波形纤维蛋白在其转基因小鼠晶状体的表达

Expression of Vimentin in the Lenses of Vimentin Transgenic Mice

目的:观察波形纤维蛋白(Vimentin)在转基因小鼠晶状体的表达,探讨波形纤维蛋白与白内障形成的关系。方法:用显微注射法将鸡的12.7kb波形纤维蛋白基因导入小鼠受精卵的雄前核内,经培育、杂交传代得到20只波形纤维蛋白转基因白内障小鼠。取4只白内障眼球及4只正常眼球,制备石蜡切片,采用ABC免疫组化方法,观察波形纤维蛋白在晶状体的表达。再分别取正常、部分混浊和完全混浊的小鼠晶状体各1只,经SDS-聚丙烯酰胺凝胶电泳(SDS-PAGE)分离蛋白,Western印迹法检测波形纤维蛋白。结果:波形纤维蛋白除在其转基因小鼠晶状体上皮细胞有表达以外,还在前部皮质及赤道部有异常的较强表达。在晶状体的尿素溶性蛋白中,经SDS-PAGE及Westem印迹法检测出57KD、与波形纤维蛋白抗体有特异反应的蛋白,在正常鼠、部分混浊和完全混浊的晶状体中,该蛋白带的灰密度(grey density)分别是15.56、34.59和33.06;波形纤维蛋白在溶于尿素的蛋白中的百分比分别是7.26％、9.05％和9.17％。结论:波形纤维蛋白的过度、异位表达可影响晶状体细胞的分化,导致白内障形成。眼科学报1999;15:199—203。

Purpose: To observe the expression of vimentin in the lenses of vimentin transgenic mice and investigate the role of vimentin in the formation of cataract. Methods; The 12.7kb chicken vimentin genes were microinjected into the male pronuclei of ICR mice. The injected embryos were transferred to the pseudopregnant recipient females and then generated several transgenic mice. Twenty mice developed cataract by cross-breeding. Expression of vimentin in 4 cataractous lenses and 4 normal lenses were detected by ABC immunohistochemical method. Usig SDS-PAGE and Western blot technique, we seperated and measured vimentin in the normal and two cataract lenses, in which one was part opacity and the other was complete opacity. Results: Besides in the epithelial cells, vimentin was overexpressed in the cells in anterior cortex and equatorial region in the lenses of transgenic mice. By SDS-PAGE and Western blot, 57KD urea-soluble protein bands were seperated in normal and cataract lenses and were reacted with vimentin antiserum specifically. In normal lens, partly and completely opaque lenses respectively, the values of average grey density of these bands were 15.56, 34.59 and 33.06 and the percentages of vimentin in urea-soluble fraction were 7. 26% , 9. 05% and 9. 17% , according to the sum of the grey level values of each band. Conclusion: Over and ectopic expression of vimentin in the Lenses of transgenic mice inhibits the lens cell differentiation and is linked to cataract formation. Eye Science 1999; 15: 199-203.