Xie Z, Kometiani P, Liu J, Li J, Shapiro J I, Askari A
Departments of Pharmacology and Medicine, Medical College of Ohio, Toledo, Ohio 43614, USA.
J Biol Chem. 1999 Jul 2;274(27):19323-8. doi: 10.1074/jbc.274.27.19323.
We showed before that in cardiac myocytes partial inhibition of Na+/K+-ATPase by nontoxic concentrations of ouabain causes hypertrophy and transcriptional regulations of growth-related marker genes through multiple Ca2+-dependent signal pathways many of which involve Ras and p42/44 mitogen-activated protein kinases. The aim of this work was to explore the roles of intracellular reactive oxygen species (ROS) in these ouabain-initiated pathways. Ouabain caused a rapid generation of ROS within the myocytes that was prevented by preexposure of cells to N-acetylcysteine (NAC) or vitamin E. These antioxidants also blocked or attenuated the following actions of ouabain: inductions of the genes of skeletal alpha-actin and atrial natriuretic factor, repression of the gene of the alpha3-subunit of Na+/K+-ATPase, activation of mitogen-activated protein kinases, activation of Ras-dependent protein synthesis, and activation of transcription factor NF-kappaB. Induction of c-fos and activation of AP-1 by ouabain were not sensitive to NAC. Ouabain-induced inhibition of active Rb+ uptake through Na+/K+-ATPase and the resulting rise in intracellular Ca2+ were also not prevented by NAC. A phorbol ester that also causes myocyte hypertrophy did not increase ROS generation, and its effects on marker genes and protein synthesis were not affected by NAC. We conclude the following: (a) ROS are essential second messengers within some but not all signal pathways that are activated by the effect of ouabain on Na+/K+-ATPase; (b) the ROS-dependent pathways are involved in ouabain-induced hypertrophy; (c) increased ROS generation is not a common response of the myocyte to all hypertrophic stimuli; and (d) it may be possible to dissociate the positive inotropic effect of ouabain from its growth-related effects by alteration of the redox state of the cardiac myocyte.
我们之前表明，在心肌细胞中，无毒浓度的哇巴因对Na+/K+-ATP酶的部分抑制会通过多种Ca2+依赖信号通路导致肥大以及生长相关标记基因的转录调控，其中许多通路涉及Ras和p42/44丝裂原活化蛋白激酶。这项工作的目的是探讨细胞内活性氧（ROS）在这些哇巴因启动的通路中的作用。哇巴因导致心肌细胞内ROS迅速产生，而细胞预先暴露于N-乙酰半胱氨酸（NAC）或维生素E可阻止这种情况。这些抗氧化剂还阻断或减弱了哇巴因的以下作用：诱导骨骼肌α-肌动蛋白和心钠素基因、抑制Na+/K+-ATP酶α3亚基基因、激活丝裂原活化蛋白激酶、激活Ras依赖的蛋白质合成以及激活转录因子NF-κB。哇巴因诱导的c-fos表达和AP-1激活对NAC不敏感。哇巴因通过Na+/K+-ATP酶诱导的对活性Rb+摄取的抑制以及由此导致的细胞内Ca2+升高也不能被NAC阻止。一种也会导致心肌细胞肥大的佛波酯不会增加ROS的产生，其对标记基因和蛋白质合成的影响不受NAC影响。我们得出以下结论：（a）ROS是哇巴因对Na+/K+-ATP酶作用激活的一些但不是所有信号通路中的必需第二信使；（b）ROS依赖的通路参与哇巴因诱导的肥大；（c）ROS产生增加不是心肌细胞对所有肥大刺激的共同反应；（d）通过改变心肌细胞的氧化还原状态，有可能将哇巴因的正性肌力作用与其生长相关作用分离。
