Department of Ophthalmology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
Department of Immunology, Laboratory Medical Immunology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
Invest Ophthalmol Vis Sci. 2023 Mar 1;64(3):27. doi: 10.1167/iovs.64.3.27.
Scleritis is a severe inflammatory ocular disorder with unknown pathogenesis. We investigated healthy sclera as well as sclera affected by noninfectious scleritis for differentially expressed proteins using a mass spectrometry approach.
We collected scleral samples of enucleated eyes due to severe noninfectious scleritis (n = 3), and control scleral tissues (n = 5), all exenterated eyes for eyelid carcinomas (n = 4), or choroidal melanoma (n = 1) without scleral invasion. Samples were prepared for the nano liquid-chromatography mass spectrometer (LC-MS), data were analyzed using proteomics software (Scaffold), and is available via ProteomeXchange (identifier PXD038727). Samples were also stained for immuno-histopathological evaluation.
Mass spectrometry identified 629 proteins within the healthy and diseased scleral tissues, whereof collagen type XII, VI, and I were the most abundantly expressed protein. Collagen type II-XII was also present. Filaggrin-2, a protein that plays a crucial role in epidermal barrier function, was found upregulated in all scleritis cases. In addition, other epithelial associated proteins were upregulated (such as keratin 33b, 34, and 85, epiplakin, transglutaminase-3, galectin 7, and caspase-14) in scleritis. Further, upregulated proteins involved in regulation of the cytoskeleton (vinculin and myosin 9), and housekeeping proteins were found (elongation factor-2 and cytoplasmic dynein 1) in our study. Upregulation of filaggrin-2 and myosin-9 was confirmed with immunohistochemistry, the latter protein showing co-localization with the endothelial cell marker ETC-related gene (ERG), indicating neovascularization in scleral tissue affected by scleritis.
We found upregulation of filaggrin-2 and signs of neovascularization in scleral tissue of patients with noninfectious scleritis. Further research, ideally including more scleritis cases, is needed to validate our findings.
巩膜炎是一种严重的眼部炎症性疾病，其发病机制尚不清楚。我们使用质谱分析方法研究了健康巩膜和非感染性巩膜炎患者的巩膜中的差异表达蛋白。
我们收集了 3 例因严重非感染性巩膜炎而摘除眼球的巩膜样本，以及 5 例正常巩膜组织（对照组）、4 例因眼睑癌而摘除眼球的巩膜样本，以及 1 例无巩膜侵犯的脉络膜黑色素瘤的巩膜样本。将样本制备用于纳升液相色谱-质谱仪（LC-MS）分析，使用蛋白质组学软件（Scaffold）进行数据分析，并可通过 ProteomeXchange（标识符 PXD038727）获取。样本还进行了免疫组织化学评估。
质谱分析在健康和患病的巩膜组织中鉴定出 629 种蛋白质，其中胶原 XII、VI 和 I 是表达最丰富的蛋白。胶原 II-XII 也存在。在所有巩膜炎病例中，发现丝聚合蛋白 2 （一种在表皮屏障功能中起关键作用的蛋白）上调。此外，还上调了其他上皮相关蛋白（如角蛋白 33b、34 和 85、表皮板蛋白、转谷氨酰胺酶-3、半乳糖凝集素 7 和半胱氨酸蛋白酶-14）。此外，还发现了细胞骨架调节蛋白（ vinculin 和肌球蛋白 9）和管家蛋白（延伸因子 2 和细胞质动力蛋白 1）的上调。丝聚合蛋白 2 和肌球蛋白 9 的上调通过免疫组织化学得到证实，后者蛋白与内皮细胞标志物 ETC 相关基因（ERG）共定位，表明巩膜组织中的新生血管化。
我们发现非感染性巩膜炎患者的巩膜中丝聚合蛋白 2 上调和新生血管化迹象。需要进一步的研究，最好包括更多的巩膜炎病例，以验证我们的发现。
