The Change of Hypertrophic Signaling during the Development of Cardiac Hypertrophy and Its Regulation by Reactive Oxygen Species

目的：分析活性氧簇(ROS)对成年大鼠心肌细胞肥大信号转导的调节作用。背景：ROS调节多种细胞类型的促有丝分裂信号转导。在新生大鼠心肌细胞中，抗氧化剂被证明可以抑制心肌肥大，ROS被认为是调节肥大信号的因素。然而，由于新生和成年心肌细胞的性质不同，这一结论可能不能反映成熟心脏的情况。方法：培养的成年大鼠心肌细胞在内皮素-1(ET-1)或苯肾上腺素(PE)的刺激下，检测细胞内ROS水平、丝裂原活化蛋白激酶(MAPKs；ERK、p38和JNK)活性和3 H-苯丙氨酸掺入。我们还观察了心肌细胞抗氧化预处理对MAPK活性和心肌肥大的影响，以分析氧化还原状态对MAPK介导的肥大信号的调节作用。结果：ET-1或PE刺激的心肌细胞ROS水平在刺激后5min达到最大值。ROS的来源可能是NADH/NADPH氧化酶，因为NADH/NADPH氧化酶抑制剂二苯基碘可抑制心肌细胞ROS的增加。ET-1或PE刺激可使细胞外信号调节激酶(ERK)活性升高。而p38和c-Jun-N末端蛋白激酶(JNK)活性在刺激后无明显变化。心肌细胞的抗氧化剂处理可抑制这些刺激引起的心肌细胞内ROS的增加，并阻断ERK的激活和随后的心肌肥大。结论：ROS介导了ET-1或PE诱导的成年大鼠心肌细胞肥大的信号转导。

OBJECTIVES : We analyzed the regulatory function of reactive oxygen species (ROS) on the hypertrophic signaling in adult rat cardiac myocytes.BACKGROUND : The ROS regulate mitogenic signal transduction in various cell types. In neonatal rat cardiac myocytes, antioxidants have been shown to inhibit cardiac hypertrophy, and ROS are suggested to modulate the hypertrophic signaling. However, the conclusion may not reflect the situation of mature heart, because of the different nature between neonatal and adult cardiac myocytes.METHODS : Cultured adult rat cardiac myocytes were stimulated with endothelin-1 (ET-1) or phenylephrine (PE), and intracellular ROS levels, the activities of mitogen-activated protein kinases (MAPKs ; ERK, p38, and JNK), and 3H-phenylalanine incorporation were examined. We also examined the effects of antioxidant pretreatment of myocytes on MAPK activities and cardiac hypertrophy to analyze the modulatory function of redox state on MAPK-mediated hypertrophic signaling.RESULTS : The ROS levels in ET-1 or PE-stimulated myocytes were maximally increased at 5 min after stimulation. The origin of ROS appears to be from NADH/NADPH oxidase, because the increase in ROS was suppressed by pretreatment of myocytes with NADH/NADPH oxidase inhibitor diphenyleneiodonium. Extracellular signal-regulated kinase (ERK) activity was increased by the stimulation of ET-1 or PE. In contrast, p38 and c-Jun-N-terminal protein kinase (JNK) activities did not change after these stimulation. Antioxidant treatment of myocytes suppressed the increase in ROS and blocked ERK activation and subsequent cardiac hypertrophy induced by these stimuli.CONCLUSIONS : These data demonstrate that ROS mediate signal transduction of cardiac hypertrophy induced by ET-1 or PE in adult rat cardiac myocytes.