TRANSIENT ADAPTATION TO OXIDATIVE STRESS IN MAMMALIAN-CELLS
TRANSIENT ADAPTATION TO OXIDATIVE STRESS IN MAMMALIAN-CELLS
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DOI：
10.1006/abbi.1995.1225
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发表时间：
1995-04-01
影响因子：
3.9
通讯作者：
DAVIES, KJA
中科院分区：
文献类型：
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作者：
WIESE, AG;PACIFICI, RE;DAVIES, KJA
我们报告了几种哺乳动物细胞系对过氧化氢（H2 O2）暴露的氧化应激的短暂适应：中国仓鼠卵巢成纤维细胞（CHO）、HA-1细胞（一种确定的CHO亚克隆）、C3 H 10 T1/2细胞（胚胎小鼠成纤维细胞）、V79细胞2（中国仓鼠肺成纤维细胞）和克隆9肝细胞（大鼠肝上皮细胞）。在用相对低的H2 O2“引发”剂量预处理后，对H2 O2挑战的抗性发生高达40倍的适应性增加，从未处理细胞的低至1.9%的细胞活力到H2 O2预处理细胞的高达76.5%的活力。对HA-1细胞的详细研究揭示了以下对H2 O2的反应模式：0.1至0.5 μ mol/10（7）细胞的极低H2 O2浓度（3 - 15 μ M）刺激细胞生长25 - 45%; 2-5 μ mol/10（7）细胞的低H2 O2浓度（120 - 150 μ M）诱导暂时的生长停滞，细胞周期从18小时延长至约26小时，和H2 O2抗性的显著适应性增加; 9至14 μ mol 1/10（7）细胞（250至400 μ M）的中等H2 O2浓度引起永久性生长停滞（即，复制或分裂能力的永久丧失），无坏死迹象; 30 μ mol/10（7）细胞或更高（大于或等于1 mM）的高H2 O2浓度导致了类坏死性细胞死亡和破坏。HA-1细胞预处理后18小时，对2-5 μ mol/10（7）（120 - 150 μ M）低H2 O2浓度的适应性反应最大，此后向基线敏感性下降，并在7天固定和染色程序以及克隆形成活力测定中观察到。瞬时适应H2 O2预处理后的4.15 μ mol/10（7）（150 μ M）涉及从头合成至少20种蛋白质，并被阻断的翻译抑制剂，放线菌酮。在HA-1细胞的18小时适应过程中，蛋白质的合成分为三个阶段：早期（0-4小时），中期（4-8小时）和晚期（8-15小时）。没有H2 O2反应蛋白的合成超过18小时后预处理，时间适应已经最大化。选择性翻译抑制的早期，中期，或晚期蛋白质显示，所有三套是必要的最大的适应性增加过氧化氢的电阻。北方印迹和酶活性分析显示，经典的抗氧化酶过氧化氢酶，谷胱甘肽过氧化物酶，磷脂氢谷胱甘肽过氧化物酶，铜，锌超氧化物歧化酶，锰超氧化物歧化酶在H2 O2适应HA-1细胞的转录或翻译没有显着增加。此外，总的H2 O2消耗能力的HA-1细胞是不变的H2 O2适应。由于抗氧化应激是由抗氧化能力和各种损伤清除和修复酶的活性决定的，瞬时适应的HA-1细胞应该提供一个很好的模型来研究损伤清除和修复酶的诱导。(C)出版社：Academic Press
We report a transient adaptation to the oxidative stress of hydrogen peroxide (H2O2) exposure in several mammalian cell lines: Chinese hamster ovary fibroblast (CHO) cells, HA-1 cells (a defined CHO subclone), C3H 10T1/2 cells (embryonic mouse fibroblasts), V79 cells 2 (Chinese hamster lung fibroblasts), and Clone 9 liver cells (rat liver epithelial cells). Up to 40-fold adaptive increases in resistance to H2O2 challenge occurred following pretreatment with relatively low H2O2 ''priming'' doses, from as little as 1.9% cell viability for untreated cells to as much as 76.5% viability for H2O2 pretreated cells. Detailed studies with HA-1 cells revealed the following pattern of responses to H2O2: very low H2O2 concentrations of 0.1 to 0.5 mu mo1/10(7) cells (3 to 15 mu M) stimulated cell growth by 25 to 45%; low H2O2 concentrations of 2-5 mu mo1/10(7) cells (120 to 150 mu M) induced a temporary growth-arrest, a lengthening of cell cycle from 18 h to approximately 26 h, and marked adaptive increases in H2O2 resistance; intermediate H2O2 concentrations of 9 to 14 mu mo1/10(7) cells (250 to 400 mu M) caused permanent growth-arrest (i.e., permanent loss of replicative or divisional competence) with no evidence of necrosis; high H2O2 concentrations of 30 mu mol/10(7) cells or greater (greater than or equal to 1 mM) caused an apoptotic-like necrotic cell death and destruction. The adaptive response to low H2O2 concentrations of 2-5 mu mol/10(7) (120 to 150 mu M) was maximal 18 h after pretreatment of HA-1 cells, declined thereafter toward baseline sensitivity, and was observed with both 7-day fix and stain procedures and clonogenic viability assays. Transient adaptation following H2O2 pretreatment of 4.15 mu mol/10(7) (150 mu M) involved the de novo synthesis of at least 20 proteins and was blocked by the translation inhibitor, cycloheximide. During the 18-h adaptation in HA-1 cells proteins were synthesized in three phases; early (0-4 h), middle (4-8 h), and late (8-15 h). No H2O2 response proteins were synthesized beyond 18 h after pretreatment, by which time adaptation had already maximized. Selective translational inhibition of the early, middle, or late proteins revealed that all three sets were necessary for a maximal adaptive increase in H2O2 resistance. Northern blot and enzyme activity analyses revealed no significant increases in transcription or translation of the classical antioxidant enzymes catalase, glutathione peroxidase, phospholipid hydroperoxide glutathione peroxidase, Cu, Zn superoxide dismutase, or Mn superoxide dismutase in H2O2-adapted HA-1 cells. Furthermore, the total H2O2-consuming capacity of HA-1 cells was unchanged by H2O2 adaptation. Since resistance to oxidative stress is determined by both antioxidant capacity and the activities of various damage removal and repair enzymes, transiently adapted HA-1 cells should provide a good model in which to study the induction of damage removal and repair enzymes. (C) 1995 Academic Press, Inc.