Ogino T, Packer L, Maguire J J
Department of Molecular and Cellular Biology, Lawrence Berkeley National Laboratory, University of California, Berkeley 94720, USA.
Free Radic Biol Med. 1997;23(3):445-52. doi: 10.1016/s0891-5849(97)00115-9.
Low-molecular weight antioxidants in rat peritoneal neutrophils undergo rapid redox recycling, so measurements were made of their initial content and subsequent changes during the respiratory burst, when superoxide formation is maximized. Endogenous vitamin E, ascorbate and total glutathione (reduced + oxidized) were not significantly changed during 30 min of respiratory burst, which was stimulated by phorbol 12-myristate 13-acetate (PMA). When de novo synthesis of glutathione was inhibited by buthionine-[S,R] sulfoximine (BSO), the glutathione content rapidly decreased in activated neutrophils but not in resting cells. The lost total glutathione was recovered neither from the incubation medium nor as a protein-bound form, which suggests that irreversible oxidation of glutathione occurs. Furthermore, the glutathione loss continues even 30 min after PMA stimulation, when the respiratory burst has almost ceased. The decrease of glutathione was prevented by added catalase, or by addition of NaN3 or KCN which inhibits myeloperoxidase (MPO). Superoxide dismutase had no protective effects. These findings suggest the involvement of an MPO-H2O2-halide system in the accelerated consumption of glutathione during the respiratory burst. Additional studies showed that neutrophil-derived chloramines found in the extracellular medium could lead to intracellular glutathione loss. Incubation of resting cells with chemically produced membrane permeable monochloramine in the presence of BSO resulted in a decrease of glutathione, whereas membrane-impermeable taurine-chloramine was less effective. We conclude that chloramines are responsible for accelerated glutathione turnover in neutrophils during the respiratory burst. Permeable extracellular chloramines derived from the respiratory burst activity, such as monochloramine, can reenter cells and react with thiols.
大鼠腹膜中性粒细胞中的低分子量抗氧化剂会经历快速的氧化还原循环，因此在呼吸爆发期间（此时超氧化物生成最大化）对其初始含量及随后的变化进行了测量。在佛波酯12 -肉豆蔻酸酯13 -乙酸酯（PMA）刺激的30分钟呼吸爆发过程中，内源性维生素E、抗坏血酸和总谷胱甘肽（还原型 + 氧化型）没有显著变化。当谷胱甘肽的从头合成被丁硫氨酸 - [S，R]亚砜胺（BSO）抑制时，活化的中性粒细胞中谷胱甘肽含量迅速下降，而静息细胞中则没有。丢失的总谷胱甘肽既没有从孵育培养基中回收，也没有以蛋白质结合形式回收，这表明谷胱甘肽发生了不可逆氧化。此外，即使在PMA刺激30分钟后呼吸爆发几乎停止时，谷胱甘肽的损失仍在继续。添加过氧化氢酶，或添加抑制髓过氧化物酶（MPO）的NaN3或KCN可防止谷胱甘肽的减少。超氧化物歧化酶没有保护作用。这些发现表明，MPO - H2O2 -卤化物系统参与了呼吸爆发期间谷胱甘肽的加速消耗。进一步的研究表明，细胞外培养基中发现的中性粒细胞衍生的氯胺可导致细胞内谷胱甘肽损失。在存在BSO的情况下，用化学合成的可透过膜的一氯胺孵育静息细胞会导致谷胱甘肽减少，而不可透过膜的牛磺酸 - 氯胺效果较差。我们得出结论，氯胺是呼吸爆发期间中性粒细胞中谷胱甘肽周转加速的原因。源自呼吸爆发活性的可透过细胞外氯胺，如一氯胺，可重新进入细胞并与硫醇反应。
