人青光眼潜在血清生物标志物候选物的免疫蛋白质组学分析。
Immunoproteomic analysis of potential serum biomarker candidates in human glaucoma.
机构信息
Department of Ophthalmology & Visual Sciences, University of Louisville School of Medicine, Louisville, Kentucky 40202, USA.
出版信息
Invest Ophthalmol Vis Sci. 2012 Dec 13;53(13):8222-31. doi: 10.1167/iovs.12-10076.
PURPOSE
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.
目的
支持免疫系统参与青光眼的证据包括血清中针对视网膜和视神经蛋白的抗体滴度增加，尽管其致病作用尚不清楚。本研究采用基于抗体的蛋白质组学方法，旨在鉴定与疾病相关的抗原作为青光眼的候选生物标志物。
方法
从 111 例原发性开角型青光眼患者和 49 例年龄匹配的无青光眼健康对照组中采集血清样本。为了高通量表征抗原，从 5 个随机选择的青光眼样本中洗脱血清 IgG，并通过线性离子阱质谱（LC-MS/MS）进行分析。然后通过特异性 ELISA 在整个样本池中（包括糖尿病视网膜病变的另一个对照组）测量选定生物标志物候选物的血清滴度。
结果
LC-MS/MS 分析 IgG 洗脱液显示出一组复杂的蛋白质，包括仅在青光眼样本中检测到的蛋白质。有趣的是，其中许多抗原与先前在青光眼供体中鉴定出的上调视网膜蛋白相对应（或表现出增加的蛋氨酸氧化）。此外，进一步的分析检测到患者血清对青光眼视网膜蛋白（或氧化应激细胞培养蛋白）的更高免疫反应性，从而表明疾病相关蛋白修饰在自身抗体产生/反应中的重要性。作为选择初始生物标志物候选物的缩小策略，我们确定了与已知在青光眼上调的视网膜蛋白重叠的血清蛋白。在所选择的 10 个候选物中的 4 个（AIF、环 AMP 反应元件结合蛋白、ephrin 型-A 受体和亨廷顿蛋白）在青光眼血清中的 ELISA 滴度更高。
结论
本研究通过对人类青光眼的免疫蛋白质组学研究鉴定的一些血清蛋白可能代表患病组织相关抗原，并可作为青光眼的候选生物标志物。
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