1-Cys peroxiredoxin overexpression protects cells against phospholipid peroxidation-mediated membrane damage
1-Cys peroxiredoxin overexpression protects cells against phospholipid peroxidation-mediated membrane damage
复制标题
DOI：
10.1073/pnas.182384499
复制
发表时间：
2002-09-03
影响因子：
11.1
通讯作者：
Fisher, AB
中科院分区：
文献类型：
--
作者：
Manevich, Y;Sweitzer, T;Fisher, AB
1-Cys peroxiredoxin（1-cysPrx）是一种新型的抗氧化酶，能在体外以谷胱甘肽为还原剂还原磷脂过氧化氢。这种酶广泛表达，并在肺中富集。绿色荧光蛋白与1-cysPrx的融合蛋白在缺乏内源性酶的肺源性细胞系（NCI-H441）中稳定表达。过表达细胞（C17或C48）降解过氧化氢和叔丁基氢过氧化物更迅速，并表现出降低的敏感性，以氧化应激测定Cr-51释放。暴露于（OH）-O-。与H441相比，过表达细胞中硫代巴比妥酸反应物质和磷脂酰胆碱氢过氧化物含量的增加较少。这种效应可通过细胞谷胱甘肽的耗尽而逆转。二苯基-1-pyrenoylphosphonium荧光，用作膜磷脂过氧化反应的实时探针，增加立即暴露于铜+-天冬氨酸，并取消预孵育的细胞与Trolox（可溶性维生素E）或Tempol（自由基清除剂）。与H441相比，C17和C48细胞中的二苯基-1-芘酰基磷鎓荧光随Cu ~（2+）-Asc暴露而增加的速率显著减弱。膜联蛋白V-Cy 3用于检测磷脂酰丝氨酸从质膜的内叶到外叶的易位。Cu ~（2+）-Asc处理H441细胞2 h内可诱导磷脂酰丝氨酸移位，而C48细胞24 h内未见移位。这些结果表明，1-cysPrx可以消除过氧化物，但另外可以减少过氧化的膜磷脂。因此，该酶可以保护细胞免受氧化剂诱导的质膜损伤，从而在细胞防御氧化应激中发挥重要作用。
1-Cys peroxiredoxin (1-cysPrx) is a novel antioxidant enzyme able to reduce phospholipid hydroperoxides in vitro by using glutathione as a reductant. This enzyme is widely expressed and is enriched in lungs. A fusion protein of green fluorescent protein with 1-cysPrx was stably expressed in a lung-derived cell line (NCl-H441) lacking endogenous enzyme. Overexpressing cells (C17 or C48) degraded H2O2 and t-butylhydroperoxide more rapidly and showed decreased sensitivity to oxidant stress as measured by Cr-51 release. On exposure to (OH)-O-. generated by Cu2+-ascorbate (Asc), overexpressing cells compared with H441 showed less increase in thiobarbituric acid-reactive substance and phosphatidylcholine hydroperoxide content. This effect was reversed by depletion of cellular glutathione. Diphenyl-1-pyrenoylphosphonium fluorescence, used as a real-time probe of membrane phospholipid peroxidation, increased immediately on exposure to Cu2+-Asc and was abolished by preincubation of cells with Trolox (a soluble vitamin E) or Tempol (a radical scavenger). The rate of diphenyl-1-pyrenoylphosphonium fluorescence increase with Cu2+-Asc exposure was markedly attenuated in C17 and C48 cells as compared with H441. Annexin V-Cy3 was used to detect phosphatidylserine translocation from the inner to outer leaflet of the plasma membrane. Cu2+-Asc treatment induced phosphatidylserine translocation within 2 h in H441 cells but none was observed in C48 cells up to 24 h. These results indicate that 1-cysPrx can scavenge peroxides but in addition can reduce peroxidized membrane phospholipids. Thus, the enzyme can protect cells against oxidant-induced plasma membrane damage, thereby playing an important role in cellular defense against oxidant stress.