title: Immunoproteomic analysis of potential serum biomarker candidates in human glaucoma.
abstract: OBJECTIVE
  Evidence supporting the immune system involvement in glaucoma includes increased titers of serum antibodies to retina and optic nerve proteins, although their pathogenic importance remains unclear. This study using an antibody-based proteomics approach aimed to identify disease-related antigens as candidate biomarkers of glaucoma.
  
  
  METHODS
  Serum samples were collected from 111 patients with primary open-angle glaucoma and an age-matched control group of 49 healthy subjects without glaucoma. For high-throughput characterization of antigens, serum IgG was eluted from five randomly selected glaucomatous samples and analyzed by linear ion trap mass spectrometry (LC-MS/MS). Serum titers of selected biomarker candidates were then measured by specific ELISAs in the whole sample pool (including an additional control group of diabetic retinopathy).
  
  
  RESULTS
  LC-MS/MS analysis of IgG elutes revealed a complex panel of proteins, including those detectable only in glaucomatous samples. Interestingly, many of these antigens corresponded to upregulated retinal proteins previously identified in glaucomatous donors (or that exhibited increased methionine oxidation). Moreover, additional analysis detected a greater immunoreactivity of the patient sera to glaucomatous retinal proteins (or to oxidatively stressed cell culture proteins), thereby suggesting the importance of disease-related protein modifications in autoantibody production/reactivity. As a narrowing-down strategy for selection of initial biomarker candidates, we determined the serum proteins overlapping with the retinal proteins known to be up-regulated in glaucoma. Four of the selected 10 candidates (AIF, cyclic AMP-responsive element binding protein, ephrin type-A receptor, and huntingtin) exhibited higher ELISA titers in the glaucomatous sera.
  
  
  CONCLUSIONS
  A number of serum proteins identified by this immunoproteomic study of human glaucoma may represent diseased tissue-related antigens and serve as candidate biomarkers of glaucoma.
authors.affiliation: Department of Ophthalmology & Visual Sciences, University of Louisville School of Medicine, Louisville, Kentucky 40202, USA. gulgun.tezel@louisville.edu
authors.forename: Gülgün
authors.initials: G
authors.lastname: Tezel
authors.forename: Ivey L
authors.initials: IL
authors.lastname: Thornton
authors.forename: Melissa G
authors.initials: MG
authors.lastname: Tong
authors.forename: Cheng
authors.initials: C
authors.lastname: Luo
authors.forename: Xiangjun
authors.initials: X
authors.lastname: Yang
authors.forename: Jian
authors.initials: J
authors.lastname: Cai
authors.forename: David W
authors.initials: DW
authors.lastname: Powell
authors.forename: Joern B
authors.initials: JB
authors.lastname: Soltau
authors.forename: Jeffrey M
authors.initials: JM
authors.lastname: Liebmann
authors.forename: Robert
authors.initials: R
authors.lastname: Ritch
mesh_terms: D000368:Aged; D000818:Animals; D001323:Autoantibodies / Q000097:blood*; D001324:Autoantigens / Q000276:immunology*; D015415:Biomarkers / Q000097:blood*; D001798:Blood Proteins / Q000276:immunology*; D002853:Chromatography, Liquid; D018920:Coculture Techniques; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005136:Eye Proteins / Q000276:immunology*; D005902:Glaucoma, Open-Angle / Q000175:diagnosis / Q000276:immunology*; D006801:Humans; D015151:Immunoblotting; D007074:Immunoglobulin G / Q000097:blood; D007429:Intraocular Pressure; D008875:Middle Aged; D018384:Oxidative Stress; D040901:Proteomics; D051381:Rats; D053719:Tandem Mass Spectrometry
chemical_list: D001323:Autoantibodies; D001324:Autoantigens; D015415:Biomarkers; D001798:Blood Proteins; D005136:Eye Proteins; D007074:Immunoglobulin G