【摘要】 实验在原代培养的新生大鼠心肌细胞中进行,检测内皮素-1(endothelin-1,ET-1)及其他药物对心肌细胞活性氧(reactiveoxygen species,ROS)产生和心肌细胞肥大的作用,以探讨ROS在ET-1诱导的心肌细胞肥大信号通路中的作用及ROS与蛋白激酶C(protein kinase C,PKC)活化的关系。细胞内ROS水平用ROS敏感的荧光探针2,7-dichlorofluorescin dictate(DCF-DA)反映,心肌细胞肥大通过细胞内RNA含量、细胞内蛋白质含量、细胞表面积大小来确定。实验结果如下:单独使用ET-1后,心肌细胞内反应ROS含量的DCF-DA荧光值比对照组增加77%,反应心肌肥大的PI荧光值、细胞内蛋白质含量、细胞表面积也分别比对照组增加128%、87%和151%。ET-1合用内皮素受体A亚型(ET_A)受体拮抗剂ABT-627、PKC抑制剂CC或过氧化氢酶后,DCF-DA的增加分别减弱62%、60%和51%,同时心肌细胞肥大也被抑制,单独使用PKC激动剂佛波醇脂(PMA)也能使DCF-DA的产生比对照组增加74%。因此,在ET-1诱导心肌细胞肥大的过程中,ET-1能够使心肌细胞产生ROS和诱导ROS依赖的心肌细胞肥大,这一作用依赖于ET_A受体的激活和PKC的活化,·ROS在ET-1诱导心肌细胞肥大中起信号传递的作用。更多还原

【Abstract】 The effects of endothelin-1 (ET-1) and other drugs on the reactive oxygen species (ROS) generation and cardiomyocytehypertrophy were examined in experiments on the cultured neonatal rat cardiomyocytes. The role of ROS on neonatal rat cardiomyocytehypertrophy induced by ET-1 was studied and the relationship of PKC activation and ROS generation was investigated. The level ofintracellular ROS was measured by the ROS-specific probe 2’,7’-dichlorofluorescin diacetate (DCF-DA). Cardiomyocyte hypertrophywas determined by the RNA content, the total protein of cells and the cell surface area. The results are as follows. The fluorescence intensityof intracellular DCF-DA increased by 77% in cultured neonatal rat cardiac myocytes treated with ET-1 (10 nmol/L) vs control group.Compared with control group, the fluorescence intensity of intracellular PI, protein content and cell surface area increased by 128%, 87%and 151% respectively (all P<0.01) in cardiac myocytes treated with ET-1 (10 mol/L). ABT-627, CC, or CAT inhibited the ET-1-inducedincrease in fluorescence intensity of intracellular DCF-DA by 62%, 60% and 51% respectively (all P<0.01), and also attenuated the cardiachypertrophy. The fluorescence intensity of intracellular DCF-DA increased by 74% (P<0.01) in myocytes treated with PMA (1μmol/L) vs control group. Therefore, in the course of cardiomyocyte hypertrophy, ET-1 increases intracellular ROS in the cultured neonatalrat cardiac myocytes and inhibits cardiomyocyte hypertrophy induced by ROS. The ET_A and PKC activation mediate the ROS productionand cardiomyocyte hypertrophy induced by ET-1. ROS is necessary in the ET-1-induced cardiomyocyte hypertrophy.更多还原