Generation and characteristics of transgenic rabbit model of retinal degeneration

转基因兔视网膜变性模型的建立及特点

基本信息

批准号：
18591913
负责人：
KONDO Mineo
金额：
$ 2.49万
依托单位：
Nagoya University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

项目摘要

Purpose of the present research was to generate a rabbit that expresses a mutant rhodopsin gene, and to characterize the progressive retinal degeneration using electroretinography(ERG), histology, and immunohistochemistry. The rabbit DNA clone that contains a rhodopsin gene was identified in a NZW rabbit BAC library. The Pro347Leu mutation of the rhodopsin gene was introduced homologous recombination in E. col. and these BACs were microinjected into fertilized eggs. Transgene-positive animals were identified by Southern blot analysis. The expression level of transgene in the retinal tissue was measured by qualitative real-time RT-PCR. Scotopic and photopic ERGs elicited by different stimulus intensities were recorded. Paraffin retinal sections were stained with hematoxylin and eosin. Rhodspin and lectin immunohistochemistory, and electronmicroscopy was also performed. Ten transgene-positive founders were identified, six of which passed transgenes to offsprings. The expression level of the transgene ranged from 7-80% for six transgenic lines, and was roughly correlated with transgene copy number and speed of retinal degeneration. In line 7, the rod ERG a-wave was nearly extinguished by 12-month-old, but the cone a-wave retained 60% of controls. Retinal histology demonstrated a progressive loss of photoreceptors and a shortening of the outer segments, and ONL at visual streak showed only 1-2 laws by 12 weeks. This is the first transgenic rabbit model of retinal degeneration. The speed of retinal degeneration was dependant on the expression level of the transgene. Because the rabbits have large eyes and are easy to handle, these animals will be a useful model to stud the pathophysiology and treatment of retinal degeneration

本研究的目的是产生一种表达突变型紫红蛋白基因的兔子，并使用电子模拟（ERG），组织学和免疫组织化学来表征进行性视网膜变性。在NZW兔子BAC文库中鉴定出含有Rhodopsin基因的兔DNA克隆。在大肠杆菌中引入了视紫红质基因的Pro347Leu突变。并将这些BAC微型注射到受精卵中。通过Southern印迹分析确定转基因阳性动物。通过定性实时RT-PCR测量视网膜组织中转基因的表达水平。记录了不同刺激强度引起的SCOTOPIC和PHOTOPIC ERG。用苏木精和曙红对石蜡视网膜切片进行染色。还进行了Rhodspin和凝集素免疫组织化学，并进行了电子显微镜检查。确定了十个转基因阳性的创始人，其中6个通过了转基因的后代。转基因的表达水平范围为六个转基因线的7-80％，与转基因拷贝数和视网膜变性速度大致相关。在第7行中，杆ERG A波几乎被12个月大的大熄灭，但锥A波保留了60％的对照。视网膜组织学表明，感光体的逐渐丧失和外部段的缩短，视觉条纹的ONL仅显示1-2周的法律。这是视网膜变性的第一个转基因兔模型。视网膜变性的速度取决于转基因的表达水平。由于兔子的眼睛很大并且易于处理，因此这些动物将是一个有用的模型，用于研究视网膜变性的病理生理学和治疗

项目成果

期刊论文数量（0）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Effects of basic fibroblast growth factor on experimental diabetic neuropathy in rats

DOI：
10.2337/db05-1160
发表时间：
2006-05-01
期刊：
DIABETES
影响因子：
7.7
作者：
Nakae, M;Kamiya, H;Nakamura, J
通讯作者：
Nakamura, J

Abnormalities of visual-evoked potentials and pupillary light reflexes in a family with autosomal dominant occult macular dystrophy.

常染色体显性隐匿性黄斑营养不良家系的视觉诱发电位和瞳孔对光反射异常。

DOI：
发表时间：
2007
期刊：
Clin Exp Ophthalmol 35
影响因子：
0
作者：
Koyasu;T;Nakamura Y.(et al);Miyata K.(et al);Kurimoto T.(et al);Nishiguchi KM.(et al);Kaneko H.(et al);Kondo M.(et al);Hood DC.(et al);Koyasu T.(et al);Koike C.(et. al.);Kikuchi M.(et. al.);Ishikawa K.(et. al.);Ikenoya K.(et. al.);Nishiguchi KM.(et. al.);Ito Y.(et. al.);Yata T.(et. al.);Okuno T.(et. al.)
通讯作者：
Okuno T.(et. al.)

Effect of axial length on laser spot size during photodynamic therapy : an experimental study in monkeys

光动力治疗期间轴长对激光光斑大小的影响：猴子的实验研究