大鼠青光眼慢性高眼压视网膜组织的二维凝胶电泳分析

Studies on retinal proteome of chronic high intraocular pressure of SD rats by two-dimensional gel electrophoresis

目的应用二维凝胶电泳对SD大鼠青光眼慢性高眼压视网膜组织的蛋白进行初步分析。方法分离大鼠青光眼慢性高眼压视网膜组织,以对侧眼视网膜组织为对照,经样本初处理后,进行二维凝胶电泳和凝胶银染色,然后分析电泳图谱,观察蛋白的分离效果,对比、分析其表达蛋白质点的差异,寻找SD大鼠视网膜中与慢性高眼压相关的蛋白质。结果用化学裂解法结合超声波粉碎,可以裂解视网膜神经组织,使其大部分蛋白质得到溶解,成功建立对大鼠青光眼慢性高眼压眼及对照眼视网膜组织进行样本处理、二维凝胶电泳的基本方法和条件,得到两组视网膜组织的二维凝胶电泳图谱。各组织蛋白在小胶上均能显示120～290个左右的斑点。两组视网膜组织的凝胶图谱呈现出9个斑点的差异。结论二维凝胶电泳可以有效分离、显示视网膜特异表达蛋白质。通过双向电泳,我们发现SD大鼠视网膜蛋白质表达与对照眼相比有质和量的差异。

Objective To apply two-dimensional （2D） gel electrophoresis to resolve specifically expressed to glaucoma in the retina of SD rats. Methods Glaucoma was induced in the left eye of 45 rats by repeated argon laser photocoagulation of the trabecular meshwork. The right eye served Retinas were isolated from glaucomatous and control eyes 35 days after photocoagulation. Profrom the retinas of glaucomatous and control eyes, were separated by using two-dimensional eteins, prepared lectrophoresis and then analyzed by 2D electrophoresis imaging analyzer. Results 2D gel electrophoresis was established for the retinal proteome analysis. Retinal neural tissue was lysed by using chemical lysis solution and ultrasonic. Using cartier ampholyte to set up pH gradient as first dimension and casting vertical 12% SDS-acylamide-bis slab as second dimension, the major retinal proteins showed maps of proteome on 2D gel clearly. There were 120 -290 spots with good distribution on the gels. The number of most different spots was 9. Retinal proteome of glaucomatous and control eyes had different expression patterns. Conclusion 2D gel electrophoresis is effective to separate specifically expressed retinal proteins. The expressed proteins in the glaucomatous retina are different in quality and quantity from that in the control eyes.