title: NADPH oxidase 4 induces cardiac arrhythmic phenotype in zebrafish.
abstract: Oxidative stress has been implicated in cardiac arrhythmia, although a causal relationship remains undefined. We have recently demonstrated a marked up-regulation of NADPH oxidase isoform 4 (NOX4) in patients with atrial fibrillation, which is accompanied by overproduction of reactive oxygen species (ROS). In this study, we investigated the impact on the cardiac phenotype of NOX4 overexpression in zebrafish. One-cell stage embryos were injected with NOX4 RNA prior to video recording of a GFP-labeled (myl7:GFP zebrafish line) beating heart in real time at 24-31 h post-fertilization. Intriguingly, NOX4 embryos developed cardiac arrhythmia that is characterized by irregular heartbeats. When quantitatively analyzed by an established LQ-1 program, the NOX4 embryos displayed much more variable beat-to-beat intervals (mean S.D. of beat-to-beat intervals was 0.027 s/beat in control embryos versus 0.038 s/beat in NOX4 embryos). Both the phenotype and the increased ROS in NOX4 embryos were attenuated by NOX4 morpholino co-injection, treatments of the embryos with polyethylene glycol-conjugated superoxide dismutase, or NOX4 inhibitors fulvene-5, 6-dimethylamino-fulvene, and proton sponge blue. Injection of NOX4-P437H mutant RNA had no effect on the cardiac phenotype or ROS production. In addition, phosphorylation of calcium/calmodulin-dependent protein kinase II was increased in NOX4 embryos but diminished by polyethylene glycol-conjugated superoxide dismutase, whereas its inhibitor KN93 or AIP abolished the arrhythmic phenotype. Taken together, our data for the first time uncover a novel pathway that underlies the development of cardiac arrhythmia, namely NOX4 activation, subsequent NOX4-specific NADPH-driven ROS production, and redox-sensitive CaMKII activation. These findings may ultimately lead to novel therapeutics targeting cardiac arrhythmia.
authors: Divisions of Molecular Medicine and Cardiology, Departments of Anesthesiology and Medicine, Cardiovascular Research Laboratories, David Geffen School of Medicine at UCLA and Los Angeles, California 90095.
authors: Yixuan
authors: Y
authors: Zhang
authors: Department of Molecular, Cell and Developmental Biology, UCLA, Los Angeles, California 90095.
authors: Hirohito
authors: H
authors: Shimizu
authors: Divisions of Molecular Medicine and Cardiology, Departments of Anesthesiology and Medicine, Cardiovascular Research Laboratories, David Geffen School of Medicine at UCLA and Los Angeles, California 90095.
authors: Kin Lung
authors: KL
authors: Siu
authors: Divisions of Molecular Medicine and Cardiology, Departments of Anesthesiology and Medicine, Cardiovascular Research Laboratories, David Geffen School of Medicine at UCLA and Los Angeles, California 90095.
authors: Aman
authors: A
authors: Mahajan
authors: Department of Molecular, Cell and Developmental Biology, UCLA, Los Angeles, California 90095.
authors: Jau-Nian
authors: JN
authors: Chen
authors: Divisions of Molecular Medicine and Cardiology, Departments of Anesthesiology and Medicine, Cardiovascular Research Laboratories, David Geffen School of Medicine at UCLA and Los Angeles, California 90095. Electronic address: hcai@mednet.ucla.edu.
authors: Hua
authors: H
authors: Cai
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified / Q000196:embryology / Q000235:genetics; D001145:Arrhythmias, Cardiac / Q000196:embryology / Q000201:enzymology* / Q000235:genetics; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000235:genetics / Q000378:metabolism; D004195:Disease Models, Animal; D004789:Enzyme Activation / Q000187:drug effects / Q000235:genetics; D004791:Enzyme Inhibitors / Q000494:pharmacology; D009200:Myocardial Contraction; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism*; D010641:Phenotype; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015027:Zebrafish / Q000196:embryology* / Q000235:genetics; D029961:Zebrafish Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D017382:Reactive Oxygen Species; D029961:Zebrafish Proteins; D019255:NADPH Oxidases; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2
keywords: Ca2+/Calmodulin-dependent Protein Kinase II (CaMKII); NADPH Oxidase; Oxidative Stress; Reactive Oxygen Species (ROS); Signal Transduction