Jeong Jae Ick, Lee Young Woo, Kim Yong Keun
Department of Neurosurgery, College of Medicine, Pusan National University, Pusan 602-739, Korea.
Neurochem Res. 2003 Aug;28(8):1201-11. doi: 10.1023/a:1024280429036.
This study was undertaken to evaluate whether chemical hypoxia-induced cell injury is a result of reactive oxygen species (ROS) generation, ATP depletion, mitochondrial permeability transition, and an increase in intracellular Ca2+, in A172 cells, a human glioma cell line. Chemical hypoxia was induced by incubating cells with antimycin A, an inhibitor of mitochondrial electron transport, in a glucose-free medium. Exposure of cells to chemical hypoxia resulted in cell death, ROS generation, ATP depletion, and mitochondrial permeability transition. The H2O2 scavenger pyruvate prevented cell death, ROS generation, and mitochondrial permeability transition induced by chemical hypoxia. In contrast, changes mediated by chemical hypoxia were not affected by hydroxyl radical scavengers. Antioxidants did not affect cell death and ATP depletion induced by chemical hypoxia, although they prevented ROS production and mitochondrial permeability transition induced by chemical hypoxia. Chemical hypoxia did not increase lipid peroxidation even when antimycin A was increased to 50 microM, whereas the oxidant t-butylhydroperoxide caused a significant increase in lipid peroxidation, at a concentration that is less effective than chemical hypoxia in inducing cell death. Fructose protected against cell death and mitochondrial permeability transition induced by chemical hypoxia. However, ROS generation and ATP depletion were not prevented by fructose. Chemical hypoxia caused the early increase in intracellular Ca2+. The cell death and ROS generation induced by chemical hypoxia were altered by modulation of intracellular Ca2+ concentration with ruthenium red, TMB-8, and BAPTA/AM. However, mitochondrial permeability transition was not affected by these compounds. These results indicate that chemical hypoxia causes cell death, which may be, in part, mediated by H2O2 generation via a lipid peroxidation-independent mechanism and elevated intracellular Ca2+. In addition, these data suggest that chemical hypoxia-induced cell death is not associated directly with ATP depletion and mitochondrial permeability transition.
本研究旨在评估在人胶质瘤细胞系A172细胞中，化学性缺氧诱导的细胞损伤是否是由活性氧（ROS）生成、ATP耗竭、线粒体通透性转换以及细胞内Ca2+增加所致。通过在无葡萄糖培养基中用线粒体电子传递抑制剂抗霉素A孵育细胞来诱导化学性缺氧。将细胞暴露于化学性缺氧会导致细胞死亡、ROS生成、ATP耗竭和线粒体通透性转换。H2O2清除剂丙酮酸可预防化学性缺氧诱导的细胞死亡、ROS生成和线粒体通透性转换。相比之下，化学性缺氧介导的变化不受羟基自由基清除剂的影响。抗氧化剂虽可预防化学性缺氧诱导的ROS产生和线粒体通透性转换，但不影响化学性缺氧诱导的细胞死亡和ATP耗竭。即使将抗霉素A增加至50μM，化学性缺氧也不会增加脂质过氧化，而氧化剂叔丁基过氧化氢在诱导细胞死亡方面比化学性缺氧效果差的浓度下，会导致脂质过氧化显著增加。果糖可预防化学性缺氧诱导的细胞死亡和线粒体通透性转换。然而，果糖不能预防ROS生成和ATP耗竭。化学性缺氧导致细胞内Ca2+早期增加。用钌红、TMB - 8和BAPTA/AM调节细胞内Ca2+浓度可改变化学性缺氧诱导的细胞死亡和ROS生成。然而，这些化合物不影响线粒体通透性转换。这些结果表明，化学性缺氧导致细胞死亡，这可能部分是由通过脂质过氧化非依赖机制产生的H2O2和细胞内Ca2+升高介导的。此外，这些数据表明化学性缺氧诱导的细胞死亡与ATP耗竭和线粒体通透性转换没有直接关联。
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