文献类型：期刊文献

中文题名：水翁花对神经细胞氧化损伤保护作用的量效值(英文)

英文题名：Effects of cleistocalyx operculatus on oxygenic injury of nerve cells

作者：卢艳花[1];杜长斌[1];吴子斌[2];周文瑜[1];魏东芝[1]

机构：[1]华东理工大学生物反应器国家重点实验室,上海市200237;[2]香港中文大学生物化学系

年份：2005

卷号：9

期号：41

起止页码：171

中文期刊名：中国临床康复

外文期刊名：Chinese Journal of Clinical Rehabilitation

收录：北大核心:【北大核心2004】；CSCD:【CSCD_E2011_2012】；

语种：中文

中文关键词：水翁花;氧自由基;神经细胞;过氧化氢

摘要：背景:水翁花为中药桃金娘科植物水翁的干燥花蕾,已有报道水翁花提取物能通过抑制Na+/K+-ATP酶的活性加强心脏的收缩功能,同时降低心脏的收缩频率,水翁花提取物是否具有抗氧化方面的生物活性? 目的:观察水翁花对过氧化氢诱导的PC12神经细胞氧化损伤保护作用的量效关系。设计:非随机对照的实验。单位:华东理工大学生物工程学院生物反应器国家重点实验室。材料:实验于2002-05／11在华东理工大学反应器国家重点实验室鲁华生物技术研究所完成。选择昆明种雄性小鼠8只。PC12神经细胞购自中国科学院上海细胞所。方法:制备神经细胞氧化损伤模型。①水翁花细胞毒性测定:在96孔微量培养板中加入PC12神经细胞,水翁花以RPMI 1640培养液稀释成 0．001,0．01,0．1,0．5,1 g／L 5个浓度,每个浓度3孔,每孔接种2×103个细胞,另设空白对照组,即无药培养液组。标准条件下培养48 h,噻唑蓝比色法测定。②PC12神经细胞预先接种于96孔板中,培养24 h贴壁,分为正常对照组(正常细胞,不加H2O2和水翁花提取物)、0,0．01, 0．1,0．5,1 g／L水翁花提取物共7个组。除正常对照组外,其余细胞用 200 μmol／L的H2O2处理,加入不同浓度的水翁花水提物,作用12 h 后用噻唑蓝法测定细胞的存活率。③氧自由基水平测定:PC12神经细胞处理同存活率测定法,采用CDCFH染色法测定细胞氧自由基水平。主要观察指标:①水翁花提取物对PC12神经细胞的毒性。②水翁花提取物对氧化损伤的PC12神经细胞存活率影响。③水翁花提取物对氧化损伤的PC12神经细胞内、细胞外氧自由基水平的影响。结果:①水翁花提取物在浓度低于0．055 g／L时,对神经细胞有保护作用,在0．055-1．00 g／L浓度时,对细胞生长几乎无任何影响。②在低于 0．10g/L的浓度下,水翁花提取物对H2O2诱导的PC12神经细胞的氧化损伤没表现出明显的保护作用。但当浓度为1．00 g／L时,对H2O2诱导的PC12神经细胞的氧化损伤表现出明显的氧化修复能力。③H2O2损伤后,PC12细胞内、外氧自由基水平显著升高,当水翁花提取物浓度为 0．01g/L时,能明显的降低氧化损伤神经细胞内、外的氧自由基水平。结论:①水翁花水提物对H2O2诱导的PC12神经细胞的氧化损伤亦有很强的保护作用。②水翁花提取物抗氧化特性与提取液的浓度比有明显的关系,当浓度为1．00g/L时,具有明显的修复能力;当浓度为 0．01 g／L时能够降低细胞内外的氧自由基水平。
BACKGROUND： Cleistocalyx operculatus is a dried alsbastrum of myrtle. It is reported that cleistocalyx operculatus extracts can improve cardiac contraction through inhibiting the activity of Na^＋/K^＋-ATPase, and decrease rate of contraction. Do eleistocalyx operculatus extracts have the biological activity of antioxidation？ OBJECTIVE： To observe the effects of cleistocalyx operculatus on oxidative injury of PC12 nerve cells induced by H2O2. DESIGN： Non-randomized controlled study. SETTING： State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology. MATERIALS： The experiment was conducted at New World Institute of Biotechnology, State Key Laboratory of Bioreactor Engineering of East China University of Science and Technology, from May to November 2002. Eight adult male Kunming mice were selected. PC12 nerve cells were supplied by Shanghai Cell Institute of the Chinese Academy of Sciences. METHODS： Model of oxygenic injury of PC12 nerve cells was established. ① Assay of cytotoxicity of cleistocalyx operculatus to PC12 cells： PC12 cells were cultured in 96-well plates. Cleistocalyx operculatus was diluted with RPMI1640 culture medium into five concentrations of 0.001, 0.01, 0.1, 0.5 and 1 g/L with 3 wells in each concentration; each well had 2×10^3 cells. Blank control group, or non-drug culture medium group, was set. Under the standard condition, cells were cultured for 48 hours and assayed with thiazole blue chromatometry. ②Assay of survival rate： PC12 cells were inoculated in 96-well plate with the density of 2×10^3 for 24-hour wall adhering, and then divided into normal control group （normal cell without H2O2 or cleistocalyx operculatus extracts）, 0, 0.01, 0.1, 0.5 and 1 g/L cleistocalyx operculatus. Cells in all groups except normal control group were treated with 200 μmol/L H2O2 at 37℃ for 3 hours, then cleistocalyx operculatus of various concentrations was added and survival rate was assayed with thiazole blue method after 12 hours. ③ Level of oxygen-derived free radicals： PC12 cells with oxygen-derived free radicals were treated in the same way as done for cell survival rate assay and measured with CDCFH staining method. MAIN OUTCOME MEASURES： ① Toxicological effects of cleistocalyx operculatus on PC12 cells.② Protective effect of cleistocalyx operculatus extracts on the survival rate of PC12 nerve cells induced by oxidation. ③ Effect of cleistocalyx operculatus extracts on intracellular and extracellular oxygen-derived free radicals in PC12 nerve cells induced by oxidative injury. RESULTS： ① When the concentration was below 0.055 g/L, cleistocalyx operculatus could protect nerve cells; however, at 0.055-1.00 g/L the effect on cell growth did not significantly differ from that of blank control group. ② At the concentration below 0.10 g/L, cleistocalyx operculatus extracts had no protective effect on the injury of PC12 nerve cells induced by H202. At 1.00 g/L, it had strong plerosis for oxidative injury of PC12 nerve cells induced by H202. ③ After H202 injury, the level of intracellularand extracellular oxygen-derived free radicals in PC12 nerve cells was increased; however, at 0.01 g/L concentration of cleistocalyx operculatus extracts, the level was lower than that in model group. CONCLUSION： ① Cleistocalyx operculatus extracts not only inhibit oxidation of membrane lipid of hepatic microsome, but also protect against oxidative injury of PC12 nerve cells induced by H2O2. ②Antioxidation of cleistocalyx operculatus extracts is related to its concentration. At 1.00 g/L, it has great capacity of oxidation plerosis, and at 0.01 g/L it can decrease the level of oxygen-derived free radicals inside and outside cells.