The Structure and Function of Retinal Pigment Epithelial Cells

视网膜色素上皮细胞的结构和功能

基本信息

批准号：
07457003
负责人：
HIROSAWA Kazushige
金额：
$ 4.48万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

项目摘要

Monoclonal antibodies were raised against chick and bovine retinal pigment epithelial (RPE) cells using mice as immune animal. Among the monoclonal antibodies, S5D8 and RPE7 were analyzed their characters by means of molecular and cellular biological methods.1) ; S5D8 : The antgen protein has molecular weight of 63kDa. The whole length of cDNA is 2510bp, open reading frame being 1599bp with 2 olyadenylation signals. Amino acid residues are 533 and their molecular size being 60.9kDa. The protein has 3 glycosylation sites. It has neither signal peptide nor membrane expanding region. The size of mRNA is about 2.9kbases. The open reading frame was transfected to cultured fibroblasts, the endoplasmic reticula of which were stained with S5D8.2) ; PRE7 : The localization of RPE7 was on the plasma membranes of RPE and Muller cells of neural retina. This result seem to suggest the protein RPE7 is involved in the supporting function of the retina. The protein has molecular weight of 40-45kDa by Western blotting. RPE7 has an open reading frame of 271 amino acid residues, 3 glycosylation sitesand is found to have a membrane expanding hydrophobic region. Homology research of this cDNA sequences revealed that RPE7 protein belongs to a novel type of immunoglobulin superfamily, one of which has been named EMMPRIN.EMMPRIN is involved in the remodeling of extrcellular matrix and plays a role in the metastasis of cancer cells. Metalloproteinase activity was assayd with cultured bovine RPE by zymography. The culture supernatant contained gelatinaseA,the activity of which became higher when anti-RPE7 antibody was added to the culture medium. It is concluded, from this result, that RPE7 plays an important role in the phagocytosis of outer segment and mobilization of RPE at some pathlogical conditions.

使用小鼠作为免疫动物，对鸡和牛视网膜色素上皮（RPE）细胞饲养单克隆抗体。在单克隆抗体中，通过分子和细胞生物学方法分析了S5D8和RPE7的特征。1）； S5D8：AntGen蛋白的分子量为63KDA。 cDNA的整个长度为2510bp，开放式阅读框架为1599bp，带有2个奥甲二苯基信号。氨基酸残基为533，其分子大小为60.9kDa。该蛋白质具有3个糖基化位点。它既没有信号肽也没有膜扩展区域。 mRNA的大小约为2.9kbase。开放式阅读框转染为培养的成纤维细胞，其内质网被S5D8.2染色）； PRE7：RPE7的定位在神经视网膜的RPE和Muller细胞的质膜上。该结果似乎表明蛋白质RPE7参与了视网膜的支撑功能。该蛋白质的分子量为40-45kDa，通过蛋白质印迹。 RPE7具有271个氨基酸残基的开放式阅读框，发现3个糖基化位点和具有膜膨胀的疏水区。该cDNA序列的同源性研究表明，RPE7蛋白属于一种新型的免疫球蛋白超家族，其中一个被称为Emmprin.emmprin参与螺旋基质的重塑，并在癌细胞转移中起作用。金属蛋白酶活性是用Zymography用培养的牛RPE测定的。培养上清液中含有明胶，当将抗RPE7抗体添加到培养基中时，其活性变得更高。从这个结果得出的结论是，在某些途径条件下，RPE7在外部段的吞噬和RPE的动员中起着重要作用。