Mitochondrial superoxide anions induced by exogenous oxidative stress determine tumor cell fate: an individual cell-based study
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Reactive oxygen species (ROS) are involved in a variety of biological phenomena and serve both deleterious and beneficial roles. ROS quantification and assessment of reaction networks are desirable but difficult because of their short half-life and high reactivity. Here, we describe a pro-oxidative model in a single human lung carcinoma SPC-A-1 cell that was created by application of extracellular H2O2 stimuli.
Modified microfluidics and imaging techniques were used to determine O2•− levels and construct an O2•− reaction network. To elucidate the consequences of increased O2•− input, the mitochondria were given a central role in the oxidative stress mode, by manipulating mitochondria-interrelated cytosolic Ca2+ levels, mitochondrial Ca2+ uptake, auto-amplification of intra-cellular ROS and the intrinsic apoptotic pathway.
Results and conclusions
Results from a modified microchip demonstrated that 1 mmol/L H2O2 induced a rapid increase in cellular O2•− levels (>27 vs. >406 amol in 20 min), leading to increased cellular oxidizing power (evaluated by ROS levels) and decreased reducing power (evaluated by glutathione (GSH) levels). In addition, we examined the dynamics of cytosolic Ca2+ and mitochondrial Ca2+ by confocal laser scanning microscopy and confirmed that Ca2+ stores in the endoplasmic reticulum were the primary source of H2O2-induced cytosolic Ca2+ bursts. It is clear that mitochondria have pivotal roles in determining how exogenous oxidative stress affects cell fate. The stress response involves the transfer of Ca2+ signals between organelles, ROS auto-amplification, mitochondrial dysfunction, and a caspase-dependent apoptotic pathway.
Key wordsIndividual cell Superoxide anion Reactive oxygen species (ROS) dynamics Intrinsic apoptotic pathway Ca2+ signaling
通过细胞外过氧化氢 (H2O2) 的刺激建立单个人 肺癌SPC-A-1 细胞的氧化压力模型。
氧自由基 (ROS) 涉及多种生物现象, 包括有益 和有害两个方面。ROS 的定量检测和反应网络的 评估结果令人期待。但ROS 半衰期很短且反应过 程很快, 因此, 我们通过多种手段克服了检测和 评估的困难。
利用改进的微流控和成像技术测定 ROS 水平, 构 建氧反应网络。通过调控线粒体胞浆Ca2+水平、 线粒体 Ca2+摄取、细胞内ROS 自扩增以及内在 凋亡途径, 确定线粒体在外源氧化压力模式中扮 演的角色。
研究结果表明1 mmol/L H2O2 引起细胞O2•−水平 的快速增加, 从而导致细胞氧化能力增加和还原 能力降低。此外, 研究还证实了内质网中储存的 Ca2+是H2O2 诱导的线粒体Ca2+爆发的主要来源。 外源氧化压力反应涉及细胞器间Ca2+信号的传 递、ROS 自身扩增、线粒体功能紊乱和半胱天冬 酶依赖性凋亡途径。线粒体在外源性氧化应激影 响细胞命运方面发挥着关键作用。
关键词单个细胞 超氧阴离子 氧自由基动力学 内源性凋亡途径 钙信号
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- Akopova OV, 2008. The role of mitochondrial permeability transition pore in transmembrane Ca2+-exchange in mitochondria. Ukr Biokhim Zh (1999), 80(3):40–47 (in Ukrainian).Google Scholar
- Liochev SI, 2013. Reactive oxygen species and the free radical theory of aging. Free Radic Biol Med, 60:1–4. https://doi.org/10.1016/j.freeradbiomed.2013.02.011 CrossRefGoogle Scholar
- Lyublinskaya OG, Zenin VV, Shatrova AN, et al., 2014. Intracellular oxidation of hydroethidine: compartmentalization and cytotoxicity of oxidation products. Free Radic Biol Med, 75:60–68. https://doi.org/10.1016/j.freeradbiomed.2014.07.008 CrossRefGoogle Scholar
- Qin Y, Pan X, Tang TT, et al., 2011. Anti-proliferative effects of the novel squamosamide derivative (FLZ) on HepG2 human hepatoma cells by regulating the cell cycle-related proteins are associated with decreased Ca2+/ROS levels. Chem Biol Interact, 193(3):246–253. https://doi.org/10.1016/j.cbi.2011.07.004 CrossRefGoogle Scholar
- Tang HY, Qin Y, Li JY, et al., 2011. The scavenging of superoxide radicals promotes apoptosis induced by a novel cell-permeable fusion protein, sTRAIL:FeSOD, in tumor necrosis factor-related apoptosis-inducing ligand-resistant leukemia cells. BMC Biol, 9: 18. https://doi.org/10.1186/1741-7007-9-18 CrossRefGoogle Scholar
- Zhao HT, Kalivendi S, Zhang H, et al., 2003. Superoxide reacts with hydroethidine but forms a fluorescent product that is distinctly different from ethidium: potential implications in intracellular fluorescence detection of superoxide. Free Radic Biol Med, 34(11):1359–1368. https://doi.org/10.1016/S0891-5849(03)00142-4 CrossRefGoogle Scholar