈0.01. A

关 键 词：角膜新生血管化 肿瘤坏死因子α 脂质体 基因疗法 

分 类 号：R686[医药卫生—骨科学]