中文摘要：

目的以H2O2诱导大鼠原代海马神经细胞建立氧化损伤细胞模型。方法取新生大鼠海马神经细胞,培养至第5天,加入终浓度为50、100、150、200和250μmol/L的H2O2,培养12、24和48 h后,观察细胞形态,通过MTT法测定细胞存活率,CCK-8法测定细胞损伤率,LDH酶标法测定上清液LDH活性,Annexin V-FITC/PI流式细胞术检测细胞凋亡率,确定出细胞损伤的最佳浓度时间。结果 H2O2可诱导神经细胞损伤且呈时间浓度依赖性。150μmol/L H2O2作用24 h时,MTT法测得细胞存活率为（70.18±4.66）%;CCK-8法测得细胞损伤率为（28.30±6.72）%;LDH酶标法测得LDH活性为（208.81±12.24）U/L;流式细胞术测得细胞早期凋亡率为（11.53±2.53）%,晚期凋亡率及坏死率为（13.75±2.22）%。结论 150μmol/L的H2O2作用24 h,可成功构建海马神经细胞氧化损伤模型。

英文摘要：

Objective To establish the oxidative cell model with hydrogen peroxide. Methods primary cultured hippocampal neurons were divided into six groups. Each group involved 6 wells of cells in 96 well plate. 100 μl experimental cells in logarithmic phase after adjusted concentration of 1 × 105/ml were vaccinated in the 96 well plate and cuhured in saturated humidity at 37 ℃ and under 5 % CO2 and 95 % air. 5 days later, 100 μl different concentrations of hydrogen peroxide were added. The final concentrations of hydrogen peroxide of groups were successively 0, 50, 100, 150, 200 and 250 μmol/ L. After 12, 24 and 48 h of treatment respectively, we make sure what is the best time and what is the most approptiate concentration of H2O2 to induce eelluar injuries by observing cell morphology through inverted microscope and scanning electron microscope, detecting the cell survival rate with MTT assay, evaluating the cell damage rate with CCK- 8 assay, assessing the cell mortality with LDH activity detection kit, examining the cell apoptosis with AnnexinV/PI formation method. Results H2O2 concentration-dependently and time-dependently promoted damage of primary cultured hippocampal neurons. Cellsviabilities in concentrations of 50, 100, 150, 200 and 250 μmol/L decreased obviously comparing with normal control group（0μmol/L hydrogen peroxide）. At the concentration of 150 μmol/L hydrogen peroxide and treated with 24 h, cell viability was （70. 18 ± 4.66）%, cell damage rate was（28.30 ±6.72）% , LDH activity was（208.81 ± 12.24） U/L; early apoptosis rate was（ 11.53 ±2.53）% and late apoptosis or necrosis rate was （ 13.75 ± 2.22） %. Conclusion under the present conditions, treating with 150 μmol/L hydrogen peroxide for 24 h can successfully establish the oxidative cell model.