Tyurina Yulia Y, Kawai Kazuaki, Tyurin Vladimir A, Liu Shang-Xi, Kagan Valerian E, Fabisiak James P
Department of Environmental and Occupational Health, Graduate School of Public Health, University of Pittsburgh, Pittsburgh, PA, USA.
Antioxid Redox Signal. 2004 Apr;6(2):209-25. doi: 10.1089/152308604322899288.
Phosphatidylserine (PS) externalization, a functional end point of apoptosis that triggers phagocytic recognition of dying cells, may be modulated by oxidative stress in biological membranes. We previously observed selective oxidation of PS during apoptosis, but the intracellular location and molecular mechanisms responsible for PS oxidation remain to be described. Peroxidation in individual classes of cellular phospholipids was monitored in whole cells and various subcellular fractions obtained from HL-60 cells undergoing apoptosis in response to tert-butyl hydroperoxide (t-BuOOH) after metabolic acylation of phospholipids with the oxidation-sensitive fluorescent fatty acid, cis-parinaric acid. Nonrandom selective oxidation of PS was observed in whole cells, as well as in plasma membrane. PS in mitochondria appeared selectively resistant to oxidation during apoptosis. All phospholipids in nuclear membranes appeared resistant to oxidation after t-BuOOH treatment. Selective PS oxidation was accompanied by cytochrome c release and PS externalization. PS oxidation and externalization were followed by caspase activation and other end points of apoptosis. HL-60 cells "loaded" with exogenous cytochrome c by mild sonication showed selective oxidation of PS in both the absence and presence of t-BuOOH. Cytochrome c/hydrogen peroxide could effectively oxidize purified PS but not phosphatidylcholine in a cell-free model system. Selective plasma membrane-based PS oxidation and subsequent externalization during oxidant-induced apoptosis may be mediated through the redox activity of cytochrome c.
磷脂酰丝氨酸（PS）外化是细胞凋亡的一个功能终点，可触发对死亡细胞的吞噬识别，它可能受到生物膜中氧化应激的调节。我们之前观察到细胞凋亡过程中PS的选择性氧化，但负责PS氧化的细胞内位置和分子机制仍有待描述。在用对氧化敏感的荧光脂肪酸顺式-紫黄质对磷脂进行代谢酰化后，使用叔丁基过氧化氢（t-BuOOH）诱导HL-60细胞凋亡，然后在全细胞和从这些细胞获得的各种亚细胞组分中监测各类细胞磷脂的过氧化情况。在全细胞以及质膜中均观察到PS的非随机选择性氧化。线粒体中的PS在细胞凋亡过程中似乎对氧化具有选择性抗性。叔丁基过氧化氢处理后，核膜中的所有磷脂似乎都对氧化具有抗性。PS的选择性氧化伴随着细胞色素c的释放和PS的外化。PS氧化和外化之后是半胱天冬酶激活及细胞凋亡的其他终点。通过温和超声处理“加载”外源性细胞色素c的HL-60细胞，在有无叔丁基过氧化氢的情况下均显示出PS的选择性氧化。在无细胞模型系统中，细胞色素c/过氧化氢可有效氧化纯化的PS，但不能氧化磷脂酰胆碱。在氧化剂诱导的细胞凋亡过程中，基于质膜的PS选择性氧化及随后的外化可能是通过细胞色素c的氧化还原活性介导的。
