Alvarez Ivan A, Lee Minsup, Eshaq Randa S, Leskova Wendy, Harris Norman R
School of Medicine, Louisiana State University Health Shreveport, 1501 Kings Hwy, Shreveport, LA 71103, USA.
Department of Molecular and Cellular Physiology, Louisiana State University Health Shreveport, 1501 Kings Hwy, Shreveport, LA 71103, USA.
Pathophysiology. 2024 Feb 6;31(1):89-99. doi: 10.3390/pathophysiology31010007.
Our purpose in this study was to identify the role played by oxidative stress in the changes to proteoglycans that occur under hyperglycemic conditions, using primary rat retinal microvascular endothelial cells (RRMEC) and cultured ophthalmic arteries. The cells and blood vessels obtained from rats were cultured in normal glucose (5.6 mM) and high glucose (25 mM) with or without N-acetylcysteine (NAC), an antioxidant. Intracellular oxidative stress was determined by measuring dihydroethidium (DHE) fluorescence and malondialdehyde (MDA)-modified protein levels. mRNA and protein levels were evaluated using quantitative real-time polymerase chain reaction and immunoblot, respectively. High glucose increased levels of glypican-1 mRNA and protein. The level of syndecan-1 mRNA also was increased, but its protein level was decreased, by high glucose. Evaluation of DHE and MDA showed that high glucose increased oxidative stress. These changes caused by high glucose were significantly reversed by NAC treatment. Matrix metalloproteinase-9 (MMP-9) levels, which increased under high glucose conditions, were suppressed by NAC treatment. Oxidative stress caused by hyperglycemia may be responsible for significant changes to the ocular endothelial glycocalyx.
在本研究中，我们的目的是利用原代大鼠视网膜微血管内皮细胞（RRMEC）和培养的眼动脉，确定氧化应激在高血糖条件下蛋白聚糖变化中所起的作用。从大鼠获取的细胞和血管在添加或不添加抗氧化剂N-乙酰半胱氨酸（NAC）的正常葡萄糖（5.6 mM）和高葡萄糖（25 mM）环境中培养。通过测量二氢乙锭（DHE）荧光和丙二醛（MDA）修饰的蛋白质水平来测定细胞内氧化应激。分别使用定量实时聚合酶链反应和免疫印迹评估mRNA和蛋白质水平。高葡萄糖增加了磷脂酰肌醇蛋白聚糖-1（glypican-1）的mRNA和蛋白质水平。高葡萄糖也增加了多功能蛋白聚糖-1（syndecan-1）的mRNA水平，但其蛋白质水平却降低了。对DHE和MDA的评估表明，高葡萄糖增加了氧化应激。NAC处理可显著逆转高葡萄糖引起的这些变化。在高葡萄糖条件下增加的基质金属蛋白酶-9（MMP-9）水平，受到NAC处理的抑制。高血糖引起的氧化应激可能是导致眼内皮糖萼发生显著变化的原因。
