Serum and Antibodies of Glaucoma Patients Lead to Changes in the Proteome, Especially Cell Regulatory Proteins, in Retinal Cells

Katharina Bell,Franz H Grus,Sebastian Funke,Norbert Pfeiffer,Demetrios Vavvas

doi:10.1371/journal.pone.0046910

Abstract

PurposePrevious studies show significantly specifically changed autoantibody reactions against retinal antigens in the serum of glaucoma and ocular hypertension (OHT) patients in comparison to healthy people. As pathogenesis of glaucoma still is unknown the aim of this study was to analyze if the serum and antibodies of glaucoma patients interact with neuroretinal cells.MethodsR28 cells were incubated with serum of patients suffering from primary open angle glaucoma (POAG), normal tension glaucoma (NTG) or OHT, POAG serum after antibody removal and serum from healthy people for 48 h under a normal or an elevated pressure of 15000 Pa (112 mmHg). RGC5 cells were additionally incubated with POAG antibodies under a normal pressure. Protein profiles of the R28 cells were measured with Seldi-Tof-MS, protein identification was performed with Maldi-TofTof-MS. Protein analysis of the RGC5 cells was performed with ESI-Orbitrap MS. Statistical analysis including multivariate statistics, variance component analysis as well as calculating Mahalanobis distances was performed.ResultsHighly significant changes of the complex protein profiles after incubation with glaucoma and OHT serum in comparison to healthy serum were detected, showing specific changes in the cells (e.g. Protein at 9192 Da (p<0.001)). The variance component analysis showed an effect of the serum of 59% on the cells. The pressure had an effect of 11% on the cells. Antibody removal led to significantly changed cell reactions (p<0.03). Furthermore, the incubation with POAG serum and its antibodies led to pro-apoptotic changes of proteins in the cells.ConclusionsThese studies show that the serum and the antibodies of glaucoma patients significantly change protein expressions involved in cell regulatory processes in neuroretinal cells. These could lead to a higher vulnerability of retinal cells towards stress factors such as an elevated IOP and eventually could lead to an increased apoptosis of the cells as in glaucoma.

Highlights

Glaucoma, a group of diseases leading to loss of retinal ganglion cells with still unknown pathogenesis, is a leading cause for blindness worldwide, as the estimated number of affected people counting nearly 7 million shows [1]
In the second experiment the analysis showed significant differences between cells incubated with ocular hypertension (OHT) serum and healthy or primary open angle glaucoma (POAG) serum, as well as cells incubated with normal tension glaucoma (NTG) serum and healthy or POAG serum
Significant differences between protein profiles of cells incubated with NTG or OHT serum were detected

Summary

Introduction

A group of diseases leading to loss of retinal ganglion cells (rgc) with still unknown pathogenesis, is a leading cause for blindness worldwide, as the estimated number of affected people counting nearly 7 million shows [1]. 4–7% of people over the age of 40 have ocular hypertension (OHT) [3] but per year only 1% develop glaucoma [4,5]. Still a major risk factor, the presumption that an elevated IOP is the only cause for glaucoma is long out-dated. Glaucoma can be added to the long list of neurodegenerative diseases with mostly unknown pathogenesis, conditions characterized by progressive nervous system dysfunction and often accompanied by the atrophy of the affected central or peripheral nervous system structures [6]

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