Ascorbate as a "redox sensor" and protector against irradiation-induced oxidative stress in 32D cl 3 hematopoietic cells and subclones overexpressing human manganese superoxide dismutase
Ascorbate as a "redox sensor" and protector against irradiation-induced oxidative stress in 32D cl 3 hematopoietic cells and subclones overexpressing human manganese superoxide dismutase
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DOI：
10.1016/j.ijrobp.2003.10.022
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发表时间：
2004-03-01
影响因子：
7
通讯作者：
Kagan, VE
中科院分区：
文献类型：
--
作者：
Epperly, MW;Osipov, AN;Kagan, VE
用途：为了确定锰超氧化物歧化酶（MnSOD）的表达增加是否通过防止活性氧（ROS）的产生来保护细胞免受辐射，使用抗坏血酸盐作为内源性自旋捕集器来检测自由基中间体的新方法。来自32 D cl 3造血细胞系或过表达MnSOD的亚克隆的细胞（2C 6）用脱氢抗坏血酸盐孵育30分钟，并照射至0至50戈伊的剂量。自由基中间体与自旋陷阱或抗坏血酸反应，测量电子自旋共振谱。将结果与辐射诱导的巯基水平变化、辐射存活曲线和8-OHdG积累（作为DNA氧化损伤的测量）进行比较。锰超氧化物歧化酶过表达的2C 6细胞维持较高水平的抗坏血酸与32 D cl 3亲本细胞相比，32 D cl 3亲本细胞的表型为（分别为5.4 +/- 0.5和2.6 +/- 0.5 nmol/10（6）细胞）和巯基（14.0 +/- 0.1和11.1 +/- 0.2 nmol/10（6）细胞）。细胞过表达MnSOD产生较低水平的活性氧比亲本32 D Cl 3细胞，证明了辐射后的抗坏血酸和GSH的支出较低。增加抗坏血酸水平保护32 D CL 3和2C 6细胞从辐射杀伤，如增加的肩膀上的生存曲线和减少DNA 8-OHdG accumulation.Conclusions：锰超氧化物歧化酶过表达保护2C 6细胞从辐射损伤清除ROS，很容易与主要的内源性抗氧化剂抗坏血酸和GSH-在未转染的造血32 D CL 3细胞。(C)2004爱思唯尔公司
Purpose: To determine whether increased expression of manganese superoxide dismutase (MnSOD) protects cells from irradiation by preventing the production of reactive oxygen species (ROS), a new approach to detecting free radical intermediates using ascorbate as an endogenous spin trap was used.Materials and Methods: Cells from the 32D cl 3 hematopoietic cell line or a subclone overexpressing MnSOD (2C6) were incubated with dehydroascorbate for 30 min and irradiated to doses from 0 to 50 Gy. Radical intermediates reacting with spin traps or ascorbate were measured by electron spin resonance spectroscopy. Results were compared to irradiation-induced changes in thiol levels, irradiation survival curves, and accumulation of 8-OHdG as a measurement of DNA oxidative damage.Results: Manganese superoxide dismutase-overexpressing 2C6 cells maintained higher levels of ascorbate (5.4 +/- 0.5 and 2.6 +/- 0.5 nmol/10(6) cells, respectively) and thiols (14.0 +/- 0.1 and 11.1 +/- 0.2 nmol/10(6) cells) compared to 32D cl 3 parent cells. Cells overexpressing MnSOD produced lower levels of ROS than did the parental 32D cl 3 cells, as evidenced by lower expenditure of ascorbate and GSH after irradiation. Increased ascorbate levels protected both 32D cl 3 and 2C6 cells from irradiation killing, as demonstrated by an increased shoulder on survival curves and decreased DNA 8-OHdG accumulation.Conclusions: Manganese superoxide dismutase overexpression protects 2C6 cells from irradiation damage by scavenging ROS that readily interact with major endogenous antioxidants-ascorbate and GSH-in nontransfected hematopoietic 32D cl 3 cells. (C) 2004 Elsevier Inc.