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International audience; Aims: Atrial Fibrillation (AF) is an arrhythmia of increasing prevalence in the aging populations of developed countries. One of the important indicators of AF is sustained atrial dilatation, highlighting the importance of mechanical overload in the pathophysiology of AF. The mechanisms by which atrial cells, including fibroblasts, sense and react to changing mechanical forces, are not fully elucidated. Here, we characterise stretch-activated ion channels (SAC) in human atrial fibroblasts and changes in SAC- presence and activity associated with AF.Methods and results: Using primary cultures of human atrial fibroblasts, isolated from patients in sinus rhythm or sustained AF, we combine electrophysiological, molecular and pharmacological tools to identify SAC. Two electrophysiological SAC- signatures were detected, indicative of cation-nonselective and potassium-selective channels. Using siRNA-mediated knockdown, we identified the cation-nonselective SAC as Piezo1. Biophysical properties of the potassium-selective channel, its sensitivity to calcium, paxilline or iberiotoxin (blockers), and NS11021 (activator), indicated presence of calcium-dependent 'big potassium channels' (BKCa). In cells from AF patients, Piezo1 activity and mRNA expression levels were higher than in cells from sinus rhythm patients, while BKCa activity (but not expression) was downregulated. Both Piezo1-knockdown and removal of extracellular calcium from the patch pipette resulted in a significant reduction of BKCa current during stretch. No co-immunoprecipitation of Piezo1 and BKCa was detected.Conclusions: Human atrial fibroblasts contain at least two types of ion channels that are activated during stretch: Piezo1 and BKCa. While Piezo1 is directly stretch-activated, the increase in BKCa activity during mechanical stimulation appears to be mainly secondary to calcium influx via SAC such as Piezo1. During sustained AF, Piezo1 is increased, while BKCa activity is reduced, highlighting differential regulation of both channels. Our data support the presence and interplay of Piezo1 and BKCa in human atrial fibroblasts in the absence of physical links between the two channel proteins.

Aims The risk of developing atrial fibrillation (AF) and its complications continues to increase, despite good progress in preventing AF-related strokes. Methods and results This article summarizes the outcomes of the 7th Consensus Conference of the Atrial Fibrillation NETwork (AFNET) and the European Heart Rhythm Association (EHRA) held in Lisbon in March 2019. Sixty-five international AF specialists met to present new data and find consensus on pressing issues in AF prevention, management and future research to improve care for patients with AF and prevent AF-related complications. This article is the main outcome of an interactive, iterative discussion between breakout specialist groups and the meeting plenary. AF patients have dynamic risk profiles requiring repeated assessment and risk-based therapy stratification to optimize quality of care. Interrogation of deeply phenotyped datasets with outcomes will lead to a better understanding of the cardiac and systemic effects of AF, interacting with comorbidities and predisposing factors, enabling stratified therapy. New proposals include an algorithm for the acute management of patients with AF and heart failure, a call for a refined, data-driven assessment of stroke risk, suggestions for anticoagulation use in special populations, and a call for rhythm control therapy selection based on risk of AF recurrence. Conclusion The remaining morbidity and mortality in patients with AF needs better characterization. Likely drivers of the remaining AF-related problems are AF burden, potentially treatable by rhythm control therapy, and concomitant conditions, potentially treatable by treating these conditions. Identifying the drivers of AF-related complications holds promise for stratified therapy.

BACKGROUND The underlying mechanisms of exercise intolerance in sickle cell anaemia (SCA) patients are complex and not yet completely understood. While latent heart failure at rest could be unmasked upon exercise, most previous studies assessed cardiac function at rest. We aimed to investigate exercise cardiovascular reserve as a potential contributor to exercise intolerance in adult SCA patients. METHODS In this observational prospective study, we compared prospectively 60 SCA patients (median age 31 years, 60% women) to 20 matched controls. All subjects underwent symptom-limited combined exercise echocardiography and oxygen uptake (VO2 ) measurements. Differences between arterial and venous oxygen content (C(a-v)O2 ) were calculated. Cardiac reserve was defined as the absolute change in cardiac index (Ci) from baseline to peak exercise. RESULTS Compared to controls, SCA patients demonstrated severe exercise intolerance (median peakVO2 , 34.3 vs. 19.7 ml/min/kg, respectively, p

Recent developments in imaging, mapping, and ablation techniques have shown that the epicardial region of the heart is a key player in the occurrence of ventricular arrhythmic events in several cardiac diseases, such as Brugada syndrome, arrhythmogenic cardiomyopathy, or dilated cardiomyopathy. At the atrial level as well, the epicardial region has emerged as an important determinant of the substrate of atrial fibrillation, pointing to common underlying pathophysiological mechanisms. Alteration in the gradient of repolarization between myocardial layers favouring the occurrence of re-entry circuits has largely been described. The fibro-fatty infiltration of the subepicardium is another shared substrate between ventricular and atrial arrhythmias. Recent data have emphasized the role of the epicardial reactivation in the formation of this arrhythmogenic substrate. There are new evidences supporting this structural remodelling process to be regulated by the recruitment of epicardial progenitor cells that can differentiate into adipocytes or fibroblasts under various stimuli. In addition, immune-inflammatory processes can also contribute to fibrosis of the subepicardial layer. A better understanding of such ‘electrical fragility’ of the epicardial area will open perspectives for novel biomarkers and therapeutic strategies. In this review article, a pathophysiological scheme of epicardial-driven arrhythmias will be proposed.

Funding Acknowledgements Type of funding sources: Public grant(s) – EU funding. Main funding source(s): CATCH ME Horizon 2020, EU-H2020-PHC-RIA (633196) CVON2014-09, RACE V: Reappraisal of Atrial Fibrillation: Interaction between hyperCoagulability, Electrical remodeling, and Vascular Destabilisation in the Progression of AF Background Atrial fibrosis is one of the most important aspects of structural remodeling in the atria and largely increases the inducibility and sustainability of AF. The main risk factors for recurrent AF, such as ageing, heart failure, and history of AF, partly enhance AF propensity by inducing atrial fibrosis. A distinction should be made between replacement fibrosis following myocyte death and reactive fibrosis, which often occurs in the absence of myocyte death. Endomysial fibrosis, a type of reactive fibrosis in between cardiomyocytes, is poorly studied as a result of limitations and labor intensiveness of traditional histochemical quantification. Purpose We examined the contributions of age, sex, AF and heart failure to the development of overall and endomysial fibrosis in the context of concurrent pathologies. Methods We developed an algorithm for automated quantification of multiple features of structural remodeling following myocardial staining with wheat germ agglutinin (WGA). We studied the type, quantity and distribution of fibrosis in left (LAA, n = 95) and right (RAA, n = 76) atrial appendage biopsies in a large European cohort of patients with varying indications for cardiac surgery. Linear mixed effect models were constructed to predict endomysial fibrosis quantity and clustering as a function of AF, heart failure, sex, age and 4 principle components that accounted for potential confounding due to other clinical characteristics. Results Persistent AF, heart failure and female sex were independently associated with endomysial fibrosis, age was not. AF and age were not associated with overall fibrosis. Clustering of endomysial fibrosis was observed in females (LAA), paroxysmal AF (RAA), persistent AF and heart failure (LAA) patients (table 1). Conclusions Female sex, AF, and heart failure are associated with the quantity and distribution of endomysial fibrosis, the effects of age are limited. summary of results Clinical parameter Overall fibrosis Endomysial fibrosis Clustering of endomysial fibrosis LAA RAA LAA RAA LAA RAA Paroxysmal AF +1.6%±1.5; p = 0.29 +2.1%±2.2; p = 0.34 -0.7μm ± 0.5; p = 0.19 +1.0μm ± 0.6; p = 0.10 -0.8%±3.6; p = 0.82 +7.4%±4.1; p = 0.04 Persistent AF +1.6%±1.5; p = 0.26 +2.2%±2.1; p = 0.30 +1.1μm ± 0.5; p = 0.04 +1.3μm ± 0.6; p = 0.03 +5.7%±0.03; p = 0.04 +6.9%±3.9, p = 0.04 Heart failure +4.8%±1.5; p = 0.01 +2.3%±2.0; p = 0.24 +2.5μm ± 0.5; p

Funding Acknowledgements Type of funding sources: Public grant(s) – EU funding. Main funding source(s): TRAIN-HEART Innovative Training Network, funded by the European Union’s Horizon 2020 research and innovation program (under the Marie Sklodowska-Curie grant agreement no. 813716) Characterizing Atrial fibrillation by Translating its Causes into Health Modifiers in the Elderly (CATCH ME), funded by the European Union’s Horizon 2020 research and innovation program (under the grant agreement no. 633196) Background Atrial fibrillation (AF) is the most common cardiac arrhythmia and is associated with heart failure (HF) and stroke. Clinical and experimental data from previous studies suggest gender differences in mechanisms and phenotypes of AF: women may have more atrial fibrosis, worse outcomes after catheter ablation, and some women carry a higher risk for thromboembolic complications than men. The molecular mechanisms underlying these differences are still poorly understood. Methods Gender-based transcriptional patterns were assessed using paired-end, directional RNA sequencing data generated from atrial tissue biopsies in 199 patients either in sinus rhythm or with paroxysmal or persistent AF as part of the CATCH-ME project. Transcript counts were compared between genders separately in the left and right atria using the DESeq2 package in R. The models were adjusted for potential sources of confounding (age, atrial fibrillation status, heart failure status and sequencing batch). Interaction models were implemented using DESeq2 to compare gender*morbidity interactions for persistent AF and HF. Significance was assessed using likelihood ratio tests comparing models with and without the interaction terms. Results with an adjusted P-value 0.05 were considered significant and utilized for subsequent downstream assessments. Differentially expressed (DE) genes were tested for enrichment of gene ontology (GO) terms and KEGG pathways using the WebGestalt toolkit. Results Transcriptome-wide profiling across the cohort identified 33 sex-differentiated genes in the left atria and 51 in the right atrial samples, with 21 of these showing bilateral differences. Interestingly, 36 (44%) of the results from these analyses were comprised of non-coding transcripts, including long non-coding RNAs (lncRNAs), antisense RNAs and pseudogenes. GO and pathway enrichment analyses for these genes revealed their involvement in critical pathways such as the complement and coagulation cascades and RNA transport. Interaction analyses between gender and AF identified two genes (MPP2 & GNAS-AS1) that were differentially transcribed in the right atria and one gene (MYL2) that was DE in the left atria by gender in persistent AF samples. A similar analysis comparing gender*HF morbidity also revealed evidence of DE. Four transcripts (HLA-DQB1-AS1, EIF1AY, UTY and ZFY-AS1) showed gender-specific differences in expression by HF status in left atria, while HLA-DQB1-AS1 was differentially regulated by gender and HF status in right atrial samples. Conclusions These RNA-seq analyses provide novel insights into gender-related differences in the transcriptional landscape of right and left adult human atrial appendages. Moreover, interaction analyses identified three genes DE in female atria in persistent AF and four DE genes in female atria in heart failure, providing a molecular anchor for the observed differences in atrial diseases phenotypes between men and women.

AimsAtrial Fibrillation (AF) is an arrhythmia of increasing prevalence in the aging population of developed countries. One of the important indicators of AF is sustained atrial dilatation, highlighting the importance of mechanical overload in the pathophysiology of AF. The mechanisms by which atrial cells, including fibroblasts, sense and react to changing mechanical forces, are not fully elucidated. Here, we characterise stretch-activated ion channels (SAC) in human atrial fibroblasts and changes in SAC-presence and -activity associated with AF.Methods and ResultsUsing primary cultures of human atrial fibroblasts, isolated from patients in sinus rhythm or sustained AF, we combine electrophysiological, molecular and pharmacological tools to identify SAC. Two electrophysiological SAC-signatures were detected, indicative of cation-nonselective and potassium-selective channels. Using siRNA-mediated knockdown, we identified the nonselective SAC as Piezo1. Biophysical properties of the potassium-selective channel, its sensitivity to calcium, paxilline and iberiotoxin (blockers), and NS11021 (activator), indicated presence of calcium-dependent ‘big potassium channels’, BKCa. In cells from AF patients, Piezo1 activity and mRNA expression levels were higher than in cells from sinus rhythm patients, while BKCa activity (but not expression) was downregulated. Both Piezo1-knockdown and removal of extracellular calcium from the patch pipette resulted in a significant reduction of BKCa current during stretch. No co-immunoprecipitation of Piezo1 and BKCa was detected.ConclusionsHuman atrial fibroblasts contain at least two types of ion channels that are activated during stretch: Piezo1 and BKCa. While Piezo1 is directly stretch-activated, the increase in BKCa activity during mechanical stimulation appears to be mainly secondary to calcium influx via SAC such as Piezo1. During sustained AF, Piezo1 is increased, while BKCa activity is reduced, highlighting differential regulation of both channels. Our data support the presence and interplay of Piezo1 and BKCa in human atrial fibroblasts in the absence of physical interactions between the two channel proteins.

Background: Early detection of atrial fibrillation (AF) and subsequent initiation of anticoagulation and rhythm control therapy markedly reduce stroke, cardiovascular death, and heart failure, but unselected ECG screening for AF is time- and resource-intensive. Combining biomarkers reflecting different biological processes may identify patients at high risk of AF, enabling targeted ECG screening. Methods: We systematically reviewed literature and patent information to define candidate biomarkers for AF detection. The top 12 biomarkers identified through this process were quantified on a high-precision, high-throughput platform in 1485 consecutive patients at risk for AF presenting acutely to hospital (median age 69 years [Q1, Q3 60,78]; 60% male). Patients had either known AF (45%) or AF ruled out by 7-day ECG-monitoring. A model simultaneously considering 7 key clinical characteristics and all biomarkers was developed in a randomly sampled discovery cohort (n=933) and validated in the remaining patients (n=552). Neural networks were also applied. Findings: Using backward elimination, a model using age, sex, body mass index (BMI), BMP10, ANG2, and FGF23 discriminated between patients with and without AF with an AUC of 0·743 [95% confidence interval (CI) 0·712-0·775]. The biomarkers represent distinct pathways relevant for atrial cardiomyopathy and AF, namely hypertrophy and fibrosis (FGF23), endothelial dysfunction (ANG2), and atrial oxidative stress and the genomic predisposition to AF (BMP10). The SHAP procedure for neural networks identified the same variables as the regression. The validation yielded an AUC of 0·719 (95%CI 0·677, 0·762), corroborated using deep neural networks (AUC 0·784 [95%CI 0·745, 0·822]). Interpretation: The combination of three simple characteristics (age, sex, BMI) and three biomarkers (BMP10, ANG2, and FGF23) robustly identifies patients with AF. Such an approach enables targeted screening for AF and provides a platform to develop personalised prevention and treatment in patients with AF. Funding Statement: This work was partially supported by the European Commission (grant agreements no. 633196 [CATCH ME]) to PKi, LF, BC, SH, SK, LM, MFS, US, and no. 116074 [BigData@Heart EU IMI] to PKi, British Heart Foundation (FS/13/43/30324 and (AA/18/2/34218) to PKi and LF), German Centre for Cardiovascular Research supported by the German Ministry of Education and Research (DZHK, via a grant to AFNET to PKi), and Leducq Foundation (14CVD01) to PKi. Declaration of Interests: LF has received institutional research grants and non-financial support from European Union, British Heart Foundation, Medical Research Council (UK), and DFG and Gilead. PKi has received additional support for research from the European Union, British Heart Foundation, Leducq Foundation, Medical Research Council (UK), and German Centre for Heart Research, from several drug and device companies active in atrial fibrillation, honoraria from several such companies. PKi and LF are listed as inventors on two patents held by University of Birmingham (Atrial Fibrillation Therapy WO 2015140571, Markers for Atrial Fibrillation WO 2016012783). PKa is an employee of Roche Diagnostics GmbH. AZ is an employee of Roche Diagnostics Intl. All other authors have reported no relationships relevant to the contents of this paper to disclose. Ethics Approval Statement: This study complied with the Declaration of Helsinki, was approved by the National Research Ethics Service Committee (IRAS ID 97753), and was sponsored by the University of Birmingham.

Objective The transition from paediatric to adult medicine involves risks of poor patient outcomes and of significant losses of patients to follow up. The research aimed to analyse the implementation in an initial cohort of patients of a new programme of transition to adult care based on a case management approach. Design A longitudinal study of the case management approach to transition, initiated in a university hospital in France in September 2016. Methods Patients with the endocrine or metabolic disease diagnosed during childhood and transferred to adult care were included. The transition programme includes three steps based on case management: liaising with paediatric services, personalising care pathways, and liaising with structures outside the hospital (general practitioners, agencies in the educational and social sector). Results The cohort included 500 patients, with malignant brain tumour (n = 56 (11%)), obesity (n = 55 (11%)), type 1 diabetes (n = 54 (11%)), or other disease (n = 335 (67%)). Their median age at transfer was 19, and the sex ratio was 0.5. At median 21 months of follow-up, 439 (88%) had a regular follow-up in or outside the hospital, 47 (9%) had irregular follow-up (absence at the last appointment or no appointment scheduled within the time recommended), 4 had stopped care on doctor’s advice, 4 had died, 3 had moved, and 3 had refused care. The programme involved 9615 case management actions; 7% of patients required more than 50 actions. Patients requiring most support were usually those affected by a rare genetic form of obesity. Conclusions Case managers successfully addressed the complex needs of patients. Over time, the cohort will provide unprecedented long-term outcome results for patients with various conditions who experienced this form of transition.

SUMMARYMetabolic disorders such as obesity are risk factors of atrial fibrillation, not only by sharing comorbidities but likely through their direct impact on atria, notably its adipogenicity. Here, we submitted mice that lack cardiac adipose tissue to a high fat diet and first studied the atrial metabolomic and lipidomic phenotypes using liquid chromatography-mass spectrometry. We found an increased consumption of free fatty acid by the beta-oxidation and an accumulation of long-chain lipids in atria of obese mice. Free fatty acid was the main substrate of mitochondrial respiration studied in the saponin-permeabilized atrial muscle. Conducted action potential recorded in atrial trabeculae was short, and ATP-sensitive potassium current was increased in perforated patch-clamp atrial myocytes of obese mice. There was histological and phenotypical evidence for an accumulation of adipose tissue in obese mice atria. Thus, an obesogenic diet transforms the energy metabolism, causes fat accumulation and induces electrical remodeling of atria myocardium.HIGHLIGHTS- Untargeted metabolomic and lipidomic analysis revealed that a high fat diet induces profound transformation of atrial energy metabolism with beta-oxidation activation and long-chain lipid accumulation.- Mitochondria respiration studied in atrial myocardial trabecula preferentially used Palmitoyl-CoA as energy substrate in obese mice.- Atria of obese mice become vulnerable to atrial fibrillation and show short action potential due to the activation of K-ATP dependent potassium current.- Adipocytes and fat molecular markers were detected in atria of obese mice together with an inflammatory profile consistence with a myocardial accumulation of fat.

Heart failure is the final common stage of most cardiopathies. Cardiomyocytes connect with others via their extremities by intercalated disk protein complexes. This planar and directional organization of myocytes is crucial for mechanical coupling and anisotropic conduction of the electric signal in the heart. One of the hallmarks of heart failure is alterations in the contact sites between cardiomyocytes. Yet no factor on its own is known to coordinate cardiomyocyte polarized organization. We report enhanced levels of an ubiquitine ligasePdzrn3in diseased hypertrophic human and mouse myocardium, which correlates with a loss of cardiomyocyte polarized elongation. We provide evidence thatPdzrn3has a causative role in heart failure. We found that cardiacPdzrn3deficiency protected against heart failure while over expression ofPdzrn3in mouse cardiomyocytes during the first weeks of life, impaired postnatal cardiomyocyte maturation leading to premature death. Our results reveal a novel signaling pathway that controls a genetic program essential for heart maturation and maintenance of overall geometry, as well as the contractile function of cardiomyocytes, and implicates PDZRN3 as a potential therapeutic target for the prevention of human heart failure.

Background Biomarkers reflecting different biological pathways have been associated with atrial fibrillation (AF). These discoveries motivate the consideration of a multiple biomarker model in the context of AF screening to improve detection. Objective We assessed a selection of clinical characteristics and biomarkers known to be associated with AF as established in literature, to identify a mechanism-based combination of markers for simplifying patient selection for screening. Methods and Results 1485 patients presenting acutely to hospital (median age [Q1, Q3] 69 [60, 78] years; 60% male; 45% with AF) with either diagnosed AF or ≥2 CHA2DS2-VASc risk factors (silent AF ruled out by 7-day ECG monitoring) were analysed. From EDTA plasma, 12 known cardiovascular biomarkers selected from published literature were quantified at a single centre on a high-throughput platform (Roche Diagnostics GMBH, DE). After adjustment for known confounders (age, sex, BMI, eGFR, heart failure, stroke/TIA, hypertension status), 6 biomarkers remained univariately associated with increased odds of AF (BMP10, ANG2, NTproBNP, IGFBP7, FGF23, CA125; see Figure). A model which simultaneously considered clinical characteristics and biomarkers was developed in a discovery cohort (n = 933) randomly sampled from all included patients (60:40 discovery:validation) and subsequently validated on the remaining patients (n = 552) using both logistic regression and machine learning methodologies for comparison. Using regression with backward statistical selection, an optimism-adjusted model of Age, Sex, BMI, BMP10, ANG2, and FGF23 was found to discriminate between patients with and without AF with an area under the ROC curve, AUC, of 0.743 [95% confidence interval, 0.712, 0.775], corroborated by machine learning (AUC 0.760 [95%CI 0.746, 0.764]). Performance was similar in the validation cohort for regression (AUC 0.719 [0.677, 0.762]) and machine learning (AUC 0.733 [95%CI 0.691, 0.775]). In a sensitivity analysis using biomarker quartiles instead of absolute values, an additional biomarker was selected: NTproBNP. Conclusion In our analysis of known markers of AF, a combination of 3 simple clinical characteristics (Age, Sex, BMI) and 3 biomarkers (BMP10, ANG2, and FGF23) robustly discriminated between patients with diagnosed AF and sinus rhythm patients with cardiovascular risks in both discovery and validation cohorts. Biomarkers implicate pathways related to inflammation (BMP10), fibrosis (FGF23) and hypoxia (ANG2), known to be associated with AF. Prospective studies can examine if AF screening with multiple biomarkers has the potential of identifying patients who could benefit from further ECG monitoring. Conflict of Interest None.

Many cardiac pathologies involve changes in tissue structure. Conventional analysis of structural features is extremely time-consuming and subject to observer bias. The possibility to determine spatial interrelations between these features is often not fully exploited. We developed a staining protocol and an ImageJ-based tool (JavaCyte) for automated histological analysis of cardiac structure, including quantification of cardiomyocyte size, overall and endomysial fibrosis, spatial patterns of endomysial fibrosis, fibroblast density, capillary density and capillary size. This automated analysis was compared to manual quantification in several well-characterized goat models of atrial fibrillation (AF). In addition, we tested inter-observer variability in atrial biopsies from the CATCH-ME consortium atrial tissue bank, with patients stratified by their cardiovascular risk profile for structural remodeling. We were able to reproduce previous manually derived histological findings in goat models for AF and AV block (AVB) using JavaCyte. Furthermore, strong correlation was found between manual and automated observations for myocyte count (r = 0.94, p p p p p

Aims Restoration of sarco/endoplasmic reticulum Ca2+ ATPase (SERCA2a) activity through gene transfer improved cardiac function in experimental and pilot studies in humans with heart failure. The AGENT-HF (NCT01966887) trial investigated the impact of adeno-associated virus (AAV1)/SERCA2a on ventricular remodelling using multimodality non-invasive cardiac imaging. Methods and results AGENT-HF was a single centre, randomized, double-blind, placebo-controlled trial in adult patients with NYHA class III–IV ischaemic or non-ischaemic heart failure and left ventricular ejection fraction ≤35%. Eligible patients were randomized to receive a single intracoronary infusion of either 1 × 1013 DNase-resistant particles of AAV1/SERCA2a or placebo. The primary endpoint was change in left ventricular end-systolic volume (LVESV), measured by cardiac computed tomography at 6 month follow-up. We planned to include 40 patients but the trial was terminated prematurely as the sponsor suspended further enrolment following neutral results of the CUPID-2 outcome trial. At the time of termination, nine patients were randomized with five patients infused with AAV1/SERCA2a and four with placebo. At 6 months, LVESV was increased in both groups compared with baseline: median (interquartile range) in AAV1/SERCA2a vs. placebo: 13 (13;14) mL vs. 3.5 (−36;36) mL, P = 0.74, with a mean difference between groups of 11.4 mL in favour of placebo. No safety issues were noted. Conclusion AGENT-HF failed to demonstrate any improvement in ventricular remodelling in response to AAV1/SERCA2a at the dose studied. However, because of premature termination, the study was underpowered to demonstrate an effect of AAV1/SERCA2a and these data should be interpreted with caution.

Background/Introduction Atrial fibrillation (AF) can be challenging to diagnose due to asymptomatic and paroxysmal presentation. Identifying prognostic factors of AF would elucidate potential mechanisms causing AF and refine screening for at risk patients. Purpose To identify the main predictors of AF and to develop a prognostic model for prevalent AF. Methods Data of 120 potential predictors were harmonised in individual patient data from 4 independent European studies. A three stage Delphi expert consensus process identified predictors based on clinical knowledge. The predictors were further reduced using statistical selection (backward elimination), and a logistic regression model was fitted. We calculated odds ratios (OR) for each of the selected predictors and evaluated model performance using the C-statistic. Results Overall, 2420 patients (mean [standard deviation] age = 62.7 [14.5] years, 35.6% female, 43.1% with AF) were included in the analysis. Thirty-one potential predictors identified from the Delphi process which had sufficient data across all datasets were modelled. Of these 14 were deemed prognostic in predicting AF (age, sex, BMI, height, hypertension, diabetes, history of coronary artery disease, left atrial volume, left ventricular end systolic diameter, abnormality on echo, tricuspid valve disease of at least moderate intensity, aldosterone-antagonists, beta-blockers and P2Y12 blockers; see Figure 1). There was a clear interaction between age and sex indicating that males are at higher risk than females early in life, while females are at increased risk of AF at older age (Figure 1). The risk prediction model combining these prognostic factors performed well (C-statistic 0.79; 95% CI 0.77–0.81). Figure 1. (a) Forest plot; (b) Interaction Conclusion(s) Our preliminary analysis identified important prognostic factors and a complex relationship between age and sex, which predicts prevalent AF, highlighting the different potential causes of AF in different patients. There is a clear need to validate these factors in external datasets and for further investigation into the molecular mechanism underlying these factors. Acknowledgement/Funding European Commission H2020 framework

Context and objectives Fibro-fatty infiltrations of the sub-epicardium is an important component of the atrial cardiomyopathy that paves the way of atrial fibrillation (AF). We previously reported that epicardium-derived cells (EPDCs) can be a source of adipocytes contributing to fat depot. Here, we examined if EPDCs can contribute also to the fibrotic remodeling of atrial subepicardium. Methods Surgical samples of human right atria were used for histological study and to harvest EPDCs. Clinical and histological data were analyzed using generalized linear models. A model atrial remodeling associated with ischemic heart failure was created in rats and in a genetic lineage tracing WT1CreERT2+/−/Rosa-tdT+/− mice to follow epicardial progenitor fate. Primary cell culture models were used to study EPDC differentiation. Flow cytometry assays and single cell sequencing were used to characterize subpopulations of EPDCs. Results In human atria, epicardium could be a thin cell monolayer or be thick in continuity with dense fibro-fatty infiltrates. The latter aspect predominated in old patient (>70 year) with valve mitral diseases or in AF, clinical conditions associated with an atrial remodeling. This was further established in rat and mouse experimental model of atrial remodeling characterized by a thickening of the epicardium observed one week following the onset of the cardiopathy and, then, followed by a dense fibrotic remodeling of the subepicardium. Cells co-expressing markers of epicardial progenitors and fibroblasts were detected in the subepicardium of diseased atria both in human and murine models. Moreover, a number of myo/fibroblasts from WT1CreERT2+/−/Rosa-tdT+/− mice were detected in diseased atria attesting of their epicardial origin. Distinct EPDC clusters were detected from cells harvested from epicardium whereas only PDGFRa-positive cells could differentiated into fibroblasts. Angiotensin-II induced the differentiation of only PDGFRa-positive cells into fibroblasts. Conclusions Activation of the epicardium is an early event during atrial remodeling that results in the triggering of differentiation of a subpopulation of epicardial progenitors into fibroblasts. Angiotensin-II is part of the signaling switch than regulate the fate of EPDC between fibroblastic or adipogenic lineage. Hence, epicardium by regulating the balance between adipose expansion and fibrosis accumulation appears as an important determinant of the atrial cardiomyopathy. Acknowledgement/Funding ANR-10-IAHU-05

Epicardium, the mesothelium covering the heart, is composed of multipotent cells and is reactivated following myocardial injury in adults. Herein, we provide evidence for activation of atrial epicardium in aged patients with diseased atria and in murine models of atrial remodeling. Epicardial activation contributed to fibro-fatty infiltration of sub-epicardium that contained a number of cells co-expressing markers of epicardial progenitors and fibroblasts. Indeed, using genetic lineage tracing of adult epicardium, we demonstrate the epicardial origin of fibroblasts within fibro-fatty infiltrates. A subpopulation of adult epicardial-derived cells (aEPDCs) expressing PDGFRα, niched in the sub-epicardium, were isolated and differentiated into myofibroblast in the presence of angiotensin-II. Furthermore, single cell RNA-seq analysis identified several clusters of aEPDCs and revealed transition from adipogenic to fibrogenic cells. In conclusion, a subset of aEPDCs, pre-programmed towards a specific cell fate, contributes to fibro-fatty infiltration of sub-epicardium of diseased atria.

The amount of adipose tissue that accumulates around the atria is associated with the risk, persistence, and severity of atrial fibrillation (AF). A strong body of clinical and experimental evidence indicates that this relationship is not an epiphenomenon but is the result of complex crosstalk between the adipose tissue and the neighbouring atrial myocardium. For instance, epicardial adipose tissue is a major source of adipokines, inflammatory cytokines, or reactive oxidative species, which can contribute to the fibrotic remodelling of the atrial myocardium. Fibro-fatty infiltrations of the subepicardium could also contribute to the functional disorganization of the atrial myocardium. The observation that obesity is associated with distinct structural and functional remodelling of the atria has opened new perspectives of treating AF substrate with aggressive risk factor management. Advances in cardiac imaging should lead to an improved ability to visualize myocardial fat depositions and to localize AF substrates.

Despite remarkable advances in antiarrhythmic drugs, ablation procedures, and stroke-prevention strategies, atrial fibrillation (AF) remains an important cause of death and disability in middle-aged and elderly individuals. Unstructured management of patients with AF sharply contrasts with our detailed, although incomplete, knowledge of the mechanisms that cause AF and its complications. Altered calcium homeostasis, atrial fibrosis and ageing, ion-channel dysfunction, autonomic imbalance, fat-cell infiltration, and oxidative stress, in addition to a susceptible genetic background, contribute to the promotion, maintenance, and progression of AF. However, clinical management of patients with AF is currently guided by stroke risk parameters, AF pattern, and symptoms. In response to this apparent disconnect between the known pathophysiology of AF and clinical management, we propose a roadmap to develop a set of clinical markers that reflect the major causes of AF in patients. Thereby, the insights into the mechanisms causing AF will be transformed into a format that can underpin future personalized strategies to prevent and treat AF, ultimately informing better patient care.

Background ECG late depolarization abnormality called epsilon wave (ɛ) is a major diagnostic criterion in ARVC/D patients according to Task Force. Objectives As ARVC/D induces progressive right ventricular enlargement, whether or not additional right precordial leads (X1, X2, V3R, V4R) may increase ECG diagnostic sensitivity at rest and/or during exercise has never been systematically investigated. Methods We studied 3 groups (G) of subjects. G1: 17 propositus patients with desmosomal mutation; G2: 17 first degree relative gene carriers; G3: 20 controls. All subjects were paired between groups for age, BSA, sex and sport practice. A 16 leads ECG including additional right precordial leads X1, X2, V3R, V4R was performed in each case both at rest and at peak stress. Recordings were amplified up to 100 mm/s and 40 mm/mV and retrospectively analyzed by two investigators blinded to patient status. Results All subjects reached a sustained power of similar strength (160 ± 50 W) on cycloergometer. At rest, by including standard (V1, V2) and additional right precordial leads, an ɛ was shown in 14/34 patients (41%, 7 in G1, 7 in G2). Among them 4 (12%, 2 in G1, 2 in G2) showed an ɛ exclusively in at least one additional lead. At peak stress, an ɛ was noticed in 19/34 patients (56%, 12 in G1, 7 in G2). Among them, 9 patients (26%, 7 in G1, 2 in G2) showed an ɛ exclusively in at least one additional lead. In other words, in 5/34 patients (15%, all in G1) not showing any ɛ at rest, peak stress unmasked a new ɛ, exclusively in at least one additional right precordial lead. No ɛ was detected in G3, neither at rest nor at stress. Conclusions By combining rest and stress test ECG, the use of additional right precordial leads X1, X2, V3R, V4R, enhances by 27% the detection of ɛ waves (major criterion), more often in propositus than in relatives, as compared to usual precordial derivations alone, and should be proposed for inclusion among ARVC/D Task Force criteria.

Background: Aging induces cardiac structural and functional changes linked to the increased deposition of extracellular matrix proteins, including OPN (osteopontin), conducing to progressive interstitial fibrosis. Although OPN is involved in various pathological conditions, its role in myocardial aging remains unknown. Methods: OPN deficient mice (OPN-/-) with their wild-type (WT) littermates were evaluated at 2 and 14 months of age in terms of cardiac structure, function, histology and key molecular markers. OPN expression was determined by reverse-transcription polymerase chain reaction, immunoblot and immunofluorescence. Luminex assays were performed to screen plasma samples for various cytokines/adipokines in addition to OPN. Similar explorations were conducted in aged WT mice after surgical removal of visceral adipose tissue (VAT) or treatment with a small-molecule OPN inhibitor agelastatin A. Primary WT fibroblasts were incubated with plasma from aged WT and OPN-/- mice, and evaluated for senescence (senescence-associated β-galactosidase and p16), as well as fibroblast activation markers (Acta2 and Fn1). Results: Plasma OPN levels increased in WT mice during aging, with VAT showing the strongest OPN induction contrasting with myocardium that did not express OPN. VAT removal in aged WT mice restored cardiac function and decreased myocardial fibrosis in addition to a substantial reduction of circulating OPN and transforming growth factor β levels. OPN deficiency provided a comparable protection against age-related cardiac fibrosis and dysfunction. Intriguingly, a strong induction of senescence in cardiac fibroblasts was observed in both VAT removal and OPN-/- mice. The addition of plasma from aged OPN-/- mice to cultures of primary cardiac fibroblasts induced senescence and reduced their activation (compared to aged WT plasma). Finally, Agelastatin A treatment of aged WT mice fully reversed age-related myocardial fibrosis and dysfunction. Conclusions: During aging, VAT represents the main source of OPN and alters heart structure and function via its profibrotic secretome. As a proof-of-concept, interventions targeting OPN, such as VAT removal and OPN deficiency, rescued the heart and induced a selective modulation of fibroblast senescence. Our work uncovers OPN’s role in the context of myocardial aging and proposes OPN as a potential new therapeutic target for a healthy cardiac aging.

There are major challenges ahead for clinicians treating patients with atrial fibrillation (AF). The population with AF is expected to expand considerably and yet, apart from anticoagulation, therapies used in AF have not been shown to consistently impact on mortality or reduce adverse cardiovascular events. New approaches to AF management, including the use of novel technologies and structured, integrated care, have the potential to enhance clinical phenotyping or result in better treatment selection and stratified therapy. Here, we report the outcomes of the 6th Consensus Conference of the Atrial Fibrillation Network (AFNET) and the European Heart Rhythm Association (EHRA), held at the European Society of Cardiology Heart House in Sophia Antipolis, France, 17-19 January 2017. Sixty-two global specialists in AF and 13 industry partners met to develop innovative solutions based on new approaches to screening and diagnosis, enhancing integration of AF care, developing clinical pathways for treating complex patients, improving stroke prevention strategies, and better patient selection for heart rate and rhythm control. Ultimately, these approaches can lead to better outcomes for patients with AF.

Purpose To determine whether left atrial (LA) strain quantification with cardiac magnetic resonance (MR) imaging feature tracking is associated with the severity of LA fibrofatty myocardial remodeling at histologic analysis. Materials and Methods This prospective case-control study was approved by the institutional review board. LA strain was evaluated with cardiac MR feature tracking between January 2014 and March 2015 in 13 consecutive patients (mean age, 61 years ± 19; nine male) with mitral regurgitation in the 24 hours before mitral valve surgery and 13 age- and sex-matched healthy control subjects. LA strain parameters were compared first between control subjects and patients and then according to atrial fibrillation and mitral regurgitation status. Associations between LA strain and histology of preoperative biopsies were reported by using receiver operating characteristic curve analysis and Spearman correlation. Results Peak longitudinal atrial strain (PLAS) was significantly lower in patients with mitral regurgitation than in healthy control subjects (P < .001). Increased LA remodeling was significantly related to altered LA strain, and the strongest association was found between PLAS and the degree of fibrofatty myocardial replacement at histologic analysis (r = -0.75, P = .017). LA end-diastolic volume was increased in patients with mitral regurgitation when compared with that in healthy volunteers (P < .001) because of volume overload; however, volume did not correlate with the histologic degree of LA fibrofatty replacement (r = -0.35, P = .330). Conclusion LA strain, especially PLAS, correlates strongly with the degree of fibrofatty replacement at histologic analysis. Such functional imaging biomarker in combination with LA volumetry could help to guide clinical decisions, since myocardial structural remodeling is a known morphologic substrate of LA dysfunction leading to atrial fibrillation with adverse outcome. © RSNA, 2017 Online supplemental material is available for this article.

Atrial fibrillation (AF) is the most common sustained arrhythmia in the general population. As an age-related arrhythmia AF is becoming a huge socio-economic burden for European healthcare systems. Despite significant progress in our understanding of the pathophysiology of AF, therapeutic strategies for AF have not changed substantially and the major challenges in the management of AF are still unmet. This lack of progress may be related to the multifactorial pathogenesis of atrial remodelling and AF that hampers the identification of causative pathophysiological alterations in individual patients. Also, again new mechanisms have been identified and the relative contribution of these mechanisms still has to be established. In November 2010, the European Union launched the large collaborative project EUTRAF (European Network of Translational Research in Atrial Fibrillation) to address these challenges. The main aims of EUTRAF are to study the main mechanisms of initiation and perpetuation of AF, to identify the molecular alterations underlying atrial remodelling, to develop markers allowing to monitor this processes, and suggest strategies to treat AF based on insights in newly defined disease mechanisms. This article reports on the objectives, the structure, and initial results of this network.

Purpose Atrial fibrillation (AF) has been associated with increased volumes of epicardial fat and atrial adipocyte accumulation. Underlying mechanisms are not well understood. This study aims to identify rapid atrial pacing (RAP)/AF-dependent changes in atrial adipocyte/adipositas-related gene expression (AARE). Methods Right atrial (RA) and adjacent epicardial adipose tissue (EAT) samples were obtained from 26 patients; 13 with AF, 13 in sinus rhythm (SR). Left atrial (LA) samples were obtained from 9 pigs (5 RAP, 4 sham-operated controls). AARE was analyzed using microarrays and RT-qPCR. The impact of diabetes/obesity on gene expression was additionally determined in RA samples (RAP ex vivo and controls) from 3 vs. 6months old ZDF rats. Results RAP in vivo of pigs resulted in substantial changes of AARE, with 66 genes being up- and 53 down-regulated on the mRNA level. Differential expression during adipocyte differentiation was confirmed using 3T3-L1 cells. In patients with AF (compared to SR), a comparable change in RA mRNA levels concerned a fraction of genes only ( RETN , IGF1 , HK2 , PYGM , LOX , and NR4A3 ). RA and EAT were affected by AF to a different extent. In patients, concomitant disease contributes to AARE changes. Conclusions RAP, and to lesser extent AF, provoke significant changes in atrial AARE. In chronic AF, activation of this gene panel is very likely mediated by AF itself, AF risk factors and concomitant diseases. This may facilitate the development of an AF substrate by increasing atrial ectopic fat and fat infiltration of the atrial myocardium.

Background Left atrium (LA) volumes and function are predictors of cardiovascular events. Because LA function cannot be assessed from cardiovascular magnetic resonance imaging (MRI) using the well-established left ventricular tagging techniques, we hypothesized that adequate feature tracking (FT) applied to conventional cine MRI data could characterize LA function accurately. Methods We studied 10 young (28 ± 7 years) and 10 elderly (64 ± 6 years) healthy subjects, as well as 20 patients with moderate to severe aortic valve stenosis (AVS; 73 ± 15 years, effective aortic valve area: 0.67 ± 0.36 cm2). MRI cine two-, three-, and four-chamber views were analyzed using a newly proposed FT method based on spatial correlation and endocardial detection resulting in: regional and global longitudinal strain and strain rate, radial motion fraction and relative velocity for the three LA motion phases including reservoir, conduit, and LA contraction. Results FT reliability was indicated by a good overlap between tracking results and manual LA endocardial borders, the low error for comparison against theoretical strains introduced in a synthetic phantom and the good inter-observer reproducibility (coefficient of variation

Aims Mutations in the KCNK3 gene, which encodes for an outward-rectifier K+ channel, have been identified in patients suffering from pulmonary arterial hypertension (PAH), and constitute the first described channelopathy in PAH. In human PAH and experimental pulmonary hypertension (PH), we demonstrated that KCNK3 expression and function are severely reduced in pulmonary vascular cells, promoting PH-like phenotype at the morphologic and haemodynamic levels. Since KCNK3 channel is also expressed in both the human and rodent heart, we aimed to elucidate the pathophysiological role of KCNK3 channel in right ventricular (RV) hypertrophy (RVH) related to PH. Methods and results Using whole-cell Patch-clamp technique, we demonstrated that KCNK3 is predominantly expressed in adult rat RV cardiomyocytes compared to the left ventricle cardiomyocytes and participates in the repolarizing phase of the RV action potential. We revealed a reduction in KCNK3 function prior to development of RVH and the rise of pulmonary vascular resistance. KCNK3 function is severely reduced in RV cardiomyocytes during the development of RVH in several rat models of PH (exposure to monocrotaline, chronic hypoxia, and Sugen/hypoxia) and chronic RV pressure overload (pulmonary artery banding). In experimental PH, we revealed a reduction in KCNK3 function before any rise in pulmonary vascular resistance and the development of RVH. KCNK3 mRNA level is also reduced in human RV tissues from PAH patients compared to non-PAH patients. In line with these findings, chronic inhibition of KCNK3 in rats with the specific inhibitor (A293) induces RV hypertrophy which is associated with the re-expression of foetal genes, RV fibrosis, RV inflammation, and subsequent loss of RV performance as assessed by echocardiography. Conclusion Our data indicate that loss of KCNK3 function and expression is a hallmark of the RV hypertrophy/dysfunction associated with PH.

International audience; The abundance of epicardial adipose tissue (EAT) is associated with atrial fibrillation (AF), the most frequent cardiac arrhythmia. However, both the origin and the factors involved in EAT expansion are unknown. Here, we found that adult human atrial epicardial cells were highly adipogenic through an epithelial-mesenchymal transition both in vitro and in vivo. In a genetic lineage tracing the WT1(CreERT2+/-)Rosa(tdT+/-)mouse model subjected to a high-fat diet, adipocytes of atrial EAT derived from a subset of epicardial progenitors. Atrial myocardium secretome induces the adipogenic differentiation of adult mesenchymal epicardium-derived cells by modulating the balance between mesenchymal Wingless-type Mouse Mammary Tumor Virus integration site family, member 10B (Wnt10b)/beta-catenin and adipogenic ERK/MAPK signaling pathways. The adipogenic property of the atrial secretome was enhanced in AF patients. The atrial natriuretic peptide secreted by atrial myocytes is a major adipogenic factor operating at a low concentration by binding to its natriuretic peptide receptor A (NPRA) receptor and, in turn, by activating a cGMP-dependent pathway. Hence, our data indicate cross-talk between EAT expansion and mechanical function of the atrial myocardium.

Atrial fibrillation (AF) is the most frequent cardiac arrhythmia in clinical practice. AF is often associated with profound functional and structural alterations of the atrial myocardium that compose its substrate. Recently, a relationship between the thickness of epicardial adipose tissue (EAT) and the incidence and severity of AF has been reported. Adipose tissue is a biologically active organ regulating the metabolism of neighbouring organs. It is also a major source of cytokines. In the heart, EAT is contiguous with the myocardium without fascia boundaries resulting in paracrine effects through the release of adipokines. Indeed, Activin A, which is produced in abundance by EAT during heart failure or diabetes, shows a marked fibrotic effect on the atrial myocardium. The infiltration of adipocytes into the atrial myocardium could also disorganize the depolarization wave front favouring micro re-entry circuits and local conduction block. Finally, EAT contains progenitor cells in abundance and therefore could be a source of myofibroblasts producing extracellular matrix. The study on the role played by adipose tissue in the pathogenesis of AF is just starting and is highly likely to uncover new biomarkers and therapeutic targets for AF.

Cardiac gene transfer is a powerful molecular tool to improve our understanding of the role of new proteins and mutants in cardiac pathophysiology. There is a need for a simple efficient myocardial gene delivery technique in order to study the physiological role of proteins in their native environment. Here we tested a new method of myocardial nonviral gene delivery, by using the combination of ultrasound energy (USE), liposomes and high pressure injections to the rat heart. Wistar rats were subjected to intra-myocardial injections of liposomes-DNA or siRNA mix. The heart was exposed after an inter-costal incision, and then injections were conducted between two sets of USE heart exposure. Ultrasound application resulted in much higher transfection efficiency (2% of left ventricle) than the liposomes-DNA alone (0.12% of left ventricle) as shown by the beta-galactosidase staining. The ultrasonic based liposomes-DNA delivery resulted in low inflammatory response, as well as in low cardiac fibrosis as shown by total collagen staining. Quantitative real time polymerase chain reaction (PCR) showed that the ultrasonic delivery resulted in cardiac specific transduction. Moreover, 23,906±2197 and 71,883±4065 calcium tolerant transfected cardiac myocytes were isolated following the delivery of a GFP plasmid or tagged siRNA, respectively. This was sufficient to perform single cell physiological measurements and biochemical experiments on homogenates. We developed an interesting safe method for local gene transfer in the heart using ultrasound and liposomes gene delivery. This method is particularly useful to study the effect of gene transfer on cardiac myocytes maintained in their normal environment in animal models.

The initiation and perpetuation of atrial fibrillation (AF) can be regarded as a complication of a progressive transformation of atrial structure and function. This transformation is the result of complex changes at the molecular, cellular and organ levels, leading to the profibrillation arrhythmic mechanisms in AF. Numerous individual and environmental factors are probably involved in this process. Therefore, progress in the diagnosis, prevention and treatment of AF requires highly integrative research from the benchtop to bedside and from specific signaling pathways and electrophysiological mechanisms to population-based studies. The European Network for Translational Research in Atrial Fibrillation was formed to provide this variety of expertise and has identified central research objectives for improvements in AF prevention and therapy.

Atrial fibrillation (AF) is the most frequent cardiac arrhythmia, which is characterized by a high risk of stroke due to increased thrombogenesis and thrombin formation in poorly contractile atria. In addition to its role in the coagulation cascade, thrombin has pleiotropic effects on cardiovascular system through the activation of protease activated receptor 1 (PAR 1). Here we examined whether thrombin contributes to the atrial myocardial remodeling that occurs during heart failure (HF) and whether direct thrombin inhibitors can prevent this remodeling process and AF progression. The study was conducted in a well established rat model of atrial dilation and susceptibility to AF associated with ischemic HF secondary to myocardial infarction (MI). The endogenous thrombin potential increased in plasma after MI, and remained elevated in HF rats. Treating MI rats for one or two months with direct thrombin inhibitors reduced the area and diameter of left atria. After two months, there was also a reduction of the duration of burst pacing induced AF and a downregulation of the expression of hypertrophic markers such as brain natriuretic peptide and β myosin heavy chain. The transcription factor NFATc3, known to be activated in cardiac hypertrophy, and the plasminogen activator inhibitor 1, which has both thrombotic and fibrogenic activities were also downregulated. Treating rats with a PAR 1 antagonist reproduced the effect of thrombin inhibitors. To study the role of thrombin on atrial myocardium in absence of other cardiac stimuli, atrial explant cultures were performed. In cultured atria, thrombin upregulated hypertrophic markers and plasminogen activator inhibitor 1 through PAR 1 and the Rho/Rho kinase pathway. These results indicate that thrombin is a potent hypertrophic factor for atrial myocardium; they point to chronic inhibition of thrombin/PAR1 pathways as a potential therapeutic option to prevent atrial remodeling and AF substrate formation. The author hereby declares no conflict of interest

Background No data on long-term outcomes of survivors of 2009 influenza A(H1N1) (A[H1N1])-associated ARDS are available. The objective of this study was to compare the 1-year outcomes of survivors of A(H1N1)-associated ARDS, according to use or no use of extracorporeal lung assist (ECLA), using its need as an ARDS severity surrogate. Methods Survivors of ARDS (12 with ECLA use vs 25 without, corresponding to 75% and 54% of the eligible patients for each group, respectively) selected from the Reseau Europeen de Ventilation Artificielle (REVA) registry had previously been healthy, with only pregnancy and/or moderate obesity (BMI ≤ 35 kg/m 2 ) as known risk factors for A(H1N1) infection. Lung function and morphology, health-related quality of life (HRQoL), and psychologic impairment were evaluated. Results At 1 year post-ICU discharge for the ECLA and no-ECLA groups, respectively, 50% and 40% reported significant exertion dyspnea, 83% and 64% had returned to work, and 75% and 64% had decreased diffusion capacity across the blood-gas barrier, despite their near-normal and similar lung function test results. For both groups, exercise test results showed diminished but comparable exercise capacities, with similar alveolar-arterial oxygen gradients at peak exercise, and CT scans showed minor abnormal findings. HRQoL assessed by the 36-Item Short-Form Health Survey was poorer for both groups than for a sex- and age-matched general population group, but without between-group differences. ECLA and no-ECLA group patients, respectively, had symptoms of anxiety (50% and 56%) and depression (28% and 28%) and were at risk for posttraumatic stress disorder (41% and 44%). Conclusions One year post-ICU discharge, a majority of survivors of A(H1N1)-associated ARDS had minor lung disabilities with diminished diffusion capacities across the blood-gas barrier, and most had psychologic impairment and poorer HRQoL than a sex- and age-matched general population group. ECLA and no-ECLA group patients had comparable outcomes. Trial registry ClinicalTrials.gov ; No.: NCT01271842 ; URL: www.clinicaltrials.gov

Atrial fibrillation (AF), the most common sustained cardiac arrhythmia in clinical practice, is often associated with progressive dilatation and remodeling of the atria which constitute the substrate of the arrhythmia. This atrial remodeling is characterized by complex structural and functional alterations of the atrial myocardium: short action potentials, heterogeneous refractory periods, dystrophic myocytes and interstitial fibrosis which act together to favor local conduction bloc, activation of ectopies and the forma-tion of microreentries of the electrical excitation. However, the underlying mechanisms of the AF substrate are not yet fully understood. The possibility of studying human atrial myocytes has led to the identification of ionic currents that contribute to the shortening of the action potential and refractory periods during AF. The down-regulation of the L-type calcium current plays a central role in this electrical remodeling. It results mainly from the dephosphorylation of calcium channels as the consequence of an excessive stimulation of atrial myocytes by neurohormones such as the atrial natriuretic factor. Abnormal trafficking and targeting of ion channels at the plasma membrane has emerged as mechanisms that can contribute to the abnormal electrical properties of the atria during AF. Fibrosis is the other feature of the AF substrate and it is favored by the atrial hemodynamic overload. Local activation of the renin-angiotensin system is involved in the extracellular matrix remodeling of the atrial myocardium. Thrombin that accumulates in dilated and fibrillating atria could be another important mediator of the myocardial structural alterations during AF. This peptide, by binding on its receptor PAR1, can modulate several signaling pathways regulating growth and survival of myocardial cells. Better understanding of pathogenic factors involved in the formation of the AF substrate is crucial for the identification of novel biomarkers and therapeutic targets that could be used to improve the diagnostic and treatment of AF.

Wound healing after angioplasty or stenting is associated with increased production of thrombin and the activation of protease-activated receptor 1 (PAR1). The aim of the present study was to examine the effects of a new selective PAR1 antagonist, 2-[5-oxo-5-(4-pyridin-2-ylpiperazin-1-yl)-penta-1,3-dienyl]-benzonitrile (F 16618), in restenosis and vascular smooth muscle cell (SMC) proliferation and migration using both in vivo and in vitro approaches. Daily oral administration of F 16618 inhibited the restenosis induced by balloon angioplasty on rat carotid artery in a dose-dependent manner. Furthermore, single intravenous administration of F 16618 during the angioplasty procedure was sufficient to protect the carotid artery against restenosis. In vitro, F 16618 inhibited the growth of human aortic SMCs in a concentration-dependent manner with maximal effects at 10 μM. At that concentration, F 16618 also prevented thrombin-mediated SMC migration. In vivo, oral and intravenous F 16618 treatments reduced by 30 and 50% the expression of the inflammatory cytokine tumor necrosis factor α (TNFα) 24 h after angioplasty. However, only acute intravenous administration prevented the induction of matrix metalloproteinase 7 expression. In contrast, F 16618 treatments had no effect on early SMC de-differentiation and transcription of monocyte chemoattractant protein-1 and interleukin-6 and late re-endothelialization of injured arteries. Furthermore, F 16618 compensated for the carotid endothelium loss by inhibiting PAR1-mediated contraction. Altogether, these data demonstrate that PAR1 antagonists such as F 16618 are a highly effective treatment of restenosis after vascular injury, by inhibition of TNFα, matrix metalloproteinase 7, and SMC migration and proliferation in addition to an antithrombotic effect.

Accumulation of atrial adipose tissue is associated with atrial fibrillation (AF). However, the underlying mechanisms remain poorly understood. We examined the relationship between the characteristics of fatty infiltrates of the atrial myocardium and the history of AF.Atrial samples, collected in 92 patients during cardiac surgery and in a sheep model of persistent AF, were subjected to a detailed histological analysis. In sections of human right atrial samples, subepicardial fatty infiltrations were commonly observed in the majority of patients. A clear difference in the appearance and fibrotic content of these fatty infiltrations was observed. Fibro-fatty infiltrates predominated in patients with permanent AF (no AF: 37 ± 24% vs. paroxysmal AF: 50 ± 21% vs. permanent AF: 64 ± 23%, P0.001). An inverse correlation between fibrotic remodelling and the amount of subepicardial adipose tissue suggested the progressive fibrosis of fatty infiltrates with permanent AF. This hypothesis was tested in a sheep model of AF. In AF sheep, an increased accumulation of peri-atrial fat depot was observed on cardiac magnetic resonance imaging and dense fibro-fatty infiltrations predominated in the left atria of AF sheep. Cellular inflammation, mainly consisting of functional cytotoxic T lymphocytes, was observed together with adipocyte cell death in human atria.Atrial fibrillation is associated with the fibrosis of subepicardial fatty infiltrates, a process in which cytotoxic lymphocytes might be involved. This remodelling of the atrial subepicardium could contribute to structural remodelling forming a substrate for AF.

International audience; Myotonic dystrophy type 1 (DM1) is the most common neuromuscular disorder and is associated with cardiac conduction defects. However, the mechanisms of cardiac arrhythmias in DM1 are unknown. We tested the hypothesis that abnormalities in the cardiac sodium current (INa) are involved, and used a transgenic mouse model reproducing the expression of triplet expansion observed in DM1 (DMSXL mouse). The injection of the class-I antiarrhythmic agent flecainide induced prominent conduction abnormalities and significantly lowered the radial tissular velocities and strain rate in DMSXL mice compared to WT. These abnormalities were more pronounced in 8-month-old mice than in 3-month-old mice. Ventricular action potentials recorded by standard glass microelectrode technique exhibited a lower maximum upstroke velocity [dV/dt]max in DMSXL. This decreased [dV/dt]max was associated with a 1.7 fold faster inactivation of INa in DMSXL myocytes measured by the whole-cell patch-clamp technique. Finally in the DMSXL mouse, no mutation in the SCN5A gene was detected and neither cardiac fibrosis nor abnormalities of expression of the sodium channel protein were observed. Therefore, alterations in the sodium current markedly contributed to electrical conduction block in DM1. This result should guide pharmaceutical and clinical research towards better therapy for the cardiac arrhythmias associated with DM1.

International audience; Aims Human epicardial adipose tissue (EAT) is a visceral and perivascular fat that has been shown to act locally on myocardium, atria, and coronary arteries. Its abundance has been linked to coronary artery disease (CAD) and atrial fibrillation. However, its physiological function remains highly debated. The aim of this study was to determine a specific EAT transcriptomic signature, depending on its anatomical peri-atrial (PA), peri-ventricular (PV), or peri-coronary location. Methods and results Samples of EAT and thoracic subcutaneous fat, obtained from 41 patients paired for cardiovascular risk factors, CAD, and atrial fibrillation were analysed using a pangenomic approach. We found 2728 significantly up-regulated genes in the EAT vs. subcutaneous fat with 400 genes being common between PA, PV, and peri-coronary EAT. These common genes were related to extracellular matrix remodelling, inflammation, infection, and thrombosis pathways. Omentin (ITLN1) was the most up-regulated gene and secreted adipokine in EAT (fold-change >12, P

Epicardial adipose tissue (EAT) is recognized as potentially involved in the pathophysiology of atrial fibrillation (AF). In addition to EAT fatty infiltration commonly observed in myocardium might be also associated with the substrate for AF. We conducted a histological study in 93 human right atrial samples obtained during cardiac surgery and in a sheep model of long-term persistent AF (PAF) induced by atrial tachypacing (24±8 weeks) (15 PAF, 11 SR). Upon Sirius Red and Harris Haematoxylin staining, human atria showed various degree of fatty infiltration starting from the epicardium (rarely from vessels), which could be associated with various degree of subepicardial fibrosis realizing in some cases a true fibro-fatty infiltration. The extent of remodeled epicardium was assessed as % of infiltrating and fibrotic epicardium±adipose tissue to total epicardial length; 44±26 % of the epicardium was remodeled. A multiple regression model (including history of AF, % adipose tissue, age, BodyMassIndex, coronary artery bypass, ejection fraction) significantly predicted the percentage of epicardial remodeling (R=0.501, p=0.003). Only AF and the % adipose tissue were significant predictors (respectively β=0.27;p=0.016 and β=-0.267;p=0.016). To further analyze the relationship between fibro-fatty infiltration and AF, a histological study was performed in LA specimen of a sheep PAF model which revealed 4 grades of subepicardial infiltrates from pure fatty to dense fibro-fatty infiltration (fig A). EAT infiltrates (316) were graded, demonstrating a shift towards more severe grades in the AF group p Conclusion: , AF is associated with the transformation of fat into fibro-fatty infiltrations suggesting that the subepicardial adipose tissue plays a role in the atrial fibrotic remodeling.

OBJECTIVES The purpose of this study was to determine the pathogenic factors and molecular mechanisms involved in fibrosis of the atria. BACKGROUND Fibrosis is an important component of the pathophysiology of atrial fibrillation, especially when the arrhythmia is associated with heart failure (HF) or atrial dilation. METHODS We used a rat model of myocardial infarction (MI) complicated by various degrees of left ventricular dysfunction and atrial dilation to study fibrosis and matrix metalloproteinase (MMP) activity in the left atrial (LA) myocardium by means of histologic, Western blot, zymographic, and immunohistologic techniques. RESULTS Three months after surgical ligature of the left coronary artery, 27 rats had a large MI, 12 were in mild HF, and 15 in severe HF. Both groups had LA enlargement at the echocardiography. Masson's trichrome and picrosirius staining of tissue sections revealed marked fibrosis at the periphery of trabeculae and also surrounding myolytic myocytes, in both mild and severe HF. In mild HF, the activity and expression of the matrilysin MMP-7 were increased (122%), whereas in severe HF, both MMP-7 (211%) and the gelatinase MMP-2 (187%) were up-regulated. There were no changes in the expression or activity of MMP inhibitors, TIMP-1, -2, and -4. Immunostaining of cryosections showed that MMP-2 was present in the interstitial spaces, whereas MMP-7 accumulated in myolytic myocytes. CONCLUSIONS Hemodynamic overload of the atria is an important pathogenic factor of fibrosis; MMP-7 appears to be involved in the early stage of this tissue remodeling process.

International audience; Background: In the diabetic heart the β-adrenergic response is altered partly by down-regulation of the β1-adrenoceptor, reducing its positive inotropic effect and up-regulation of the β3-adrenoceptor, increasing its negative inotropic effect. Statins have clinical benefits on morbidity and mortality in diabetic patients which are attributed to their “pleiotropic” effects. The objective of our study was to investigate the role of statin treatment on β-adrenergic dysfunction in diabetic rat cardiomyocytes.Methods: β-adrenergic responses were investigated in vivo (echocardiography) and ex vivo (left ventricular papillary muscles) in healthy and streptozotocin-induced diabetic rats, who were pre-treated or not by oral atorvastatin over 15 days (50 mg.kg-1.day-1). Micro-array analysis and immunoblotting were performed in left ventricular homogenates. Data are presented as mean percentage of baseline ± SD.Results: Atorvastatin restored the impaired positive inotropic effect of β-adrenergic stimulation in diabetic hearts compared with healthy hearts both in vivo and ex vivo but did not suppress the diastolic dysfunction of diabetes. Atorvastatin changed the RNA expression of 9 genes in the β-adrenergic pathway and corrected the protein expression of β1-adrenoceptor and β1/β3-adrenoceptor ratio, and multidrug resistance protein 4 (MRP4). Nitric oxide synthase (NOS) inhibition abolished the beneficial effects of atorvastatin on the β-adrenoceptor response.Conclusions: Atorvastatin restored the positive inotropic effect of the β-adrenoceptor stimulation in diabetic cardiomyopathy. This effect is mediated by multiple modifications in expression of proteins in the β-adrenergic signaling pathway, particularly through the NOS pathway.

Functional overreaching (F-OR) induced by heavy load endurance training programs has been associated with reduced heart rate values both at rest and during exercise. Because this phenomenon may reflect an impairment of cardiac response, this research was conducted to test this hypothesis. Thirty-five experienced male triathletes were tested (11 control and 24 overload subjects) before overloading (Pre), immediately after overloading (Mid), and after a 2-wk taper period (Post). Physiological responses were assessed during an incremental cycling protocol to volitional exhaustion, including catecholamines release, oxygen uptake (V̇o2), arteriovenous O2 difference, cardiac output (Q̇), and systolic (SBP) and diastolic blood pressure (DBP). Twelve subjects of the overload group developed signs of F-OR at Mid (decreased performance with concomitant high perceived fatigue), while 12 others did not [acute fatigue group (AF)]. V̇o2max was reduced only in F-OR subjects at Mid. Lower Q̇ and SBP values with greater arteriovenous O2 difference were reported in F-OR subjects at all exercising intensities, while no significant change was observed in the control and AF groups. A concomitant decrease in epinephrine excretion was reported only in the F-OR group. All values returned to baseline at Post. Following an overload endurance training program leading to F-OR, the cardiac response to exhaustive exercise is transiently impaired, possibly due to reduced epinephrine excretion. This finding is likely to explain the complex process of underperformance syndrome experienced by F-OR endurance athletes during heavy load programs.

Background Mutations in the SCN5A gene, encoding the α subunit of the cardiac Na + channel, Na v 1.5, can result in several life-threatening arrhythmias. Objective To characterize a distal truncating SCN5A mutation, R1860Gfs*12, identified in a family with different phenotypes including sick sinus syndrome, atrial fibrillation (AF), atrial flutter, and atrioventricular block. Methods Patch-clamp and biochemical analyses were performed in human embryonic kidney 293 cells transfected with wild-type (WT) and/or mutant channels. Results The mutant channel expressed alone caused a 70% reduction in inward sodium current (I Na ) density compared to WT currents, which was consistent with its partial proteasomal degradation. It also led to a negative shift of steady-state inactivation and to a persistent current. When mimicking the heterozygous state of the patients by coexpressing WT and R1860Gfs*12 channels, the biophysical properties of I Na were still altered and the mutant channel α subunits still interacted with the WT channels. Since the proband developed paroxysmal AF at a young age, we screened 17 polymorphisms associated with AF risk in this family and showed that the proband carries at-risk polymorphisms upstream of PITX2 , a gene widely associated with AF development. In addition, when mimicking the difference in resting membrane potentials between cardiac atria and ventricles in human embryonic kidney 293 cells or when using computer model simulations, R1860Gfs*12 induced a more drastic decrease in I Na at the atrial potential. Conclusion We have identified a distal truncated SCN5A mutant associated with gain- and loss-of-function effects, leading to sick sinus syndrome and atrial arrhythmias. A constitutively higher susceptibility to arrhythmias of atrial tissues and genetic variability could explain the complex phenotype observed in this family.

Atrial fibrillation (AF) can be triggered by myocardial infarction (MI) or heart failure (HF). The blockade of the ultra-rapid outward rectifier potassium current IKur is an attractive goal to control the repolarization of cardiac atria. F17727 was evaluated in patch-clamp assays, in rat post-MI and in HF in rabbit and dog. F17727 blocked human IKur (IC50=1.8µM) and did not impact IKr, IKs, INa. In rats 2 months after coronary artery ligation, the left atria surface increased by 50-150%; transœsophagial electrical bursts induced AF which lasted 5-21s in vehicle-MI rats but lower than 5s in sham. An acute iv treatment by F17727 (2.5 and 5mg/kg) significantly reduced AF induction at the level of sham. In rabbits, 2-week left atria tachypacing up to 1200bpm triggered electrical and structural remodeling; electrical stimuli induced AF. An acute iv treatment by F17727 (racemate, 2.5mg/kg) reduced AF (mean spontaneous and induced AF duration of 4±4s and 82±49s) when compared to vehicle (165±75s and 261±45s resp...

Cardiomyocyte calcium homeostasia is essential to cardiomyocyte excitation and contraction. Calcium enters through voltage-dependant calcium channels (ICal) and activates sarcoplasmic reticulum (SR) membrane ryanodine receptors (RyRs) triggering a massive release of stored Ca2+ essential to actine-myosine cross-bridging and sarcomere shortening. Calcium is than reabsorbed by the SR through SERCA and expelled from the cell by sodium/calcium exchanger (NCX). Some acquired heart diseases cause calcium homeostasia dysfunction: (1) heart failure causes reduced SERCA calcium uptake and porous RyRs. The ensuing calcium overload leads to NCX over-activation and arrhythmia triggering “late after-depolarizations”-generating cytosolic sodium increase. (2) Bordering infarct zones have disorganized RyR coupling and reduced SERCA activity responsible for potentially pro-arrhythmogenic situations. (3) Atrial fibrillation, through lessened ICal activity, leads to shortened action potential and contractile dysfunction. Some congenital heart diseases stem from abnormalities in the expression of calcium homeostasia agents: Some forms of long QT syndromes are linked to slow inactivation of ICal and persistent phase 2 plateau, with a heightened risk of ventricular arrhythmia. Heterogeneous distribution of Ito current potassium channel amplified by failures in the inward ICal depolarizing currents creates an epicardial-endocardial repolarization gradient also responsible for some forms of Brugada and short QT syndromes. Calcium leaks due to RyR abnormalities or of its regulation proteins can activate LAD triggering NCX over activation causing Polymorphic ventricular tachycardia. Some congenital atrial ventricular blocks are caused by impeded calcium channel activities of foetus hearts due to anti-bodies of lupus-affected mothers.

Atrial fibrillation (AF) is a common complication of heart failure. The aim of the present study was to investigate the effects of a new pure docosahexaenoic acid derivative called F 16915 in experimental models of heart failure-induced atria dysfunction. The atrial dysfunction-induced AF was investigated (1) in a dog model of tachypacing-induced congestive heart failure and (2) in a rat model of heart failure induced by occlusion of left descending coronary artery and 2 months reperfusion. F 16915 (5 g/day for 4 weeks) significantly reduced the mean duration of AF induced by burst pacing in the dog model (989 +/- 111 s in the vehicle group to 79 +/- 59 s with F 16915, P < 0.01). This dose of F 16915 also significantly reduced the incidence of sustained AF (5/5 dogs in the vehicle group versus 1/5 with F 16915, P < 0.05). In the rat model, the percentage of shortening fraction in the F 16915 group (100 mg/kg p.o. daily) was significantly restored after 2 months (32.6 +/- 7.4 %, n = 9 vs 17.6 +/- 3.4 %, n = 9 in the vehicle group, P < 0.01). F 16915 also reduced the de-phosphorylation of connexin43 from atria tissue. The present results show that treatment with F 16915 reduced the heart dilation, resynchronized the gap junction activity, and reduced the AF duration in models of heart failure. Thus, F 16915 constitutes a promising new drug as upstream therapy for the treatment of AF in patients with heart failure.

OBJECTIVES The aim of the present study was to determine if myocytes can die by apoptosis in fibrillating and dilated human atria. BACKGROUND The cellular remodeling that occurs during atrial fibrillation (AF) may reflect a degree of dedifferentiation of the atrial myocardium, a process that may be reversible. METHODS We examined human right atrial myocardium specimens (n = 50) for the presence of apoptotic myocytes. We used immunohistochemical and Western blotting analysis to examine the expression of a final effector of programmed cell death, caspase-3 (CASP-3) and of regulatory proteins from the BCL-2 family. RESULTS Sections from atria in AF contained a high percentage of large myocytes with a disrupted sarcomeric apparatus replaced by glycogen granules (64.4 ± 6.3% vs. 12.2 ± 5.8%). These abnormal myocytes, which also predominated in atria from hearts with decreased left ventricular ejection fraction (42.3 ± 10.1%), contained large nuclei, most of which were TUNEL positive, indicating a degree of DNA breakage. None of these abnormal myocytes expressed the proliferative antigen Ki-67. A small percentage of the enlarged nuclei (4.2 ± 0.8%) contained condensed chromatin and were strongly TUNEL positive. Both the pro- and activated forms of CASP-3 were detected in diseased myocardial samples, which also showed stronger CASP-3 expression than controls. Expression of the antiapoptotic BCL-2 protein was decreased in diseased atria, whereas that of the proapoptotic BAX protein remained unchanged. CONCLUSIONS In fibrillating and dilated atria, apoptotic death of myocytes with myolysis contributes to cellular remodeling, which may not be entirely reversible.

The risk of developing atrial fibrillation (AF) is increased in obese patients. This has been well established by several epidemiological studies. For instance, the long-term follow-up of the Framingham Heart Study reported that every unit increase in body mass index (BMI) raises AF risk by 4% [(1