bluuebunny/crossref_metadata_2025 huggingface dataset:
Factors associated with major adverse cardiovascular events among Arabs and Jews in Israel: a population-based cohort study
Authors: Kalter-Leibovici O , Agay N , Benderly M , Abu-Saad K , Kimron L , Ziv A
Published: October 2024

Introduction

Arabs in Israel are at higher risk of cardiovascular mortality; however, there are no data on factors associated with disparities in major adverse cardiovascular events (MACE) incidence among Arabs and Jews.

Purpose

To study factors associated with MACE incidence among Arabs and Jews in Israel.

Materials and Methods

This is a cohort study carried out in urban localities of Hadera District in Israel. The study sample included 1,100 participants, stratified in equal numbers by ethnicity (Arab and Jewish), sex, and five 10-year age group (mean age: 51 years, range: 26-77). Data collected via in-person interviews conducted between 2001-2006 included socio-demographic characteristics and health-related behaviors (smoking, physical activity, and diet). Data on baseline diabetes, hypertension, dyslipidemia, and cardiovascular disease were collected from interviews and participants' health records. Information on MACE occurring during follow-up was determined based on hospital discharge diagnoses, recorded in the Israel Ministry of Health hospital admission database. MACE was defined as an acute coronary, cerebrovascular or peripheral artery event, or invasive procedure for its treatment. Factors associated with the time to first MACE were tested in Cox proportional hazards models. Sensitivity analysis included death from any cause as a competing risk, using Fine and Gray’s model.

Results

During a median follow-up period of 20 years, there were 221 first MACE during follow-up; 91 among Jewish participants and 130 among Arab participants, hazard ratio (HR) Arab vs. Jewish: 1.59; 95% confidence interval (CI) 1.22, 2.09, adjusted for age and sex. Arab ethnicity was no longer significantly associated with MACE, following further adjustment for years of education; HR: 1.32; 95%CI 0.95, 1.84. The HR (95%CI) associated with ethnicity after additional adjustment for health-related behaviors, body weight and baseline chronic morbidity was 1.02 (95%CI: 0.73, 1.42). A subgroup analysis among 950 participants, who did not have a baseline diagnosis of cardiovascular disease, produced similar results. These participants experience 147 first MACE during follow-up. The age and sex-adjusted HR (95%CI) associated with Arab ethnicity was 1.72 (1.23, 2.40), compared to 1.08 (0.72, 1.64) in a fully adjusted model. Competing risk analysis did not materially change these results.

Conclusions

Arabs in Israel are at higher risk for MACE. Lower socioeconomic position and higher prevalence of unhealthy lifestyle and cardiovascular risk factors may explain the excess risk of MACE among Arabs. Improving cardiovascular health among Arabs should consider social determinants of health and cultural aspects.
Global burden of high-risk cardiovascular patients without prior myocardial infarction or stroke: VESALIUS-REAL - preliminary results from Germany
Authors: Chan Q , Ochs A , Dhalwani N , O Kelly J , Duxbury M , Neasham D , Shanno E , Maskell J , Paiva Da Silva Lima G , Avcil S , Chui C , Budoff M , Laufs U , Cegla J , Wong I
Published: October 2024

Background

Clinical trial data has shown that low-density lipoprotein cholesterol (LDL-C) lowering with PCSK9-inhibitors reduced cardiovascular (CV) endpoints in patients with established atherosclerotic cardiovascular disease (ASCVD). Intervening early in patients with high CV risk prior to myocardial infarction (MI) or stroke, while perceived to be less urgent, may result in larger public health benefits. The effect of evolocumab in this population is investigated in a phase-3 clinical trial (NCT03872401: Effect of Evolocumab in Patients at High Cardiovascular Risk Without Prior Myocardial Infarction or Stroke [VESALIUS-CV]). This observational study examines the global burden of VESALIUS-CV-like population in the REAL-World ("VESALIUS-REAL") in eight regions (Germany, Hong Kong, Japan, South Korea, Sweden, Taiwan, UK and US) using a unified protocol across multiple databases. The multi-database study is ongoing, and the current analysis focuses on characteristics of VESALIUS-CV like patients in Germany.

Methods

Eligibility criteria are shown in the Figure. Data were obtained from the German Disease Analyzer (2013-2022), a representative claims database from Germany. Criteria were aligned with the clinical trial, and real-world definitions were created using diagnosis and procedure codes.

Results

A total of 195,621 patients were included in the present analysis (Table). At baseline, the median age was 71 years (Q1-Q3: 64-79) and 48% of patients were women. Approximately two-thirds of patients had atherosclerosis (48.5% had coronary artery disease, 13.8% had cerebrovascular disease and 13.8% had peripheral artery disease) and 33.7% had high-risk diabetes mellitus (DM), defined as DM with microvascular complications or chronic insulin use. The median baseline LDL-C was 139 (Q1-Q3: 113-166) mg/dL [3.6 (2.9 - 4.3) mmol/L] and 110,867 patients (57%) had an LDL-C ≥130 mg/dL (≥3.4 mmol/L). Overall, 69% of VESALIUS-CV like patients were receiving no background LLT.

Conclusions

Despite having atherosclerosis and/or high-risk DM and LDL-C exceeding target thresholds, a large proportion of VESALIUS-CV like patients were not on any LLT in Germany. Final results from VESALIUS-REAL will be able to report if this treatment gap is also observed in other countries.
Reassessing pharmacological trials with positive composite outcome driven by reduction in nonfatal myocardial infarction: a systematic review and meta-analysis
Authors: Sun H , Li L , Yang Q , Zhou X
Published: October 2024

Background

Nonfatal myocardial infarction (MI) inadequately predicts cardiovascular and all-cause mortality in contemporary RCTs. The perceived efficacy of new treatments could be overstated if positivity in composite outcome is primarily attributed to reduction in nonfatal MI.

Purpose

This study seeks to assess pharmacological RCTs with positive results influenced by nonfatal MI reductions and scrutinize the associated clinical guideline recommendations.

Methods

Pharmacological RCTs focusing on mortality and nonfatal MI published from 2000 to 2023 were searched in PubMed and Web of Science, with subsequent citation tracking in Web of Science and Google Scholar for candidate RCTs referenced in English-language clinical guidelines. Main outcomes included: 1) relative risk reduction in the composite outcome excluding nonfatal MI; 2) relevant guideline recommendations based on the supporting trials. The impact of nonfatal MI on composite outcome was assessed by using the leave-one-out method. Meta-analysis utilized a random effects model.

Results

After reviewing 19,825 records, we identified 8 RCTs where positive composite outcomes were driven by reductions in non-fatal MI (Figure 1A). These were divided into three therapeutic groups: anti-thrombotic (3 trials), lipid-lowering (3 trials), and anti-inflammatory (2 trials). In guidelines citing these RCTs, lipid-lowering trials (IMPROVE-IT, FOURIER, CLEAR Outcomes; the latter has not been cited yet; a total of 60 recommendations across 17 guidelines; Figure 2) outperformed anti-thrombotic (27 recommendations across 15 guidelines) and anti-inflammatory trials (3 recommendations in 3 guidelines), with significantly higher Class I (45%) and IIa (33.3%) recommendations, and fewer Class IIb (21.7%) recommendations, compared to anti-thrombotic (0%, 22.2%, 66.7%) and anti-inflammatory (0%, 0%, 100%) trials (Figure 1B). In a meta-analysis of all trials on combined intensive lipid-lowering on top of maximally-tolerated statins (the ODYSSEY OUTCOMES, IMPROVE-IT, and FOURIER trials), the exclusion of nonfatal MI data sustained the negative findings (HR 0.94, 95% CI: 0.88 to 1.01).

Conclusions

Despite the overestimation of efficacy in certain anti-thrombotic, anti-inflammatory, and lipid-lowering RCTs due to the inclusion of nonfatal MI, lipid-lowering trials, particularly those supporting combined intensive therapy with statins, still receive disproportionately favorable guideline recommendations. This underscores the need to reassess the strength of evidence for these trials in guidelines, advocating for a more equitable and evidence-based approach to cardiovascular guideline recommendations.
Indoxyl sulfate deteriorates cardiac electrophysiological function and increases the likelihood of arrhythmias in human ventricular myocardial cells
Authors: Pfeuffer O , Koertl T , Hankowitz N , Riedl D , Stengel L , Maier L S , Streckfuss-Boemeke K , Sossalla S
Published: October 2024

Background

Approximately 10% of the world population is suffering from chronic kidney disease (CKD). In these patients, an increased mortality and morbidity, predominantly derived from cardiovascular diseases, can be observed.

Purpose

The impact of CKD on myocardial cell mechanisms is only partially understood. This study aims to investigate the impact of the uremic toxin indoxyl sulfate (IS) on human ventricular myocardial cells.

Methods

Human left ventricular myocardium was obtained from patients with severe aortic stenosis who underwent surgical valve replacement. In addition, human ventricular induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) obtained from donors without any known cardiovascular disease were used. Before measurements were conducted, isolated single primary human cardiomyocytes were incubated for 15 minutes with an IS concentration of 120 mg/l (highest measured IS-level in dialysis patients). Further, iPSC-CM were incubated for one week with IS 45 mg/l and IS 120 mg/l, with IS 45 mg/l representing the average dialysis patients IS-level. Epifluorescence and confocal microscopy were performed to investigate Ca2+hemostasis and patch clamp measurements assessed cardiac action potentials and the occurrence of arrythmias.

Results

Experiments were conducted using human ventricular myocardium from 10 patients with a mean left ventricular ejection fraction of 51.5±12% and a mean glomerular filtration rate of 66.3±22.7 ml/min. Compared to the control group, higher systolic Ca2+ transient amplitudes were observed after acute incubation with IS 120 mg/l in single primary human cardiomyocytes. Measurements after 7 days of IS incubation (IS 45 mg/l and 120 mg/l) in iPSC-CM confirmed these previous results by showing higher Ca2+ transient amplitudes (Figure 1). Additionally, cells showed a diminished diastolic Ca2+ level compared to controls. An increased Ca2+ spark frequency in IS-treated iPSC-CM versus the control group was unveiled by confocal microscopy measurements. While an increased number of delayed afterdepolarizations could be observed, IS seemed to have no effect on cardiac action potential duration in iPSC-CM.

Conclusion

For the first time, we could reveal the impact of IS on cellular electrophysiology of primary human myocardium cells and iPSC-CM and identify IS as a proarrhythmogenic trigger. Therefore, IS may be of relevance in explaining the high cardiovascular mortality in end-stage CKD patients. To improve the outcome of these patients, new therapeutic approaches will be necessary since IS, a protein-bound uremic toxin, is only insufficiently removed by dialysis.
The H222P-Lamin mutation induces heart failure via impaired mitochondrial calcium uptake in human cardiac laminopathy
Authors: Hulot J , Seguret M , Jouve C , Deshayes L , Ruiz-Velasco A , Pereira C , Wahbi K , Muchir A , Bonne G
Published: October 2024

Background

Mutations in LMNA gene, which encodes lamins A/C, cause a variety of diseases, called laminopathies. Some mutations are particularly associated with the occurrence of dilated cardiomyopathy. The pathological mechanisms are still unclear limiting the development of specific therapies.

Purpose

Here, we focused on a mutation in the LMNA gene (c.665A>C, p.His222Pro), associated with the Emery-Dreyfuss Muscular dystrophy. We used LMNA H222P mutant human induced pluripotent stem cells (hiPSCs) and CRISPR/Cas9 corrected isogenic control hiPSCs to better characterize the cardiac phenotype associated with the mutation.

Methods

LMNA H222P mutant and the isogenic control hiPSCs clones were differentiated into cardiomyocytes (CMs). Immunofluorescence staining was performed on hiPSC-CMs to quantify their sarcomere organization (SarcOrgScore) using a Matlab code. Ring-shaped cardiac organoids were generated to compare the contractile properties of the two clones. Calcium transients in mutant and corrected hiPSC-CMs were measured by live confocal imaging. Mitochondrial respiration was measured by Seahorse.

Results

hiPSC-CMs were generated from the LMNA H222P mutant and the corrected hiPSCs with no difference in the differentiation yield (proportion of troponin-positive cells: 92.74% for LMNA H222P vs. 89.38% for Ctrl-iso1, p=0.1038). The nuclei of LMNA H222P mutant hiPSC-CMs showed morphological abnormalities, typically observed with Lamin A/C mutations. hiPSC-CMs displayed well-formed sarcomeres and their organization was similar between the two cell lines. However, 3D cardiac organoids generated with LMNA H222P hiPSC-CMs showed an impaired contractility compared to control organoids. Calcium transient recordings in LMNA H222P mutant hiPSC-CMs showed a significantly higher calcium transient amplitude with a significantly slower calcium re-uptake. Transcriptomic analyses suggested a global mitochondrial dysfunction and in particular an impaired mitochondrial calcium uptake with a significantly decreased expression of the mitochondrial calcium uniporter (MCU). This decrease in MCU expression was confirmed by Western blot and was accompanied by an increased MICU1 : MCU ratio, as well as an increased PDH Ser232 and Ser300 phosphorylation, indicating a decreased mitochondrial calcium uptake in the LMNA H222P mutant hiPSC-CMs. Finally, measurement of mitochondrial respiration using Seahorse showed lower basal and maximal respiration in LMNA H222P hiPSC-CMs.

Conclusions

LMNA H222P mutant hiPSC-CMs exhibit contractile dysfunction associated with mitochondrial dysfunction with impaired MCU complex activity, decreased mitochondrial calcium homeostasis, and reduced mitochondrial energy production.
Extracellular vesicles characterization in patients with hypertrophic cardiomyopathy
Authors: Ruscica M , Rizzuto A S , Macchi C , Baroni M , Gnan E , Pandolfi A , Fichtner I , Paganini S , Giusti I , Dolo V , Corsini A , Faggiano A , Calcagnino M , Carugo S
Published: October 2024

Background

Hypertrophic cardiomyopathy (HCM) is a genetic disorder, diagnosed according to the presence of phenotypical traits of the myocardium. A specific biomarker which can associate with the severity of HCM is lacking. Extracellular Vesicles (EVs), nanoparticles released by cells into biological fluids, hold promise as accessible diagnostic tools, as their abundance and molecular composition can reflect the severity of cardiac diseases (Rizzuto 2024). We aim at characterising plasma-derived EVs isolated from 28 HCM patients and 18 healthy volunteers (CTR).

Methods

The whole cohort underwent transthoracic echocardiography, whereas magnetic resonance and exome sequencing was performed on HCM patients. EVs were isolated via ultracentrifugation from plasma of both groups. Quantitative and qualitative assessments of EVs were performed by nanoparticle tracking analysis, transmission electron microscopy, Western blot, targeted (Olink) and untargeted (nLC-MS/MS) proteomics and FACS analysis. Plasma-derived EV subpopulations were characterised according to their cellular origin. Data are expressed as median and interquartile range (25th, 75th).

Results

Most patients were male (68% HCM and 66% CTR) with a median age of 58 (49-69) years (HCM) and 47 (44-52) (CTR). Among HCM patients, missense mutations in the MYH7 gene were the most identified. The median maximum wall thickness (WTMax) in HCM was 16 mm (15-19) vs 8 mm (7-9) in CTR. The isolated EVs expressed tetraspanins (CD9, CD63 and CD81), Alix and β-integrin and showed an intact phospholipid bilayer of 100 nm in size. EV concentration was reduced in HCM vs CTRL, respectively 2.8*109 EV/ml/cell count (2*109-4*109) and 4.6*10⁹ EV/ml/cell count (3*109-6*109). Among 15 plasma-derived EV subpopulations studied, those released from platelets (CD41a) and activated platelets (CD62P and CD40L) were increased in HCM patients vs CTR by 1.7 fold. Negative associations were found between E/e’, parameter of diastolic function, and cardiomyocyte-derived EVs (r= -0.2658), and between left ventricle ejection fraction and platelet-derived EVs (r=-0.2189); a positive correlation was found between WTMax and EVs derived from activated endothelial cells (r=0.2330). The proteomic analysis of EVs shows an enrichment in proteins related to platelets adhesion and inflammation in HCM patients.

Conclusions

HCM patients present a peculiar phenotypic pattern of EVs that may associate with diagnostic clinical features of HCM.
Investigation of the usefulness of collagen peptide in intramyocardial injection of human iPS cell-derived cardiomyocytes
Authors: Iida J , Kotani K , Murata K , Wusiman M , Mandai Y , Hiraoka Y , Minatoya K , Masumoto H
Published: October 2024

Background

Injecting human induced pluripotent stem (iPS) cell-derived cardiomyocytes directly into the heart muscle holds promise as a treatment for heart failure. Achieving better engraftment is crucial for maximizing therapeutic effectiveness. Collagen peptide (CP) is a form of hydrolyzed gelatin known for its resistance to solidification. By adjusting its concentration, the viscosity of CP can be easily managed, potentially influencing the retention and engraftment efficiency of transplanted cells.

Purpose

We investigated the impact of different CP concentrations as a solvent for intramyocardial injection of human iPS cell-derived cardiomyocytes.

Methods

Initially, we examined the leakage of CP solution from the injection site using chicken meat and ex vivo rat heart. Subsequently, an in vivo experiment was conducted using beating rat myocardium injected with CP solution and Indian ink to assess CP diffusion in dynamic heart tissue. Finally, we performed intramyocardial injections of human iPS cell-derived cardiomyocytes onto a myocardial infarction model in immunosuppressed rats (1.0×107 cells/rat), followed by echocardiographic evaluation of cardiac function four weeks post-transplantation (N=3-6).

Results

There was less leakage from the injection site at lower CP concentration (10%) compared to higher concentration (20%), particularly in chicken or ex vivo rat heart tissue under static, non-beating conditions (chicken; 10% vs. 20% CP: 3.2±0.7% vs. 11.6±0.8%, P=0.01 / rat heart; 1.5±0.9% vs. 6.8±1.1%, P=0.0072). However, when examining fluid leakage from the cross-section of ex vivo rat hearts, we observed significantly higher leakage, regardless of CP concentration (10% CP: 45.7±2.7%, 20% CP: 59.1±8.1%), suggesting that CP diffusion within myocardial tissues had a greater influence on local retention than leakage from the injection site. Subsequently, in an experiment using beating rat myocardium and Indian ink, we observed significant suppression of diffusion in tissues treated with CP solution, with the 20% concentration showing a tendency to remain at the injection site (0% CP: 24.5±6.2 mm2, 10% CP: 6.9±7.1 mm2, 20% CP: 10.6±3.3 mm2, P=0.005). This indicates that 20% CP, rather than 10% CP, may be more suitable for local retention in tissues experiencing intra-tissue pressure, such as the beating myocardium. Finally, in vivo experiment using a myocardial infarction model of rat, there was no significant difference in left ventricular ejection fraction (LVEF) when cardiomyocytes were transplanted with 10% CP (0％ CP: 56.1±8.9%, 10%CP: 61.9±5.2%, P=0.92). However, there was a trend towards improved LVEF with the 20% CP solution (0% CP: 56.1±8.9%, 20%CP: 68.8±7.9%, P=0.13),

Conclusion

Twenty-percent CP may be more beneficial for treating myocardial ischemia due to enhanced retention and engraftment of transplanted cardiomyocytes.
Macrophage-T cell hybrid membrane-camouflaged biomimetic nanoparticles ameliorate autoimmune myocarditis via suppressing macrophage pyroptosis by target gene silencing
Authors: Xiong Y , Zhang Z , Yao Y
Published: October 2024

Background

Current management of myocarditis mainly relies on conventional approaches and immunosuppressive therapies. However, these strategies often yield limited efficacy. Our previous research revealed the pivotal role of macrophage pyroptosis in myocarditis pathogenesis. Therefore, targeted intervention to inhibit macrophage pyroptosis may provide new insights into myocarditis treatment.

Purpose

We previously identified interferon regulatory factor 1 (IRF1) as the key modulator of macrophage pyroptosis in myocarditis. In this study, we synthesized a nano-delivery platform consisting of a macrophage-T cell hybrid membrane-coated zeolitic imidazolate framework-8 (ZIF-8) encapsulating IRF1-small interfering RNA (siRNA@ZIF@HM). We aimed to evaluate its targeting capability and therapeutic efficacy for macrophage pyroptosis in myocarditis.

Methods

The fabrication of siRNA@ZIF@HM was validated using transmission electron microscopy (TEM) and dynamic light scattering (DLS). Cellular uptake, cytotoxicity, endolysosome escape, IRF1 silencing, and anti-pyroptotic effect were assessed using mouse bone marrow-derived macrophages and the mouse macrophage cell line J774A.1 and RAW264.7. An experimental autoimmune myocarditis (EAM) mouse model was induced in Balb/c mice for in vivo investigations. Pharmacokinetics and targeting capability were determined with ex vivo fluorescence imaging and immunofluorescence staining. Subsequently, mice were randomly allocated into control, EAM + siRNA, EAM + siRNA@ZIF, and EAM + siRNA@ZIF@HM groups to assess the therapeutic efficacy. Additionally, the biodistribution and biosafety of siRNA@ZIF@HM were also evaluated.

Results

TEM, DLS, and element mapping confirmed the successful fabrication of siRNA@ZIF@HM, with a diameter of approximately 130 nm. The nanoparticle exhibited pH responsiveness and membrane markers of macrophage and T lymphocytes (Figure 1). It demonstrated high targeting specificity for pro-inflammatory macrophages and myocarditis. Immunofluorescence microscopy revealed successful endolysosome escape of IRF1-siRNA in macrophages. Consequently, the nanoparticle significantly silenced IRF1 protein expression and suppressed macrophage pyroptosis both in vivo and in vitro, resulting in the amelioration of autoimmune myocarditis (Figure 2). Furthermore, the nanoparticle showed good biocompatibility with no significant changes observed in other organs.

Conclusions

The pH-responsive, macrophage-T cell hybrid membrane-coated biomimetic nanoparticle demonstrates promising capability for targeted siRNA delivery to myocardial inflammation sites, particularly pro-inflammatory macrophages. Additionally, targeting IRF1-mediated macrophage pyroptosis represents a potential therapeutic strategy for myocarditis treatment.Figure 1Figure 2
Human epicardial adipose tissue-derived factors promote atrial endothelial dysfunction: role of pro-inflammatory cytokines and AT1R/NADPH oxidases/SGLT2 pro-oxidant pathway
Authors: Fakih W , Alhelou C , Mroueh A , Kindo M , Mommerot A , Mazzucotelli J P , Pieper M P , Ohlmann P , Morel O , Schini-Kerth V , Jesel L
Published: October 2024

Introduction

Epicardial adipose tissue (EAT) has been suggested to contribute to left atrium dysfunction via paracrine mechanisms to induce endothelial dysfunction, pro-arrhythmogenic and pro-remodeling responses thereby favoring atrial fibrillation (AF) onset. Recently, sodium-glucose co-transporter2 inhibitors (SGLT2i) have been shown to reduce AF incidence, EAT volume, differentiation and inflammation, however, the underlying mechanisms are unknown.

Purpose

This study investigates the impact of human EAT-derived mediators on atrial endothelial cell function and, if so, to characterize the role of SGLT2 using the inhibitor empagliflozin (EMPA).

Methods

Epicardial and subcutaneous adipose tissues (SAT) were collected from 70 cardiac patients at a university hospital. Conditioned medium (CM, 24 h) from fats were applied to porcine atrial endothelial cells (AECs, first passage) for 24 h. Histology of tissues was assessed by haematoxylin and eosin staining, protein expression by Western blot analysis or immunofluorescence staining, mRNA levels by RT-qPCR, formation of reactive oxygen species (ROS) and nitric oxide (NO) by fluorescent probes, and pro-inflammatory cytokine levels by ELISA.

Results

Compared to SAT, EATs were more vascularized with increased infiltration of M1-like macrophages, expression of proinflammatory cytokines (IL-1ß, IL-6, TNF-α), markers of ECs VCAM-1 and eNOS, fibrosis TGF-β, NADPH oxidases NOX-1 and SGLT1/2. These effects were associated with a pro-oxidant response that was inhibited by neutralizing Abs directed against IL-1ß or IL-6, and by inhibition of either NOS, NADPH oxidases, ACE, AT1R or SGLT2. Levels of SGLT2 and ROS in EATs were higher in AF compared to non-AF patients and progressively increasing with age. CM of EATs showed higher concentrations of IL-1ß, IL-6, TNF-α and MCP-1 than that of SAT. Exposure of AECs to CM of EATs increased the level of oxidative stress that was positively correlated to the level of pro-inflammatory cytokines, and reduced basal and bradykinin-induced NO formation that was prevented by EMPA. It also resulted in upregulation of p53 and SGLT2 expression, and nuclear translocation of NF-kB by CM of the top quartile inflamed EATs but not by the lowest quartile of EATs or SATs, prevented by EMPA.

Conclusion

The findings indicate that compared to SATs, the EATs showed higher in situ expression and release of pro-inflammatory cytokines that contributed to oxidative stress involving the AT1R/NADPH oxidases/SGLT2 pro-oxidant pathway. CMs of inflamed EATs induced AECs dysfunction, oxidative stress and activation of NF-kB involving pro-inflammatory cytokines and SGLT2. Thus, SGLT2i appear as an interesting strategy to decrease EAT inflammation and improve endothelial function contributing to prevent cardiac remodeling and fibrotic responses.
Pulsed field ablation using additional mapping and substrate modification to treat atrial fibrillation - one year outcome comparison
Authors: Mergani M , Mannion J , Healy L , Ramsamy K , Egan S , Boles U
Published: October 2024

Background

Pulsed field ablation (PFA) is a relatively new non-thermal form of ablation employing irreversible cell electroporation as an alternative to radio-frequency ablation. Conventional use of PFA relies on fluoroscopy. Electro-anatomical and voltages mapping (EAM) may potentially provide refinement of PFA via ablation of non-PV targets and accurate localisation of catheter placement. Currently there is limited data on the efficacy of the use of voltages criteria of EAM in conjunction PFA in the treatment of atrial fibrillation.

Objective

The aim of our study was to determine the effect on atrial arrhythmia recurrence with the use of EAM with PFA in patients requiring atrial fibrillation ablation.

Methods

This single centre retrospective observational analysis included one hundred patients with symptomatic AF (paroxysmal and persistent) undergoing ablation for the first time. Fifty patients underwent PFA-PVI alone and fifty patients underwent PFA-PVI with EAM. Data were obtained on patient characteristics, outcomes, procedure details and complications up to one year. A blanking period of three months was used post-ablation. For rhythm monitoring we utilised ECGs and holters at three, six, nine and twelve months and twenty-four hour holter monitoring at six and nine months. We also used remote monitoring in between these periods to record any symptomatic episodes atrial arrhythmias.

Results

Baseline patient characteristics are represented in Table 1. Despite having no significant difference in category of AF between groups (Paroxysmal, persistent, longstanding persistent) the mapping group were significantly older (68.2 years ± 10.2 vs 60.1 ± 11.11; p=0.01), longer duration of documented atrial fibrillation (45.9 ± 50 vs 28.6 ± 38 months; p=0.038), higher risk for CV events with CHA2DS2-VASc score (2.2 ± 1.5 vs 1.42 ± 1.2; p=0.04), however, procedure times were greater in the PVI and mapping group (72.8 ± 6.2 vs 41.1 ± 17 minutes; p=0.03).

Successful PVI was achieved acutely in all patients upon voltage guided mapping validation and most importantly 1 year follow up revealed comparable outcome in both groups (Fig 1).

Conclusion

Patients who underwent PFA with mapping were older, greater co-morbidity and longer duration of AF burden. Yet, the addition of EAM with PFA did demonstrate equivalent achievement of sinus rhythm and was relatively high as higher than 80 % at one year follow up. This study may suggest a preference to using EAM in patients who may be deemed to be older and have a greater co-morbidity. It emerges as a great hope to safer approach and successful intervention to reduce AF burden.mapping vs PVI onlyGraph showing one year follow up
Daily Subcutaneous injections of Relaxin is a therapy for heart failure with preserved ejection fraction
Authors: Salama G , Dschietzig T B D , Gabris-Weber B W , Mcmahon B M , Mauro A M , Romero G R
Published: October 2024

Background and Significance: Relaxin (RLX), a hormone of pregnancy has engendered a great deal of excitement as a therapy for cardiovascular diseases. Clinical trials treated patients with acute decompensated Heart Failure (HF) with RLX (i.v. 30µg/kg/day-2 days) and reported a 37% decrease in mortality during a 6-months follow up. This led the FDA to declare RLX a ‘break-through’ therapy. However, a larger trial failed to meet the sought-after end points. Here, we propose that periodic subQ RLX pulses are more effective. We measured the pharmacokinetics of serum RLX following subQ injections of RLX in Sprague Dawley rats. Further, we treated rats with confirmed HFpEF with daily subQ RLX (2-weeks) to reverse the diastolic dysfunction (DD), arrhythmia and fibrotic profile.

Methods

Sprague-Dawley (250 g, n=20, either sex) received 2-subQ injections 12 hrs apart of RLX (from Relaxera) or the vehicle. Serial blood samples were drawn following each injection to measure [RLX]. ZSF1 rats (9-weeks old, either sex) were placed on a high fat diet (HFD) and serial echos tracked the DD. Once severe DD was established (25-35 weeks of HFD), rats received daily subQ injections of RLX (100µg/kg) or vehicle. After 2-weeks, hearts were perfused to optically map action potentials and Ca2+ transients, and analyze the arrhythmia profile. IF and Westerns were used to measure changes in fibrosis (collagen 1), Nav1.5, connexin 43, Wnt1 and β-catenin.

Results

After an injection, RLX reached a peak in ~60 min, fell to 50% in 5-6 hours. Injections of 0, 30, 100 or 500 µg/kg yielded peak levels of 0, 11.26±3.52, 58.33±16.10, and 209.42±29.04 ng/ml and residual levels after 24-hrs of 0, 4.9, 45.1 and 156 pg/ml, respectively. The 30µg/kg injections had no effect and 100 µg/kg increased Nav1.5 (25%), Cx43 (30%) and β-catenin (90%). The 500 µg/kg injections increased Nav1.5 and Cx43 but upregulated β-catenin, only slightly. In ZSF1 HFpEF rats that received the vehicle (n=12), a premature stimulus (at 30 ms) elicited arrhythmia in 67% of the hearts. RLX injections reversed DD, suppressed sustained atrial and ventricular arrhythmias (n=0/12) but not self-terminating arrhythmias, lasting < 10sec. RLX improved the conduction velocity (CV), particularly at fast rates (100ms) from 0.74 to 0.9 m/s (p<0.001, n=12). RLX significantly increased Cx43 expression, Nav1.5 and β-catenin at intercalated disks. RLX reduced collagen deposition in HFpEF rats, to normal levels and increased Wnt1 compared to control HFpEF rats.

Conclusions

The ZSF1 diabetic rat on a HFD recapitulates human HFpEF, high blood pressure, fibrosis, DD, AF and VF. SubQ RLX injections reversed DD i.e. enlarged Left Atrium, E/e’ ratio, fibrosis, and the arrhythmia vulnerability by increasing CV, Cx43, and Nav1.5. Daily subQ RLX injections were highly effective as a therapy for HFpEF.Echo parameterHFpEF + RLX
The STRIPAK associated kinase MST4 is a novel mediator in heart failure pathogenesis
Authors: Leye M , Eden M , Hahn J , Luzarowski M , Tannert C , Voelschow B , Martini S , Sossalla S , Lucena-Porcel C , Frey N
Published: October 2024

Heart failure still poses substantial challenges in understanding its underlying molecular pathways. We recently found the mammalian STE20-like kinase 4 (MST4) significantly upregulated in human failing hearts (5.7-fold in DCM and 3.8-fold ICM) as well as in heart failure animal models (i.e. 2.5-fold in Calsarcin-1-KO, 2.5-fold in MLP-KO). MST4 is a member of striatin interacting phosphatases and kinases complexes (STRIPAK), an emerging group of multiprotein complexes responsible for integrating cellular signalling pathways. We provide the first evidence of functional STRIPAK complexes containing MST4 by co-immunoprecipitation and interactomics experiments in cardiomyocytes. Moreover, we found MST4 to interact with several intercalated disc proteins like beta-Catenin or Desmin.

Adenoviral overexpression of MST4 in neonatal rat ventricular cardiomyocytes (NRVCM) results in a significant increase in cell size (+13%). This was not accompanied by the activation of fetal genes such as NPPA, NPPB or RCAN1.4, but rather by an increase in phosphorylation of Protein kinase B (PKB/Akt, 3.3-fold). Akt is usually associated with physiological hypertrophy, indicating that MST4 may be involved in protective pathways in cardiomyocytes. In support of this notion, MST4 overexpression reduced apoptotic activity (cleavage of Caspase 3 -69%, Caspase 7 -80%, Parp1 -27%) and improved both contractility (fractional shortening cell +23%, fractional shortening sarcomere +30%, time to max. contraction cell -10%, time to max. contraction sarcomere -12%) and relaxation parameters (time to max. relaxation velocity cell -12%) in isolated adult rat ventricular cardiomyocytes.

In order to identify cardiac targets of MST4, we performed a comprehensive analysis of the phosphoproteome of NRVCM overexpressing MST4 compared with control-virus +/- pharmacological inhibitor at two time points. We identified more than 70,000 peptides including more than 20,000 phosphopeptides in more than 4,000 protein groups. Using linear modelling with a reduced data set containing regulated features selected by thresholds for p-value and fold change, we observed a consistent MST4 effect and a number of enriched gene ontology terms. Here, we found several potential MST4 targets and effects on protein expression levels associated with intercalated disc proteins. In human heart samples, MST4 was also localized to the intercalated disc region by immunostaining. Interesting proteins in that context that might help explain some of the observed cardioprotective MST4 effects are Connexin 43, Desmin, Ryanodine receptor, Phospholemman, Junctophilin2, Myozap or Frizzled 1.

Taken together, our data imply that MST4 upregulation in human heart failure serves an adaptive role which may offer translational opportunities.Graphical Abstract
A novel murine model of stress-induced cardiomyopathy achieved by chemogenetic stimulation of the locus coeruleus-norepinephrine system
Authors: Bengs S , Ministrini S , Liberale L , Duss S , Floriou-Servou A , Portmann A , Rossi A , Mikail N , Puspitasari Y M , Montecucco F , Luescher T F , Bohacek J , Camici G C , Gebhard C
Published: October 2024

Background

Takotsubo syndrome (TTS) is a stress-induced acute cardiomyopathy characterized by transient left ventricular (LV) dysfunction without obstructive coronary artery disease. TTS is often underdiagnosed and undertreated and typically occurs in middle-aged women who recover from initial symptoms within a few weeks. Despite the apparent recovery, TTS patients have a worse cardiovascular prognosis compared to healthy individuals. To date, the pathophysiology underlying TTS remains unclear.

So far, preclinical models barely reflect the complete clinical picture of TTS observed in humans. Moreover, these models display several limitations, e.g., high mortality or disregarding the involvement of the central nervous system. Consequently, much of the current obscurity concerning the pathophysiology of TTS is due to the lack of appropriate preclinical models.

Purpose

The present study aimed to assess whether chemogenetic activation of the locus coeruleus-(LC)-norepinephrine-(NE) system in vivo can induce LV dysfunction similar to that observed in patients with TTS. For this purpose, we used the designer excitatory receptor activated exclusively by designer drug (DREADD) system.

Methods

11-13 weeks-old female Cre-dependent DREADD mice were genetically modified to express a codon-improved Cre recombinase under the dopamine-beta-hydroxylase-(DBH) promoter. For this purpose, a designed excitatory receptor exclusively activated by designer drugs or a control vector was stereotactically delivered to the LC using adenoviral transfection. Restricted expression of DBH-positive noradrenergic neurons to the LC was validated using pupillometry. LC-NE was activated by administering low-dose clozapine 0.03 mg/kg/day intraperitoneally. Serial anatomical and functional assessments of the cardiac phenotype in these mice were performed at baseline, the end of stress induction, and after ten days and three months of the recovery phase using echocardiography and cardiac magnetic resonance imaging.

Results

Targeted stimulation of the LC-NE system resulted in apical LV wall motion abnormality in 70–83% of mice. A complete recovery of LV wall motion abnormality was observed after the recovery phase.

Conclusions

Chemogenetic stimulation of the LC-NE system in mice mimics LV wall abnormalities similar to those observed in patients with TTS and may, therefore, be a novel preclinical model to investigate the pathophysiology of TTS.
Reinterpretation of genetic testing for Cardiomyopathies: Less is more
Authors: Manzoni M , Sommariva E , Pizzamiglio F , Mushtaq S , Bonalumi G , Farina S , Volpe A , Biondi M L , Novelli V
Published: October 2024

Background

In the last years, the concept of reinterpretation of genetic testing has been considered as part of the diagnostic workflow. However, this process requires time, expert curators and new pieces of evidence to evaluate, as functional studies, segregation and allele frequency data.

Last year, the European Society of Cardiology (ESC) published new guidelines for the management of cardiomyopathies. This document includes a precise list of genes associated with the different types of Cardiomyopathies and their level of contribution, according to ClinGen evaluation: definitive/strong, moderate, limited, and refuted/disputed. This document poses a real support in the reinterpretation of tests that report variants in genes not clearly correlated with these phenotypes.

Purpose

To evaluate the impact of the new ESC indications regarding genetic testing, we reconsidered all genetic results performed in our Clinic in patients with confirmed or suspicious Cardiomyopathy.

Methods

From 2017 to 2023 we analyzed 296 probands eligible for Cardiomyopathy genetic testing, in particular: 105 patients were tested for Arrhythmogenic Cardiomyopathy (ACM), 113 for Hypertrophic Cardiomyopathy (HCM) and 78 for Dilated Cardiomyopathy (DCM), using a custom sequencing panel including 44 genes (11 genes for ACM, 34 for DCM and 29 for HCM).

Results

Comparison analysis with the genes reported from the ESC guidelines, considering only the definitive, moderate, and strong genes, showed that 24 genes are not more associated with the phenotype and then not clinically actionable. In this scenario, we reconsidered 12 variants (8%), previously classified as VUS, changing the referral status of 8 patients, from uncertain to negative.

Conclusions

These results highlight the importance of variant reinterpretation as a fundamental step of genetic testing, posing the importance of the selection of the genes to analyze, in order to improve the management of the patients.
A novel murine wire injury based infectious endocarditis model aggravates aortic valve stenosis
Authors: Bartsch B , Nazir M L , Altrogge M , Fester L , Lambertz J , Coburn M , Nickenig G , Zimmer S , Weisheit C K
Published: October 2024

Background

With mortality rates up to 40% under optimal medical treatment, infectious endocarditis (IE) remains one of the deadliest infectious diseases worldwide. In the early stages of IE, diagnosis and treatment initiation are often difficult and research is limited due to unreliable models. Most murine IE models rely on permanent prosthetic wire placement and i.v. bacterial injection, thus making it inadequate to study IE pathophysiology with the aim to reflect the course of disease in humans.

Purpose

To develop a reproducible murine IE model, based on aortic valve injury preceding intravenous S. aureus bacterial challenge.

Methods

Using a wire-injury model we induced endothelial lesions. 72h after wire injury (WI), we performed intravenous bacterial challenge using S. aureus. Echocardiography was performed to confirm bacterial vegetations and assess valvular function. Cross sections of valvular leaflets were prepared for scanning electron microscopy (SEM). Macrophage, neutrophil and S. aureus-specific immunofluorescence staining was performed and visualized by immunofluorescence microscopy. Bacterial cultivation was performed from peripheral blood and valve tissue. Systemic immune response was analyzed using flow cytometry.

Results

WI induced endothelial damage in all mice verified using scanning electron microscopy imaging. The combination of wire injury and injection of 10^5 and 10^6 colonies-forming-units (CFU) S. aureus reliably caused IE validated via in vivo echocardiography as well as S.aureus immunofluorescence staining and SEM imaging. Mice undergoing bacterial challenge responded with significant neutrophilia in the blood. Using 10^6 CFU S. aureus was associated with an increased mortality, liver and kidney abscess formation due to systemic bacterial spreading compared to 10^5 CFU. Peak velocity across the aortic valve and aortic valve leaflet thickness as markers for aortic valve stenosis were increased in IE compared to WI only mice. Aortic regurgitation was more prevalent after bacterial challenge mediating increased left ventricular volumes, meanwhile ejection fraction was not altered. Immunofluorescence staining revealed a pro-inflammatory milieu including increased macrophage and neutrophil infiltration in IE compared to WI only mice.

Conclusion

In vivo echocardiography and ex vivo histological staining revealed reliable IE induction using our new model with S. aureus concentrations of 10^5 CFU. The additional bacterial challenge causes a more severe aortic valve stenosis compared to WI only mice, potentially due to an observed increase in valvular inflammation.Fig 1 Scanning electron microscopy after
Myocardial SGLT2 expression and cardiac remodeling in patients with severe aortic stenosis: the BIO-AS Study
Authors: Paolisso P , Scisciola L , Belmonte M , Gallinoro E , Delrue L , Casselman F , Van Praet V , Torella M , Marfella R , De Feo M , Bartunek J , Paolisso G , Barbato E , Barbieri M , Vanderheyden M
Published: October 2024

Background

Cardiac remodeling plays a major role in the prognosis of patients with aortic stenosis (AS) and could impact the benefits of aortic valve replacement. Blood and tissue biomarkers have a differential pattern and expression level in patients with AS, which may retain a pathophysiological role in cardiac remodeling and metabolism. However, the data available are limited and contradictory.

Aim

Our study aimed to evaluate the expression of sodium-glucose cotransporter 2 (SGLT2) gene and protein in patients with severe AS stratified in high gradient (HG) and low flow-low gradient (LF-LG) AS and its association with cardiac functional impairments.

Methods

Gene expression and protein levels of main biomarkers of cardiac fibrosis (Galectin-3, sST2, Serpin-4, PINP, PICP, Collagen, TGF-b), inflammation (GDF-15, IL-6, NF-kB), oxidative stress (SOD1, SOD2), and cardiac metabolism (NHE, PPAR-a, PPAR-g, GLUT1, and GLUT4) were evaluated in blood samples and heart biopsies of 45 patients with AS.

Results

Our study showed SGLT2 gene and protein hyper-expression in patients with LF-LG AS, compared to controls and HG AS (p<0.05). These differences remained significant even after adjusting for age, gender, body mass index, history of diabetes mellitus, arterial hypertension, and coronary artery disease. SGLT2 gene expression was positively correlated with: i) TGF-b (r=0.72, p<0.001) and collagen (r=0.73, p<0.001) as markers of fibrosis; ii) NF-kB (r=0.36, p<0.01) and myocardial IL-6 (r=0.68, p<0.001) as markers of inflammation: iii) SOD2 (r=-0.38, p<0.006) as a marker of oxidative stress; iv) GLUT4 (r= 0.33, p<0.02) and PPAR-a (r=0.36, p<0.01) as markers of cardiac metabolism (Figure 1).

Conclusion

In patients with LF-LG AS, SGLT2 gene and protein were hyper-expressed in cardiomyocytes and associated with myocardial fibrosis, inflammation, and oxidative stress. The hyper-expression of the SGLT2 gene and protein in patients with LF-LG might pave the way for using SGLT2-Is in this subset population, with the potential to improve the outcomes of AVR and reduce the likelihood of re-hospitalization within one year.Figure 1
Catheter ablation increases cerebral blood flow and cognitive function in elderly patients with atrial fibrillation
Authors: Manita M , Asahi T , Higa N , Zaima S , Arakaki Y , Tabata K
Published: October 2024

Background

Atrial fibrillation (AF) has been considered to have an increased risk for cognitive impairment independent of a stroke and a silent micro-infarction. The mechanism of this interaction is unclear. One of the potential explanations is brain hypoperfusion due to a reduced cardiac output. To date, there are no reports focusing on the cerebral blood flow and cognitive impairment in patients undergoing AF ablation.

Purpose

The purpose of this study was to investigate whether catheter ablation of AF would increase cerebral blood flow and restore cognitive impairment.

Methods

AF patients aged 75 years or older with mild or more cognitive impairment were selected. The assessment of cognitive function using Mini-Mental State Examination (MMSE) and Hasegawa's Dementia Scale-Revised (HDS-R), and measurement of cerebral blood flow using 99mTc-ECD SPECT imaging were examined before and 3 months after ablation.

Results

A total of 25 elderly patients (mean age 83, 14 males and 11 females) were included in this study. Of them, 23 patients had persistent AF. Twenty-three patients (92%) maintained sinus rhythm after ablation. There was a significant improvement in MMSE score (from 24.6±4.5 to 25.9±4.0, p<0.05) and HDS-R (from 23.2±5.5 to 24.8±5.1, p<0.05) after ablation. Cerebral blood flow also significantly increased after ablation (right lobe: from 34.4±4.6 to 38.5±4.4 ml/100g/min, p<0.01, left lobe: from 35.3±9.7 to 38.8±3.7 ml/100g/min, p<0.01). Overall average cerebral blood flow increased by 10.1±9.4% after ablation. There was a positive correlation between the changes in MMSE score and the changes in cerebral blood flow (r=0.65, p<0.05).

Conclusion

AF ablation in elderly patients increased cerebral blood flow and cognitive function. AF ablation would be one of the therapeutic options to improve cognitive function in elderly patients with cognitive impairment.