Quantitative Protein Analysis of Intraocular Fluids from Patients with Idiopathic Epiretinal Membranes

Abstract

Purpose: : To perform an in-depth quantitative protein analysis of aqueous and vitreous from eyes with idiopathic epiretinal membranes (iERM) by mass spectrometry (MS) and multiplex bead assays.

Methods: : Aqueous and vitreous was collected from 24 patients with iERM during surgery. Samples of 8 patients were labeled with 4-plex iTRAQ reagents and analyzed by 2-dimensional liquid chromatography MS on an LTQ Orbitrap XL mass spectrometer. With Luminex multiplex assays 29 different growth factors and cytokines were analyzed in aqueous and vitreous of 16 patients with iERM and compared to the aqueous of 16 cataract patients.

Results: : A total of 303 proteins were identified in the aqueous and vitreous from 8 patients with iERM by MS. Grouping these proteins according to involvement in biological processes, the majority was found to be involved in the classical and alternative pathway of the complement activation (n=27), in proteolysis (n=26) and in cell adhesion (n=28). iTRAQ relative quantitation revealed that there was minimal variation of the protein content in the two ocular compartments (3.96% of proteins significantly differentially regulated). Analysis of 29 cytokines and growth factors by multiplex bead assay revealed that comparing aqueous and vitreous in eyes with iERM only PDGF-AA was significantly higher in the vitreous. Comparing aqueous of eyes with iERM to cataract showed that MCP-1 was significantly higher in the iERM group.

Conclusions: : The combined results from the mass spectrometric and multiplex bead analyses revealed that the protein content is similar between the vitreous and aqueous of eyes with iERM. In addition, our results showed similar levels of cytokines and growth factors in the aqueous when comparing eyes with iERM to eyes with cataract suggesting that both disease entities have a similar protein composition in the aqueous.

Keywords: proteomics