Chronic ocular pathologies such as cataracts and glaucoma are emerging as an important problem for public health due to the changes in lifestyle and longevity. These age-related ocular diseases are largely mediated by oxidative stress. Small extracellular vesicles (sEVs) are involved in cell-to-cell communication and transport. There is an increasing interest about the function of small extracellular vesicles (sEVs) in the eye. However, the proteome content and characterization of sEVs released by ocular cells under pathological conditions are not yet well known. Here, we aimed to analyze the protein profile of sEVs and the intracellular protein content from two ocular cell lines (lens epithelial cells and retinal ganglion cells) exposed to oxidative stress to identify altered proteins that could serve as potential diagnostic biomarkers. The protein content was analyzed by quantitative mass spectrometry-based proteomics. Validation was performed by WB and ELISA using cell extracts and aqueous humor from cataract and glaucoma patients. After data analysis, 176 and 7 dysregulated proteins with an expression ratio≥1.5 were identified in lens epithelial cells’ protein extract and sEVs, respectively, upon oxidative stress induction. In retinal ganglion cells, oxidative stress induction resulted in the dysregulation of 1033 proteins in cell extracts and 9 proteins in sEVs. In addition, by WB and ELISA, the dysregulation of proteins was mostly confirmed in aqueous humor samples from cataract or glaucoma patients in comparison to ICL individuals, with RAD23B showing high glaucoma diagnostic ability. Importantly, this work expands the knowledge of the proteome characterization of cataracts and glaucoma and provides new potential diagnostic glaucoma biomarkers.

中文翻译：

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通过深入的蛋白质组学揭示氧化应激、眼部疾病和小细胞外囊泡的联系，以识别新的青光眼生物标志物


由于生活方式和寿命的变化，白内障和青光眼等慢性眼部疾病正在成为公共卫生的重要问题。这些与年龄相关的眼部疾病主要由氧化应激介导。小细胞外囊泡 （sEV） 参与细胞间通讯和运输。人们对眼中小细胞外囊泡 （sEV） 的功能越来越感兴趣。然而，在病理条件下，眼细胞释放的 sEVs 的蛋白质组含量和特征尚不清楚。在这里，我们旨在分析暴露于氧化应激的两种眼细胞系 （晶状体上皮细胞和视网膜神经节细胞） 的 sEVs 蛋白谱和细胞内蛋白质含量，以确定可能作为潜在诊断生物标志物的改变蛋白质。通过基于定量质谱的蛋白质组学分析蛋白质含量。使用 WB 和 ELISA 使用白内障和青光眼患者的细胞提取物和房水进行验证。经过数据分析，在氧化应激诱导后，晶状体上皮细胞的蛋白提取物和 sEVs 中分别鉴定出 176 个和 7 个表达比≥为 1.5 的失调蛋白。在视网膜神经节细胞中，氧化应激诱导导致细胞提取物中 1033 种蛋白质和 sEVs 中 9 种蛋白质失调。此外，通过 WB 和 ELISA，与 ICL 个体相比，白内障或青光眼患者的房水样本中蛋白质失调大多得到证实，RAD23B 表现出较高的青光眼诊断能力。重要的是，这项工作扩展了白内障和青光眼蛋白质组特征的知识，并提供了新的潜在诊断青光眼生物标志物。