摘要
[目的]探讨a-硫辛酸(a-lipoic acid,LA)对外源性促氧化剂H2O2诱导中国仓鼠肺细胞(Chinese hamster lung cells,CHL)的活性氧(Reactive oxygen species,ROS)水平以及DNA氧化损伤的影响。[方法]①胞质内活性氧检测:用H2O2作外源性促氧化剂诱导细胞活性氧升高,以2’,7’-二氯荧光素二乙酸酯(2’,7’-dichlorofluorescin- diacetate,DCFH-DA)作为荧光信号标记物捕获胞质内活性氧,设置两种反应体系:体系Ⅰ为H2O2+LA,作用20min;体系Ⅱ为Fe2++LA,分别作用2、4、8h;借助流式细胞仪检测荧光信号强度。②单细胞凝胶电泳(Single cell gel electrophoresis, SCGE):用H2O2染毒细胞,诱发DNA氧化损伤,同时在反应体系中加入不同浓度LA反应2h,单细胞凝胶电泳分析DNA氧化损伤情况。[结果]ROS:①反应体系Ⅰ:H2O2引发细胞内活性氧骤升,但LA使活性氧水平剂量依赖性降低。②反应体系Ⅱ:Fe2+诱导胞质内活性氧升高,LA能抑制活性氧,降低荧光信号强度。100μmol／L Fe2+诱导细胞活性氧升高,并随时间增加而增强;LA能降低并持续抑制100pmol／I．Fe2-’诱导的活性氧升高,但未表现出明显的浓度依赖性。SCGE: H2O2造成细胞DNA断裂损伤,彗星率、彗星尾长、尾部DNA含量、olive尾矩等指标值均升高;加入LA后使上述各指标值剂量依赖性下降。[结论]LA能清除活性氧,抑制Fe2+诱导的活性氧升高,表现出剂量-反应关系和时间-效应关系;可能的机制为直接清除活性氧、阻断Fe2+介导的Fenton反应。LA发挥抗氧化作用,剂量依赖性降低H2O2诱导的DNA氧化损伤。
[ Objective ] To study the effects of α-Lipoic acid （ LA ） on reactive oxygen species （ ROS ） level and DNA oxidative damage in Chinese hamster lung cell. [ Methods ] ROS was detected by flow cytometer. Use 2＇, 7＇-dichlorofluorescin-diacetate （ DCFH-DA ） as fluorescent probe. Reaction system Ⅰ ： H2O2＋LA, incubated for 20 min. Reaction system Ⅱ ： Fe^2＋＋LA, incubated for 2, 4, 8 h respectively. Single cell gel electrophoresis （ SCGE ）： oxidative damage of cells were induced by H2O2. Certain concentrations of LA were added into reaction system, incubated for 2 h, and then DNA damage was detected with SCGE assay. [ Results ] （ 1 ）ROS： Reaction system Ⅰ： H2O2 heightened ROS level, but LA degraded it and showed the dose-response relationship. ROS increased after cells were incubated with Fe^2＋, LA also cut down ROS level. 100 μmol/L Fe^2＋ significantly elevated ROS, Although LA inhibited this effect, there is no conspicuous doseresponse relationship was observed. （ 2 ）SCGE： H2O2 caused DNA damage. The indices of DNA damage cells, Tail DNA, Tail length, Olive tail moment（ OTM ）were all increased. LA could inhibit these increasing indices dose-dependently. [ Conclusion ] LA scavenged ROS, inhibited ROS level induced by Fe^2＋, dose-response and time-reaction relationship were observed. LA protected cells from DNA damage caused by H2O2.
出处
《环境与职业医学》
CAS
北大核心
2007年第2期180-182,189,共4页
Journal of Environmental and Occupational Medicine
