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| 咖啡酸对氧化应激诱导的PC12细胞5-脂氧酶激活的抑制作用 | |
| 引用本文： | 李成檀,张丽慧,赵建波,方三华,张纬萍,魏尔清. 咖啡酸对氧化应激诱导的PC12细胞5-脂氧酶激活的抑制作用[J]. 中国药理学与毒理学杂志, 2011, 25(1): 45-51. DOI: 10.3867/j.issn.1000-3002.2011.01.009 |
| 作者姓名： | 李成檀 张丽慧 赵建波 方三华 张纬萍 魏尔清 |
| 作者单位： | 1. 杭州师范大学医学神经生物学市级重点实验室,基础医学部药理学教研室,浙江,杭州,310036;浙江大学医学院药理学系,浙江,杭州,310058 2. 杭州师范大学医学神经生物学市级重点实验室,基础医学部药理学教研室,浙江,杭州,310036 3. 浙江大学医学院药理学系,浙江,杭州,310058 |
| 基金项目： | 国家自然科学基金资助项目(30772561); 浙江省自然科学基金资助项目(Y204210); 杭州市科技计划项目(20080333B21)~~ |
| 摘 要： | 目的探讨咖啡酸是否通过抑制氧化应激诱导的5-脂氧酶(5-LOX)激活而减轻细胞损伤。方法稳定转染绿荧光蛋白(GFP)-5-LOX的PC12细胞,预先给予咖啡酸0.001~10μmol.L-1和对照药MK886,30min后观察缺氧缺糖/恢复(OGD/R)及过氧化氢(H2O2)160μmol.L-1处理后的变化。MTT法和碘化丙啶染色法分析细胞存活率和死亡率;荧光显微镜观察OGD 2 h/R2 h和H2O2处理40 min时5-LOX的核膜移位;ELISA法测定OGD 2 h/R 3 h时5-LOX代谢产物的生成;DCF法检测OGD 2 h/R 0.5 h细胞内活性氧(ROS)的产生。结果 OGD 2 h/R 24 h GFP-5-LOX转染和GFP-转染PC12细胞的存活率分别为(63.1±6.6)%和(70.7±6.9)%;H2O2处理24 h细胞存活率分别为(62.5±7.7)%和(75.7±9.5)%。在GFP-5-LOX转染的PC12细胞中,咖啡酸和对照药MK886可使OGD/R细胞存活率从(63.1±6.6)%分别增加到(87.3±2.0)%和(89.9±6.3)%,细胞坏死率从(31.4±1.5)%降低到(10.1±2.0)%和(11.7±1.3)%(P<0.01);使H2O2处理细胞存活率从(62.51±7.65)%增加到(92.59±4.02)%和(75.31±6.60)%;使OGD/R细胞CysLTs的生成从261.1±33.7降低到108.5±16.7和(90.6±19.5)ng.g-1蛋白(P<0.01)。此外,咖啡酸抑制OGD/R诱导PC12细胞的ROS产生,IC50值为8.021μmol.L-1;抑制OGD/R诱导的GFP-5-LOX转染的PC12细胞5-LOX核膜移位,IC50值为0.974μmol.L-1;抑制H2O2诱导的GFP-5-LOX转染的PC12细胞5-LOX核膜移位,IC50值为0.501μmol.L-1;MK886无上述作用。结论咖啡酸可抑制氧化应激诱导的PC12细胞5-LOX激活,对缺血损伤的PC12细胞具有保护作用。 |
| 关 键 词： | 咖啡酸 氧化应激 PC12细胞 5-脂氧酶 |
| 收稿时间： | 2010-01-14 |
Inhibitory effect of caffeic acid on 5-lipoxygenase activation induced by oxidative stress in PC12 cells |
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| LI Cheng-tan,ZHANG Li-hui,ZHAO Jian-bo,FANG San-hua,ZHANG Wei-ping,WEI Er-qing. Inhibitory effect of caffeic acid on 5-lipoxygenase activation induced by oxidative stress in PC12 cells[J]. Chinese Journal of Pharmacology and Toxicology, 2011, 25(1): 45-51. DOI: 10.3867/j.issn.1000-3002.2011.01.009 | |
| Authors: | LI Cheng-tan ZHANG Li-hui ZHAO Jian-bo FANG San-hua ZHANG Wei-ping WEI Er-qing |
| Affiliation: | (1. Key Laboratory of Neurobiology and Department of Pharmacology, School of Basic Medicine, Hangzhou Normal University, Hangzhou 310036, China; 2. Department of Pharmacology, School of Medicine, Zhejiang University, Hangzhou 310058, China) |
| Abstract: | OBJECTIVE To determine whether caffeic acid attenuates cell injury by inhibiting oxidative stress-induced 5-lipoxygenase (5-LOX) activation. METHODS In PC12 cells steadily expressed by green fluorescence protein (GFP)-5- LOX, caffeic acid or the 5 LOX activating protein inhibitor MK886 0.001-10 μmol·L-1 were added in cells for 30 min. The changes were observed after exposure to 2- h oxygen-glucose deprivation/2-h recovery (OGD/R) and hydrogen peroxide (H2O2) 160 μmol·L-1. Cell viability and cell death were assessed by MTT reduction assay and propidium iodide staining after treatment for 24 h. 5- LOX translocation to nuclear envelope was determined by fluorescence microscopy after 2-h OGD/2-h R or 40-min exposure to (H2O2). 5- LOX metabolites were evaluated by ELISA after 2-h OGD/3- h recovery, and production of reactive oxygen spices (ROS) was measured by DCF assay after 2-h OGD and 0.5-h recovery. RESULTS Cell viabilities were (63.1±6.6)% and (70.7±6.9)% after OGD 2 h/R 24 h; (62.5±7.7)% and (75.7±9.5)% after H2O2 treatment 24 h in GFP-5-LOX transfected and GFP transfected PC12 cells, respectively. In GFP-5- LOX transfected PC12 cells, caffeic acid and MK886 attenuated reduction of cell viability from (63.1±6.6)% to (87.3±2.0) % and (89.9±6.3)%, and reduced rate of necrotic cells from (31.4±1.5) % to (10.1±2.0)% and (11.7±1.3)% after OGD treatment. Caffeic acid and MK886 attenuated reduction in cell viability induced by H2O2 from (62.51±7.65)% to (92.59±4.02)% and (75.31±6.60)%. Both agents also reduced production of 5-LOX metabolites from 261.1±33.7 to 108.5±16.7 and (90.6±19.5)ng·g-1 protein after OGD/R in GFP-5- LOX transfected PC12 cells. Meanwhile, caffeic acid inhibited production of reactive oxygen species (ROS) after OGD/R in wild-type PC12 cells; IC50 was 8.021 μmol·L-1. Caffeic acid also inhibited 5- LOX translocation to the nuclear envelope induced by OGD/R and H2O2 in GFP-5-LOX transfected PC12 cells, respectively; IC50 was 0.974 and 0.501 μmol·L-1. However, MK886 did not show these effects. CONCLUSION Caffeic acid inhibits oxidative stress-induced 5-LOX activation in a concentration-dependent manner, and exerts a protective effect on ischemic-like injury in PC12 cells. |
| Keywords: | caffeic acid oxidative stress PC12 cells 5-lipoxygenase |
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