Proteome Analysis of Human Vitreous Fluid From Eyes With Diabetic Retinopathy

Abstract

Abstract: : Purpose: To create a map that shows protein expression in the human vitreous from eyes with proliferative diabetic retinopathy (PDR), a proteome survey was performed. Methods: Vitreous and serum samples were obtained from eyes with PDR (28 cases) macular hole (23cases). We adopted IgG removal procedures in sample preparation in advance of 2-D PAGE with several samples of MH and PDR. The expressed proteins in the samples from each patient were separated by two-dimensional polyacrylamide gel electrophoresis (2-D PAGE). The protein spots were visualized by silver staining, and the protein expression patterns were analyzed by a computer- associated image analysis system. Some protein spots of concern were excised from the 2-D gels, digested in situ with trypsin, and analyzed by mass spectrometry (Q-TOF mass spectrometer). Results: The patterns of vitreous protein expression profile were similar to those of corresponding serum protein. More than 600 spots were detected on 2-D gels of vitreous samples form eyes with PDR, where 100 spots were successfully analyzed; the spots corresponded to peptide fragments of 27 proteins. Enolase and catalase were identified among 4 spots that were detected. They were found neither in MH vitreous samples nor in PDR serum samples. Conclusions: A map of protein expression was made in human vitreous from eyes with PDR using proteome analysis. In PDR, many proteins were detected probably due to blood-ocular barrier dysfunction and/or the production in the eye.

Keywords: diabetic retinopathy • molecular biology • protein purification and characterization