CAJ | 학술논문

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)、0,0.01,0.1,0.5,1g/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0.10 g/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0.01 g/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1.00 g/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. 揹景:水翁花為中藥桃金孃科植物水翁的榦燥花蕾,已有報道水翁花提取物能通過抑製Na+/K+-ATP酶的活性加彊心髒的收縮功能,同時降低心髒的收縮頻率,水翁花提取物是否具有抗氧化方麵的生物活性?目的:觀察水翁花對過氧化氫誘導的PC12神經細胞氧化損傷保護作用的量效關繫.設計:非隨機對照的實驗.單位:華東理工大學生物工程學院生物反應器國傢重點實驗室.材料:實驗于2002-05/11在華東理工大學反應器國傢重點實驗室魯華生物技術研究所完成.選擇昆明種雄性小鼠8隻.PC12神經細胞購自中國科學院上海細胞所.方法:製備神經細胞氧化損傷模型.①水翁花細胞毒性測定:在96孔微量培養闆中加入PC12神經細胞,水翁花以RPMI 1640培養液稀釋成0.001,0.01,0.1,0.5,1 g/L 5箇濃度,每箇濃度3孔,每孔接種2×103箇細胞,另設空白對照組,即無藥培養液組.標準條件下培養48 h,噻唑藍比色法測定.②PC12神經細胞預先接種于96孔闆中,培養24 h貼壁,分為正常對照組(正常細胞,不加H2O2和水翁花提取物)、0,0.01,0.1,0.5,1g/L水翁花提取物共7箇組.除正常對照組外,其餘細胞用200 μmol/L的H2O2處理,加入不同濃度的水翁花水提物,作用12 h後用噻唑藍法測定細胞的存活率.③氧自由基水平測定:PC12神經細胞處理同存活率測定法,採用CDCFH染色法測定細胞氧自由基水平.主要觀察指標:①水翁花提取物對PC12神經細胞的毒性.②水翁花提取物對氧化損傷的PC12神經細胞存活率影響.③水翁花提取物對氧化損傷的PC12神經細胞內、細胞外氧自由基水平的影響.結果:①水翁花提取物在濃度低于0.055 g/L時,對神經細胞有保護作用,在0.055～1.00 g/L濃度時,對細胞生長幾乎無任何影響.②在低于0.10 g/L的濃度下,水翁花提取物對H2O2誘導的PC12神經細胞的氧化損傷沒錶現齣明顯的保護作用.但噹濃度為1.00 g/L時,對H2O2誘導的PC12神經細胞的氧化損傷錶現齣明顯的氧化脩複能力.③H2O2損傷後,PC12細胞內、外氧自由基水平顯著升高,噹水翁花提取物濃度為0.01 g/L時,能明顯的降低氧化損傷神經細胞內、外的氧自由基水平.結論:①水翁花水提物對H2O2誘導的PC12神經細胞的氧化損傷亦有很彊的保護作用.②水翁花提取物抗氧化特性與提取液的濃度比有明顯的關繫,噹濃度為1.00 g/L時,具有明顯的脩複能力;噹濃度為0.01 g/L時能夠降低細胞內外的氧自由基水平.
배경:수옹화위중약도금낭과식물수옹적간조화뢰,이유보도수옹화제취물능통과억제Na+/K+-ATP매적활성가강심장적수축공능,동시강저심장적수축빈솔,수옹화제취물시부구유항양화방면적생물활성?목적:관찰수옹화대과양화경유도적PC12신경세포양화손상보호작용적량효관계.설계:비수궤대조적실험.단위:화동리공대학생물공정학원생물반응기국가중점실험실.재료:실험우2002-05/11재화동리공대학반응기국가중점실험실로화생물기술연구소완성.선택곤명충웅성소서8지.PC12신경세포구자중국과학원상해세포소.방법:제비신경세포양화손상모형.①수옹화세포독성측정:재96공미량배양판중가입PC12신경세포,수옹화이RPMI 1640배양액희석성0.001,0.01,0.1,0.5,1 g/L 5개농도,매개농도3공,매공접충2×103개세포,령설공백대조조,즉무약배양액조.표준조건하배양48 h,새서람비색법측정.②PC12신경세포예선접충우96공판중,배양24 h첩벽,분위정상대조조(정상세포,불가H2O2화수옹화제취물)、0,0.01,0.1,0.5,1g/L수옹화제취물공7개조.제정상대조조외,기여세포용200 μmol/L적H2O2처리,가입불동농도적수옹화수제물,작용12 h후용새서람법측정세포적존활솔.③양자유기수평측정:PC12신경세포처리동존활솔측정법,채용CDCFH염색법측정세포양자유기수평.주요관찰지표:①수옹화제취물대PC12신경세포적독성.②수옹화제취물대양화손상적PC12신경세포존활솔영향.③수옹화제취물대양화손상적PC12신경세포내、세포외양자유기수평적영향.결과:①수옹화제취물재농도저우0.055 g/L시,대신경세포유보호작용,재0.055～1.00 g/L농도시,대세포생장궤호무임하영향.②재저우0.10 g/L적농도하,수옹화제취물대H2O2유도적PC12신경세포적양화손상몰표현출명현적보호작용.단당농도위1.00 g/L시,대H2O2유도적PC12신경세포적양화손상표현출명현적양화수복능력.③H2O2손상후,PC12세포내、외양자유기수평현저승고,당수옹화제취물농도위0.01 g/L시,능명현적강저양화손상신경세포내、외적양자유기수평.결론:①수옹화수제물대H2O2유도적PC12신경세포적양화손상역유흔강적보호작용.②수옹화제취물항양화특성여제취액적농도비유명현적관계,당농도위1.00 g/L시,구유명현적수복능력;당농도위0.01 g/L시능구강저세포내외적양자유기수평.
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 cleistocalyx 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 estabPC12 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 2×103 cells. Blank control group, or non-drug culture medium group, was set. Under the standard condition, cells were cultured for 48 hours and ascells were inoculated in 96-well plate with the density of 2×103 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 asfree 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.fect of cleistocalyx operculatus extracts on intracellular and extracellular oxygen-derived free radicals in PC12 nerve cells induced by oxidative injury.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 extracts had no protective effect on the injury of PC12 nerve cells induced by H2O2. At 1.00 g/L, it had strong plerosis for oxidative injury of PC12 and 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.dation of membrane lipid of hepatic microsome, but also protect against oxcleistocalyx 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.