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X. Gu, X. Yuan, J. S. Crabb, K. Shadrach, J. G. Hollyfield, J. W. Crabb; Comparison of Proteins in Dry and Wet AMD Bruch's Membrane. Invest. Ophthalmol. Vis. Sci. 2009;50(13):2343.
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Proteomic differences between dry and wet AMD Bruch’s membrane were sought for insights into the molecular mechanisms contributing to disease progression and visual loss in age-related macular degeneration (AMD).
Bruch’s membrane trephine samples were isolated from the macular region of 16 dry AMD, 8 wet and 25 normal control donor eyes. Protein was extracted in SDS, digested with trypsin, peptides were labeled with iTRAQ tags, individual AMD samples were combined with pooled control samples in equal amounts. Peptides were analyzed by LC MS/MS and proteins identified using the Mascot search engine and the Swiss-Protein sequence database. Relative protein quantification from iTRAQ labeling utilized code written in the statistical program R. Bioinformatic pathway analysis was performed with Ingenuity Pathways Analysis 5.0.
Proteomic analyses comparing dry and wet AMD tissues yielded relative quantitation for over 900 proteins, 236 of which were found in ≥ 3 tissues from both dry and wet donors. From these proteins, several were detected as more abundant in wet than in dry AMD tissues, including protein S100-A9, complement C9 and C3, vitronectin, fibrinogen β, and IgG3 heavy chain. Proteins more abundant in wet AMD tissues by virtue of being reduced in dry tissues included annexin-A6, IgA heavy chain, serum amyloid P and caveolin-1. Proteins more abundant in dry AMD tissues included alpha-crystalline A, alpha-crystalline B, and galectin-3. Proteins reduced in wet AMD tissues relative to dry AMD and control tissues included housekeeping enzymes, a histone and CD9 antigen.
Quantitative analysis has identified several proteomic differences between dry and wet AMD Bruch’s membrane. The abundance of complement and antibody components in wet AMD Bruch’s membrane is consistent with inflammatory processes contributing to advanced AMD pathology.
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