酸性肽对神经元凋亡的抑制(英文)  

Inhibition of neuron apoptosis by acidic peptide

作　　者：安玉会[1] 毛红丽[1] 马红霞[1] 郭茂锋[1] 

机构地区：[1]郑州大学医学院生物化学与分子生物学教研室,河南省郑州市450052

出　　处：《中国组织工程研究与临床康复》2007年第17期3457-3460,F0003,共5页Journal of Clinical Rehabilitative Tissue Engineering Research

摘　　要：背景:高浓度的一氧化氮能引起神经细胞的凋亡。因此抑制一氧化氮引起的细胞凋亡就能达到预防和治疗老年痴呆病的目的。目的:观察酸性肽能否抑制一氧化氮引起的神经元凋亡。设计:对照观察细胞和分子实验。材料:实验于2003-05/2005-05在郑州大学生物活性肽研究所第二实验室和郑州大学基础医学院生物化学与分子生物学教研室细胞培养室完成。新生的24h内SD雄性大鼠,由河南省动物中心提供(410117)。方法:对新生的SD大鼠海马神经元进行原代培养。取培养至第11天的神经细胞,用不同剂量的酸性肽预处理6h,再加入终浓度为50μmmol/L的亚硝基铁氰化钠,继续培养24h,收集细胞进行实验。每次实验均分为以下5组:正常对照组、亚硝基铁氰化钠处理组、亚硝基铁氰化钠+0.0375mg/mL酸性肽组、亚硝基铁氰化钠+0.075mg/mL酸性肽组,亚硝基铁氰化钠+0.15mg/mL酸性肽组。用噻唑蓝法测定细胞的存活率,用免疫组织化学法对神经丝蛋白进行染色,再用吖啶橙荧光染色法显示细胞凋亡的形态。用琼脂糖凝胶电泳法分析凋亡细胞的DNA梯带。用WesternBlot和吸光度扫描分析Bcl-2蛋白和Bax蛋白的表达水平。主要观察指标:①噻唑蓝法比色法检测细胞存活率的实验结果。②细胞凋亡的核型观察结果。③细胞凋亡的DNA电泳分析。④Bcl-2和Bax蛋白的WesternBlot分析结果。结果:①亚硝基铁氰化钠处理组的神经元存活率为58.9%,亚硝基铁氰化钠+0.037mg/mL酸性肽处理组为70.0%,亚硝基铁氰化钠+0.075mg/mL酸性肽处理组为72.8%,亚硝基铁氰化钠+0.15mg/mL酸性肽处理组为75.3%。②细胞凋亡的核型观察结果:亚硝基铁氰化钠处理组呈现细胞凋亡的明显特征。不同浓度的酸性肽+亚硝基铁氰化钠共同处理组海马神经元的细胞核接近正常对照组的细胞核形态。③细胞凋亡的DNA电泳分析结果表明,仅亚硝基铁氰化钠处理组�BACKGROUND： Excessive nitric oxide （NO） release can cause the occurrence and development of brain injury and senile dementia due to the apoptosis induction role of NO at high concentration to nerve cells. Therefore one strategy to prevent and treat senile dementia is inhibiting the apoptosis induced by NO. OBJECTIVE ： To observe whether acidic peptide will inhibit the neuron apoptosis caused by NO DESIGN ： An cell and molecule observation experiment by comparisons SETTING ： Department of Biochemistry and Molecular Biology of Basic Medical College in Zhengzhou University and the Second Laboratory of Biological Active Peptide Institute in Zhengzhou University. MATERIALS ： The experiment was performed between May 2003 and May 2005, in the Second Laboratory of Biological Active Peptide Institute in Zhengzhou University and the call culture room of Department of Biochemistry and Molecular Biology of Basic Medical College in Zhengzhou University. The newborn SD male rats within 24 hours after birth were provided by the Animal Center of Henan Province （410117）. METHODS ： On day 11 of primary cultures, hippocampus neurons of the newborn SD rats were pretreated with different dosages of acidic peptide for six hours. Sodium nitroprusside （SNP） of 50 p.mol/L final concentration was added to the cells which were incubated for another 24 hours. Cells were collected and adopted in this experiment of five different groups, namely normal control group, group treated with SNP, group of SNP plus 0.037 5 mg/mL acidic peptide, group of SNP plus 0.075 mg/mL acidic peptide, group of SNP plus 0.15 mg/mL acidic peptide. The cell＇s survival rate was measure by methyl thiazolyl （MTT） method; The neurofllament protein was stained with the method of immunohisto- chemistry. The shape of apoptosis was display with acridine orange fluorescent stain. Then DNA ladder zone of apoptosis cells was analyzed with the method of agarose gel electrophoresis. Western Blot and absorbance scan were used to determine the expres

关 键 词：细胞凋亡 神经元 细胞培养技术 westem免疫杂交 

分 类 号：R338.2[医药卫生—人体生理学]