Paller M S, Neumann T V
Department of Medicine, University of Minnesota, Minneapolis.
Kidney Int. 1991 Dec;40(6):1041-9. doi: 10.1038/ki.1991.312.
To study the importance of oxygen free radical production by and injury to proximal tubule epithelial cells, an in vitro model was established. Rat renal proximal tubule epithelial cells in primary culture were subjected to normoxic conditions or 60 minutes of hypoxia and 30 minutes of reoxygenation. Under normoxic conditions, these cells produced superoxide radical, hydrogen peroxide, and hydroxyl radical. During hypoxia and reoxygenation, there was an increase in the production of these reactive oxygen species, detected in the extracellular medium, of 252, 226, and 45 percent, respectively. The production rate of superoxide radical was most markedly increased in the first five minutes of reoxygenation. Studies employing 2,7-dichlorofluorescein which fluoresces when oxidized by peroxides revealed a seven-fold increase in cellular fluorescence in cells studied after hypoxia and reoxygenation compared with control cells. That increased production of reactive oxygen species played a role in cellular injury was demonstrated by an increase in lipid peroxidation during hypoxia and reoxygenation, as well as substantial injury during hypoxia and reoxygenation which could be largely prevented by the addition of superoxide dismutase, catalase, dimethylthiourea, or deferoxamine to the cells. These studies demonstrate that proximal tubule epithelial cells produce reactive oxygen species in increased amounts during hypoxia and reoxygenation, and that these reactive oxygen species are injurious to the cells under these conditions.
为研究近端肾小管上皮细胞产生氧自由基及受其损伤的重要性，建立了一个体外模型。原代培养的大鼠肾近端肾小管上皮细胞分别置于常氧条件下，或经历60分钟缺氧及30分钟复氧。在常氧条件下，这些细胞产生超氧阴离子自由基、过氧化氢和羟自由基。在缺氧和复氧期间，细胞外培养基中检测到的这些活性氧的产生分别增加了252%、226%和45%。超氧阴离子自由基的产生速率在复氧的前五分钟增加最为明显。使用2,7 - 二氯荧光素（被过氧化物氧化时会发出荧光）的研究显示，与对照细胞相比，缺氧和复氧后的细胞中细胞荧光增加了7倍。缺氧和复氧期间脂质过氧化增加，以及缺氧和复氧期间的大量损伤（通过向细胞中添加超氧化物歧化酶、过氧化氢酶、二甲基硫脲或去铁胺可在很大程度上预防）证明了活性氧产生增加在细胞损伤中起作用。这些研究表明，近端肾小管上皮细胞在缺氧和复氧期间产生的活性氧数量增加，并且在这些条件下这些活性氧对细胞具有损伤作用。
