Lin T, Yang M S
Department of Biology, Hong Kong Baptist University, Kowloon, Hong Kong, PR China.
Toxicology. 2007 Jun 3;235(1-2):1-10. doi: 10.1016/j.tox.2007.03.002. Epub 2007 Mar 12.
Glutathione (GSH) is one of the most important antioxidants in mammalian cells. It also plays an important role in chemical detoxification. Some evidence showed that polycyclic aromatic hydrocarbons, such as benzo[a]pyrene (B[a]P [50-32-8]), could increase GSH content as a defense mechanism against oxidative stress as well as to promote its detoxification. However, there has been very little study on clarifying the role GSH plays in antioxidation and detoxification actions. Therefore, the present study aims to analyze intracellular glutathione metabolism in the human hepatoma cells (HepG2) upon exposure to B[a]P. Exposure of the cells to B[a]P (1-100 microM) for 24 h did not cause significant cell death in this cell line. By selecting the sublethal concentration of 10 microM, B[a]P caused a significant increase in GSH and a small (13%) but significant decrease in glutathione reductase activity. However, there was no change in the activity of glutathione peroxidase, and no detectable increase in reactive oxygen species (ROS) production. Treatment with B[a]P caused up to 1.5 folds increase in gamma-glutamylcysteine synthatase (gamma-GCS) activity over control. Buthioneine sulfoximine (BSO), an inhibitor of gamma-GCS, could suppress GSH increase in a dose-dependent manner. Assessment of the oxidative state of the cells indicated that the increase in GSH caused the cells to become more reduced. Thus, the results concluded that cells were not suffering from oxidative stress at 24 h after treatment with 10 microM B[a]P. Upon analyzing the activities of detoxification enzymes, there was an increase in the activity of CYP1A subfamily monooxygenases and glutathione S-transferase. Both changes occurred prior to the changes in gamma-GCS activity and the increase in GSH. In summary, results of the present study demonstrate that B[a]P caused an activation of detoxification enzymes. The increase in intracellular GSH level was due to activation of gamma-GCS activities. Oxidative stress may not be an important risk factor for B[a]P (at 10 microM of up to 24 h) induced injury.
谷胱甘肽（GSH）是哺乳动物细胞中最重要的抗氧化剂之一。它在化学解毒过程中也发挥着重要作用。一些证据表明，多环芳烃，如苯并[a]芘（B[a]P [50-32-8]），可增加GSH含量，作为对抗氧化应激的防御机制，并促进其解毒。然而，关于阐明GSH在抗氧化和解毒作用中所起作用的研究很少。因此，本研究旨在分析人肝癌细胞（HepG2）在暴露于B[a]P后细胞内谷胱甘肽的代谢情况。将细胞暴露于1-100微摩尔的B[a]P 24小时，在该细胞系中未引起明显的细胞死亡。通过选择10微摩尔的亚致死浓度，B[a]P导致GSH显著增加，谷胱甘肽还原酶活性小幅（13%）但显著下降。然而，谷胱甘肽过氧化物酶的活性没有变化，活性氧（ROS）的产生也没有可检测到的增加。用B[a]P处理导致γ-谷氨酰半胱氨酸合成酶（γ-GCS）活性比对照增加高达1.5倍。γ-GCS的抑制剂丁硫氨酸亚砜胺（BSO）可以剂量依赖性方式抑制GSH的增加。对细胞氧化状态的评估表明，GSH的增加使细胞变得更加还原。因此，结果得出结论，在用10微摩尔B[a]P处理24小时后，细胞没有遭受氧化应激。在分析解毒酶的活性时，CYP1A亚家族单加氧酶和谷胱甘肽S-转移酶的活性增加。这两种变化都发生在γ-GCS活性变化和GSH增加之前。总之，本研究结果表明，B[a]P导致解毒酶的激活。细胞内GSH水平的增加是由于γ-GCS活性的激活。氧化应激可能不是B[a]P（10微摩尔，长达24小时）诱导损伤的重要风险因素。
