Characterisation of the vitreous proteome in proliferative diabetic retinopathy

Background: Diabetes can lead to serious microvascular complications such as proliferative diabetic retinopathy (PDR), which results in severe vision loss. The diabetes-induced alterations in the vitreous protein composition in diabetic patients with PDR may be responsible for the presence of PDR. The vitreous humour can be utilised in a variety of studies aimed toward the discovery of new targets for the treatment or prevention of PDR and the identification of novel disease mechanisms. The aim of this study was to compare the protein profile of vitreous humour from diabetic patients with PDR with that of vitreous humour from normal human eyes donated for corneal transplant. Results: Vitreous humour from type 2 diabetic patients with PDR (n = 10) and from normal human eyes donated for corneal transplant (n = 10) were studied. The comparative proteomic analysis was performed using two-dimensional fluorescence difference gel electrophoresis (2-D DIGE). Differentially produced proteins (abundance ratio > 2 or < -2, p < 0.01) were identified by matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS) and MALDI-TOF tandem mass spectrometry. A total of 1242 protein spots were detected on the 2-D master gel of the samples, and 57 spots that exhibited statistically significant variations were successfully identified. The spots corresponded to peptide fragments of 29 proteins, including 8 proteins that increased and 21 proteins that decreased in PDR. Excluding the serum proteins from minor vitreous haemorrhage, 19 proteins were found to be differentially produced in PDR patients compared with normal subjects; 6 of these proteins have never been reported to be differentially expressed in PDR vitreous: N(G),N(G)-dimethylarginine dimethylaminohydrolase 1 (DDAH 1), tubulin alpha-1B chain, gamma-enolase, cytosolic acyl coenzyme A thioester hydrolase, malate dehydrogenase and phosphatidylethanolamine-binding protein 1 (PEBP 1). The differential production of pigment epithelium-derived factor (PEDF) and clusterin was confirmed by Western blot analysis. Conclusions: These data provide an in-depth analysis of the human vitreous proteome and reveal protein alterations that are possibly involved in the pathogenesis of PDR. Further investigation of these special proteins may provide potential new targets for the treatment or the prevention of PDR.