褪黑素对心肌细胞抗过氧化氢损伤的实验研究

目的 :观察 H2 O2 对心肌细胞内活性氧 （ROS）和游离钙 （Ca2 + ]i）的影响及褪黑素 （MT）的保护作用。方法 :酶消化法原代分离培养乳鼠心肌细胞 ,分别用荧光探针 DCFH- DA和 Fluo- 3- AM标记 ,H2 O2 和 MT处理后 ,流式细胞仪检测细胞荧光强度的变化。结果 :低浓度的 H2 O2 （10 0 μm ol/ L）随着时间的延长可使心肌细胞内的 ROS呈上升趋势 ,30 m in,6 0 m in与 0 min或对照组相比均明显增高 （P<0 .0 1） ;MT干预后上升趋势均明显减弱 ,在 30 ,6 0 min两组均差异显著 （P<0 .0 1）。 Ca2 + ]i 亦呈显著上升趋势 ,30 min时与 0 min或对照组相比有差异 （P<0 .0 5 ） ,6 0min时与 30 m in,0 min或对照组相比差异显著 （P<0 .0 1） ;MT干预后其上升明显减少 ,两者差异显著 （P<0 .0 1）。结论 :MT有很好的抗氧化效果 ,能起到保护心肌的作用。

Effects of melatonin on cultured rat cardiomyocytes anti-injury mediated by H_2O_2

AIM: To investigate the role of H2O2 in myocardial intracellular reactive oxygen species (ROS) and free calcium（［Ca2+］i）and the effects of melatonin (MT) on them. METHODS: The fluorescent probe DCFH-DA was used to detect intracellular levels of ROS and Fluo-3-AM to detect ［Ca2+］i. Cultured rat ventricular myocytes were divided into 4 groups: Control group, H2O2 group, MT group and MT+H2O2 group. Cells were incubated with DCFH-DA or Fluo-3-AM prior to the exposure to 100 μmol/L H2O2 and/or MT. The changes in cell fluorescence were measured by flow cytometer (FCM) at different time points. RESULTS: Compared with the control group, H2O2 at low concentration(100 μmol/L) caused significant time-dependent increase in intracellular ROS and ［Ca2+］i (ROS:30 min,742±326 vs 200±130, P<0.01， 60 min,1630±421 vs 198±130, P<0.01; ［Ca2+］i :30 min,126±114 vs 121±99， P<0.05, 60 min,844±356 vs 121±106, P<0.01). However, those changes were significantly attenuated in MT+H2O2 group (ROS:30 min,742±326 vs 223±125, P<0.01, 60 min, 1630±421 vs 317±211， P<0.01; ［Ca2+］i ： 60 min,844±356 vs 439±261， P<0.01). CONCLUSION: Intracellular ROS and ［Ca2+］i increase when cardiomyocytes are challenged by 100 μmol/L H2O2 . MT has very good antioxidant effect and can be used to protect cardiomyocytes.