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Posts Tagged ‘Atrial fibrillation’


Renal Function Biomarker, β-trace protein (BTP) as a Novel Biomarker for Cardiac Risk Diagnosis in Patients with Atrial Fibrilation

Curator: Aviva Lev-Ari, PhD, RN

Original Research | November 2013

β-Trace Protein and Prognosis in Patients With Atrial Fibrillation Receiving Anticoagulation Treatment

Juan Antonio Vílchez, BSc Pharm, PhD; Vanessa Roldán, MD, PhD; Sergio Manzano-Fernández, MD, PhD; Hermógenes Fernández, MD; Francisco Avilés-Plaza, MD, PhD; Pedro Martínez-Hernández, BSc Pharm, PhD; Vicente Vicente, MD, PhD; Mariano Valdés, MD, PhD; Francisco Marín, MD, PhD; Gregory Y. H. Lip, MD

From the University of Birmingham Centre for Cardiovascular Sciences (Drs Apostolakis and Lip), City Hospital, Birmingham, England; and the Division of Cardiovascular Medicine (Drs Sullivan and Olshansky), University of Iowa Hospitals and Clinics, Iowa City, IA.

Correspondence to: Gregory Y. H. Lip, MD, University of Birmingham Centre for Cardiovascular Sciences, City Hospital, Dudley Rd, Birmingham, B18 7QH, England; e-mail: g.y.h.lip@bham.ac.uk

Abstract

Background:  Atrial fibrillation (AF) is associated with a high risk of mortality and morbidity and it commonly coexists with chronic kidney disease. A biomarker of renal function, β-trace protein (BTP), has been implicated in the progression of cardiovascular disease. The aim of our study was to evaluate the association of BTP with adverse cardiovascular events, bleeding, and mortality in patients with AF.

Methods:  In a consecutive cohort of patients with nonvalvular AF receiving anticoagulation treatment, plasma BTP was determined using an automated nephelometer BN ProSpec System (Siemens) and related to estimated glomerular filtration rate (eGFR). We recorded adverse cardiovascular events (stroke, acute coronary syndrome, and acute pulmonary edema), major bleeding, and mortality.

Results:  We included 1,279 patients (48.6% men), aged 76 years (IQR, 71-81 years), who were followed up for 996 days (IQR, 802-1,254 days). During the follow-up, there were 150 cardiovascular events (annual rate, 3.99%), 57 embolisms (annual rate, 1.54%), and 114 major bleeding events (annual rate, 3.04%), and 161 patients died (annual rate, 4.32%). BTP levels were inversely associated with eGFR (P < .001). High BTP concentrations were significantly associated with embolic events (hazard ratio [HR], 4.64 [1.98-10.86]; P < .001), composite adverse cardiovascular events (HR, 1.93 [1.31-2.85]; P = .001), and mortality (HR, 2.08 [1.49-2.90]; P < .001), even after adjusting for CHAD2DS2-VASc (congestive heart failure, hypertension, age ≥ 75 years [doubled], diabetes mellitus, stroke [doubled], vascular disease, age 65 to 74 years, sex category) score and renal function. High BTP was associated with major bleeding events (HR, 1.88 [1.18-3.00]; P = .008), even after adjusting for the HAS-BLED (hypertension, abnormal renal/liver function, stroke, bleeding history or redisposition, labile international normalized ratio, elderly [> 65 years], drugs/alcohol concomitantly) score.

Conclusions:  We suggest that BTP, a proposed renal damage biomarker, may be a novel predictor of adverse cardiovascular events, major bleeding, and mortality in patients with AF. BTP may help refine clinical risk stratification in these patients.

SOURCE

http://journal.publications.chestnet.org/article.aspx?articleid=1730537

Editorials | November 2013

Predicting the Quality of Anticoagulation During Warfarin Therapy:The Basis for an Individualized Approach

Giuseppe Boriani, MD, PhD

From the Institute of Cardiology, Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna.

Correspondence to: Giuseppe Boriani, MD, PhD, Institute of Cardiology, Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Via Massarenti 9, 40138 Bologna, Italy; e-mail: giuseppe.boriani@unibo.it

Chest. 2013;144(5):1437-1438. doi:10.1378/chest.13-1285

In medicine, there is an emerging tendency toward individualized medicine, that is, an approach to medicine based on available evidence, but enriched by the awareness of the inherent limitations of any “one size fits all” approach. As a matter of fact, diseases show individual differences with regard to onset and course, and individuals show different responses to drugs and interventions, thus suggesting the rationale for an individualized approach to disease treatments, able to predict individual responses. The most sophisticated approach to individualization and tailoring of medicine is personalized medicine, a broad and rapidly advancing field of health care that is informed by each person’s unique clinical, genetic, genomic, and environmental information.1 Treatment with vitamin K antagonists (VKAs) has been one of the traditional settings for individualization of treatment. The concept of personalized medicine specifically applies to warfarin dosing, a setting where knowledge of the complex polymorphic variants in the gene encoding cytochrome 2C9 (CYP2C9) and of the genetic variants in the gene encoding vitamin K epoxide reductase complex 1 (VKORC 1) may help to predict the interindividual variability in warfarin pharmacokinetics and pharmacodynamics, as well as warfarin-associated events and costs.2 However, it is still uncertain and unproven whether management of warfarin dosing guided by pharmacogenetics may improve patient outcomes.3

Biomarker Can Predict Events in Afib Patients

Published: Nov 6, 2013 | Updated: Nov 7, 2013

By Todd Neale, Senior Staff Writer, MedPage Today
Reviewed by Zalman S. Agus, MD; Emeritus Professor, Perelman School of Medicine at the University of Pennsylvania and Dorothy Caputo, MA, BSN, RN, Nurse Planner

Beta-trace protein (BTP), a biomarker that has been associated with both kidney damage and an increased cardiovascular risk, may help identify high-risk atrial fibrillation patients, researchers found.

Among patients with atrial fibrillation who were on stable oral anticoagulant therapy, high plasma levels of the protein were associated with significantly elevated risks of embolic events, adverse cardiovascular events, death, and major bleeding, according to Gregory Lip, MD, of the University of Birmingham in England, and colleagues.

Also, adding information about BTP levels modestly improved the predictive ability of models that included two established risk scores — CHAD2DS2-VASc and HAS-BLED — as indicated by higher C-statistics, they reported in the Nov. 5 issue of CHEST.

“This raises the possibility that BTP may help refine the clinical risk stratification for thrombotic or hemorrhagic events and mortality in these patients,” they wrote.

BTP has been proposed has a marker of renal damage, and it has also been associated with inflammation, atherogenesis, angina, vasomotor reactivity, and hypertension. Previous studies have also identified a relationship between BTP and the progression of cardiovascular disease.

In the current study, Lip and colleagues explored whether BTP levels were related to outcomes in 1,279 patients with nonvalvular atrial fibrillation who were on stable oral anticoagulant therapy with an international normalized ratio (INR) of 2.0 to 3.0. Their average age was 76.

The median estimated glomerular filtration rate at baseline was 71.28 mL/min/1.73 m2; BTP levels and renal function were inversely related (P<0.001).

The BTP cut-offs with the best sensitivity and specificity for predicting each of the endpoints varied — 0.561 mg/L for adverse cardiovascular events, 0.556 mg/L for embolic events, 0.670 mg/L for mortality, and 0.573 mg/L for major bleeds.

During a median follow-up of 2.7 years, cardiovascular events occurred at a rate of 3.99% per year, embolisms at 1.54% per year, deaths at 4.32% per year, and major bleeds at 3.04% per year.

After adjustment for renal function and the CHAD2DS2-VASc risk score — which incorporates congestive heart failure, hypertension, age, diabetes, stroke, vascular disease, and sex — a BTP level above the cutoff was associated with increased risks of cardiovascular events (HR 1.93, 95% CI 1.31-2.85), embolic events (HR 4.64, 95% CI 1.98-10.86), and mortality (HR 2.08, 95% CI 1.49-2.90).

Also, after adjustment for the HAS-BLED risk score — which takes into account hypertension, abnormal renal and liver function, stroke, bleeding history or predisposition, labile INR, age over 65, and concomitant use of drugs and alcohol — a high BTP level was associated with a greater risk of major bleeding (HR 1.88, 95% CI 1.18-3.00).

“We suggest that BTP, a proposed renal damage biomarker, may be a novel predictor of adverse cardiovascular events, major bleeding, and mortality in patients with atrial fibrillation,” the authors wrote.

They acknowledged some limitations of the analysis, however, including possible selection bias because all of the patients were on stable oral anticoagulant therapy, the measurement of renal function and BTP levels at a single time point only, and the exclusion of patients with end-stage renal disease.

SOURCE

http://www.medpagetoday.com/Cardiology/Arrhythmias/42751

These are promising early results, but the data include plenty of limitations. As the article notes, the researchers themselves acknowledge that their work only looked at patients on a regular oral anticlotting drug at a certain point in time. Further research must include a broader class of patients to determine if BTP can be a reliable biomarker to help identify atrial fibrillation patients with an added risk of other health problems.

As hard as it might be to spot atrial fibrillation patients at risk of more problems, doctors struggle to definitively identify the condition in the first place and apply targeted treatments. The med tech industry, meanwhile, is trying to fill the gap. Topera, a 2013 Fierce 15 winner, recently won U.S. and EU approval for a 3-D device and mapping tool designed to better detect cardiac rhythm problems such as atrial fibrillation in order to enable more targeted and accurate treatment. In late August, St. Jude Medical ($STJ) snatched up Endosense, which makes a cutting-edge irrigated ablation catheter designed to treat atrial fibrillation, and rival companies are developing or promoting electrophysiology treatments and other devices for the condition.

 SOURCE

From: FierceBiomarkers <editors@fiercebiomarkers.com>
Reply-To: <editors@fiercebiomarkers.com>
Date: Wednesday, November 13, 2013 10:31 AM
To: AvivaLev-Ari@alum.berkeley.edu
Subject: | 11.13.13 | Investigators flag new biomarkers for atrial fib

Articles related to Diagnosis of Atrial Fibrilation published on this Open Access Online Scientific Journal include the following:

Genetic Analysis of Atrial Fibrillation, Larry H Bernstein, MD, FCAP  and Aviva-Lev Ari, PhD, RN

https://pharmaceuticalintelligence.com/2013/10/27/genetic-analysis-of-atrial-fibrillation/

Cardiac Contractility & Myocardium Performance: Ventricular Arrhythmiasand Non-ischemic Heart Failure – Therapeutic Implications for Cardiomyocyte Ryanopathy (Calcium Release-related Contractile Dysfunction) and Catecholamine Responses
Justin Pearlman, MD, PhD, FACC, Larry H Bernstein, MD, FCAP and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/08/28/cardiac-contractility-myocardium-performance-ventricular-arrhythmias-and-non-ischemic-heart-failure-therapeutic-implications-for-cardiomyocyte-ryanopathy-calcium-release-related-contractile/

Oxidized Calcium Calmodulin Kinase and Atrial Fibrillation, Larry H. Bernstein, MD, FCAP and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/10/26/oxidized-calcium-calmodulin-kinase-and-atrial-fibrillation/

Genetics of Conduction Disease: Atrioventricular (AV) Conduction Disease (block): Gene Mutations – Transcription, Excitability, and Energy Homeostasis, Aviva Lev-Ari, PhD, RN

https://pharmaceuticalintelligence.com/2013/04/28/genetics-of-conduction-disease-atrioventricular-av-conduction-disease-block-gene-mutations-transcription-excitability-and-energy-homeostasis/

On Devices and On Algorithms: Prediction of Arrhythmia after Cardiac Surgery and ECG Prediction of an Onset of Paroxysmal Atrial Fibrillation, Justin D. Pearlman, MD, PhD, FACC and Aviva Lev-Ari, PhD, RN

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Genetic Analysis of Atrial Fibrillation

Author and Curator: Larry H Bernstein, MD, FCAP  

and 

Curator: Aviva-Lev Ari, PhD, RN

This article is a followup of the wonderful study of the effect of oxidation of a methionine residue in calcium dependent-calmodulin kinase Ox-CaMKII on stabilizing the atrial cardiomyocyte, giving protection from atrial fibrillation.  It is also not so distant from the work reviewed, mostly on the ventricular myocyte and the calcium signaling by initiation of the ryanodyne receptor (RyR2) in calcium sparks and the CaMKII d isoenzyme.

We refer to the following related articles published in pharmaceutical Intelligence:

Oxidized Calcium Calmodulin Kinase and Atrial Fibrillation
Author: Larry H. Bernstein, MD, FCAP and Curator: Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/10/26/oxidized-calcium-calmodulin-kinase-and-atrial-fibrillation/

Jmjd3 and Cardiovascular Differentiation of Embryonic Stem Cells

Author: Larry H. Bernstein, MD, FCAP and Curator: Aviva Lev-Ari, PhD, RN

https://pharmaceuticalintelligence.com/2013/10/26/jmjd3-and-cardiovascular-differentiation-of-embryonic-stem-cells/

Contributions to cardiomyocyte interactions and signaling
Author and Curator: Larry H Bernstein, MD, FCAP  and Curator: Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/10/21/contributions-to-cardiomyocyte-interactions-and-signaling/

Cardiac Contractility & Myocardium Performance: Therapeutic Implications for Ryanopathy (Calcium Release-related Contractile Dysfunction) and Catecholamine Responses
Editor: Justin Pearlman, MD, PhD, FACC, Author and Curator: Larry H Bernstein, MD, FCAP, and Article Curator: Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/08/28/cardiac-contractility-myocardium-performance-ventricular-arrhythmias-and-non-ischemic-heart-failure-therapeutic-implications-for-cardiomyocyte-ryanopathy-calcium-release-related-contractile/

Part I. Identification of Biomarkers that are Related to the Actin Cytoskeleton
Curator and Writer: Larry H Bernstein, MD, FCAP
https://pharmaceuticalintelligence.com/2012/12/10/identification-of-biomarkers-that-are-related-to-the-actin-cytoskeleton/

Part II: Role of Calcium, the Actin Skeleton, and Lipid Structures in Signaling and Cell Motility
Larry H. Bernstein, MD, FCAP, Stephen Williams, PhD and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/08/26/role-of-calcium-the-actin-skeleton-and-lipid-structures-in-signaling-and-cell-motility/

Part IV: The Centrality of Ca(2+) Signaling and Cytoskeleton Involving Calmodulin Kinases and Ryanodine Receptors in Cardiac Failure, Arterial Smooth Muscle, Post-ischemic Arrhythmia, Similarities and Differences, and Pharmaceutical Targets
Larry H Bernstein, MD, FCAP, Justin Pearlman, MD, PhD, FACC and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/09/08/the-centrality-of-ca2-signaling-and-cytoskeleton-involving-calmodulin-kinases-and-ryanodine-receptors-in-cardiac-failure-arterial-smooth-muscle-post-ischemic-arrhythmia-similarities-and-differen/

Part VI: Calcium Cycling (ATPase Pump) in Cardiac Gene Therapy: Inhalable Gene Therapy for Pulmonary Arterial Hypertension and Percutaneous Intra-coronary Artery Infusion for Heart Failure: Contributions by Roger J. Hajjar, MD
Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/08/01/calcium-molecule-in-cardiac-gene-therapy-inhalable-gene-therapy-for-pulmonary-arterial-hypertension-and-percutaneous-intra-coronary-artery-infusion-for-heart-failure-contributions-by-roger-j-hajjar/

Part VII: Cardiac Contractility & Myocardium Performance: Ventricular Arrhythmias and Non-ischemic Heart Failure – Therapeutic Implications for Cardiomyocyte Ryanopathy (Calcium Release-related Contractile Dysfunction) and Catecholamine Responses
Justin Pearlman, MD, PhD, FACC, Larry H Bernstein, MD, FCAP and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/08/28/cardiac-contractility-myocardium-performance-ventricular-arrhythmias-and-non-ischemic-heart-failure-therapeutic-implications-for-cardiomyocyte-ryanopathy-calcium-release-related-contractile/

Part VIII: Disruption of Calcium Homeostasis: Cardiomyocytes and Vascular Smooth Muscle Cells: The Cardiac and Cardiovascular Calcium Signaling Mechanism
Justin Pearlman, MD, PhD, FACC, Larry H Bernstein, MD, FCAP and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/09/12/disruption-of-calcium-homeostasis-cardiomyocytes-and-vascular-smooth-muscle-cells-the-cardiac-and-cardiovascular-calcium-signaling-mechanism/

Part IX: Calcium-Channel Blockers, Calcium Release-related Contractile Dysfunction (Ryanopathy) and Calcium as Neurotransmitter Sensor
Justin Pearlman, MD, PhD, FACC, Larry H Bernstein, MD, FCAP and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/09/16/calcium-channel-blocker-calcium-as-neurotransmitter-sensor-and-calcium-release-related-contractile-dysfunction-ryanopathy/

Part X: Synaptotagmin functions as a Calcium Sensor: How Calcium Ions Regulate the fusion of vesicles with cell membranes during Neurotransmission
Larry H Bernstein, MD, FCAP and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/09/10/synaptotagmin-functions-as-a-calcium-sensor-how-calcium-ions-regulate-the-fusion-of-vesicles-with-cell-membranes-during-neurotransmission/

The material presented is very focused, and cannot be found elsewhere in Pharmaceutical Intelligence with respedt to genetics and heart disease.  However, there are other articles that may be of interest to the reader.

Volume Three: Etiologies of Cardiovascular Diseases – Epigenetics, Genetics & Genomics

Curators: Larry H Bernstein, MD, FCAP and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/biomed-e-books/series-a-e-books-on-cardiovascular-diseases/volume-three-etiologies-of-cardiovascular-diseases-epigenetics-genetics-genomics/

PART 3.  Determinants of Cardiovascular Diseases: Genetics, Heredity and Genomics Discoveries

3.2 Leading DIAGNOSES of Cardiovascular Diseases covered in Circulation: Cardiovascular Genetics, 3/2010 – 3/2013

The Diagnoses covered include the following – relevant to this discussion

  • MicroRNA in Serum as Bimarker for Cardiovascular Pathologies: acute myocardial infarction, viral myocarditis, diastolic dysfunction, and acute heart failure
  • Genomics of Ventricular arrhythmias, A-Fib, Right Ventricular Dysplasia, Cardiomyopathy
  • Heredity of Cardiovascular Disorders Inheritance

3.2.1: Heredity of Cardiovascular Disorders Inheritance

The implications of heredity extend beyond serving as a platform for genetic analysis, influencing diagnosis,

  1. prognostication, and
  2. treatment of both index cases and relatives, and
  3. enabling rational targeting of genotyping resources.

This review covers acquisition of a family history, evaluation of heritability and inheritance patterns, and the impact of inheritance on subsequent components of the clinical pathway.

SOURCE:   Circulation: Cardiovascular Genetics.2011; 4: 701-709.  http://dx.doi.org/10.1161/CIRCGENETICS.110.959379

3.2.2: Myocardial Damage

3.2.2.1 MicroRNA in Serum as Biomarker for Cardiovascular Pathologies: acute myocardial infarction, viral myocarditis,  diastolic dysfunction, and acute heart failure

Increased MicroRNA-1 and MicroRNA-133a Levels in Serum of Patients With Cardiovascular Disease Indicate Myocardial Damage
Y Kuwabara, Koh Ono, T Horie, H Nishi, K Nagao, et al.
SOURCE:  Circulation: Cardiovascular Genetics. 2011; 4: 446-454   http://dx.doi.org/10.1161/CIRCGENETICS.110.958975

3.2.2.2 Circulating MicroRNA-208b and MicroRNA-499 Reflect Myocardial Damage in Cardiovascular Disease

MF Corsten, R Dennert, S Jochems, T Kuznetsova, Y Devaux, et al.
SOURCE: Circulation: Cardiovascular Genetics. 2010; 3: 499-506.  http://dx.doi.org/10.1161/CIRCGENETICS.110.957415

3.2.4.2 Large-Scale Candidate Gene Analysis in Whites and African Americans Identifies IL6R Polymorphism in Relation to Atrial Fibrillation

The National Heart, Lung, and Blood Institute’s Candidate Gene Association Resource (CARe) Project
RB Schnabel, KF Kerr, SA Lubitz, EL Alkylbekova, et al.
SOURCE:  Circulation: Cardiovascular Genetics.2011; 4: 557-564   http://dx.doi.org/10.1161/CIRCGENETICS.110.959197

 Weighted Gene Coexpression Network Analysis of Human Left Atrial Tissue Identifies Gene Modules Associated With Atrial Fibrillation

N Tan, MK Chung, JD Smith, J Hsu, D Serre, DW Newton, L Castel, E Soltesz, G Pettersson, AM Gillinov, DR Van Wagoner and J Barnard
From the Cleveland Clinic Lerner College of Medicine (N.T.), Department of Cardiovascular Medicine (M.K.C., D.W.N.), and Department of Thoracic & Cardiovascular Surgery (E.S., G.P., A.M.G.); and Department of Cellular & Molecular Medicine (J.D.S., J.H.), Genomic Medicine Institute (D.S.), Department of Molecular Cardiology (L.C.), and Department of Quantitative Health Sciences (J.B.), Cleveland Clinic Lerner Research Institute, Cleveland, OH
Circ Cardiovasc Genet. 2013;6:362-371; http://dx.doi.org/10.1161/CIRCGENETICS.113.000133
http://circgenetics.ahajournals.org/content/6/4/362   The online-only Data Supplement is available at http://circgenetics.ahajournals.org/lookup/suppl/doi:10.1161/CIRCGENETICS.113.000133/-/DC1

Background—Genetic mechanisms of atrial fibrillation (AF) remain incompletely understood. Previous differential expression studies in AF were limited by small sample size and provided limited understanding of global gene networks, prompting the need for larger-scale, network-based analyses.

Methods and Results—Left atrial tissues from Cleveland Clinic patients who underwent cardiac surgery were assayed using Illumina Human HT-12 mRNA microarrays. The data set included 3 groups based on cardiovascular comorbidities: mitral valve (MV) disease without coronary artery disease (n=64), coronary artery disease without MV disease (n=57), and lone AF (n=35). Weighted gene coexpression network analysis was performed in the MV group to detect modules of correlated genes. Module preservation was assessed in the other 2 groups. Module eigengenes were regressed on AF severity or atrial rhythm at surgery. Modules whose eigengenes correlated with either AF phenotype were analyzed for gene content. A total of 14 modules were detected in the MV group; all were preserved in the other 2 groups. One module (124 genes) was associated with AF severity and atrial rhythm across all groups. Its top hub gene, RCAN1, is implicated in calcineurin-dependent signaling and cardiac hypertrophy. Another module (679 genes) was associated with atrial rhythm in the MV and coronary artery disease groups. It was enriched with cell signaling genes and contained cardiovascular developmental genes including TBX5.

Conclusions—Our network-based approach found 2 modules strongly associated with AF. Further analysis of these modules may yield insight into AF pathogenesis by providing novel targets for functional studies. (Circ Cardiovasc Genet. 2013;6:362-371.)

Key Words: arrhythmias, cardiac • atrial fibrillation • bioinformatics • gene coexpression • gene regulatory networks • genetics • microarrays

Introduction

trial fibrillation (AF) is the most common sustained car­diac arrhythmia, with a prevalence of ≈1% to 2% in the general population.1,2 Although AF may be an isolated con­dition (lone AF [LAF]), it often occurs concomitantly with other cardiovascular diseases, such as coronary artery disease (CAD) and valvular heart disease.1 In addition, stroke risk is increased 5-fold among patients with AF, and ischemic strokes attributed to AF are more likely to be fatal.1 Current antiarrhythmic drug therapies are limited in terms of efficacy and safety.1,3,4 Thus, there is a need to develop better risk pre­diction tools as well as mechanistically targeted therapies for AF. Such developments can only come about through a clearer understanding of its pathogenesis.

Family history is an established risk factor for AF. A Danish Twin Registry study estimated AF heritability at 62%, indicating a significant genetic component.5 Substantial progress has been made to elucidate this genetic basis. For example, genome-wide association studies (GWASs) have identified several susceptibil­ity loci and candidate genes linked with AF. Initial studies per­formed in European populations found 3 AF-associated genomic loci.6–9 Of these, the most significant single-nucleotide polymor-phisms (SNPs) mapped to an intergenic region of chromosome 4q25. The closest gene in this region, PITX2, is crucial in left-right asymmetrical development of the heart and thus seems promising as a major player in initiating AF.10,11 A large-scale GWAS meta-analysis discovered 6 additional susceptibility loci, implicating genes involved in cardiopulmonary development, ion transport, and cellular structural integrity.12

Differential expression studies have also provided insight into the pathogenesis of AF. A study by Barth et al13 found that about two-thirds of the genes expressed in the right atrial appendage were downregulated during permanent AF, and that many of these genes were involved in calcium-dependent signaling pathways. In addition, ventricular-predominant genes were upregulated in right atrial appendages of sub­jects with AF.13 Another study showed that inflammatory and transcription-related gene expression was increased in right atrial appendages of subjects with AF versus controls.14 These results highlight the adaptive responses to AF-induced stress and ischemia taking place within the atria.

Despite these advances, much remains to be discovered about the genetic mechanisms of AF. The AF-associated SNPs found thus far only explain a fraction of its heritability15; furthermore, the means by which the putative candidate genes cause AF have not been fully established.9,15,16 Additionally, previous dif­ferential expression studies in human tissue were limited to the right atrial appendage, had small sample sizes, and provided little understanding of global gene interactions.13,14 Weighted gene coexpression network analysis (WGCNA) is a technique to construct gene modules within a network based on correla­tions in gene expression (ie, coexpression).17,18 WGCNA has been used to study genetically complex diseases, such as meta­bolic syndrome,19 schizophrenia,20 and heart failure.21 Here, we obtained mRNA expression profiles from human left atrial appendage tissue and implemented WGCNA to identify gene modules associated with AF phenotypes.

Methods

Subject Recruitment

From 2001 to 2008, patients undergoing cardiac surgery at the Cleveland Clinic were prospectively screened and recruited. Informed consent for research use of discarded atrial tissues was ob­tained from each patient by a study coordinator during the presur­gical visit. Demographic and clinical data were obtained from the Cardiovascular Surgery Information Registry and by chart review. Use of human atrial tissues was approved by the Institutional Review Board of the Cleveland Clinic.

Table S1: Clinical definitions of cardiovascular phenotype groups

Criterion Type Mitral Valve (MV) Disease Coronary Artery Disease (CAD) Lone Atrial Fibrillation (LAF)
Inclusion Criteria Surgical indication – Surgical indication – History of atrial fibrillation
mitral valve repair or replacement coronary artery bypass graft
Surgical indication
– MAZE procedure
Preserved ejection fraction (≥50%)
Exclusion Criteria Significant coronary artery disease: Significant mitral valve disease: Significant
coronary artery
– Significant (≥50%) stenosis – Documented echocardiography disease:
 in at least finding of – Significant
one coronary artery  mitral regurgitation (≥3) or (≥50%) stenosis in
via cardiac catheterization mitral stenosis at least one
– History of revascularization – History of mitral valve coronary artery via
(percutaneous coronary intervention or coronary artery bypass graft surgery)  repair or replacement cardiac catheterization
– History of revascularization
(percutaneous coronary intervention or coronary artery bypass graft surgery)
Significant valvular heart disease:
-Documented echocardiography finding of valvular regurgitation (≥3) or stenosis
-History of valve repair or replacement

RNA Microarray Isolation and Profiling

Left atria appendage specimens were dissected during cardiac surgery and stored frozen at −80°C. Total RNA was extracted using the Trizol technique. RNA samples were processed by the Cleveland Clinic Genomics Core. For each sample, 250-ng RNA was reverse tran­scribed into cRNA and biotin-UTP labeled using the TotalPrep RNA Amplification Kit (Ambion, Austin, TX). cRNA was quantified using a Nanodrop spectrophotometer, and cRNA size distribution was as­sessed on a 1% agarose gel. cRNA was hybridized to Illumina Human HT-12 Expression BeadChip arrays (v.3). Arrays were scanned using a BeadArray reader.

Expression Data Preprocessing

Raw expression data were extracted using the beadarray package in R, and bead-level data were averaged after log base-2 transformation. Background correction was performed by fitting a normal-gamma deconvolution model using the NormalGamma R package.22 Quantile normalization and batch effect adjustment with the ComBat method were performed using R.23 Probes that were not detected (at a P<0.05 threshold) in all samples as well as probes with relatively lower vari­ances (interquartile range ≤log2[1.2]) were excluded.

The WGCNA approach requires that genes be represented as sin­gular nodes in such a network. However, a small proportion of the genes in our data have multiple probe mappings. To facilitate the representation of singular genes within the network, a probe must be selected to represent its associated gene. Hence, for genes that mapped to multiple probes, the probe with the highest mean expres­sion level was selected for analysis (which often selects the splice isoform with the highest expression and signal-to-noise ratio), result­ing in a total of 6168 genes.

Defining Training and Test Sets

Currently, no large external mRNA microarray data from human left atrial tissues are publicly available. To facilitate internal validation of results, we divided our data set into 3 groups based on cardiovascular comorbidities: mitral valve (MV) disease without CAD (MV group; n=64), CAD without MV disease (CAD group; n=57), and LAF (LAF group; n=35). LAF was defined as the presence of AF without concomitant structural heart disease, according to the guidelines set by the European Society of Cardiology.1 The MV group, which was the largest and had the most power for detecting significant modules, served as the training set for module derivation, whereas the other 2 groups were designated test sets for module reproducibility. To mini­mize the effect of population stratification, the data set was limited to white subjects. Differences in clinical characteristics among the groups were assessed using Kruskal–Wallis rank-sum tests for con­tinuous variables and Pearson x2 test for categorical variables.

Weight Gene Coexpression Network Analysis

WGCNA is a systems-biology method to identify and characterize gene modules whose members share strong coexpression. We applied previously validated methodology in this analysis.17 Briefly, pair-wise gene (Pearson) correlations were calculated using the MV group data set. A weighted adjacency matrix was then constructed. I is a soft-thresholding pa­rameter that provides emphasis on stronger correlations over weaker and less meaningful ones while preserving the continuous nature of gene–gene relationships. I=3 was selected in this analysis based on the criterion outlined by Zhang and Horvath17 (see the online-only Data Supplement).

Next, the topological overlap–based dissimilarity matrix was com­puted from the weighted adjacency matrix. The topological overlap, developed by Ravasz et al,24 reflects the relative interconnectedness (ie, shared neighbors) between 2 genes.17 Hence, construction of the net­work dendrogram based on this dissimilarity measure allows for the identification of gene modules whose members share strong intercon-nectivity patterns. The WGCNA cutreeDynamic R function was used to identify a suitable cut height for module identification via an adap­tive cut height selection approach.18 Gene modules, defined as branches of the network dendrogram, were assigned colors for visualization.

Network Preservation Analysis

Module preservation between the MV and CAD groups as well as the MV and LAF groups was assessed using network preservation statis­tics as described in Langfelder et al.25 Module density–based statistics (to assess whether genes in each module remain highly connected in the test set) and connectivity-based statistics (to assess whether con­nectivity patterns between genes in the test set remain similar com­pared with the training set) were considered in this analysis.25 In each comparison, a Z statistic representing a weighted summary of module density and connectivity measures was computed for every module (Zsummary). The Zsummary score was used to evaluate module preserva­tion, with values ≥8 indicating strong preservation, as proposed by Langfelder et al.25 The WGCNA R function network preservation was used to implement this analysis.25

Table S2: Network preservation analysis between the MV and CAD groups – size and Zsummary scores of gene modules detected.

Module Module Size

ZSummary

Black 275 15.52
Blue 964 44.79
Brown 817 12.80
Cyan 119 13.42
Green 349 14.27
Green-Yellow 215 19.31
Magenta 239 15.38
Midnight-Blue 83 15.92
Pink 252 23.31
Purple 224 16.96
Red 278 17.30
Salmon 124 13.84
Tan 679 28.48
Turquoise 1512 44.03


Table S3: Network preservation analysis between the MV and LAF groups – size and Zsummary scores of gene modules detected

Module Module Size ZSummary
Black 275 13.14
Blue 964 39.26
Brown 817 14.98
Cyan 119 11.46
Green 349 14.91
Green-Yellow 215 20.99
Magenta 239 18.58
Midnight-Blue 83 13.87
Pink 252 19.10
Purple 224 8.80
Red 278 16.62
Salmon 124 11.57
Tan 679 28.61
Turquoise 1512 42.07

Clinical Significance of Preserved Modules

Principal component analysis of the expression data for each gene module was performed. The first principal component of each mod­ule, designated the eigengene, was identified for the 3 cardiovascular disease groups; this served as a summary expression measure that explained the largest proportion of the variance of the module.26 Multivariate linear regression was performed with the module ei-gengenes as the outcome variables and AF severity (no AF, parox­ysmal AF, persistent AF, permanent AF) as the predictor of interest (adjusting for age and sex). A similar regression analysis was per­formed with atrial rhythm at surgery (no AF history, AF history in sinus rhythm, AF history in AF rhythm) as the predictor of interest. The false discovery rate method was used to adjust for multiple com­parisons. Modules whose eigengenes associated with AF severity and atrial rhythm were identified for further analysis.

In addition, hierarchical clustering of module eigengenes and se­lected clinical traits (age, sex, hypertension, cholesterol, left atrial size, AF state, and atrial rhythm) was used to identify additional module–trait associations. Clusters of eigengenes/traits were detected based on a dissimilarity measure D, as given by

D=1−cor(Vi,Vj),i≠j                                                                              (3)

where V=the eigengene or clinical trait.

Enrichment Analysis

Gene modules significantly associated with AF severity and atrial rhythm were submitted to Ingenuity Pathway Analysis (IPA) to determine enrichment for functional/disease categories. IPA is an application of gene set over-representation analysis; for each dis-ease/functional category annotation, a P value is calculated (using Fisher exact test) by comparing the number of genes from the mod­ule of interest that participate in the said category against the total number of participating genes in the background set.27 All 6168 genes in the current data set served as the background set for the enrichment analysis.

Hub Gene Analysis

Hub genes are defined as genes that have high intramodular connectivity17,20

Alternatively, they may also be defined as genes with high module membership21,25

Both definitions were used to identify the hub genes of modules associated with AF phenotype.

To confirm that the hub genes identified were themselves associ­ated with AF phenotype, the expression data of the top 10 hub genes (by intramodular connectivity) were regressed on atrial rhythm (ad­justing for age and sex). In addition, eigengenes of AF-associated modules were regressed on their respective (top 10) hub gene expres­sion profiles, and the model R2 indices were computed.

Membership of AF-Associated Candidate Genes From Previous Studies

Previous GWAS studies identified multiple AF-associated SNPs.8,9,12,15,28 We selected candidate genes closest to or containing these SNPs and identified their module locations as well as their clos­est within-module partners (absolute Pearson correlations).

Sensitivity Analysis of Soft-Thresholding Parameter

To verify that the key results obtained from the above analysis were robust with respect to the chosen soft-thresholding parameter (I=3), we repeated the module identification process using I=5. The eigen-genes of the detected modules were computed and regressed on atrial rhythm (adjusting for age and sex). Modules significantly associated with atrial rhythm in ≥2 groups of data set were compared with the AF phenotype–associated modules from the original analysis.

Results

Subject Characteristics

Table 1 describes the clinical characteristics of the cardiac surgery patients who were recruited for the study. Subjects in the LAF group were generally younger and less likely to be a current smoker (P=2.0×10−4 and 0.032, respectively). Subjects in the MV group had lower body mass indices (P=2.7×10−6), and a larger proportion had paroxysmal AF compared with the other 2 groups (P=0.033).

Table 1. Clinical Characteristics of Study Subjects

Characteristics

MV Group (n=64)

CAD Group (n=57)

LAF Group (n=35)

P Value*

Age, median y (1st–3rd quartiles)

60 (51.75–67.25)

64 (58.00–70.00)

56 (45.50–60.50)

2.0×10−4

Sex, female (%) 19 (29.7) 6 (10.5)

7 (20.0)

0.033

BMI, median (1st–3rd quartiles)

25.97 (24.27–28.66)

29.01 (27.06–32.11)

29.71 (26.72–35.10)

2.7×10−6

Current smoker (%) 29 (45.3) 35 (61.4)

12 (21.1)

0.032

Hypertension (%) 21 (32.8) 39 (68.4)

16 (45.7)

4.4×10−4

AF severity (%)
No AF 7 (10.9) 7 (12.3)

0 (0.0)

0.033

Paroxysmal 19 (29.7) 10 (17.5)

7 (20.0)

Persistent 30 (46.9) 26 (45.6)

15 (42.9)

Permanent 8 (12.5) 14 (24.6)

13 (37.1)

Atrial rhythm at surgery (%)
No AF history in sinus rhythm 7 (10.9) 7 (12.3)

0 (0)

0.065

AF history in sinus rhythm 28 (43.8) 16 (28.1)

11 (31.4)

AF History in AF rhythm 29 (45.3) 34 (59.6)

24 (68.6)

Gene Coexpression Network Construction and Module Identificationsee document at  http://circgenetics.ahajournals.org/content/6/4/362

A total of 14 modules were detected using the MV group data set (Figure 1), with module sizes ranging from 83 genes to 1512 genes; 38 genes did not share similar coexpression with the other genes in the network and were therefore not included in any of the identified modules

Figure 1. Network dendrogram (top) and colors of identified modules (bottom).

Figure 1. Network dendrogram (top) and colors of identified modules (bottom). The dendrogram was constructed using the topological overlap matrix as the similarity measure. Modules corresponded to branches of the dendrogram and were assigned colors for visualization.

Network Preservation Analysis Revealed Strong Preservation of All Modules Between the Training and Test Sets

All 14 modules showed strong preservation across the CAD and LAF groups in both comparisons, with Z [summary]  scores of >10 in most modules (Figure 2). No major deviations in the Z [summary] score distributions for the 2 comparisons were noted, indicating that modules were preserved to a similar extent across the 2 groups

Figure 2. Preservation of mod-ules between mitral valve (MV) and coronary artery disease

Figure 2. Preservation of mod­ules between mitral valve (MV) and coronary artery disease (CAD) groups (left), and MV and lone atrial fibrillation (LAF) groups (right). A Zsummary sta­tistic was computed for each module as an overall measure of its preservation relating to density and connectivity. All modules showed strong pres­ervation in both comparisons with Zsummary scores >8 (red dot­ted line).

Regression Analysis of Module Eigengene Profiles Identified 2 Modules Associated With AF Severity and Atrial Rhythm

Table IV in the online-only Data Supplement summarizes the proportion of variance explained by the first 3 principal components for each module. On average, the first principal component (ie, the eigengene) explained ≈18% of the total variance of its associated module. For each group, the mod­ule eigengenes were extracted and regressed on AF severity (with age and sex as covariates). The salmon module (124 genes) eigengene was strongly associated with AF severity in the MV and CAD groups (P=1.7×10−6 and 5.2×10−4, respec­tively); this association was less significant in the LAF group (P=9.0×10−2). Eigengene levels increased with worsening AF severity across all 3 groups, with the greatest stepwise change taking place between the paroxysmal AF and per­sistent AF categories (Figure 3A). When the module eigen-genes were regressed on atrial rhythm, the salmon module eigengene showed significant association in all groups (MV: P=1.1×10−14; CAD: P=1.36×10−6; LAF: P=2.1×10−4). Eigen-gene levels were higher in the AF history in AF rhythm cat­egory (Figure 3B).

Table S4: Proportion of variance explained by the principal components for each module.

Dataset
Group

Principal
Component

Black

Blue

Brown

Cyan

Green

Green-
Yellow

Magenta

Mitral

1

20.5% 22.2% 20.1% 21.8% 21.4% 22.8% 19.6%

2

4.1% 3.6% 4.8% 5.7% 4.5% 5.9% 3.9%

3

3.4% 3.1% 3.8% 4.4% 3.9% 3.7% 3.7%

CAD

1

12.5% 18.6% 7.1% 16.8% 12.2% 20.3% 12.8%

2

6.0% 5.5% 5.0% 7.0% 5.5% 6.1% 6.4%

3

4.9% 4.1% 4.4% 6.5% 4.8% 4.4% 4.8%

LAF

1

14.0% 16.6% 11.7% 14.3% 14.7% 20.8% 20.2%

2

8.9% 8.5% 7.6% 9.3% 7.3% 11.1% 6.9%

3

6.5% 6.3% 5.5% 8.2% 6.1% 5.3% 6.2%

Dataset
Group

Principal
Component

Midnight- Blue

Pink

Purple

Red

Salmon

Tan

Turquoise

Mitral

1

28.5% 22.6% 18.7% 20.5% 22.3% 19.0% 25.8%

2

4.6% 6.0% 4.7% 4.1% 6.9% 4.0% 3.5%

3

4.2% 4.2% 4.2% 3.5% 4.0% 3.6% 3.3%

CAD

1

23.4% 17.1% 15.5% 15.0% 18.0% 14.6% 18.2%

2

7.4% 8.6% 6.0% 6.4% 7.2% 5.8% 6.6%

3

5.1% 5.4% 5.3% 5.4% 6.2% 5.1% 4.5%

LAF

1

23.5% 18.4% 12.0% 15.9% 16.9% 13.7% 16.5%

2

7.9% 8.5% 9.8% 9.4% 9.5% 9.1% 9.6%

3

6.7% 7.0% 6.6% 6.0% 6.9% 6.8% 6.3%

Figure 3. Boxplots of salmon module eigengene expression levels with respect to atrial fibrillation (AF) severity (A) and atrial rhythm (B).

Figure 3. Boxplots of salmon module eigengene expression levels with respect to atrial fibrillation (AF) severity (A) and atrial rhythm (B).
A, Eigengene expression correlated positively with AF severity, with the largest stepwise increase between the paroxysmal AF and per­manent AF categories. B, Eigengene expression was highest in the AF history in AF rhythm category in all 3 groups. CAD indicates coro­nary artery disease; LAF, lone AF; and MV, mitral valve.

The regression analysis also revealed statistically significant associations between the tan module (679 genes) eigengene and atrial rhythm in the MV and CAD groups (P=5.8×10−4 and 3.4×10−2, respectively). Eigengene levels were lower in the AF history in AF rhythm category compared with the AF history in sinus rhythm category (Figure 4); this trend was also observed in the LAF group, albeit with weaker statistical evidence (P=0.15).

Figure 4. Boxplots of tan module eigengene expression levels with respect to atrial rhythm.

Figure 4. Boxplots of tan module eigengene expression levels with respect to atrial rhythm.
Eigengene expression levels were lower in the atrial fibrillation (AF) history in AF rhythm category compared with the AF history in sinus rhythm category. CAD indicates coronary artery disease; LAF, lone AF; and MV, mitral valve

Hierarchical Clustering of Eigengene Profiles With Clinical Traits

Hierarchical clustering was performed to identify relation­ships between gene modules and selected clinical traits. The salmon module clustered with AF severity and atrial rhythm; in addition, left atrial size was found in the same cluster, sug­gesting a possible relationship between salmon module gene expression and atrial remodeling (Figure 5A). Although the tan module was in a separate cluster from the salmon module, it was negatively correlated with both atrial rhythm and AF severity (Figure 5B).

Figure 5. Dendrogram (A) and correlation heatmap (B) of module eigengenes and clinical traits.

Figure 5. Dendrogram (A) and correlation heatmap (B) of module eigengenes and clinical traits

A, The salmon module eigengene but not the tan module eigengene clustered with atrial fibrillation (AF) severity, atrial rhythm, and left atrial size. B, AF severity and atrial rhythm at surgery correlated positively with the salmon module eigengene and negatively with the tan module eigengene. Arhythm indicates atrial rhythm at surgery; Chol, cholesterol; HTN, hypertension; and LASize, left atrial size.

IPA Enrichment Analysis of Salmon and Tan Modules

The salmon module was enriched in genes involved in cardio­vascular function and development (smallest P=4.4×10−4) and organ morphology (smallest P=4.4×10−4). In addition, the top disease categories identified included endocrine system disor­ders (smallest P=4.4×10−4) and cardiovascular disease (small­est P=2.59×10−3).

The tan module was enriched in genes involved in cell-to-cell signaling and interaction (smallest P=8.9×10−4) and cell death and survival (smallest P=1.5×10−3). Enriched disease categories included cancer (smallest P=2.2×10−4) and cardio­vascular disease (smallest P=4.5×10−4).

see document at  http://circgenetics.ahajournals.org/content/6/4/362

Hub Gene Analysis of Salmon and Tan Modules

We identified hub genes in the 2 modules based on intramod-ular connectivity and module membership. For the salmon module, the gene RCAN1 exhibited the highest intramodular connectivity and module membership. The top 10 hub genes (by intramodular connectivity) were significantly associated with atrial rhythm, with false discovery rate–adjusted P values ranging from 1.5×10−5 to 4.2×10−12. These hub genes accounted for 95% of the variation in the salmon module eigengene.

In the tan module, the top hub gene was CPEB3. The top 10 hub genes (by intramodular connectivity) correlated with atrial rhythm as well, although the statistical associations in the lower-ranked hub genes were relatively weaker (false discovery rate–adjusted P values ranging from 1.1×10−1 to 3.4×10−4). These hub genes explained 94% of the total varia­tion in the tan module eigengene.

The names and connectivity measures of the hub genes found in both modules are presented in Table 2.

Table 2. Top 10 Hub Genes in the Salmon (Left) and Tan (Right) Modules as Defined by Intramodular Connectivity and Module Membership

Salmon Module

Tan Module

Gene

IMC

Gene

MM

Gene

IMC

Gene

MM

RCAN1 8.2

RCAN1

0.81

CPEB3

43.3

CPEB3

0.85
DNAJA4 7.7

DNAJA4

0.81

CPLX3

42.4

CPLX3

0.84
PDE8B 7.7

PDE8B

0.80

NEDD4L

40.8

NEDD4L

0.83
PRKAR1A 6.9

PRKAR1A

0.77

SGSM1

40.7

SGSM1

0.82
PTPN4 6.7

PTPN4

0.75

UCKL1

39.0

UCKL1

0.81
SORBS2 6.0

FHL2

0.69

SOSTDC1

37.2

SOSTDC1

0.79
ADCY6 5.7

ADCY6

0.69

PRDX1

35.5

RCOR2

0.78
FHL2 5.7

SORBS2

0.68

RCOR2

35.4

EEF2K

0.77
BVES 5.4

DHRS9

0.67

NPPB

35.3

PRDX1

0.76
TMEM173 5.3

LAPTM4B

0.65

LRRN3

34.6

MMP11

0.76

A visualiza­tion of the salmon module is shown using the Cytoscape tool (Figure 6). A full list of the genes in the salmon and tan mod­ules is provided in the online-only Data Supplement.

Figure 6. Cytoscape visualization of genes in the salmon module.
Nodes representing genes with high intramodu-lar connectivities, such as RCAN1 and DNAJA4, appear larger in the network. Strong connections are visualized with darker lines, whereas weak connections appear more translucent

Figure 6. Cytoscape visualization of genes in the salmon module.

Membership of AF-Associated Candidate Genes From Previous Studies

The tan module contained MYOZ1, which was identified as a candidate gene from the recent AF meta-analysis. PITX2 was located in the green module (n=349), and ZFHX3 was located in the turquoise module (n=1512). The locations of other can­didate genes (and their closest partners) are reported in the online-only Data Supplement.

Sensitivity Analysis of Key Results

We repeated the WGCNA module identification approach using a different soft-thresholding parameter (β=5). One mod­ule (n=121) was found to be strongly associated with atrial rhythm at surgery across all 3 groups of data set, whereas another module (n=244) was associated with atrial rhythm at surgery in the MV and CAD groups. The first module over­lapped significantly with the salmon module in terms of gene membership, whereas most of the second modules’ genes were contained within the tan module. The top hub genes found in the salmon and tan modules remained present and highly connected in the 2 new modules identified with the dif­ferent soft-thresholding parameter.

Discussion

To our knowledge, our study is the first implementation of an unbiased, network-based analysis in a large sample of human left atrial appendage gene expression profiles. We found 2 modules associated with AF severity and atrial rhythm in 2 to 3 of our cardiovascular comorbidity groups. Functional analy­ses revealed significant enrichment of cardiovascular-related categories for both modules. In addition, several of the hub genes identified are implicated in cardiovascular disease and may play a role in AF initiation and progression.

In our study, WGCNA was used to construct modules based on gene coexpression, thereby reducing the net-work’s dimensionality to a smaller set of elements.17,21 Relating modulewise changes to phenotypic traits allowed statistically significant associations to be detected at a lower false discovery rate compared with traditional differential expression studies. Furthermore, shared functions and path­ways among genes in the modules could be inferred via enrichment analyses.

We divided our data set into 3 groups to verify the repro­ducibility of the modules identified by WGCNA; 14 modules were identified in the MV group in our gene network. All were strongly preserved in the CAD and LAF groups, suggesting that gene coexpression patterns are robust and reproducible despite differences in cardiovascular comorbidities.

The use of module eigengene profiles as representative summary measures has been validated in a number of studies.20,26 Additionally, we found that the eigengenes accounted for a significant proportion (average 18%) of gene expression variability in their respective modules. Regression analysis of the module eigengenes found 2 modules associated with AF severity and atrial rhythm in ≥2 groups of data set. The association between the salmon module eigengene and AF severity was statistically weaker in the LAF group (adjusted P=9.0×10−2). This was probably because of its significantly smaller sample size compared with the MV and CAD groups. Despite this weaker association, the relationship between the salmon module eigengene and AF severity remained consistent among the 3 groups (Figure 3A). Similarly, the lack of statistical significance for the association between the tan module eigengene and atrial rhythm at surgery in the LAF group was likely driven by the smaller sample size and (by definition) lack of samples in the no AF category.

A major part of our analysis focused on the identifica­tion of module hub genes. Hubs are connected with a large number of nodes; disruption of hubs therefore leads to wide­spread changes within the network. This concept has powerful applications in the study of biology, genetics, and disease.29,30 Although mutations of peripheral genes can certainly lead to disease, gene network changes are more likely to be motivated by changes in hub genes, making them more biologically inter­esting targets for further study.17,29,31 Indeed,

  • the hub genes of the salmon and tan modules accounted for the vast majority of the variation in their respective module eigengenes, signaling their importance in driving gene module behavior.

The hub genes identified in the salmon and tan modules were significantly associated with AF phenotype overall. It was noted that this association was statistically weaker for the lower-ranked hub genes in the tan module. This highlights an important aspect and strength of WGCNA—to be able to capture module-wide changes with respect to disease despite potentially weaker associations among individual genes.

The implementation of WGCNA necessitated the selection of a soft-thresholding parameter 13. Unlike hard-thresholding (where gene correlations below a certain value are shrunk to zero), the soft-thresholding approach gives greater weight to stronger correlations while maintaining the continuous nature of gene–gene relationships. We selected a 13 value of 3 based on the criteria outlined by Zhang and Horvath.17 His team and other investigators have demonstrated that module identifica­tion is robust with respect to the 13 parameter.17,19–21 In our data, we were also able to reproduce the key findings reported with a different, larger 13 value, thereby verifying the stability of our results relating to 13.

The salmon module (124 genes) was associated with both AF phenotypes; furthermore, IPA analysis of its gene con­tents suggested enrichment in cardiovascular development as well as disease. Its eigengene increased with worsening AF severity, with the largest stepwise change occurring between the paroxysmal AF and persistent AF categories (Figure 3). Hence,

  • the gene expression changes within the salmon mod­ule may reflect the later stages of AF pathophysiology.

The top hub gene of the salmon module was RCAN1 (reg­ulator of calcineurin 1). Calcineurin is a cytoplasmic Ca2+/ calmodulin-dependent protein phosphatase that stimulates cardiac hypertrophy via its interactions with NFAT and L-type Ca2+ channels.32,33 RCAN1 is known to inhibit calcineurin and its associated pathways.32,34 However, some data suggest that RCAN1 may instead function as a calcineurin activator when highly expressed and consequently potentiate hypertrophic signaling.35 Thus,

  • perturbations in RCAN1 levels (attribut­able to genetic variants or mutations) may cause an aberrant switching in function, which in turn triggers atrial remodeling and arrhythmogenesis.

Other hub genes found in the salmon module are also involved in cardiovascular development and function and may be potential targets for further study.

  • DNAJA4 (DnaJ homolog, subfamily A, member 4) regulates the trafficking and matu­ration of KCNH2 potassium channels, which have a promi­nent role in cardiac repolarization and are implicated in the long-QT syndromes.36

FHL2 (four-and-a-half LIM domain protein 2) interacts with numerous cellular components, including

  1. actin cytoskeleton,
  2. transcription machinery, and
  3. ion channels.37

FHL2 was shown to enhance the hypertrophic effects of isoproterenol, indicating that

  • FHL2 may modulate the effect of environmental stress on cardiomyocyte growth.38
  • FHL2 also interacts with several potassium channels in the heart, such as KCNQ1, KCNE1, and KCNA5.37,39

Additionally, blood vessel epicardial substance (BVES) and other members of its family were shown to be highly expressed in cardiac pacemaker cells. BVES knockout mice exhibited sinus nodal dysfunction, suggesting that BVES regulates the development of the cardiac pacemaking and conduction system40 and may therefore be involved in the early phase of AF development.

The tan module (679 genes) eigengene was negatively correlated with atrial rhythm in the MV and CAD groups (Figure 4); this may indicate a general decrease in gene expres­sion of its members in fibrillating atrial tissue. IPA analysis revealed enrichment in genes involved in cell signaling as well as apoptosis. The top-ranked hub gene, cytoplasmic polyade-nylation element binding protein 3 (CPEB3), regulates mRNA translation and has been associated with synaptic plasticity and memory formation.41 The role of CPEB3 in the heart is currently unknown, so further exploration via animal model studies may be warranted.

Natriuretic peptide-precursor B (NPPB), another highly interconnected hub gene, produces a precursor peptide of brain natriuretic peptide, which

  • regulates blood pressure through natriuresis and vasodilation.42

(NPPB) gene variants have been linked with diabetes mellitus, although associations with cardiac phenotypes are less clear.42 TBX5 and GATA4, which play important roles in the embryonic heart development,43 were members of the tan module. Although not hub genes, they may also contribute toward developmental sus­ceptibility of AF. In addition, TBX5 was previously reported to be near an SNP associated with PR interval and AF in separate large-scale GWAS studies.12,28 MYOZ1, another candidate gene identified in the recent AF GWAS meta-analysis, was found to be a member as well; it associates with proteins found in the Z-disc of skeletal and cardiac muscle and may suppress calcineurin-dependent hypertrophic signaling.12

Some, but not all, of the candidate genes found in previous GWAS studies were located in the AF-associated modules. One possible explanation for this could be the difference in sample sizes. The meta-analysis involved thousands of indi­viduals, whereas the current study had <100 in each group of data set, which limited the power to detect significant differ­ences between levels of AF phenotype even with the module-wise approach. Additionally, transcription factors like PITX2 are most highly expressed during the fetal phase of develop­ment. Perturbations in these genes (attributable to genetic variants or mutations) may therefore initiate the development of AF at this stage and play no significant role in adults (when we obtained their tissue samples).

Limitations in Study

We noted several limitations in this study. First, no human left atrial mRNA data set of adequate size currently exists publicly. Hence, we were unable to validate our results with an external, independent data set. However, the network pres­ervation assessment performed within our data set showed strong preservation in all modules, indicating that our findings are robust and reproducible.

Although the module eigengenes captured a significant pro­portion of module variance, a large fraction of variability did remain unaccounted for, which may limit their use as repre­sentative summary measures.

We extracted RNA from human left atrial appendage tis­sue, which consists primarily of cardiomyocytes and fibro­blasts. Atrial fibrosis is known to occur with AF-associated remodeling.44 As such, the cardiomyocyte to fibroblast ratio is likely to change with different levels of AF severity, which in turn influences the amount of RNA extracted from each cell type. Hence, true differences in gene expression (and coexpression) within cardiomyocytes may be confounded by changes in cellular composition attributable to atrial remod­eling. Also, there may be significant regional heterogeneity in the left atrium with respect to structure, cellular composi­tion, and gene expression,45 which may limit the generaliz-ability of our results to other parts of the left atrium.

All subjects in the study were whites to minimize the effects of population stratification. However, it is recognized that the genetic basis of AF may differ among ethnic groups.9 Thus, our results may not be generalizable to other ethnicities.

Finally, it is possible for genes to be involved in multiple processes and functions that require different sets of genes. However, WGCNA does not allow for overlapping modules to be formed. Thus,

  • this limits the method’s ability to character­ize such gene interactions.

Conclusions

In summary, we constructed a weighted gene coexpression network based on RNA expression data from the largest collection of human left atrial appendage tissue specimens to date. We identified 2 gene modules significantly associated with AF severity or atrial rhythm at surgery. Hub genes within these modules may be involved in the initiation or progression of AF and may therefore be candidates for functional stud­ies.

Refererences

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2. Lemmens R, Hermans S, Nuyens D, Thijs V. Genetics of atrial fibrilla­tion and possible implications for ischemic stroke. Stroke Res Treat. 2011;2011:208694.

3. Wann LS, Curtis AB, January CT, Ellenbogen KA, Lowe JE, Estes NA III, et al; ACCF/AHA/HRS. 2011 ACCF/AHA/HRS focused update on the management of patients with atrial fibrillation (Updating the 2006 Guideline): a report of the American College of Cardiology Foundation/ American Heart Association Task Force on Practice Guidelines. J Am Coll Cardiol. 2011;57:223–242.

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5. Christophersen IE, Ravn LS, Budtz-Joergensen E, Skytthe A, Haunsoe S, Svendsen JH, et al. Familial aggregation of atrial fibrillation: a study in Danish twins. Circ Arrhythm Electrophysiol. 2009;2:378–383.

6. Gudbjartsson DF, Arnar DO, Helgadottir A, Gretarsdottir S, Holm H, Sig-urdsson A, et al. Variants conferring risk of atrial fibrillation on chromo­some 4q25. Nature. 2007;448:353–357.

7. Ellinor PT, Lunetta KL, Glazer NL, Pfeufer A, Alonso A, Chung MK, et al. Common variants in KCNN3 are associated with lone atrial fibrillation. Nat Genet. 2010;42:240–244.

8. Benjamin EJ, Rice KM, Arking DE, Pfeufer A, van Noord C, Smith AV, et al. Variants in ZFHX3 are associated with atrial fibrillation in individuals of European ancestry. Nat Genet. 2009;41:879–881.

9. Sinner MF, Ellinor PT, Meitinger T, Benjamin EJ, Kääb S. Genome-wide association studies of atrial fibrillation: past, present, and future. Cardio-vasc Res. 2011;89:701–709.

10. Clauss S, Kääb S. Is Pitx2 growing up? Circ Cardiovasc Genet. 2011;4:105–107.

11. Kirchhof P, Kahr PC, Kaese S, Piccini I, Vokshi I, Scheld HH, et al. PITX2c is expressed in the adult left atrium, and reducing Pitx2c expres­sion promotes atrial fibrillation inducibility and complex changes in gene expression. Circ Cardiovasc Genet. 2011;4:123–133.

12. Ellinor PT, Lunetta KL, Albert CM, Glazer NL, Ritchie MD, Smith AV, et al. Meta-analysis identifies six new susceptibility loci for atrial fibrillation. Nat Genet. 2012;44:670–675.

13. Barth AS, Merk S, Arnoldi E, Zwermann L, Kloos P, Gebauer M, et al. Reprogramming of the human atrial transcriptome in permanent atrial fi­brillation: expression of a ventricular-like genomic signature. Circ Res. 2005;96:1022–1029.

Continues to 45.  see

http://circgenetics.ahajournals.org/content/6/4/362

CLINICAL PERSPECTIVE

Atrial fibrillation is the most common sustained cardiac arrhythmias in the United States. The genetic and molecular mecha­nisms governing its initiation and progression are complex, and our understanding of these mechanisms remains incomplete despite recent advances via genome-wide association studies, animal model experiments, and differential expression studies. In this study, we used weighted gene coexpression network analysis to identify gene modules significantly associated with atrial fibrillation in a large sample of human left atrial appendage tissues. We further identified highly interconnected genes (ie, hub genes) within these gene modules that may be novel candidates for functional studies. The discovery of the atrial fibrillation-associated gene modules and their corresponding hub genes provide novel insight into the gene network changes that occur with atrial fibrillation, and closer study of these findings can lead to more effective targeted therapies for disease management.

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Oxidized Calcium Calmodulin Kinase and Atrial Fibrillation

Author: Larry H. Bernstein, MD, FCAP

and

Curator: Aviva Lev-Ari, PhD, RN

 

Introduction

This is a review of a recent work from the laboratory of Mark E. Anderson and associates at the University of Iowa.  WE have covered the role of CaMKII in calcium signaling and myocardiocyte contraction, as well as signaling in smooth muscle, skeletal muscle, and nerve transmission.  There are tissue specific modus operandi, partly related to the ryanogen receptor, and also related to tissue specific isoenzymes of CaMKII.  There is much ground that has been traversed in exploring these mechanisms, most recently, the discoverey of hormone triggering by the release from vesicles at the nerve muscle junction, and much remains open to investigation.  The recently published work by Mark E. Anderson and associates in Mannheim and Heidelberg, Germany, clarifies the relationship between the oxidized form of CaMKII and the triggering of atrial fibrillation. The following studies show:

  1. Ang II infusion increased the susceptibility of mice to AF induction by rapid right atrial pacing and established a framework for us to test the hypothesized role of ox-CaMKII in promoting AF. ox-CaMKII is critical for AF.
  2. Estalished a critical role of ox-CaMKII in promoting AF
  3. Ang II induced increases in ROS production seen in WT atria were absent in atria from MsrA TG mice suggesting that MsrA sensitive targets represent an important component of Ang II mediated atrial oxidation.
  4. The protection from AF in MsrA TG mice appeared to be independent of pressor effects that are critical for the proarrhythmic actions.
  5. These findings suggest that NADPH oxidase dependent ROS and elevated ox-CaMKII drive Ang II  -pacing-induced AF and that
  6. targeted antioxidant therapy, by MsrA over-expression, can reduce or prevent AF in Ang -II-infused mice.  
  7. Atrial myocytes from Ang II treated WT mice showed a significant (p<0.05) increase in spontaneous Ca2+ sparks compared to atrial myocytes from saline treated control mice
  8. In contrast to findings in WT mice, the atrial myocytes isolated from Ang II treated MM-VV mice did not show an increase in Ca2+ sparks compared to saline treated MM-VV mice
  9. These data to suggest that  in ox–the proarrhythmic effects of Ang I I infusion depend upon an increaseCaMKII, sarcoplasmic reticulum Ca2+ leak and DADs.
  10. Enhanced CaMKII-mediated phosphorylation of serine 2814 on RyR2 is associated with an increased susceptibility to acquired arrhythmias, including AF
  11. Proarrhythmic actions of ox-CaMKII require access to RyR2 serine 2814.
  12. Mutant S2814A knock-in mice (lacking serine 2814) were highly resistant to Ang II mediated AF
  13. AC3-I mice with transgenic myocardial expression of a CaMKII inhibitory peptide were also resistant to the proarrhythmic effects of Ang II infusion on pacing-induced AF
  14. S2814A, AC3-I and WT mice, all developed similar BP increases and cardiac hypertrophy in response to Ang II, indicating that these mice were not resistant to the hemodynamic effects of Ang II, but were nevertheless protected from AF.
  15. selectively targeted antioxidant therapies could be effective in preventing or reducing AF 
  16. half of patients enrolled in the Mode Selection Trial (MOST) with sinus node dysfunction had a history of AF
  17. Ang II and diabetes-induced CaMKII oxidation caused sinus node dysfunction by increased pacemaker cell death and fibrosis
  18.  ox-CaMKII increases susceptibility for AF via increased diastolic sarcoplasmic reticulum Ca2+ release
  19. clinical association between sinus node dysfunction and AF might have a mechanistic basis because sinus node dysfunction and AF are downstream consequences of elevated ox-CaMKII.

We refer to the following related articles published in pharmaceutical Intelligence:

Contributions to cardiomyocyte interactions and signaling
Author and Curator: Larry H Bernstein, MD, FCAP  and Curator: Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/10/21/contributions-to-cardiomyocyte-interactions-and-signaling/

Cardiac Contractility & Myocardium Performance: Therapeutic Implications for Ryanopathy (Calcium Release-related Contractile Dysfunction) and Catecholamine Responses
Editor: Justin Pearlman, MD, PhD, FACC, Author and Curator: Larry H Bernstein, MD, FCAP, and Article Curator: Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/08/28/cardiac-contractility-myocardium-performance-ventricular-arrhythmias-and-non-ischemic-heart-failure-therapeutic-implications-for-cardiomyocyte-ryanopathy-calcium-release-related-contractile/

Part I. Identification of Biomarkers that are Related to the Actin Cytoskeleton
Curator and Writer: Larry H Bernstein, MD, FCAP
https://pharmaceuticalintelligence.com/2012/12/10/identification-of-biomarkers-that-are-related-to-the-actin-cytoskeleton/

Part II: Role of Calcium, the Actin Skeleton, and Lipid Structures in Signaling and Cell Motility
Larry H. Bernstein, MD, FCAP, Stephen Williams, PhD and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/08/26/role-of-calcium-the-actin-skeleton-and-lipid-structures-in-signaling-and-cell-motility/

Part IV: The Centrality of Ca(2+) Signaling and Cytoskeleton Involving Calmodulin Kinases and Ryanodine Receptors in Cardiac Failure, Arterial Smooth Muscle, Post-ischemic Arrhythmia, Similarities and Differences, and Pharmaceutical Targets
Larry H Bernstein, MD, FCAP, Justin Pearlman, MD, PhD, FACC and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/09/08/the-centrality-of-ca2-signaling-and-cytoskeleton-involving-calmodulin-kinases-and-ryanodine-receptors-in-cardiac-failure-arterial-smooth-muscle-post-ischemic-arrhythmia-similarities-and-differen/

Part VI: Calcium Cycling (ATPase Pump) in Cardiac Gene Therapy: Inhalable Gene Therapy for Pulmonary Arterial Hypertension and Percutaneous Intra-coronary Artery Infusion for Heart Failure: Contributions by Roger J. Hajjar, MD
Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/08/01/calcium-molecule-in-cardiac-gene-therapy-inhalable-gene-therapy-for-pulmonary-arterial-hypertension-and-percutaneous-intra-coronary-artery-infusion-for-heart-failure-contributions-by-roger-j-hajjar/

Part VII: Cardiac Contractility & Myocardium Performance: Ventricular Arrhythmias and Non-ischemic Heart Failure – Therapeutic Implications for Cardiomyocyte Ryanopathy (Calcium Release-related Contractile Dysfunction) and Catecholamine Responses
Justin Pearlman, MD, PhD, FACC, Larry H Bernstein, MD, FCAP and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/08/28/cardiac-contractility-myocardium-performance-ventricular-arrhythmias-and-non-ischemic-heart-failure-therapeutic-implications-for-cardiomyocyte-ryanopathy-calcium-release-related-contractile/

Part VIII: Disruption of Calcium Homeostasis: Cardiomyocytes and Vascular Smooth Muscle Cells: The Cardiac and Cardiovascular Calcium Signaling Mechanism
Justin Pearlman, MD, PhD, FACC, Larry H Bernstein, MD, FCAP and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/09/12/disruption-of-calcium-homeostasis-cardiomyocytes-and-vascular-smooth-muscle-cells-the-cardiac-and-cardiovascular-calcium-signaling-mechanism/

Part IX: Calcium-Channel Blockers, Calcium Release-related Contractile Dysfunction (Ryanopathy) and Calcium as Neurotransmitter Sensor
Justin Pearlman, MD, PhD, FACC, Larry H Bernstein, MD, FCAP and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/09/16/calcium-channel-blocker-calcium-as-neurotransmitter-sensor-and-calcium-release-related-contractile-dysfunction-ryanopathy/

Part X: Synaptotagmin functions as a Calcium Sensor: How Calcium Ions Regulate the fusion of vesicles with cell membranes during Neurotransmission
Larry H Bernstein, MD, FCAP and Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/09/10/synaptotagmin-functions-as-a-calcium-sensor-how-calcium-ions-regulate-the-fusion-of-vesicles-with-cell-membranes-during-neurotransmission/

Oxidized CaMKII Triggers Atrial Fibrillation

Running title: Purohit et al.; oxCaMKII and AF

Anil Purohit, Adam G. Rokita, Xiaoqun Guan, Biyi Chen, Olha M. Koval, Niels Voigt, Stefan Neef, Thomas Sowa, Zhan Gao, Elizabeth D. Luczak, Hrafnhildur Stefansdottir, Andrew C. Behunin, Na Li, Ramzi N. El Accaoui, Baoli Yang, Paari Dominic Swaminathan, Robert M. Weiss, Xander H. T. Wehrens, Long-Sheng Song, Dobromir Dobrev, Lars S. Maier and Mark E. Anderson

1Dept of Internal Medicine, Division of Cardiovascular Medicine and Cardiovascular Research Center, Carver College of Medicine, University of Iowa, Iowa City, IA; 2Institute of Pharmacology, Faculty of Medicine, University Duisburg-Essen, Essen, Germany, and Division of Experimental Cardiology, Medical Faculty Mannheim, University of Heidelberg, Mannheim, Germany; 3Cardiology and Pneumology, German Heart Center, University Hospital Goettingen, Goettingen, Germany; 4Dept of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, TX; 5Dept of Obstetrics and Gynecology; 6Dept of Molecular Physiology and Biophysics, University of Iowa, Iowa City, IA
Circulation Sept 12, 2013;

http://circ.ahajournals.org/content/early/2013/09/12/CIRCULATIONAHA.113.003313
http://circ.ahajournals.org/content/suppl/2013/09/12/CIRCULATIONAHA.113.003313.DC1.html

http://dx.doi.org/10.1161/CIRCULATIONAHA.113.003313

Journal Subject Codes: Basic science research:[132] Arrhythmias – basic studies, Etiology:[5] Arrhythmias, clinical electrophysiology, drugs

 Abstract

Background—Atrial fibrillation is a growing public health problem without adequate therapies. Angiotensin II (Ang II) and reactive oxygen species (ROS) are validated risk factors for atrial fibrillation (AF) in patients, but the molecular pathway(s) connecting ROS and AF is unknown. The Ca2+/calmodulin-dependent protein kinase II (CaMKII) has recently emerged as a ROS activated proarrhythmic signal, so we hypothesized that oxidized CaMKII􀄯(ox-CaMKII) could contribute to AF.  Methods and Results—We found ox-CaMKII was increased in atria from AF patients compared to patients in sinus rhythm and from mice infused with Ang II compared with saline. Ang II treated mice had increased susceptibility to AF compared to saline treated WT mice, establishing Ang II as a risk factor for AF in mice. Knock in mice lacking critical oxidation sites in CaMKIId (MM-VV) and mice with myocardial-restricted transgenic over-expression of methionine sulfoxide reductase A (MsrA TG), an enzyme that reduces ox-CaMKII, were resistant to AF induction after Ang II infusion. Conclusions—Our studies suggest that CaMKII is a molecular signal that couples increased ROS with AF and that therapeutic strategies to decrease ox-CaMKII may prevent or reduce AF.

Key words: atrial fibrillation, calcium/calmodulin-dependent protein kinase II, angiotensin II, reactive oxygen species, arrhythmia (mechanisms)

Introduction

Atrial fibrillation (AF) is the most common sustained  arrhythmia. AF produces lifestyle-limiting symptoms and increases the risk of stroke and death,1 but current therapies have limited efficacy. The renin-angiotensin-system is upregulated in cardiovascular disease and elevated Angiotensin II (Ang II) favors AF.2,3 Ang II activates NADPH oxidase, leading to increased ROS and fibrillating atria are marked by increased reactive oxygen species (ROS).4,5 We recently identified the multifunctional Ca2+ and calmodulin-dependent protein kinase II (CaMKII) as a ROS sensor6 and proarrhythmic signal.7 Oxidation of critical methionines (281/282) in the CaMKII regulatory domain lock CaMKII into a constitutively active, Ca2+ and calmodulinin-dependent conformation that is associated with cardiovascular disease.8 Based on this information, we asked if oxidized CaMKII (ox-CaMKII) could be a biomarker and proarrhythmic signal for connecting increased atrial ROS to AF. We found that ox-CaMKII was increased in atrial tissue from patients with AF compared to patients in sinus rhythm, and in atrial tissue from Ang II-infused, compared to saline-infused, mice. We used a validated mouse model of AF induction by rapid right atrial pacing9,10 and found that mice with prior Ang II infusion were at significantly higher risk of AF compared to vehicle-infused mice. We tested AF induction in Ang II and vehicle-infused mice with genetically engineered resistance to CaMKII oxidation by knock-in replacement of methionines 281/282 with valines in CaMKIId (MM-VV), the isoform associated with cardiovascular disease11-14 or by myocardial-targeted antioxidant therapy by transgenic over-expression of methionine sulfoxide reductase A (MsrA), an enzyme that reduces ox-CaMKII.15,16  Collectively, our results support a view that Ang II promotes AF induction by increasing ROS, ox-CaMKII, CaMKII activity, sarcoplasmic reticulum Ca2+ leak and delayed after-depolarizations (DADs). Our findings provide novel insights into a ROS and Ang II-dependent mechanism of AF by linking oxidative stress to dysfunctional intracellular Ca2+ signaling via ox-CaMKII and identify a potential new approach for treating AF by targeted antioxidant therapy.

Methods

Human samples and immunodetection of ox-CaMKII.

The human samples were provided by the Georg-August-University Goettingen and the University of Heidelberg after approval by the local ethics committee of the Georg-August-University Göttingen and the Medical Faculty Mannheim, University of Heidelberg (#2011-216N-MA). 

Right atrial appendage tissue samples were obtained from patients undergoing thoracotomy with sinus rhythm or with AF (Table 1) as published previously.17 For immunostaining experiments a total of 9 samples were studied including 5 patients with sinus rhythm and 4 patients with AF ( Table 1A). For immunob lotting a total of 51 samples were studied including 25 patients with SR and 26 patients with AF (Table 1B). The pat ei nt charts were reviewed by the authors to obtain relevant clinical information.

Mouse Models and Experimental Methods

All mice used in the study were available to us in C57Bl6 background. All experiments were performed in male mice 8-12 weeks of age. In total we studied 262 mice. Numbers for each experimental group are provided in the figures or figure legends. See Supplemental Material for detailed methods.

Statistics

Data are presented as mean ± SEM. P values were assessed with a Student’s t-test (2-tailed), ANOVA or two-way ANOVA, as appropriate, for continuous data. The effect of Ang II compared to saline on ox-CaMKII, CaMKII, and ox-CaMKII/CaMKII ratio was tested within each mouse genotype (strain) and compared among the four genotypes using the two-way analysis of variance (ANOVA). The factors that were tested in the ANOVA model were genotype (WT, MM-VV, p47-/- and MsrA TG), treatment (Ang II versus saline), and genotype treatment interaction effect. A significant genotype treatment interaction (*) indicated that the effect of Ang II (versus saline) differed significantly among the strains. Post hoc comparisons after ANOVA were performed using the Bonferroni test. Discrete variables were analyzed by Fisher’s exact test.

Results

Oxidized CaMKII is increased in AF

Patients with AF have increased atrial CaMKII activity18,19 and high circulating levels of serum markers for oxidative stre ss. 4, 5 We first obtained right atrial tissue from patients undergoing cardiac surgery (Table 1) and measured ox-CaMKII using a validated antiserum against oxidized Met 281/282 in the CaMKII regulatory domains.6 These pilot immunofluorescence studies on atrial tissue samples made available upon consent by patients with AF or normal sinus rhythm (Table 1A) showed significantly (p<0.05) higher (~2.5 fold) ox-CaMKII levels in patients with AF (Figure 1A and B). Based on these initial findings, we measured ox-CaMKII in atrial tissue from a larger cohort of patients (Table 1B; for complete gels see supplementary Figure 1) in sinus rhythm (N = 25) or AF (N = 26) using Western blots, and confirmed that AF patients have significantly elevated expression of ox-CaMKII, while there was no difference in total CaMKII (Figure 1C-F). The patient characteristics in the two groups (Table 1) were similar in terms of age, presence of hypertension, diabetes and left ventricular ejection fraction, recognized risk factors for AF.20 The subgroup of AF patients that were not treated with angiotensin converting enzyme inhibitor (ACE-i) or angiotensin receptor blockers (ARB) showed the highest levels of ox-CaMKII and total CaMKII (Supplementary Figure 1A and B). Taken together, these findings showed a positive association between AF and increased expression of atrial ox-CaMKII and a loss of this association in AF patients treated with ACE-i or ARBs.

Ang II treatment enhances AF susceptibility

  

To test the hypothesis that ox-CaMKII contributes to AF we developed a mouse model of AF by infusing wild type (WT) mice with Ang II (2000 ng/kg/min) or an equal volume of normal saline via osmotic mini-pumps for three weeks. We previously established that this dose of Ang II caused a significant increase in atrial ox-CaMKII7 and resulted in serum Ang II levels similar to those measured in heart failure patients.21
In order to test if Ang II treatment can promote AF we performed burst pacing in the right atrium of anesthetized mice, using an established method ( Figure 2A-C). 10 Mice treated wit Ang II showed significantly higher AF induction rates compared to saline treated mice (64% [9/14] versus 18% [2/14], p=0.018 Fisher’s exact test) (Figure 2D). Ang II is known to contribute to hypertension, left ventricular hypertrophy and heart failure, all established clinical risk factors for AF.20 Therefore, we measured blood pressure (BP) by tail-cuff and assessed left ventricular size and systolic function by echocardiography. As expected, Ang II treatment significantly increased systolic BP (Figure 2E; p<0.01) and left ventricular mass (Figure 2F; p<0.001). Ang II treated mice maintained a normal left ventricular ejection fraction, similar to saline-infused control mice (Figure 2G). These data showed that Ang II infusion increased the susceptibility of mice to AF induction by rapid right atrial pacing and established a framework for us to test the hypothesized role of ox-CaMKII in promoting AF. ox-CaMKII is critical for AF.
In order to test if ox-CaMKII was required for AF induction in our model we used oxidation resistant knock in MM-VV mice (Supplementary Figure 2).22 CaMKII with the MM-VV mutation is resistant to oxidative activation but retains normal Ca2+ and calmodulin dependent activation and is capable of transitioning into a Ca2+ and calmodulin independent enzyme after threonine 287 autophosphorylation.6 The MM-VV mice were significantly resistant to AF induction after Ang II infusion, compared to WT controls (Figure 3A), suggesting that ox-CaMKII is required for increased AF susceptibility in Ang II infused mice. WT mice treated with Ang II showed significantly higher (~2.7 fold; 95% confidential interval, CI: 1.4, 5.1) ) levels of mice. When indexed to total CaMKII levels (Supplementary Figure 3A and B) this increase in ox-CaMKII was much higher (~14. 2 fold; 95% confidential interval, CI: 1.4, 5.1)  in Ang II treated WT mice (figure 4C).  The residual increase in ox–CaMKII in the -MM-VV mice likely results from expression of atrial ox-CaMKII compared to saline treated mice. As expected, Ang II infusion increased ox-CaMKII less in -MM-VV (~2.1 fold; 95% CI: 1.1, 4.0) than in control WT.  ox-CaMKII was much higher (~14.2 fold; 95% CI: 5.9, 34.5) in Ang II treated WT mice.
CaMKIILI, a myocardial CaMKII isoform not affected by the MM-VV mutation.23 However, despite the greater increase in ox-CaMKII in WT compared to MM-VV mice, Ang II-related ROS production was increased in both WT and MM-VV mice to a similar degree (Supplementary Figure 4). Interestingly, Ang II treated WT mice showed a significant decrease in total CaMKII levels (Supplementary Figure 3A and B) suggesting feedback inhibition of total CaMKII expression.
Atrial lysates from MM-VV mice showed significantly less Ca2+ and calmodulin-independent activity after Ang II treatment, but retained WT level CaMKII activity increases in response to isoproterenol (Supplementary Figure 2A). At 8 weeks MM-VV mice had body weight (Supplementary Figure 2B) and BP (Figure 3B) that were similar to WT mice, suggesting CaMKIIį methionine 281/282 oxidation did not affect basal BP or developmentally appropriate growth. CaMKII is known to regulate the chronotropic response to stress and mice with CaMKII inhibition have a smaller increase in heart rate with isoproterenol treatment compared to controls.24 Isolated Langendorff-perfused hearts from WT and MM-VV mice had similar resting heart rates (Supplementary Figure 2C) and comparable heart rate increases after isoproterenol treatment (Supplementary Figure 2D), suggesting that CaMKII dependent physiological heart rate increases do not require CaMKIIį methionine oxidation. L-type Ca2+ currents were similar in MM-VV and WT mice, and L-type Ca2+ current facilitation, a CaMKII-dependent phenotype, was also preserved in MM-VV mice.25,26 KN-93, a small molecule CaMKII inhibitor,27 significantly reduced facilitation in WT and -MM-VV mice (Supplementary Figure 5). MM-VV mice and WT controls showed similar increases in systolic BP (Figure 3B) and heart weight (Figure 3C) or left ventricular mass estimated by echocardiography after Ang II infusion ( Supplementary Figure 6), suggesting that -ox-CaMK IIį is dispensable for hypertensive and myocardial hypertrophic actions of Ang II. Taken together, these findings indicate loss of methionines 281/282 in CaMKIIį selectively reduce the pro-arrhythmic actions of Ang II in a pacing-induced model of AF.

NADPH oxidase and MsrA regulate ox-CaMKII and AF susceptibility.

  •  Ang II increases intracellular ROS in myocardium by activating NADPH oxidase and
  • p47-/-mice28, lacking functional NADPH oxidase, are resistant to Ang II dependent increases in ROS and ox-CaMKII.6
  • Atrial lysates from Ang II treated p47-/- mice did not show an increase in ox-CaMKII (Figure 4), and
  • the p47-/- mice were also resistant to Ang II-mediated increases in AF
However, there were similar increases in BP (Figure 3B) effects of Ang II. This was observed with MsrA TG and WT mice (Figure 3A), showing similar increases in BP (Figure 3B), overall heart weight (Figure 3C) and estimated left ventricular mass (Supplementary Figure 6) after Ang II treatment compared to WT controls. ox-CaMKII is reduced by MsrA15 and transgenic mice with myocardial-delimited MsrA overexpression (MsrA TG) have increased atrial MsrA protein (Supplementary Figure 3C) and
  • are resistant to ROS induced myocardial injury.16

We found that Ang II treated MsrA TG mice showed decreased AF induction compared to Ang II-treated WT mice (Figure 3A) and

  • had similar atrial ox-CaMKII expression compared to saline treated controls (Figure 4).
  • Ang II induced increases in ROS production seen in WT atria were absent in atria from MsrA TG mice (Supplementary Figure 4),
suggesting that MsrA sensitive targets represent an important component of Ang II mediated atrial oxidation. The protection from AF in MsrA TG mice appeared to be independent of pressor effects that are critical for the proarrhythmic actions. Taken together, these findings suggest that
  • NADPH oxidase dependent ROS and elevated ox-CaMKII drive Ang II  -pacing-induced AF and that
  • targeted antioxidant therapy, by MsrA over-expression, can reduce or prevent AF in Ang -II-infused mice.

Ang II increases Ca2+ sparks and triggered action potentials

CaMKII contributes to increased sarcoplasmic reticulum Ca2+ leak in mice with a RyR2 mutation modeled after a human arrhythmia syndrome, catecholaminergic polymorphic ventricular tachycardia,9 in a goat model of AF and in atrial myocytes isolated from patients with AF.18,29 Atrial myocytes from patients with AF
  • show increased CaMKII activity and increased CaMKII-dependent ryanodine receptor phosphorylation at serine 2814.29
  •  CaMKII inhibition with KN-93 reduced the open probability of single RyR2 channels and
  • prevented the increased frequency of sarcoplasmic reticulum Ca2+ sparks in atrial myocardium biopsied from AF patients.18,29
Based on this knowledge, we asked if increased RyR2 Ca2+ leak also contributed to the mechanism of AF in WT Ang II infused mice and measured diastolic Ca2+ sparks, a marker of RyR2 Ca2+ leak.30
  • Atrial myocytes from Ang II treated WT mice showed a significant (p<0.05) increase in spontaneous Ca2+ sparks compared to atrial myocytes from saline treated control mice (Figure 5A and B).
Other Ca2+ spark parameters and sarcoplasmic reticulum Ca2+ content were not different between the saline and Ang II treated WT mice (Supplementary Figure 7). In contrast to findings in WT mice,
  • the atrial myocytes isolated from Ang II treated MM-VV mice did not show an increase in Ca2+ sparks compared to saline treated MM-VV mice (Figure 5A and B).
  • A significantly greater proportion of atrial myocytes isolated from Ang II treated WT mice showed DADs, compared to atrial myocytes from saline treated mice (Figure 5C and D, p=0.03; Fisher’s exact test).
  • atrial myocytes from Ang II infused MM-VV mice did not show a significant increase in DADs compared to the atrial myocytes from saline treated MM-VV mice.

We interpret these data to suggest that the proarrhythmic effects of Ang I I infusion depend upon an increase in ox–CaMKII, sarcoplasmic reticulum Ca2+ leak and DADs.

Mice with CaMKII-resistant RyR2 are protected from AF after Ang II infusion

Enhanced CaMKII-mediated phosphorylation of serine 2814 on RyR2 is associated with an increased susceptibility to acquired arrhythmias, including AF.31 Based on our findings

  • that atrial myocytes from Ang II infused WT mice developed more Ca2+ sparks than atrial myocytes from saline-infused mice,

we hypothesized that the proarrhythmic actions of ox-CaMKII require access to RyR2 serine 2814. We tested this hypothesis by treating mutant S2814A knock-in mice (lacking serine 2814)9 with Ang II or saline and performing right atrial burst pacing.

  • The S2814A mice were highly resistant to Ang II mediated AF (Figure 6A). Similarly,
  • AC3-I mice with transgenic myocardial expression of a CaMKII inhibitory peptide32 were also resistant to the proarrhythmic effects of Ang II infusion on pacing-induced AF (Figure 6A). S2814A,

AC3-I and WT mice, all developed similar BP increases (Figure 6B) and cardiac hypertrophy (Figure 6C) in response to Ang II, indicating that

  • these mice were not resistant to the hemodynamic effects of Ang II, but were nevertheless protected from AF.

 Discussion

AF usually develops in patients with underlying structural heart disease, such as left ventricular hypertrophy, coronary artery disease, valve disease and congestive heart failure.20 Elevated ROS is a common feature of these conditions.33 The dose of Ang II used in our model produces a fourfold increase in plasma Ang II compared to saline controls,7 similar to increases in Ang II observed in heart failure patients evidence of elevated ROS in structural heart disease, clinical trials with antioxidants have generally been unsatisfactory.34-36 One potential obstacle to developing effective antioxidant therapies is lack of detailed understanding of molecul ra pathways that are affected by ROS. The renin-angiotensin-system is one of the best understood pathways that contributes to ROS production in AF patients.37 In the current study, we created a model of AF by infusing mice with Ang II for three weeks and assembled a cohort of genetically altered mice to rigorously test a novel molecular pathway that links oxidative stress to AF (Figure 7). Our current study provides strong evidence that CaMKII is a critical ROS sensor for transducing increased ROS into enhanced AF susceptibility in mice and suggests that atrial ox-CaMKII could contribute to AF in patients.

CaMKII and increased ROS are now widely recognized to contribute to cardiac arrhythmias.8,38,39 Recent studies suggest that patients with persistent AF have elevated markers of oxidative stress in serum4 and depleted levels of atrial glutathione.40 Under increased oxidative stress CaMKII is activated by oxidation of methionines (M281/282),6 which lock it into a constitutively active conformation, suggesting a possible role for ox-CaMKII as a ROS activated proarrhythmic signal in AF.39 Our laboratory recently demonstrated that

  • ox-CaMKII plays a major role in sinus node dysfunction,7,22
  • adverse post-myocardial infarct remodeling6 and
  • cardiac rupture16.

In the current study, we investigated the role of ox-CaMKII in AF. Human atria (Figure 1) and Ang II treated WT mouse atria showed significantly elevated ox-CaMKII (Figure 4).

  • Atrial myocytes from Ang II treated WT mice had a higher frequency of spontaneous Ca2+ sparks and DADs compared to controls (Figure 5).

Based on these findings we hypothesized that oxidation of methionines 281/282 on CaMKII į causes diastolic sarcoplasmic reticulum Ca2+ leak and DADs, both cellular AF triggers. However, resistant to oxidative activation,22

  • Ang II, the myocardial CaMKII a recently developed knock-in mouse (MM-VV) where CaMKII isoform implicated in myocardial disease,1,2 13 treatment
  • did not increase Ca2+ and calmodulin independent CaMKII activity (Supplementary Figure 2A), Ca2+ sparks (Figure 5A and B), DADs (Figure 5C and D) or enhance AF susceptibility in MM-VV mice (Figure 3A).

It is important to note that the MM-VV mutant form of CaMKIIį selectively ablates the response to oxidation while retaining other aspects of CaMKII molecular physiology, such as

  • activation by Ca2+ and calmodulin and
  • constitutive activation by threonine 287 autophosphorylation.6

Thus, the residual AF observed in Ang II infused MM-VV mice could be a result of non-oxidation-dependent mechanisms for CaMKIIį activation in our model. We found that atrial tissue from AF patients treated with ACE-i or ARBs did not show elevated ox-CaMKII, suggesting that Ang II stimulation oxidizes CaMKII in human atria and that ox-CaMKII independent pathways are operative in AF patients. AF in patients is more complex than AF in our Ang II infused mice. In particular, patients present with variable chronicity, tissue and structural changes. In contrast the triggers for our mice are uniform (i.e. Ang II infusion and rapid right atrial pacing) and result in a similar, modest degree of hypertrophy. We interpret the data showing that an increase in ox-CaMKII in AF patients is reduced or eliminated by clinical antagonist drugs that reduce Ang II signaling to validate our findings in mice that Ang II increases ox-CaMKII. However, we suppose that the presence of AF in patients on ACE-i or ARBs means that other pathways also result in AF. Our sample is not powered to ask if AF resistance to Ang II antagonist drugs represents later stage disease, but this is our hypothesis. Furthermore, CaMKII can be activated independently of oxidation, although oxidation appears to be the primay r pathway for activating CaMKII during Ang II infusion. Thus, it is unknown if CaMKII is also important for AF progression in the group of patients treated by Ang II antagonist drugs who exhibit normal levels of ox -CaMKII.

Although we did not see higher total CaMKII in AF patients (as compared with patients in sinus rhythm), the sub-group of AF patients who were not treated with ACE-i or ARBs did show significantly elevated CaMKII levels, supporting prior studies that reported elevated CaMKII activity in AF18,19.  In contrast to the situation in patients, total CaMKII expression was reduced in mice after sub-acute Ang II infusion. While the mechanism(s) for the variable response of CaMKII expression in mice and patients is unclear, the change in expression in mice and in humans in response to manipulation of the Ang II pathway supports the idea that CaMKII is a fundamental component of Ang II signaling. The relatively small number of patient samples is not powered for analysis of AF subtypes, but human AF may transition from paroxysmal to persistent and permanent (chronic) forms.41 In contrast, our mouse model is simpler because it is triggered by a single upstream event (i.e. Ang II infusion) and elicited in a highly controlled environment by rapid atrial pacing. The resistance of MM-VV mice to AF provides new evidence that oxidative activation of CaMKII delta (d) is important for initiation of AF, while the finding that ox-CaMKII is elevated in atrial tissue from AF patients and particularly in AF patients naive to Ang II antagonist therapies suggests this pathway may also participate in human AF.

Thus, our findings in MM-VV mice provide strong, mechanistic evidence that ox-CaMKII plays a critical role in proarrhythmic responses to Ang II. Our studies showed that mice deficient in NADPH oxidase (p47-/-) and mice expressing increased MsrA are also resistant to AF (Figure 3A), suggesting that

  • selectively targeted antioxidant therapies could be effective in preventing or reducing AF.
  • Half of patients enrolled in the Mode Selection Trial (MOST) with sinus node dysfunction had a history of AF48,

but a clear mechanistic link between increased risk of AF and sinus node dysfunction is unknown. In recent studies we showed that Ang II and diabetes-induced CaMKII oxidation caused sinus node dysfunction by increased pacemaker cell death and fibrosis,7 while MM-VV mice are resistant to sinus node dysfunction evoked by hyperglycemia.22 Here we provide evidence that

  • ox-CaMKII increases susceptibility for AF via increased diastolic sarcoplasmic reticulum Ca2+ release, showing that
  • the proarrhythmic actions of ox-CaMKII may occur in cardiomyocytes by increasing sarcoplasmic reticulum Ca2+ leak or by enhanced cell death.

Our findings suggest that the clinical association between sinus node dysfunction and AF might have a mechanistic basis because sinus node dysfunction and AF are downstream consequences of elevated ox-CaMKII.

Selected References

1. Benjamin EJ, Wolf PA, D’Agostino RB, Silbershatz H, Kannel WB, Levy D. Impact of atrial fibrillation on the risk of death: the Framingham Heart Study. Circulation. 1998;98:946-952.
2. Khatib R, Joseph P, Briel M, Yusuf S, Healey J. Blockade of the renin-angiotensinaldosterone system (RAAS) for primary prevention of non-valvular atrial fibrillation: A systematic review and meta analysis of randomized controlled trials. Int J Cardiol. 2013;165:17-24.

4. Shimano M, Shibata R, Inden Y, Yoshida N, Uchikawa T, Tsuji Y, Murohara T. Reactive oxidative metabolites are associated with atrial conduction disturbance in patients with atrial
fibrillation. Heart Rhythm. 2009;6:935-940.
5. Neuman RB, Bloom HL, Shukrullah I, Darrow LA, Kleinbaum D, Jones DP, Dudley SC. Oxidative stress markers are associated with persistent atrial fibrillation. Clin Chem.
2007;53:1652-1657.
 6. Erickson JR, Joiner M-LA, Guan X, Kutschke W, Yang J, Oddis CV, Bartlett RK, Lowe JS, O’Donnell SE, Aykin-Burns N, Zimmerman MC, Zimmerman K, Ham A-JL, Weiss RM, Spitz DR, Shea MA, Colbran RJ, Mohler PJ, Anderson ME. A dynamic pathway for calciumin-dependent activation of CaMKII by methionine oxidation. Cell. 2008;133:462-474.

7. Swaminathan PD, Purohit A, Soni S, Voigt N, Singh MV, Glukhov AV, Gao Z, He BJ, Luczak ED, Joiner M-LA, Kutschke W, Yang J, Donahue JK, Weiss RM, Grumbach IM, Ogawa M, Chen P-S, Efimov I, Dobrev D, Mohler PJ, Hund TJ, Anderson ME. Oxidized CaMKII
causes cardiac sinus node dysfunction in mice. J Clin Invest. 2011;121:3277-3288.

8. Erickson JR, He BJ, Grumbach IM, Anderson ME. CaMKII in the cardiovascular system: sensing redox states. Physiol Rev. 2011;91:889-915.
9. Chelu MG, Sarma S, Sood S, Wang S, van Oort RJ, Skapura DG, Li N, Santonastasi M, Müller FU, Schmitz W, Schotten U, Anderson ME, Valderrábano M, Dobrev D, Wehrens XHT. Calmodulin kinase II-mediated sarcoplasmic reticulum Ca2+ leak promotes atrial fibrillation in mice. J Clin Invest. 2009;119:1940-1951.
15. Moskovitz J, Bar-Noy S, Williams WM, Requena J, Berlett BS, Stadtman ER. Methionine sulfoxide reductase (MsrA) is a regulator of antioxidant defense and lifespan in mammals. Proc Natl Acad Sci USA. 2001;98:12920-12925.
16. He BJ, Joiner M-LA, Singh MV, Luczak ED, Swaminathan PD, Koval OM, Kutschke W, Allamargot C, Yang J, Guan X, Zimmerman K, Grumbach IM, Weiss RM, Spitz DR, Sigmund CD, Blankesteijn WM, Heymans S, Mohler PJ, Anderson ME. Oxidation of CaMKII determines the cardiotoxic effects of aldosterone. Nat Med. 2011;17:1610-1618.
18. Neef S, Dybkova N, Sossalla S, Ort KR, Fluschnik N, Neumann K, Seipelt R, Schöndube FA, Hasenfuss G, Maier LS. CaMKII-dependent diastolic SR Ca2+ leak and elevated diastolic Ca2+ levels in right atrial myocardium of patients with atrial fibrillation. Circ Res. 2010;106:1134-1144.

19. Tessier S, Karczewski P, Krause EG, Pansard Y, Acar C, Lang-Lazdunski M, Mercadier JJ, Hatem SN. Regulation of the transient outward K+ current by Ca2+/calmodulin-dependent protein kinases II in human atrial myocytes. Circ Res. 1999;85:810-819.
22. Luo M, Guan X, Luczak ED, Lang D, Kutschke W, Gao Z, Yang J, Glynn P, Sossalla S, Swaminathan PD, Weiss RM, Yang B, Rokita AG, Maier LS, Efimov IR, Hund TJ, Anderson ME. Diabetes increases mortality after myocardial infarction by oxidizing CaMKII. J Clin Invest. 2013;123:1262-1274.
24. Wu Y, Gao Z, Chen B, Koval OM, Singh MV, Guan X, Hund TJ, Kutschke W, Sarma S, Grumbach IM, Wehrens XHT, Mohler PJ, Song L-S, Anderson ME. Calmodulin kinase II is required for fight or flight sinoatrial node physiology. Proc Natl Acad Sci USA. 2009;106:5972-5977.
25. Dzhura I, Wu Y, Colbran RJ, Balser JR, Anderson ME. Calmodulin kinase determines calcium-dependent facilitation of L-type calcium channels. Nat Cell Biol. 2000;2:173-177.
26. Koval OM, Guan X, Wu Y, Joiner ML, Gao Z, Chen B, Grumbach IM, Luczak ED, Colbran RJ, Song LS, Hund TJ, Mohler PJ, Anderson ME. CaV1.2 -subunit coordinates CaMKII triggered cardiomyocyte death and afterdepolarizations. Proc Natl Acad Sci USA. 2010;107:4996–5000.
44. Anderson ME. Multiple downstream proarrhythmic targets for calmodulin kinase II: moving beyond an ion channel-centric focus. Cardiovasc Res. 2007;73:657-666.

46. Chang HY, Lin YJ, Lo LW, Chang SL, Hu YF, Li CH, Chao TF, Yin WH, Chen SA. Sinus node dysfunction in atrial fibrillation patients: the evidence of regional atrial substrate remodelling. Europace. 2013;15:205-211.
47. Lee JMS, Kalman JM. Sinus node dysfunction and atrial fibrillation: two sides of the same coin? Europace. 2013;15:161-162.

Table 1. Summary of patient characteristics.
A. Patient characteristics for immunofluorescence studies in Figure 1A and B. B. Patient characteristics for immunoblotting experiments in Figure 1C-F.
http://dx.doi.org/10.1161/CIRCULATIONAHA.113.003313

Figures and/or Legends

The source of all the figures is from the circulation article – including supplementary.  Obtaining the images and presenting them in a cropped form was difficult.

http://circ.ahajournals.org/content/early/2013/09/12/CIRCULATIONAHA.113.003313
http://circ.ahajournals.org/content/suppl/2013/09/12/CIRCULATIONAHA.113.003313.DC1.html

http://dx.doi.org/10.1161/CIRCULATIONAHA.113.003313

Figure 1. ox-CaMKII is increased in atria from patients with Atrial Fibrillation (AF).
A. Representative immunofluorescence images using antiserum against ox-CaMKII in fixed sections of right atrial tissue from patients with sinus rhythm (SR) or AF. B. Image  quantification showing significantly higher ox-CaMKII in patients with AF compared to SR (*p<0.05, Student’s t-test). C. Representative immunoblots with ox-CaMKII antiserum in right atrial tissue homogenates from patients in SR or AF. D. Quantification of immunoblots showing significantly higher ox-CaMKII expression in patients with AF compared to SR (*p<0.05, Student’s t-test). The % value indicates the mean ox-CaMKII/GAPDH ratio as normalized to the mean ox-CaMKII/GAPDH ratio in the SR group. E. CaMKII antiserum in right atrial tissue homogenates from patients in SR or AF. F. Quantification of immunoblots showing similar total CaMKII expression in patients with AF and SR (p=0.3, Student’s t-tes )t . The % value indicates the mean CaMKII/GAPDH ratio as normalized to the me na CaMKII/GAPDH ratio in the SR group. The numerals shown in the bars indicate the sample size in each group, here and in subsequent figures.

Figure 2. Ang II treatment increases AF inducibility in WT mice.
A. Representative atrial (A-EGM) and ventricular (V-EGM) intracardiac electrograms and lead II surface ECG immediately after burst pacing show AF or SR in WT mice treated with Ang II or saline for 3 weeks. B. Contrasting R-R interval variability in AF and SR (C). Blue bars indicate calculated values from lead II ECGs shown in panel A. D. Higher AF inducibility in the Ang II treatment group (*p<0.05, Fisher’s exact test). E. Increase in systolic blood pressure (sBP) in WT mice after 3 

Figure 3. CaMKII oxidation is critical to Ang II mediated AF.
A. MM-VV, p47-/- and MsrA TG mice were resistant to Ang II mediated AF (*p<0.05 versus Ang II treated MM-VV, p47-/- and MsrA TG mice, Fisher’s exact test). B. All mice in panel A (WT, MM-VV, p47-/- and MsrA TG) showed a pressor response to Ang II. C. Ang II treatment induced cardiac hypertrophy as assessed by heart weight normalized to body weight (all comparisons versus saline controls from each genotype after 3 weeks of Ang II treatment(p< 0.05) (**p<0.01, Student’s t-test). The numerals shown in the graph indicate the number of mice in each group. F. Significantly higher echocardiographically estimated left ventricular (LV) mass in Ang II treated mice compared to saline controls (***p<0.001, Student’s t-test). G. Similar LV ejection fraction (LVEF) in Ang II and saline treated mice.  (** p<0.01 and ***p<0.001, Student’s t-test).

Figure 4. – ox-CaMKII in atria after Ang II or saline treatment
A. Atrial lys ate immunoblots from WT, MM-VV, p47 -/- and MsrA TG mice treated with Ang II or saline for 3 weeks and probed with an antiserum for ox-CaMKII. For quantification, ox-CaMKII bands were normalized to the total protein loading as assessed with Coomassie staining of the membrane. B. Increase in ox-CaMKII with Ang II treatment expressed as relative to the saline treated group. From each genotype 4 saline treated mice were used as controls. *p<0.05, for WT Ang II versus WT saline (*), in all other genotypes Ang II versus saline p>0.05; in addition, p=0.02 for WT Ang II versus MsrA TG Ang II and p=0.05 for MM-VV Ang II versus MsrA TG Ang II. C. Fold change in ox-CaMKII (over total CaMKII) in Ang II as relative to saline treated mice of the same genotype. From each genotype 4 saline treated mice were used as controls. ***p<0.001 versus WT saline, *p<0.05 versus MM-VV saline, #p<0.05 versus MsrA TG saline. WT Ang II versus p47-/- Ang II, P = 0.001, WT Ang II versus MsrA TG Ang II, P<0.0001, MM-VV Ang II versus MsrA TG Ang II, P=0.001. Data were analyzed using two-way ANOVA (for treatment and genotype) with Bonferroni post-hoc comparisons.

Figure 5. Ang II promotes Ca2+ sparks and DADs.
A. Representative examples of Ca2+ sparks in atrial myocytes from Ang II and saline treated WT and MM-VV mice. B. Summary of Ca2+ spark frequency data in atrial myocytes from Ang II treated mice compared to saline treated mice (*p<0.05 versus saline; Student’s t-test); WT saline (N=23 cells from 5 mice), WT Ang II (N=30 cells from 4 mice), MM-VV saline (N=36 cells from 4 mice) and MM-VV Ang II (N=28 cells from 4 mice). C. Examples of stimulated action potentials and a spontaneous, DAD triggered action potential. D. Higher incidence of DADs in atrial myocytes from Ang II treated WT mice ( *p<0.05 versus saline, Fisher’s exact test) but not in Ang II treated MM-VV mice compared to saline controls. Numerals show cells with DADs/total cells studied for each group.

Figure 6. CaMKII activation and RyR2 serine 2814 are required for AF in Ang II infused mice.
A. AC3-I and S2814A mice were treated with Ang II for 3 weeks and then burst paced to induce AF. AC3-I and S2814A mice were resistant to Ang II mediated AF promotion compared to WT Ang II treated mice (*p<0.05 versus all, Fisher’s Exact test, N=number of mice tested in each group). B. AC3-I and S2814A mice show similar systolic blood pressure (sBP) elevation after treatment with Ang II. Final sBP measurements were performed on three consecutive days prior to AF induction as shown in panel A. The numerals in the graph indicate the number of mice in each group. C. Ang II treatment causes similar cardiac hypertrophy in AC3-I and S2814A mice compared to saline controls (***p<0.001 versus AC3-I saline and **p=0.01 versus S2814A saline).

Figure 7. Schematic to illustrate the proposed mechanism of AF in Ang II infused mice.
Ang II binding activates NADPH oxidase (NOX) to increase reactive oxygen species (ROS), leading to oxidation of methionines 281/282 in CaMKII (ox-CaMKII). Elevated ox-CaMKII phosphorylates serine 2814 on RyR2, causing enhanced diastolic Ca2+ leak that promotes AF triggering DADs. Genetically modified mice were used to test key steps of the proposed pathway.

Additional Comments

This paper might be considered and compared with other papers in this series.

I Contributions to cardiomyocyte interactions and signaling

Author and Curator: Larry H Bernstein, MD, FCAP and  Curator: Aviva Lev-Ari, PhD, RN
https://pharmaceuticalintelligence.com/2013/10/21/contributions-to-cardiomyocyte-interactions-and-signaling/
This is a review of left ventricular cardiac hypertrophy and interaction with heparin-binding EGF,  based on work in the laboratory of Richard Lee, at Brigham and Women Hospital, Harvard Medical School, and MIT, titled…

Cardiomyocyte hypertrophy and degradation of connexin43 through spatially restricted autocrine/paracrine heparin-binding EGF

J Yoshioka, RN Prince, H Huang, SB Perkins, FU Cruz, C MacGillivray, DA Lauffenburger, and RT Lee *Cardiovascular Division, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA; and Biological Engineering Division, MIT, Cambridge, MA
PNAS 2005; 302(30):10622-10627.  http://pnas.org/cgi/doi/10.1073/pnas.0501198102

Growth factor signaling can affect tissue remodeling through autocrine/paracrine mechanisms. Recent evidence indicates that EGF receptor transactivation by heparin-binding EGF (HB-EGF) contributes to hypertrophic signaling in cardiomyocytes. Here, we show that HB-EGF operates in a spatially restricted circuit in the extracellular space within the myocardium, revealing the critical nature of the local microenvironment in intercellular signaling. This highly localized microenvironment of HB-EGF signaling demonstrated with 3D morphology, consistent with predictions from a computational model of EGF signaling. HB-EGF secretion by a given cardiomyocyte in mouse left ventricles led to cellular hypertrophy and reduced expression of connexin43 in the overexpressing cell and in immediately adjacent cells but not in cells farther away.

!!.  Ca2+/calmodulin δ Dependent Protein Kinase Modulates Cardiac Ryanodine Receptor Phosphorylation and Sarcoplasmic Reticulum Ca2+ Leak in Heart Failure.

Xun Ai, JW Curran, TR Shannon, DM Bers and SM Pogwizd.   
Circ Res. 2005;97:1314-1322  http://dx.doi.org/10.1161/01.RES.0000194329.41863.89
http://circres.ahajournals.org/content/97/12/1314

This contribution is unique in establishing a relationship between Ca2+ sparks in abnormal release from sarcoplasmic reticulum via the ryanodine receptor (RyR2) in contractile dysfunction and arrhythmogenesis in heart failure.  This is based on decreased transient amplitude and SR Ca2+ load with increased Na+/Ca++ exchange, and in nonischemic heart failure in a rabbit model.  In this case – with HF, expression of RyR2 and FK-506 binding protein 12.6 (FKBP12.6) were reduced, whereas inositol trisphosphate receptor (type 2) and Ca/calmodulin–dependent protein kinase II (CaMKII) expression were increased 50% to 100%.  In this study, the arrhythmogenesis appears to be ventricular.

Contractile dysfunction in HF is caused by diminished sarcoplasmic reticulum (SR) Ca load that could arise from enhanced activity of Na/Ca exchange (NCX), reduced SR Ca ATPase (SERCA) function, and increased diastolic SR Ca leak via ryanodine receptors (RyR), all of which we have demon¬strated to occur in our arrhythmogenic rabbit model of nonis-chemic HF. HF is also associated with a nearly 50% incidence of sudden cardiac death from ventricular tachycardia (VT) that degenerates to ventricular fibrillation (VF). In 3D cardiac mapping studies in our HF rabbit model, we showed that spontaneously occurring VT initiates by nonreentrant mechanisms associated with delayed afterdepolarizations. These arise from spontaneous SR Ca release that activates a transient inward current (Iti) carried primarily by NCX.2 Thus abnormal SR Ca release via RyR may contribute to both contractile dysfunction and arrhythmogenesis.

Abnormal release of Ca from sarcoplasmic reticulum (SR) via the cardiac ryanodine receptor (RyR2) may contribute to contractile dysfunction and arrhythmogenesis in heart failure (HF). We previously demonstrated decreased Ca transient amplitude and SR Ca load associated with increased Na/Ca exchanger expression and enhanced diastolic SR Ca leak in an arrhythmogenic rabbit model of nonischemic HF. Here we assessed expression and phosphorylation status of key Ca handling proteins and measured SR Ca leak in control and HF rabbit myocytes. With HF, expression of RyR2 and FK-506 binding protein 12.6 (FKBP12.6) were reduced, whereas inositol trisphosphate receptor (type 2) and Ca/calmodulin–dependent protein kinase II (CaMKII) expression were increased 50% to 100%. The RyR2 complex included more CaMKII (which was more activated) but less calmodulin, FKBP12.6, and phosphatases 1 and 2A. The RyR2 was more highly phosphorylated by both protein kinase A (PKA) and CaMKII. Total phospholamban phosphorylation was unaltered, although it was reduced at the PKA site and increased at the CaMKII site. SR Ca leak in intact HF myocytes (which is higher than in control) was reduced by inhibition of CaMKII but was unaltered by PKA inhibition. CaMKII inhibition also increased SR Ca content in HF myocytes. Our results suggest that CaMKII-dependent phosphorylation of RyR2 is involved in enhanced SR diastolic Ca leak and reduced SR Ca load in HF, and may thus contribute to arrhythmias and contractile dysfunction in HF. (Circ Res. 2005;97:1314-1322.)

Key Words: ryanodine receptor -CaMKII -phosphorylation -heart failure -arrhythmia

III.  The Fire From Within: The Biggest Ca2+ Channel Erupts and Dribbles  – Mark E. Anderson

Circ Res. 2005;97:1213-1215  http://dx.doi.org/10.1161/01.RES.0000196744.62327.36
http://circres.ahajournals.org/content/97/12/1213

Mark E. Andserson makes the point that CaMKII(δ) is the biggest calcium signaling channel, and it is pluripotent in the heart muscle.

The multifunctional Ca2+ and calmodulin (CaM)-dependent protein kinase II (CaMKII) is a serine threonine kinase that is abundant in heart where it phosphorylates Ca2+i homeostatic proteins. It seems likely that CaMKII plays an important role in cardiac physiology because these target proteins significantly overlap with the more extensively studied serine threonine kinase, protein kinase A (PKA), which is a key arbiter of catecholamine responses in heart. However, the physiological functions of CaMKII remain poorly understood, whereas the potential role of CaMKII in signaling myocardial dysfunction and arrhythmias has become an area of intense focus. CaMKII activity and expression are upregulated in failing human hearts and in many animal models of structural heart disease. CaMKII inhibitory drugs can pre-vent cardiac arrhythmias and suppress afterdepolarizations that are a probable proximate focal cause of arrhythmias in heart failure.

Cardiac contraction is initiated when Ca2+ current (ICa), through sarcolemmal L-type Ca2+ channels (LTCC), triggers RyR opening by a Ca2+-induced Ca2+ release (CICR) mechanism. LTCCs “face off” with RyRs across a highly ordered cytoplasmic cleft that delineates a kind of Ca2+ furnace during each CICR-initiated heart beat (Figure). CICR has an obvious need to function reliably, so it is astounding to consider how this feed forward process is intrinsically unstable. The increased instability of CICR in heart failure is directly relevant to arrhythmias initiated by afterdepolarizations. RyRs partly rely on a collaboration of Ca2+-sensing proteins in the SR lumen to grade their opening probability and the amount of SR Ca2+ release to a given ICa stimulus.

LTCCs and RyRs form the protein machinery for initiating contraction in cardiac and skeletal muscle, but in cardiac muscle communication between these proteins occurs without a requirement for physical contact. PKA is preassociated with LTCCs and RyRs, and PKA-dependent phosphorylation increases LTCC8 and RyR9opening. The resultant increase in Ca2+i is an important reason for the positive inotropic response to cathecholamines. The multifunctional Ca2+/calmodulin-dependent protein kinase II (CaMKII) is activated by increased Ca2+I, and so catecholamine stimulation activatesCaMKII in addition to PKA. In contrast to PKA, which is tightly linked to inotropy, CaMKII inhibition does not cause a reduction in fractional shortening during acute cate-cholamine stimulation in mice.

The key clinical phenotypes of contractile dysfunction and electrical instability in heart failure involve problems with Ca2+i homeostasis. Broad changes in Ca2+I-handling proteins can occur in various heart failure models, but in general heart failure is marked by a reduction in the capacity for SR Ca2+ uptake, enhanced activity of the sarcolemmal Na+-Ca2+ exchanger, and reduction in CICR-coordinated SR Ca2+ release. On the other hand, the opening probability of individual LTCCs is increased in human heart failure.

The Marks group pioneered the concept that RyRs are hyperphosphorylated by PKA in patients with heart failure and showed that successful therapies, ranging from beta blockers to left ventricular assist devices, reduce RyR phosphorylation in step with improved mechanical function. They have developed a large body of evidence in patients and in animal models that PKA phosphorylation of Ser2809 on cardiac RyRs destabilizes binding of FK12.6 to RyRs and promotes increased RyR opening that causes an insidious Ca2+ leak. This leak is potentially problematic because it can reduce SR Ca2+ content (to depress inotropy), engage pathological Ca2+-dependent transcriptional programs (to promote myocyte hypertrophy), and activate arrhythmia-initiating af-terdepolarizations (to cause sudden death).

 

CIRCULATIONAHA_oxCaMKII_AF_AR_Image_520 CIRCULATIONAHA_oxCaMKII_AF_AR_Image_523
CIRCULATIONAHA_oxCaMKII_AF_AR_Image_539    CIRCULATIONAHA_oxCaMKII_AF_AR_Image_538
CIRCULATIONAHA_oxCaMKII_AF_AR_Image_535                  CIRCULATIONAHA_oxCaMKII_AF_AR_Image_534
CIRCULATIONAHA_oxCaMKII_AF_AR_Image_522       CIRCULATIONAHA_oxCaMKII_AF_AR_Image_496

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Orthotopic Heart Transplant (OHT): Effects of Autonomic Innervation / Denervation on Atrial Fibrillation (AF) Genesis and Maintenance

Author and Curator: Larry H. Bernstein, MD, FCAP

and

Curator: Aviva Lev-Ari, PhD, RN

Sympathetic stimulation increases heart rate (positive chronotropy), inotropy and conduction velocity (positive dromotropy), whereas parasympathetic stimulation of the heart has opposite effects.

Noheria A, Patel SM, Mirzoyev S, Madhavan M, Friedman PA, Packer DL, Daly RC, Kushwaha SS, Edwards BS, Asirvatham SJ.

Division of Cardiology, Cedars-Sinai Medical Center, Los Angeles, California.
Pacing Clin Electrophysiol. 2013 Jun;36(6):741-7. http://dx.doi.org/10.1111/pace.12102. Epub 2013 Feb 25.

http://www.cvphysiology.com/Blood%20Pressure/ANS-medulla.gif
ANS- autonomic innervation of heart

The medulla, located in the brainstem above the spinal cord, is the primary site in the brain for regulating sympathetic and parasympathetic (vagal) outflow to the heart and blood vessels. The nucleus tractus solitarius (NTS) of the medulla receives sensory input from different systemic and central receptors (e.g., baroreceptors and chemoreceptors).
The heart is innervated by vagal and sympathetic fibers. The right vagus nerve primarily innervates the SA node, whereas the left vagus innervates the AV node; however, there can be significant overlap in the anatomical distribution. Atrial muscle is also innervated by vagal efferents, whereas the ventricular myocardium is only sparsely innervated by vagal efferents. Sympathetic efferent nerves are present throughout the atria (especially in the SA node) and ventricles, including the conduction system of the heart.
Cardiac function is altered by neural activation. Sympathetic stimulation increases heart rate (positive chronotropy), inotropy and conduction velocity (positive dromotropy), whereas parasympathetic stimulation of the heart has opposite effects.  Sympathetic and parasympathetic effects on heart function are mediated by beta-adrenoceptors and muscarinic receptors, respectively.
The overall effect of sympathetic activation is to increase cardiac output, systemic vascular resistance (both arteries and veins), and arterial blood pressure. Enhanced sympathetic activity is particularly important during exercise, emotional stress, and during hemorrhagic shock.
The actions of autonomic nerves are mediated by the release of neurotransmitters that bind to specific cardiac receptors and vascular receptors. These receptors are coupled to signal transduction pathways that evoke changes in cellular function.

                                         Sympathetic                      Parasympathetic

Heart

Chronotropy (rate)

+ + +                                     − − −

Inotropy (contractility)

+ + +                                      − 1

 Lusitropy (relaxation)                              
                                             + + +                                     –  1 
Dromotropy (conduction velocity)

                                              + +                                       − − −

Vessels

Arterial constriction    + + +                                    0

Venous constriction      + + +                                    0

Relative magnitude of responses indicated by number of + or – signs.
1 More pronounced in atria than ventricles.

CV Physiology: Autonomic Innervation of the Heart and Vasculature
http://www.cvphysiology.com/Blood%20Pressure/BP008.htm

Ablation Therapy for Cardiac Arrhythmias

By Richard N. Fogoros, M.D., About.com Guide Updated November 18, 2011
The most common form of ablation is done during a specialized form of cardiac catheterization, performed by a type of doctor known as a cardiac electrophysiologist (heart rhythm specialist). These procedures are sometimes called “trans-catheter ablations.”
During trans-catheter ablation procedures, specialized electrode catheters are positioned inside the heart, and the cardiac electrical system is mapped, showing the abnormal electrical pathways that are often responsible for producing the rapid heart rate. If these abnormal pathways are identified, the tip of the catheter (a tube) is placed on the abnormal pathway and the pathway is ablated (eliminated). The ablation itself is accomplished by transmitting some form of energy through the catheter (heat energy, freezing energy, or microwave energy), in order to damage the tissue at the tip of the catheter.

Decreased postoperative atrial fibrillation following cardiac transplantation: the significance of autonomic denervation.

BACKGROUND:  Endocardial ablation approaches have been proposed to targeting the retroatrial cardiac ganglia to treat atrial fibrillation (AF) . The potential value using this approach is unknown. Disruption of the autonomic inputs with orthotropic heart transplant (OHT) provides a unique opportunity to study the effects of autonomic innervation on AF genesis and maintenance.
The investigators hypothesized that due to denervation, the risk of postoperative AF would be lower following OHT compared to surgical maze even though both groups get isolation of the pulmonary veins.
METHODS:  We reviewed 155 OHTs (mean age 52 ± 11 years, 72% males) and used 1:1 age-, sex-, and date-of-surgery-matched two control groups from patients undergoing surgical maze or only coronary artery bypass grafting (CABG). Using conditional logistic regression we compared the odds of AF within 2 weeks following OHT versus controls.
RESULTS: Postoperative AF occurred in 10/155 (6.5%) OHT patients.
  1. The conditional odds of postoperative AF were lower for OHT as compared to controls (vs maze: odds ratio [OR] 0.27 [95% confidence interval (CI) 0.13-0.57], vs CABG: OR 0.38 [0.17-0.81], P = 0.003; and
  2. on additional adjustment for left atrial enlargement, vs maze: OR 0.28 [0.13-0.60], vs CABG: OR 0.14 [0.04-0.47], P = 0.0009).
CONCLUSIONS:
Risk of postoperative AF is significantly lower with OHT as in comparison to surgical maze. As both surgeries entail isolation of the pulmonary veins but
  • only OHT causes disruption of autonomic innervation,
this observation supports a mechanistic role of autonomic nervous system in AF. The benefit of targeting the cardiac autonomic system to treat AF needs further investigation.

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Imbalance of Autonomic Tone: The Promise of Intravascular Stimulation of Autonomics

Aviva Lev-Ari, PhD, RN

https://pharmaceuticalintelligence.com/2012/09/02/imbalance-of-autonomic-tone-the-promise-of-intravascular-stimulation-of-autonomics/

Renal Sympathetic Denervation: Updates on the State of Medicine

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https://pharmaceuticalintelligence.com/2012/12/31/renal-sympathetic-denervation-updates-on-the-state-of-medicine/

On Devices and On Algorithms: Prediction of Arrhythmia after Cardiac Surgery and ECG Prediction of an Onset of Paroxysmal Atrial Fibrillation

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https://pharmaceuticalintelligence.com/2013/05/07/on-devices-and-on-algorithms-arrhythmia-after-cardiac-surgery-prediction-and-ecg-prediction-of-paroxysmal-atrial-fibrillation-onset/

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Sympathetic (red) and parasympathetic (blue) n...

Sympathetic (red) and parasympathetic (blue) nervous system Русский: Аанатомия иннервации вегетативной нервной системы. Системы: симпатическая (красным) и парасимпатическая (синим) Українська: Аанатомія іннервації вегетативної нервової системи. Симпатична (червоним) та парасимпатична (синім) гілки Polski: Układ autonomiczny: czerwony – sympatyczny, niebieski – parasympatyczny. (Photo credit: Wikipedia)

Scheme of atrial fibrillation (top) and sinus ...

Scheme of atrial fibrillation (top) and sinus rhythm (bottom). The purple arrow indicates a P wave, which is lost in atrial fibrillation. (Photo credit: Wikipedia)

English: A graphical representation of the Ele...

English: A graphical representation of the Electrical conduction system of the heart showing the Sinoatrial node, Atrioventricular node, Bundle of His, Purkinje fibers, and Bachmann’s bundle (Photo credit: Wikipedia)

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On Devices and On Algorithms: Arrhythmia after Cardiac Surgery Prediction and ECG Prediction of Paroxysmal Atrial Fibrillation Onset

Author, and Content Consultant to e-SERIES A: Cardiovascular Diseases: Justin Pearlman, MD, PhD, FACC

and

Article Curator: Aviva Lev-Ari, PhD, RN

Cleveland Clinic research spurs a device that could predict arrhythmia after cardiac surgery

April 30, 2013 9:03 am by  |

ECG

Heart doctors at the Cleveland Clinic  hope to give doctors a way to tell which patients might develop arrhythmia after cardiac surgery.

Atrial fibrillation (AFIB) is one of the most common complications of heart surgeries, and also occurs as a complication of elevated alcohol use, high blood pressure, valve disease or thyroid disease. Atrial fibrillation consists of the round parts of the Valentine heart (the atria) shivering chaotically instead of beating rhythmically. Atrial fibrillation is a common arrhythmia, eventually affecting 20% of adults. There are 3 varieties: paroxysmal (intermittent), persistent (continual) and permanent (unremitting).  When AFIB lasts longer than 24-48 hours the risk of forming a blood clot in the atria rises, which in turn can cause a stroke or a heart attack. AFIB often results in fast heart rates which may cause low blood pressure and its possible consequences (organ injury, heart attack). Also, prolonged fast rates weaken the heart (reversible rate-related cardiomyopathy), which can persist for months after regaining target ranges for the heart rates (target for rate control is 60-80/minute instead of the fast rates of 100-180/min that are common with untreated AFIB).

A scoring system (CHADS2) can predict who may suffer from a stroke due to AFIB that lasts >24-48 hours, and in particular, who may benefit from longterm anticoagulation (blood thinners to interfere with clot formation). A pill-in-the-pocket can stop AFIB within hours.  Amiodarone, a highly toxic medication (10% long-term uses face side effects of serious damage to liver, lung, thyroid or eyes), is often prescribed “off-label” (without FDA endorsement) because it is 70% effective in preventing AFIB recurrence, and it has less anticontractility (weakening of the strength of heart beats) than most other rhythm medications. Then next most effective medication for suppression of AFIB long-term is sotalol, which reduces the strength of heart contraction (may not be tolerated by patients with severe heart failure) and it prolongs QT interval of repolarization after each heartbeat, a risk factor for a deadly rhythm called torsades de pointes. Interventional cures (“AFIB ablation”) have been developed to prevent recurrences.

Predicting AFIB may have several benefits: (1) potentially, earlier use of pill-in-the-pocket could prevent episodes rather that wait for them to occur, get noticed, and then treated, as only ~50% of AFIB episodes are noticed by the patient, according to electrographic monitor reports; (2) surrogate endpoint (prediction of onset) may offer useful guidance as to sufficiency of a suppressive therapy to enable lower dosing of toxic treatments; (3)  surrogate endpoint (prediction of onset) may offer useful guidance as to sufficient lowering of alohol intake, sufficient control of blood pressure, sufficient control of thyroid abnormalities, and other prevention opportunities; (4) surrogate endpoints may facilitate AFIB ablation.

Work done in the lab of Dr. C. Allen Bashour indicated that most patients who experience atrial fibrillation after heart surgery show clues beforehand in the form of subtle changes in their ECG readings that aren’t detected with the way they’re monitored now.

Rindex Medical is commercializing a tool that would enable physicians to predict which patients will experience AF so they can receive prophylactic treatment before it occurs.

“Right now they basically guess, or treat everyone prophylactically,” said co-founder Alex Arrow. “Some clinicians say they have an intuition about who will get it, but it’s mostly guesswork.”

Rindex’s A-50 AF Prediction System uses algorithms developed at the Clinic to analyze a patient’s ECG signals through 17 steps and produce a score, from 1 to 100, of how likely that patient is to experience AF. Arrow said the final product will be a touch-screen monitor that displays a score and tracks the score over a nine-hour period.

The Redwood City, California, company has been issued the first of its patents for the device and the exclusive license from Cleveland Clinic to develop the technology. Self-funded by Arrow and co-founders Denis Hickey and Lucas Fairfield, Rindex has a working prototype and is making progress on preparations for its 510(k) application. Arrow said the company shouldn’t need to raise a series A until it’s ready for a clinical trial.

Many other research groups have explored ways to predict AF in its various forms from natriuretic peptides to ECG changes, but no method has been established as reliably for this purpose.

Read more: http://medcitynews.com/2013/04/cleveland-clinic-research-spurs-a-device-that-could-predict-arrhythmias-after-cardiac-surgery/#ixzz2ScbxIyW0

http://medcitynews.com/2013/04/cleveland-clinic-research-spurs-a-device-that-could-predict-arrhythmias-after-cardiac-surgery/?goback=%2Egde_1503357_member_237204073

Dec 13, 2012

ECG predicts atrial fibrillation onset

Atrial fibrillation (AF), the most common cardiac arrhythmia, is categorized by different forms. One sub-type is paroxysmal AF (PAF), which refers to episodes of arrhythmia that generally terminate spontaneously after no more than a few days. Although the underlying causes of PAF are still unknown, it’s clear that predicting the onset of PAF would be hugely beneficial, not least because it would enable the application of treatments to prevent the loss of sinus rhythm.

Many research groups are tackling the issue of predicting the onset of PAF. Now, however, researchers in Spain have developed a method that assesses the risk of PAF at least one hour before its onset. To date, the approach has not only successfully discriminated healthy individuals and PAF patients, but also distinguished patients far from and close to PAF onset (Physiol. Meas. 33 1959).

“The ability to assess the risk of arrhythmia at least one hour before its onset is clinically relevant,” Arturo Martinez from the University of Castilla-La Mancha told medicalphysicsweb. “Our method assesses the P-wave feature time course from single-lead long-term ECG recordings. Using a single ECG lead reduces the computational burden, paving the way for a real-time system in future.”

Analysing sinus rhythm

If the heart is beating normally, the sinus rhythm observed on an ECG will contain certain generic features, such as a P-wave that reflects the atrial depolarization and a large characteristic R peak flanked by two minima representing the depolarization of the heart’s right and left ventricles. If an irregular heart beat is suspected, an ECG will be used and typical findings include the absence of a P-wave.

“We hypothesized that different stages of AF could be identified when analysing long-term recordings extracted from patients prone to AF,” commented Martinez. “Our method differs to others in that we also use just one single lead to detect small differences in features from the P-wave time course.”

P for paroxysmal

Martinez and his collaborators, Raul Alcaraz and Jose Rieta, studied 24-hour Holter ECG recordings from 24 patients in whom PAF had been detected for the first time. For each patient, the longest sinus rhythm interval in the recording was selected, and the two hours preceding the onset of PAF were analysed. These readings were compared with those from 28 healthy individuals. In all cases, only the trace from the V1 ECG lead was considered.

A major challenge for the researchers was to extract the P-wave from the baseline noise. To overcome this, they used an automatic delineator algorithm based on a phasor transform that determines the precise time point relating to the onset, peak and offset of the P-wave. The authors described this algorithm in a previous research paper (Physiol. Meas. 31 1467).

“All of the recordings in our study were visually supervised by expert cardiologists who corrected the P-wave fiducial points when needed,” said Martinez. “Even in the presence of noise, which generated an incredible amount of P-wave distortion, our delineator provided location errors lower than 8 ms.”

In order to assess which time course features might be useful to predict the onset of PAF, the researchers analysed a number of variables. First, they examined factors representing the duration of the P-wave (Pdur), such as the distance between the P-wave onset and peak (Pini) and the distance between the P-wave peak and its offset (Pter). They then studied factors relating P- to R-waves, such as the distance between the two waves’ peaks (PRk) and, finally, beat-to-beat P-wave factors, such as the distance between two consecutive P-wave onset points (PPon).

“The most remarkable trends were provided by the features measuring P-wave duration,” report the authors in their paper. “Pduridentified appropriately 84.21% of all the analysed patients, obtaining a discriminant accuracy of 90.79% and 83.33% between healthy subjects and PAF patients far from PAF and close to PAF, respectively. The metrics related to the PR interval showed the most limited ability to identify patient groups.”

About the author

Jacqueline Hewett is a freelance science and technology journalist based in Bristol, UK.

http://medicalphysicsweb.org/cws/article/research/51820

Original Article

Physiol Meas. 2010 Nov;31(11):1467-85. doi: 10.1088/0967-3334/31/11/005. Epub 2010 Sep 24.

Application of the phasor transform for automatic delineation of single-lead ECG fiducial points.

Martínez AAlcaraz RRieta JJ.

Source

Innovation in Bioengineering Research Group, University of Castilla La Mancha, Spain. arturo.martinez@uclm.es

Abstract

This work introduces a new single-lead ECG delineator based on phasor transform. The method is characterized by its robustness, low computational cost and mathematical simplicity. It converts each instantaneous ECG sample into a phasor, and can precisely manage P and T waves, which are of notably lower amplitude than the QRS complex. The method has been validated making use of synthesized and real ECG sets, including the MIT-BIH arrhythmia, QT, European ST-T and TWA Challenge 2008 databases. Experiments with the synthesized recordings reported precise detection and delineation performances in a wide variety of ECGs, with signal-to-noise ratios of 10 dB and above. For real ECGs, the QRS detection was characterized by an average sensitivity of 99.81% and positive predictivity of 99.89%, for all the analyzed databases (more than one million beats). Regarding delineation, the maximum localization error between automatic and manual annotations was lower than 6 ms and its standard deviation was in agreement with the accepted tolerances for expert physicians in the onset and offset identification for QRS, P and T waves. Furthermore, after revising and reannotating some ECG recordings by expert cardiologists, the delineation error decreased notably, becoming lower than 3.5 ms, on average, and reducing by a half its standard deviation. This new proposed strategy outperforms the results provided by other well-known delineation algorithms and, moreover, presents a notably lower computational cost.

SOURCES:

Original Database

MIT-BIH Polysomnographic Database

This database is described in

Ichimaru Y, Moody GB. Development of the polysomnographic database on CD-ROM. Psychiatry and Clinical Neurosciences 53:175-177 (April 1999).

Please cite this publication when referencing this material, and also include the standard citation for PhysioNet:

Goldberger AL, Amaral LAN, Glass L, Hausdorff JM, Ivanov PCh, Mark RG, Mietus JE, Moody GB, Peng C-K, Stanley HE. PhysioBank, PhysioToolkit, and PhysioNet: Components of a New Research Resource for Complex Physiologic Signals. Circulation 101(23):e215-e220 [Circulation Electronic Pages; http://circ.ahajournals.org/cgi/content/full/101/23/e215]; 2000 (June 13).

The MIT-BIH Polysomnographic Database is a collection of recordings of multiple physiologic signals during sleep. Subjects were monitored in Boston’s Beth Israel Hospital Sleep Laboratory for evaluation of chronic obstructive sleep apnea syndrome, and to test the effects of constant positive airway pressure (CPAP), a standard therapeutic intervention that usually prevents or substantially reduces airway obstruction in these subjects. The database contains over 80 hours’ worth of four-, six-, and seven-channel polysomnographic recordings, each with an ECG signal annotated beat-by-beat, and EEG and respiration signals annotated with respect to sleep stages and apnea. For further information, see Signals and Annotations.

The database consists of 18 records, each of which includes 4 files:

Sleep/apneaannotations Beatannotations Signals Header View waveforms *
slp01a.st slp01a.ecg slp01a.dat slp01a.hea
slp01b.st slp01b.ecg slp01b.dat slp01b.hea
slp02a.st slp02a.ecg slp02a.dat slp02a.hea
slp02b.st slp02b.ecg slp02b.dat slp02b.hea
slp03.st slp03.ecg slp03.dat slp03.hea
slp04.st slp04.ecg slp04.dat slp04.hea
slp14.st slp14.ecg slp14.dat slp14.hea
slp16.st slp16.ecg slp16.dat slp16.hea
slp32.st slp32.ecg slp32.dat slp32.hea
slp37.st slp37.ecg slp37.dat slp37.hea
slp41.st slp41.ecg slp41.dat slp41.hea
slp45.st slp45.ecg slp45.dat slp45.hea
slp48.st slp48.ecg slp48.dat slp48.hea
slp59.st slp59.ecg slp59.dat slp59.hea
slp60.st slp60.ecg slp60.dat slp60.hea
slp61.st slp61.ecg slp61.dat slp61.hea
slp66.st slp66.ecg slp66.dat slp66.hea
slp67x.st slp67x.ecg slp67x.dat slp67x.hea

(*) You may follow these links to view the signals and st annotations using either WAVE (under Linux, SunOS, or Solaris) or WVIEW (under MS-Windows). To do so successfully, you must have configured your browser to use wavescript (for WAVE) or wvscript (for WVIEW) as a helper application, as described in the WAVE User’s Guide(see the section titled WAVE and the Web) and in Setting up WVSCRIPT.

Andrew Walsh observed that the calibration originally provided for the BP signal of record slp37 is incorrect (since it yielded negative BPs). slp37.hea now contains an estimated BP calibration that yields more plausible BPs; these should not be regarded as accurate, however, since there is no independent calibration standard available for this recording.

SOURCE:
Original Article
Proc Inst Mech Eng H. 2010;224(1):27-42.

Finding events of electrocardiogram and arterial blood pressure signals via discrete wavelet transform with modified scales.

Ghaffari AHomaeinezhad MRAkraminia MDavaeeha M.

Source

Cardiovascular Research Group, Department of Mechanical Engineering, K. N. Toosi University of Technology, Tehran, Iran.

Abstract

A robust electrocardiogram (ECG) wave detection-delineation algorithm that can be applied to all ECG leads is developed in this study on the basis of discrete wavelet transform (DWT). By applying a new simple approach to a selected scale obtained from DWT, this method is capable of detecting the QRS complex, P-wave, and T-wave as well as determining parameters such as start time, end time, and wave sign (upward or downward). In the proposed method, the selected scale is processed by a sliding rectangular window of length n and the curve length in each window is multiplied by the area under the absolute value of the curve. In the next step, an adaptive thresholding criterion is conducted on the resulted signal. The presented algorithm is applied to various databases including the MIT-BIH arrhythmia database, European ST-T database, QT database, CinC Challenge 2008 database as well as high-resolution Holter data gathered in the DAY Hospital. As a result, the average values of sensitivity and positive prediction Se = 99.84 per cent and P+ = 99.80 per cent were obtained for the detection of QRS complexes with an average maximum delineation error of 13.7, 11.3, and 14.0 ms for the P-wave, QRS complex, and T-wave respectively. The presented algorithm has considerable capability in cases of a low signal-to-noise ratio, high baseline wander, and in cases where QRS complexes and T-waves appear with abnormal morphologies. Especially, the high capability of the algorithm in the detection of the critical points of the ECG signal, i.e. the beginning and end of the T-wave and the end of the QRS complex was validated by the cardiologist and the maximum values of 16.4 and 15.9 ms were recognized as absolute offset error of localization respectively. Finally, in order to illustrate an alternative capability of the algorithm, it is applied to all 18 subjects of the MIT-BIH polysomnographic database and the end-systolic and end-diastolic points of the blood pressure waveform were extracted and values of sensitivity and positive prediction Se = 99.80 per cent and P+ = 99.86 per cent were obtained for the detection of end-systolic, end-diastolic pulses.

http://www.ncbi.nlm.nih.gov/pubmed/20225455

Original Article

A robust wavelet-based multi-lead electrocardiogram delineation algorithm

  • a Department of Mechanical Engineering, K.N. Toosi University of Technology, Tehran, Iran
  • b CardioVascular Research Group (CVRG), Iran
  • c Non-invasive Cardiac Electrophysiology Laboratory, DAY Hospital, Tehran, Iran

Abstract

A robust multi-lead ECG wave detection-delineation algorithm is developed in this study on the basis of discrete wavelet transform (DWT). By applying a new simple approach to a selected scale obtained from DWT, this method is capable of detecting QRS complex, P-wave and T-wave as well as determining parameters such as start time, end time, and wave sign (upward or downward). First, a window with a specific length is slid sample to sample on the selected scale and the curve length in each window is multiplied by the area under the absolute value of the curve. In the next step, a variable thresholding criterion is designed for the resulted signal. The presented algorithm is applied to various databases including MIT-BIH arrhythmia database, European ST-T Database, QT Database, CinC Challenge 2008 Database as well as high resolution Holter data of DAY Hospital. As a result, the average values of sensitivity and positive predictivity Se = 99.84% and P+ = 99.80% were obtained for the detection of QRS complexes, with the average maximum delineation error of 13.7 ms, 11.3 ms and 14.0 ms for P-wave, QRS complex and T-wave, respectively. The presented algorithm has considerable capability in cases of low signal-to-noise ratio, high baseline wander, and abnormal morphologies. Especially, the high capability of the algorithm in the detection of the critical points of the ECG signal, i.e. the beginning and end of T-wave and the end of the QRS complex was validated by cardiologists in DAY hospital and the maximum values of 16.4 ms and 15.9 ms were achieved as absolute offset error of localization, respectively.

Abbreviations

  • ACL, area-curve length;
  • ECG, electrocardiogram;
  • DWT, discrete wavelet transform;
  • QTDB, QT database;
  • MITDB, MIT-BIH arrhythmia database; 
  • TWADB, T-wave alternans database;
  • CSEDB, common standards for electrocardiography database;
  • EDB, European ST-T database;
  • P+, positive predictivity (%);
  • Se,sensitivity (%);
  • FIR, finite-duration impulse response;
  • LE, location error;
  • CHECK#0, procedure of evaluating obtained results using MIT annotation files;
  • CHECK#1, procedure of evaluating obtained results consulting with a control cardiologist;
  • CHECK#2, procedure of evaluating obtained results consulting with a control cardiologist and also at least with 3 residents

Keywords

  • ECG delineation;
  • Discrete wavelet transform;
  • Variable threshold;
  • Validation

Figures and tables from this article:

Full-size image (14 K)
Fig. 1. FIR filter-bank implementation to generate discrete wavelet transform based on à trous algorithm.
Full-size image (12 K)
Fig. 2. Graphical representation of the logic of the proposed simple transformation for detecting onset and offset edges. In case I, both area and curve length are minimum, (ACLI < ACLII ≤ ACLIII).
Full-size image (58 K)
Fig. 3. The flow-chart of the proposed wavelet-aided electrocardiogram delineation algorithm (rectangle: operation, ellipse: result).
Full-size image (113 K)
Fig. 4. An excerpted segment from a total delineated ECG. Delineated (a) P-waves, (b) QRS complexes and (c) T-waves. (Circles: edges of event, triangles: peak of events, Partition A: lead I, Partition B: lead II).
Full-size image (76 K)
Fig. 5. Procedure of detecting and delineating of P and T-waves using ACL signal between two successive QRS complexes. (a) Simultaneously depiction of ACL, original ECG and the corresponding selected DWT scale, (b) QRS delineation, and (c) P and T-waves delineation.
SOURCE:

Volume 31, Issue 10, December 2009, Pages 1219–1227

http://www.sciencedirect.com/science/article/pii/S1350453309001647

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Sustained Cardiac Atrial Fibrillation: Management Strategies by Director of the Arrhythmia Service and Electrophysiology Lab at The Johns Hopkins Hospital   https://pharmaceuticalintelligence.com/2012/10/16/sustained-cardiac-atrial-fibrillation-management-strategies-by-director-of-the-arrhythmia-service-and-electrophysiology-lab-at-the-johns-hopkins-hospital/

Cardiac Arrhythmias: A Risk for Extreme Performance Athletes                                                                                                                                                       https://pharmaceuticalintelligence.com/2012/08/08/cardiac-arrhythmias-a-risk-for-extreme-performance-athletes/

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Importance of Omega-3 Fatty Acids in Reducing Cardiovascular Disease

Reporter and Curator: Dr. Sudipta Saha, Ph.D.

 

UPDATED on 7/24/2018

Omega-3 fats Supplements Effect on Cardiovascular Health: EPA and DHA has little or no effect on Mortality or Cardiovascular Health

Reporter: Aviva Lev-Ari, PhD, RN

https://pharmaceuticalintelligence.com/2018/07/24/omega-3-fats-supplements-effect-on-cardiovascular-health-epa-and-dha-has-little-or-no-effect-on-mortality-or-cardiovascular-health/

 

The available evidence for cardiovascular effects of n-3 polyunsaturated fatty acid (PUFA) consumption has been reviewed here, focusing on long chain (seafood) n-3 PUFA, including their principal dietary sources, effects on physiological risk factors, potential molecular pathways and bioactive metabolites, effects on specific clinical endpoints, and existing dietary guidelines. Major dietary sources include fatty fish and other seafood. n-3 PUFA consumption lowers plasma triglycerides, resting heart rate, and blood pressure and might also improve myocardial filling and efficiency, lower inflammation, and improve vascular function. Experimental studies demonstrate direct anti-arrhythmic effects, which have been challenging to document in humans. n-3 PUFA affect a myriad of molecular pathways, including alteration of physical and chemical properties of cellular membranes, direct interaction with and modulation of membrane channels and proteins, regulation of gene expression via nuclear receptors and transcription factors, changes in eicosanoid profiles, and conversion of n-3 PUFA to bioactive metabolites. In prospective observational studies and adequately powered randomized clinical trials, benefits of n-3 PUFA seem most consistent for coronary heart disease mortality and sudden cardiac death. Potential effects on other cardiovascular outcomes are less-well-established, including conflicting evidence from observational studies and/or randomized trials for effects on nonfatal myocardial infarction, ischemic stroke, atrial fibrillation, recurrent ventricular arrhythmias, and heart failure. Research gaps include the relative importance of different physiological and molecular mechanisms, precise dose-responses of physiological and clinical effects, whether fish oil provides all the benefits of fish consumption, and clinical effects of plant-derived n-3 PUFA. Overall, current data provide strong concordant evidence that n-3 PUFA are bioactive compounds that reduce risk of cardiac death. National and international guidelines have converged on consistent recommendations for the general population to consume at least 250 mg/day of long-chain n-3 PUFA or at least 2 servings / week of oily fish.

Source References:

http://content.onlinejacc.org/article.aspx?articleid=1146941

http://www.ncbi.nlm.nih.gov/pubmed/17047219

http://www.ncbi.nlm.nih.gov/pubmed/18614744

http://www.ncbi.nlm.nih.gov/pubmed/19364995

http://www.ncbi.nlm.nih.gov/pubmed/16172267

Other articles related to this topic were published on this Open Access Online Scientific Journal, including the following:

Reversal of Cardiac mitochondrial dysfunction

Larry H Bernstein, MD, FACP, RN 04/14/2013

https://pharmaceuticalintelligence.com/2013/04/14/reversal-of-cardiac-mitochondrial-dysfunction/

Can resolvins suppress acute lung injury?

Larry H Bernstein, MD, FACB, RN 03/06/2013

https://pharmaceuticalintelligence.com/2013/03/06/can-resolvins-suppress-acute-lung-injury/

Calcium (Ca) supplementation (>1400 mg/day): Higher Death Rates from all Causes and Cardiovascular Disease in Women

Aviva Lev-Ari, PhD., RN 02/19/2013

https://pharmaceuticalintelligence.com/2013/02/19/calcium-ca-supplementation-1400-mgday-higher-death-rates-from-all-causes-and-cardiovascular-disease-in-women/

Endothelial Function and Cardiovascular Disease

Larry H Bernstein, MD, FCAP, Pathologist, Contributor, RN 10/25/2012

https://pharmaceuticalintelligence.com/2012/10/25/endothelial-function-and-cardiovascular-disease/

Mediterranean Diet is BEST for patients with established Heart Disorders

Aviva Lev-Ari, PhD, RN 10/15/2012

https://pharmaceuticalintelligence.com/2012/10/15/mediterranean-diet-is-best-for-patients-with-established-heart-disorders/

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Xarelto (Rivaroxaban): Anticoagulant Therapy gains FDA New Indications and Risk Reduction for: (DVT) and (PE), while in use for Atrial fibrillation increase in Gastrointestinal (GI) Bleeding Reported

Reporter: Aviva Lev-Ari, PhD, RN

 

UPDATED on 8/17/2018

NOAC’s Brain Bleed Risk Outside Afib May Be Dose-Dependent

Higher risk seen only with higher rivaroxaban doses in meta-analysis

by Ashley Lyles, MedPage Today Intern 

The findings indicate the following risk of intracranial hemorrhage versus aspirin:

  • 10 mg of rivaroxaban taken once per day or 5 mg taken two times a day (three trials, OR 1.43, 95% CI 0.93-2.21)
  • 5 mg of apixaban twice daily (one trial, OR 0.84, 95% CI 0.38-1.88)

The study also showed that 15 mg to 20 mg of rivaroxaban each day was linked with an increased risk of fatal bleeding (two trials, OR 2.37, 95% CI 1.30-4.29). On the other hand, 10 mg of rivaroxaban each day or 5 mg taken twice a day (three trials, OR 1.47, 95% CI 0.72-2.97) and 5 mg of apixaban taken twice per day (one trial, OR 0.66, 95 % CI 0.19-2.35) were not linked with an increased risk.

Increased risk of major bleeding compared with aspirin was seen with 15 mg to 20 mg dose of rivaroxaban each day (two trials, OR 2.64, 95% CI 1.68-4.16) and a 10 mg dose of rivaroxaban once a day or 5 mg twice per day (three trials, OR 1.56, 95% CI 1.31-1.85).

Primary Source

JAMA Neurology

Source Reference: Huang W, et al “Association of intracranial hemorrhage risk with non–vitamin k antagonist oral anticoagulant use vs aspirin use a systematic review and meta-analysis” JAMA Neurology 2018; DOI: 10.1001.

SOURCE

https://www.medpagetoday.com/cardiology/strokes/74552?xid=nl_mpt_cardiodaily_2018-08-17&eun=g99985d0r&utm_source=Sailthru&utm_medium=email&utm_campaign=AHAWeekly_081718&utm_term=AHA%20Cardiovascular%20Daily%20-%20Active%20Users%20180%20days

 

UPDATED on 10/9/2017

Xarelto Flop in Stroke Prevention Trial; Syncope Device; Workout by Watching Hockey, Theater?

Recent developments of interest in cardiovascular medicine

  • by Crystal Phend,Senior Associate Editor, MedPage TodayOctober 09, 2017

https://www.medpagetoday.com/Cardiology/Prevention/68421

Rivaroxaban (Xarelto) flopped for preventing recurrent strokes and increased bleeding compared with aspirin in top-line results from the phase III NAVIGATE ESUS trial, Bayer and Janssen announced. (Genetic Engineering and Biotechnology News)

Xarelto (Rivaroxaban): Anticoagulant Therapy gains FDA New Indications and Risk Reduction for: (DVT) and (PE), while in use for Atrial fibrillation, increase in Gastrointestinal (GI) Bleeding Reported compared with Coumadin

Rivaroxaban Gains FDA Indications For The Treatment And Prevention Of DVT And PE

The FDA today expanded the indication for rivaroxaban (Xarelto, Johnson & Johnson) to include the treatment of deep vein thrombosis (DVT) and pulmonary embolism (PE) and to reduce the risk of recurrent DVT and PE.

The oral anticoagulant is already approved to reduce the post-surgical risk of DVT and PE  after hip and knee replacement surgery and to reduce the risk of stroke in people with atrial fibrillation. The new indication was granted under the FDA’s priority review program.

“Xarelto is the first oral anti-clotting drug approved to treat and reduce the recurrence of blood clots since the approval of warfarin nearly 60 years ago,” said Richard Pazdur,  director of the FDA’s Office of Hematology and Oncology Products, in an FDA press release.

Here is the FDA press release:

FDA expands use of Xarelto to treat, reduce recurrence of blood clots
The U.S. Food and Drug Administrationtoday expanded the approved use of Xarelto (rivaroxaban) to include treating deep vein thrombosis (DVT) or pulmonary embolism (PE), and to reduce the risk of recurrent DVT and PE following initial treatment.Blood clots occur when blood thickens and clumps together. DVT is a blood clot that forms in a vein deep in the body. Most deep vein blood clots occur in the lower leg or thigh. When a blood clot in a deep vein breaks off and travels to an artery in the lungs and blocks blood flow, it results in a potentially deadly condition called PE.Xarelto is already FDA-approved to reduce the risk of DVTs and PEs from occurring after knee or hip replacement surgery (July 2011), and to reduce the risk of stroke in people who have a type of abnormal heart rhythm called non-valvular atrial fibrillation (November 2011).

The FDA reviewed Xarelto’s new indication under the agency’s priority review program, which provides an expedited six-month review for drugs that offer major advances in treatment or that provide treatment when no adequate therapy exists.

“Xarelto is the first oral anti-clotting drug approved to treat and reduce the recurrence of blood clots since the approval of warfarin nearly 60 years ago,” said Richard Pazdur, M.D., director of the Office of Hematology and Oncology Products in the FDA’s Center for Drug Evaluation and Research.

Other drugs approved by FDA to treat or reduce the risk of blood clots include Lovenox (enoxaparin), generic versions of enoxaparin, Arixtra (fondaparinux), Fragmin (dalteparin), Coumadin (warfarin), and heparin.

The safety and effectiveness of Xarelto for the new indications were evaluated in three clinical studies. A total of 9,478 patients with DVT or PE were randomly assigned to receive Xarelto, a combination of enoxaparin and a vitamin K antagonist (VKA), or a placebo. The studies were designed to measure the number of patients who experienced recurrent symptoms of DVT, PE or death after receiving treatment.

Results showed Xarelto was as effective as the enoxaparin and VKA combination for treating DVT and PE. About 2.1 percent of patients treated with Xarelto compared with 1.8 percent to 3 percent of patients treated with the enoxaparin and VKA combination experienced a recurrent DVT or PE. Additionally, results from a third study showed extended Xarelto treatment reduced the risk of recurrent DVT and PE in patients. About 1.3 percent of patients treated with Xarelto compared with 7.1 percent of patients receiving placebo experienced a recurrent DVT or PE.

The major side effect observed with Xarelto is bleeding, similar to other anti-clotting drugs.

Xarelto is marketed by Raritan, N.J.-based Janssen Pharmaceuticals Inc.

For more information:

FDA: Office of Hematology and Oncology Products

FDA: Approved Drugs: Questions and Answers

The FDA, an agency within the U.S. Department of Health and Human Services, protects the public health by assuring the safety, effectiveness, and security of human and veterinary drugs, vaccines and other biological products for human use, and medical devices. The agency also is responsible for the safety and security of our nation’s food supply, cosmetics, dietary supplements, products that give off electronic radiation, and for regulating tobacco products.

 

SOURCE:

http://www.forbes.com/sites/larryhusten/2012/11/02/rivaroxaban-gains-fda-indications-for-the-treatment-and-prevention-of-dvt-and-pe/?goback=%2Egde_2069447_member_181862591

Cardiac Atrial Fibrillation

ATLANTA, Georgia — Patients with atrial fibrillation receiving anticoagulant therapy are more likely to experience gastrointestinal (GI) bleeding when treated with rivaroxaban than when treated with warfarin, according to a new analysis of data from ROCKET AF.

Christopher Nessel, MD, from research and development at Johnson & Johnson in Raritan, New Jersey, reported the findings here at CHEST 2012: American College of Chest Physicians Annual Meeting.

“Compared with warfarin, the risk of GI bleeding is increased with rivaroxaban, but the incidence of life-threatening or fatal GI bleeding is lower,” Dr. Nessel told Medscape Medical News. “A careful benefit/risk assessment is needed prior to prescribing rivaroxaban for high-risk patients,” he added.

The analysis examined the incidence and outcomes of GI hemorrhage in 14,264 patients with nonvalvular atrial fibrillation enrolled in ROCKET AF.

The patients were randomized to either rivaroxaban or dose-adjusted warfarin. All GI bleeding events were recorded during treatment and for 2 days after the last dose was administered. Severity of bleeding was defined by a corresponding drop in hemoglobin or transfusion of more than 2 units of red cells.

The composite principal safety end point for GI bleeding events (upper GI, lower GI, and rectal bleeding) occurred more frequently in the 394 patients receiving rivaroxaban than in the 290 receiving warfarin (3.61% vs 2.60% per year; hazard ratio [HR], 1.39; 95% confidence interval [CI], 1.19 to 1.61). Major bleeding was more frequent with rivaroxaban than with warfarin (2.00% vs 1.24% per year; HR, 1.61; 95% CI, 1.30 to 1.99), as was clinically relevant nonmajor bleeding (1.75% vs 1.39% per year; HR, 1.26; 95% CI, 1.20 to 1.55).

Patients who experienced major GI bleeding were more likely to have experienced GI bleeding in the past, to have mild anemia, to have a lower creatinine clearance, to be previous or current smokers, and to be older than patients who did not experience a GI bleeding during the trial (n = 13,552). They were also less likely to be female and to have previously experienced a stroke or transient ischemic attack.

The incidence of severe bleeding (transfusion of at least 4 units) was similar in the rivaroxaban and warfarin groups (49 vs 47). Six patients developed fatal bleeding: 1 in the rivaroxaban group and 5 in the warfarin group.

Data May Give Clinicians Pause When Considering Rivaroxaban

“The data presented extend the observations from the ROCKET AF clinical study,” Dr. Nessel said. “Specifically, the analyses identified characteristics of nonvalvular atrial fibrillation patients that may predispose them to the occurrence of GI hemorrhage. The data also indicated that the overall fatality rates for bleeds of this nature are very low.”

Independent commentator James Wisler, MD, from the division of cardiovascular disease at Duke University Medical Center in Durham, North Carolina, pointed out that this study underscores the importance of critically evaluating these newer anticoagulants when considering their use in a given patient.

“The decision regarding which anticoagulant to use for a given patient is complex, and risks and benefits need to be considered thoughtfully,” he told Medscape Medical News. He added that the results of this study might give some physicians pause about initiating a newer anticoagulant, such as rivaroxaban, in a given patient with atrial fibrillation and an unfavorable risk profile, such as those with a previous GI bleed.

“While the previously published results from ROCKET AF suggested that the risk profiles were similar between rivaroxaban and warfarin, these results demonstrate that there is indeed a subpopulation of patients who may be better served with warfarin than rivaroxaban,” he explained.

According to Dr. Wisler, both this analysis and the initial ROCKET AF study demonstrate that rivaroxaban is associated with fewer episodes of severe or fatal bleeding events, despite the increase in major and clinically relevant nonmajor bleeding observed in the specific subgroup of this study. “Currently, it is unclear why this discrepancy exists,” he added.

He recommends that clinicians take a careful patient history to assess bleeding risk factors when considering the initiation of a newer anticoagulant such as rivaroxaban.

“While perhaps more convenient and efficacious, certain patient populations, such as that evaluated in this study, may receive net harm from these newer agents,” he said.

SOURCE:

CHEST 2012: American College of Chest Physicians Annual Meeting. Presented October 22, 2012.

http://journal.publications.chestnet.org/issue.aspx?journalid=99&issueid=25283

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