Differential proteomics study in serum of patients with primary open-angle glaucoma and pseudoexfoliation glaucoma.

Abstract

Purpose: Comparative proteomics analysis in the sera between patients with primary open-angle glaucoma (POAG), pseudoexfoliation glaucoma (PEXG) and control subjects, with the aim of identifying candidate biomarkers of glaucoma and provide clues to pathogenic mechanisms associated to the disease and to identify potential pharmacological targets.

Methods: Equalization of serum proteins with ProteoMiner, two-dimensional fluorescent difference gel electrophoresis (2D-DIGE), MALDI-TOF/TOF, nano-liquid-chromatography tandem mass spectrometry (nLC-MS-MS), functional analysis (Ingenuity® Systems, www.ingenuity.com), and enzyme-linked immunosorbent assay (ELISA).

Results: A total of 149 protein spots displaying significant changes in abundance were selected for identification purposes by specifying a threshold of Gain Ratio ≥ 0.05. We established the identity of 118 spots resulting in the identification of 35 distinct proteins. The proteomic workflow implemented revealed that these proteins were altered in patients with glaucoma relative to healthy controls. The functional analysis identified the signalling network of these proteins correlated to an immunological and inflammatory pathway. The top-17-ranked proteins of the 35 panel were determined by ELISA on sera samples (non-equalized) from newly recruited subjects (POAG, PEXG and control), and confirmed their alteration in individual protein levels.

Conclusions: Overall, this work identifies a panel of candidates for glaucoma biomarkers that they are part of a network linked to regulating immune and inflammatory-related processes. This study provides a new basis to validate the identified proteins as biomarkers of glaucoma in a large-scale-multiplexed screening in serum. The data from this study offers new perspectives in the discovery of glaucoma biomarkers in serum of POAG and PEXG patients.

Keywords: 663 proteomics • 658 protein purification and characterization • 555 immunomodulation/immunoregulation