Peng Peng, Yu Yaoyao, Ma Wenhui, Lyu Shanmei, Ma Li, Liu Ting, Dong Yanling, Wei Chao
Shandong First Medical University and Shandong Academy of Medical Sciences, Jinan, Shandong, China; State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, Eye Institute of Shandong First Medical University, Qingdao, China.
State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, Eye Institute of Shandong First Medical University, Qingdao, China.
Exp Eye Res. 2023 May;230:109457. doi: 10.1016/j.exer.2023.109457. Epub 2023 Mar 21.
Corneal endothelial decompensation (CED) is the major cause of the long-term graft failure, but the underlying mechanisms remain unclear. The purpose of this study was to characterize the proteomic profile in CED aqueous humor (AH) after penetrating keratoplasty (PKP). We collected AH samples (n = 6/group) from CED patients underwent PKP and cataract patients, respectively. The label-free quantitative proteomic analysis was performed to identify the differentially-expressed proteins (DEPs). The biological functions of DEPs were evaluated using Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genome (KEGG) analysis. The protein-protein interaction (PPI) network construction was employed to distinguish the hub proteins of DEPs, and the selected proteins were validated by parallel reaction monitoring (PRM). The human peripheral blood mononuclear cells (PBMCs) were adopted to investigate the effect of biglycan (BGN) on inflammatory response, and the subsequent outcomes of inflammation on human corneal endothelial cells (HCECs). A total of 174 DEPs were identified in CED AH of patients underwent PKP, including 102 up-regulated proteins and 72 down-regulated proteins. Bioinformatics analysis revealed the significant enrichment of cytokine-mediated signaling pathway and extracellular matrix (ECM) organization in the up-regulated proteins, as well as the alterations of cellular components, including the increase of collagen and complement component C1 complex, and reduction in extracellular exosomes. A hub protein cluster of 15 proteins was determined by Molecular Complex Detection (MCODE), including FN1, BGN, COMP, COL11A1, COLA3A1, and COL1A1. Moreover, BGN promoted pro-inflammatory cytokine (such as TNF-α, IL-1β and IL-6) production in PBMCs through NF-κB signaling pathway, which subsequently resulted in HCECs death. These findings provided a systemic protein profile of AH in CED patients after corneal transplantation, with the alterations implicated in cytokine-mediated signaling, ECM, complement system, and exsomes. The identified proteins and signaling pathways probably paved the novel insight into understanding the pathogenesis of the disease.
角膜内皮失代偿（CED）是长期移植失败的主要原因，但其潜在机制仍不清楚。本研究的目的是表征穿透性角膜移植术（PKP）后CED房水（AH）中的蛋白质组学特征。我们分别从接受PKP的CED患者和白内障患者中收集AH样本（每组n = 6）。进行无标记定量蛋白质组学分析以鉴定差异表达蛋白（DEP）。使用基因本体论（GO）分析和京都基因与基因组百科全书（KEGG）分析评估DEP的生物学功能。采用蛋白质-蛋白质相互作用（PPI）网络构建来区分DEP的枢纽蛋白，并通过平行反应监测（PRM）验证所选蛋白。采用人外周血单核细胞（PBMC）研究双糖链蛋白聚糖（BGN）对炎症反应的影响，以及炎症对人角膜内皮细胞（HCEC）的后续影响。在接受PKP的患者的CED AH中总共鉴定出174种DEP，包括102种上调蛋白和72种下调蛋白。生物信息学分析显示，上调蛋白中细胞因子介导的信号通路和细胞外基质（ECM）组织显著富集，以及细胞成分的改变，包括胶原蛋白和补体成分C1复合物的增加，以及细胞外囊泡的减少。通过分子复合物检测（MCODE）确定了一个由15种蛋白质组成的枢纽蛋白簇，包括纤连蛋白1（FN1）、双糖链蛋白聚糖（BGN）、软骨寡聚基质蛋白（COMP）、11型胶原α1链（COL11A1）、3型胶原α1链（COLA3A1）和1型胶原α1链（COL1A1）。此外，BGN通过NF-κB信号通路促进PBMC中促炎细胞因子（如TNF-α、IL-1β和IL-6）的产生，随后导致HCEC死亡。这些发现提供了角膜移植后CED患者AH的系统蛋白质谱，其改变与细胞因子介导的信号传导、ECM、补体系统和外泌体有关。鉴定出的蛋白质和信号通路可能为理解该疾病的发病机制提供了新的见解。
