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目的 探讨氧化损伤时心肌细胞一氧化氮 (NO)生成的变化和褪黑素 (MT)的保护作用。方法 分离培养原代心肌细胞 ,用 NO试剂盒检测细胞培养液中 NO含量 ,硫代巴比妥酸法测定细胞中丙二醛 (MDA)含量 ,荧光探针 DCFH- DA标记细胞 ,用激光共聚焦显微镜观察心肌细胞内总的自由基水平。结果 与对照组相比 ,1 0 0 μmol/ L的 H2 O2 作用 60 min可使培养液中 NO浓度、心肌细胞中 MDA含量和心肌细胞内总的自由基水平明显增加 (分别为 2 2 .4± 2 .6vs97.3± 2 .4nmol/ L、1 .2 3± 0 .0 5vs3.65± 0 .2 0 nmol/ 1 0 6 cells和 86± 6.9vs742± 1 3.4,P<0 .0 1 ) ;加入 MT干预后 ,H2 O2 引起的 NO、MDA和细胞内总的自由基水平的升高被明显抑制 ,两组相比显著差异 (P<0 .0 1 )。结论 H2 O2 造成心肌细胞氧化损伤时 ,心肌细胞合成的 NO增多并参与了损伤机制 ,MT不仅能使心肌细胞内总的自由基水平下降 ,也能抑制 NO的过度增加 ,从而起到保护心肌细胞的作用。
Objective To investigate the changes of nitric oxide (NO) production and the protective effect of melatonin (MT) on myocardial cells during oxidative injury. Methods Primary cardiomyocytes were isolated and cultured. NO content in cell culture medium was detected by NO kit. Malondialdehyde (MDA) content was measured by thiobarbituric acid method. DCFH-DA labeled cells were labeled with laser Focusing microscope to observe the total free radical levels in cardiomyocytes. Results Compared with the control group, NO concentration in the culture medium, MDA content in cardiomyocytes and total free radical levels in myocardial cells were significantly increased after H2O2 was added at 100 μmol / L for 60 min (respectively, 22.4 ± 2 .6 vs 97.3 ± 2 .4 nmol / L, 1.2 3 ± 0. 0 5 vs 3.65 ± 0.2 0 nmol / 1 0 6 cells and 86 ± 6.9 vs 742 ± 1 3.4, P <0.01 ); The increase of NO, MDA and the total free radicals in cells induced by H2O2 was significantly inhibited by the addition of MT, with a significant difference between the two groups (P <0.01). Conclusions Oxidative injury of cardiomyocytes induced by H2O2 can increase NO synthesis and participate in the mechanism of injury. MT can not only decrease the total free radical levels but also inhibit the excessive increase of NO in cardiomyocytes, thus protecting cardiomyocytes Role.