The augmentation of O-GlcNAcylation reduces glyoxal-induced cell injury by attenuating oxidative stress in human retinal microvascular endothelial cells.
The augmentation of O-GlcNAcylation reduces glyoxal-induced cell injury by attenuating oxidative stress in human retinal microvascular endothelial cells.
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O-GlcNAcylation 的增强通过减轻人视网膜微血管内皮细胞的氧化应激来减少乙二醛诱导的细胞损伤
DOI：
10.3892/ijmm.2015.2319
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发表时间：
2015-10
影响因子：
5.4
通讯作者：
Wang F
中科院分区：
文献类型：
--
作者：
Liu GD;Xu C;Feng L;Wang F
最近有报道称，人视网膜微血管内皮细胞（HRECs）中O-linked β-N-acetyl glucosamine （O-GlcNAc）修饰(一种简单的细胞内丝氨酸（Ser)/苏氨酸（Thr）连接的单糖）与糖尿病视网膜病变（DR）有关。在o - glcn酰化过程中，O-GlcNAc被添加到Ser和Thr残基上。活性氧（ROS）的生成是晚期糖基化终末产物（AGE）损伤的特征之一，也是DR最重要的关键致病因素，因此本研究旨在探讨o - glcnac酰化与ROS生成之间的关系，以确定o - glcnac酰化是否通过生成ROS来减轻细胞损伤。为此，将HRECs分为4组：正常葡萄糖（5 mM）处理的HRECs，乙二醛（500µM）处理的HRECs，乙二醛处理的HRECs还使用200µM PUGNAc处理，乙二醛处理的HRECs感染O-GlcNAc转移酶（OGT） siRNA。我们检测到乙二醛处理的HRECs中O-GlcNAc水平升高，ROS生成增加。通过细胞ROS染色和测定抗氧化基因、超氧化物歧化酶（SOD）和谷胱甘肽过氧化物酶（GPX）的表达水平来测定细胞氧化还原状态。PUGNAc处理后的o - glcnac酰化水平升高显著降低了ROS的产生（P<0.01），提高了SOD和GPX的表达水平，而OGT siRNA感染后的o - glcnac酰化水平降低，加剧了ROS的产生（P<0.01），降低了抗氧化基因的表达。o - glcn酰化对HRECs活力的影响显著（P<0.01），特别是过氧化氢（H2O2）处理的HRECs。PUGNAc治疗可减少乙二醛诱导的细胞凋亡，转染OGT siRNA可增加HREC细胞凋亡；流式细胞术和线粒体膜电位测定证实了这些结果。o - glcnac酰化的增加通过减少ROS的产生，增加抗氧化基因的表达，阻止线粒体膜电位的耗散，防止HREC凋亡，对HRECs发挥细胞保护作用。由此可见，o - glcn酰化在DR的早期发育过程中发挥了作用。
It has recently been reported that O-linked β-N-acetyl glucosamine (O-GlcNAc) modification (a simple intracellular serine (Ser)/threonine (Thr)-linked monosaccharide) in human retinal microvascular endothelial cells (HRECs) is related to diabetic retinopathy (DR). During O-GlcNAcylation, O-GlcNAc is added to Ser and Thr residues. As the generation of reactive oxygen species (ROS) is one of the characteristics of advanced glycation end product (AGE) injury, and the most important key pathogenic factor of DR, in the present study, we aimed to investigate the association between O-GlcNAcylation and ROS generation in order to ascertain whether O-GlcNAcylation mitigates cellular injury through the generation of ROS. For this purpose, HRECs were divided into 4 groups as follows: HRECs treated with normal glucose (5 mM), HRECs treated with glyoxal (500 µM), glyoxal-treated HRECs also treated with 200 µM PUGNAc, and glyoxal-treated HRECs infected with O-GlcNAc transferase (OGT) siRNA. We detected increased O-GlcNAc levels and increased ROS production in the glyoxal-treated HRECs. The cellular redox status was determined by cellular ROS staining and by measuring the expression levels of the antioxidant genes, superoxide dismutase (SOD) and glutathione peroxidase (GPX). While the augmentation of O-GlcNAcylation following treatment with PUGNAc significantly attenuated the production of ROS (P<0.01) and increased the expression levels of SOD and GPX, the reduction of O-GlcNAcylation following infection with OGT siRNA, exacerbated the production of ROS (P<0.01) and decreased the expression of antioxidant genes. The effects of O-GlcNAcylation on the viability of HRECs were significant (P<0.01), particularly in the hydrogen peroxide (H2O2)-treated HRECs. Treatment with PUGNAc reduced glyoxal-induced cell apoptosis and transfection with OGT siRNA increased HREC apoptosis; these results were confirmed by flow cytometry and by the assessment of mitochondrial membrane potential. The augmentation of O-GlcNAcylation exerted cytoprotective effects on the HRECs by reducing the generation of ROS, increasing the expression of antioxidant genes, preventing the dissipation of mitochondrial membrane potential and preventing HREC apoptosis. Therefore, it can be concluded that O-GlcNAcylation plays a role in the early developmental process of DR.