Abstract
A quantitative proteomics analysis of the macular Bruch membrane/choroid complex was pursued for insights into the molecular mechanisms of age-related macular degeneration (AMD). Protein in trephine samples from the macular region of 10 early/mid-stage dry AMD, six advanced dry AMD, eight wet AMD, and 25 normal control post-mortem eyes was analyzed by LC MS/MS iTRAQ (isobaric tags for relative and absolute quantitation) technology. A total of 901 proteins was quantified, including 556 proteins from ≥3 AMD samples. Most proteins differed little in amount between AMD and control samples and therefore reflect the proteome of normal macular tissues of average age 81. A total of 56 proteins were found to be elevated and 43 were found to be reduced in AMD tissues relative to controls. Analysis by category of disease progression revealed up to 16 proteins elevated or decreased in each category. About 60% of the elevated proteins are involved in immune response and host defense, including many complement proteins and damage-associated molecular pattern proteins such as α-defensins 1–3, protein S100s, crystallins, histones, and galectin-3. Four retinoid processing proteins were elevated only in early/mid-stage AMD, supporting a role for retinoids in AMD initiation. Proteins uniquely decreased in early/mid-stage AMD implicate hematologic malfunctions and weakened extracellular matrix integrity and cellular interactions. Galectin-3, a receptor for advanced glycation end products, was the most significantly elevated protein in advanced dry AMD, supporting a role for advanced glycation end products in dry AMD progression. The results endorse inflammatory processes in both early and advanced AMD pathology, implicate different pathways of progression to advanced dry and wet AMD, and provide a new database for hypothesis-driven and discovery-based studies of AMD.
Highlights
A quantitative proteomics analysis of the macular Bruch membrane/choroid complex was pursued for insights into the molecular mechanisms of age-related macular degeneration (AMD)
About 90% of proteins showed no significant quantitative difference between AMD and control tissues, indicating that the determined proteome largely reflects that of normal macular Bruch membrane/choroid complex
In contrast to wet AMD, we found no complement proteins elevated in advanced dry AMD tissues, but histone H2B type 1-J and histone H4 were uniquely elevated, perhaps as components of so-called “neutrophil extracellular traps” [50] formed during cell death processes [49]
Summary
A quantitative proteomics analysis of the macular Bruch membrane/choroid complex was pursued for insights into the molecular mechanisms of age-related macular degeneration (AMD). We have proposed that oxidative protein modifications are among the catalysts of AMD [7], and a host of elevated oxidative modifications have been reported in AMD Bruch membrane, drusen, retina, RPE, and plasma [7,8,9,10,11,12,13,14] Two of these modifications, namely carboxymethyllysine and carboxyethylpyrrole (CEP), stimulate neovascularization in vivo [15, 16], and immunization with CEP-adducted protein has been shown to induce a dry AMD-like phenotype in mice [17]. Many other factors have been reported to contribute to this complex disease, including for example cigarette smoking [18], cumulative light exposure [19], lipofuscin/retinoid toxicity [20], advanced glycation end products (8 –10), and microbial infection [21]
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