title: NADPH oxidase 2 mediates angiotensin II-dependent cellular arrhythmias via PKA and CaMKII.
abstract: BACKGROUND
  Angiotensin II (Ang II) signaling has been implicated in cardiac arrhythmogenesis, which involves induction of reactive oxygen species (ROS). It was shown that Ang II can activate Ca/Calmodulin kinase II (CaMKII) by oxidation via a NADPH oxidase 2 (NOX2)-dependent pathway leading to increased arrhythmic afterdepolarizations. Interestingly, cAMP-dependent protein kinase A (PKA) which regulates similar targets as CaMKII has recently been shown to be redox-sensitive as well.
  
  
  OBJECTIVE
  This study aims to investigate the distinct molecular mechanisms underlying Ang II-related cardiac arrhythmias with an emphasis on the individual contribution of PKA vs. CaMKII.
  
  
  RESULTS
  Isolated ventricular cardiac myocytes from rats and mice were used. Ang II exposure resulted in increased NOX2-dependent ROS generation assessed by expression of redox-sensitive GFP and in myocytes loaded with ROS indicator MitoSOX. Whole cell patch clamp measurements showed that Ang II significantly increased peak Ca and Na current (ICa and INa) possibly by enhancing steady-state activation of ICa and INa. These effects were absent in myocytes lacking functional NOX2 (gp91phox(-/-)). In parallel experiments using PKA inhibitor H89, the Ang II effects on peak INa and ICa were also absent. In contrast, genetic knockout of CaMKIIδ (CaMKIIδ(-/-)) did not influence the Ang II-dependent increase in peak ICa and INa. On the other hand, Ang II enhanced INa inactivation, increased late INa and induced diastolic SR (sarcoplasmic reticulum) Ca leak (confocal Ca spark measurements) in a CaMKIIδ-, but not PKA-dependent manner. Surprisingly, only the increase in diastolic SR Ca leak was absent in gp91phox(-/-)myocytes suggesting that Ang II regulates INa inactivation in a manner dependent on CaMKII- but not on NOX2. Finally, we show that Ang II increased the propensity for cellular arrhythmias, for which PKA and CaMKII contribute, both dependent on NOX2.
  
  
  CONCLUSIONS
  Ang II activates PKA and CaMKII via NOX2, which results in disturbed Na and Ca currents (via PKA) and enhanced diastolic SR Ca leakage (via CaMKII). Oxidative activation of PKA and CaMKII via NOX2 may represent important pro-arrhythmogenic pathways in the setting of increased Ang II stimulation, which may be relevant for the treatment of arrhythmias in cardiac disease.
authors: Dept. Internal Medicine II, University Hospital Regensburg, Regensburg, Germany; Clinic for Cardiology & Pneumology, Georg-August-University Göttingen, and DZHK (German Centre for Cardiovascular Research), partner site Göttingen, Germany.
authors: Stefan
authors: S
authors: Wagner
authors: Clinic for Cardiology & Pneumology, Georg-August-University Göttingen, and DZHK (German Centre for Cardiovascular Research), partner site Göttingen, Germany.
authors: Christian
authors: C
authors: Dantz
authors: Clinic for Cardiology & Pneumology, Georg-August-University Göttingen, and DZHK (German Centre for Cardiovascular Research), partner site Göttingen, Germany.
authors: Hannah
authors: H
authors: Flebbe
authors: Clinic for Cardiology & Pneumology, Georg-August-University Göttingen, and DZHK (German Centre for Cardiovascular Research), partner site Göttingen, Germany.
authors: Azadeh
authors: A
authors: Azizian
authors: Dept. Internal Medicine II, University Hospital Regensburg, Regensburg, Germany; Cardiovascular Division, King's College London British Heart Foundation Centre, UK.
authors: Can Martin
authors: CM
authors: Sag
authors: Clinic for Cardiology & Pneumology, Georg-August-University Göttingen, and DZHK (German Centre for Cardiovascular Research), partner site Göttingen, Germany.
authors: Susanne
authors: S
authors: Engels
authors: Clinic for Cardiology & Pneumology, Georg-August-University Göttingen, and DZHK (German Centre for Cardiovascular Research), partner site Göttingen, Germany.
authors: Johanna
authors: J
authors: Möllencamp
authors: Clinic for Cardiology & Pneumology, Georg-August-University Göttingen, and DZHK (German Centre for Cardiovascular Research), partner site Göttingen, Germany.
authors: Nataliya
authors: N
authors: Dybkova
authors: Clinic for Cardiology & Pneumology, Georg-August-University Göttingen, and DZHK (German Centre for Cardiovascular Research), partner site Göttingen, Germany.
authors: Towhidul
authors: T
authors: Islam
authors: Cardiovascular Division, King's College London British Heart Foundation Centre, UK.
authors: Ajay M
authors: AM
authors: Shah
authors: Dept. Internal Medicine II, University Hospital Regensburg, Regensburg, Germany. Electronic address: lars.maier@ukr.de.
authors: Lars S
authors: LS
authors: Maier
mesh_terms: D000200:Action Potentials / Q000187:drug effects; D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D001145:Arrhythmias, Cardiac / Q000201:enzymology* / Q000473:pathology*; D020746:Calcium Channels, L-Type / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D015640:Ion Channel Gating / Q000187:drug effects; D008562:Membrane Glycoproteins / Q000378:metabolism*; D051379:Mice; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000378:metabolism*; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000378:metabolism; D015222:Sodium Channels / Q000378:metabolism
chemical_list: D020746:Calcium Channels, L-Type; D008562:Membrane Glycoproteins; D017382:Reactive Oxygen Species; D015222:Sodium Channels; D000804:Angiotensin II; C488842:Cybb protein, mouse; C500593:Cybb protein, rat; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; D017868:Cyclic AMP-Dependent Protein Kinases; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2
keywords: Angiotensin II; Calcium/calmodulin-dependent protein kinase II; Excitation–contraction coupling; NAD(P)H oxidase; Protein kinase A; Reactive oxygen species