摘要
Oxidative stress-related damage to retinal pigment epithelial (RPE) cells is an important feature in the development of age-related macular degeneration. Iron-catalysed intralysosomal production of hydroxyl radicals is considered a major pathogenic factor, leading to lipofuscin formation with ensuing depressed cellular autophagic capacity, lysosomal membrane permeabilization and apoptosis. Previously, we have shown that cultured immortalized human RPE (ARPE-19) cells are extremely resistant to exposure to bolus doses of hydrogen peroxide and contain considerable amounts of the iron-binding proteins metallothionein (MT), heat-shock protein 70 (HSP70) and ferritin (FT). According to previous findings, autophagy of these proteins depresses lysosomal redox-active iron. The aim of this study was to investigate whether up- or downregulation of these proteins would affect the resistance of ARPE-19 cells to oxidative stress. Methods
The sensitivity of ARPE-19 cells to H2O2 exposure was tested following upregulation of MT, HSP70 and/or FT by pretreatment with ZnSO4, heat shock or FeCl3, as well as siRNA-mediated downregulation of the same proteins. Results
Upregulation of MT, HSP70 and FT did not improve survival following exposure to H2O2. This was interpreted as existence of an already maximal protection. Combined siRNA-mediated attenuation of both FT chains (H and L), or simultaneous downregulation of all three proteins, made the cells significantly more susceptible to oxidative stress confirming the importance of iron-binding proteins. Conclusion
The findings support our hypothesis that the oxidative stress resistance exhibited by RPE cells may be explained by a high autophagic influx of iron-binding proteins that would keep levels of redox-active lysosomal iron low.
摘要译文
视网膜色素上皮氧化应激相关的损害（RPE）细胞是在年龄相关性黄斑变性的发展的一个重要特征铁催化溶酶体内产生羟基自由基被认为是一个重要的致病因子，从而形成脂褐素与随之而来的沮丧细胞自噬能力，溶酶体膜通透性和apoptosisPreviously，ltured永生人RPE（ARPE-19）的细胞暴露极耐推注剂量的过氧化氢和含有大量的金属硫蛋白的铁结合蛋白（MT）热休克蛋白70（HSP70）和铁蛋白（FT）根据以前的研究结果，这些蛋白质的噬按压溶酶体氧化还原活性铁本研究的目的是调查上调或这些蛋白质的下调是否会影响ARPE-19细胞对氧化应激的抗性方法ARPE-19细胞和H的敏感性2 O 2曝光了测试下列MT，HSP70和/或FT的上调预处理与硫酸锌，热休克或氯化铁，以及相同的蛋白质siRNA介导下调结果MT的上调，HSP70和FT并没有提高生存率暴露于H 2 O 2这被解释为既FT链的一个已经最大protectionCombined siRNA-介导衰减的存在（H和L），或所有三种蛋白的同时下调，制成确认铁结合蛋白的重要性细胞显著氧化应激更敏感ř假设，即由RPE细胞表现出的氧化应激抗性可由铁结合蛋白的高自噬涌入，将保持氧化还原活性的溶酶体铁低的水平来解释
Markus Karlsson[1,*] and Tino Kurz[2]. Attenuation of iron-binding proteins in ARPE-19 cells reduces their resistance to oxidative stress[J]. Acta Ophthalmologica, 2016,94(6): 556-564