bluuebunny/crossref_metadata_2025 huggingface dataset:
Discrimination and calibration are insensitive metrics for comparing the performance of cardiovascular risk prediction models
Authors: Liang J , Jackson R , Wells S , Poppe K
Published: October 2024

Background/Introduction

Accurate cardiovascular disease (CVD) risk prediction models can facilitate the identification and cost-effective treatment of high-risk individuals. Discrimination and calibration are the most commonly recommended metrics for evaluating model performance. However, both are population-level measures, and it remains unclear whether these metrics are sufficiently sensitive to assess the performance of models designed for use with individual patients.

Purpose

Using a real-world dataset, we aimed to compare CVD risk estimates for example individuals (individual-level measures) and discrimination and calibration metrics (population-level measures) from two CVD risk prediction models; one with "age" as the only predictor and one with multiple predictors.

Methods

We constructed a cohort of almost all New Zealanders without CVD or heart failure, alive and aged 30–79 years in 2014, with follow-up linkage to hospitalisations and mortality until 2018 (N = 2,098,359). We derived two sets of sex-specific Cox regression models to predict 5-year CVD risk. The age-only models included "age" as the only predictor, and the full models used nine pre-defined routinely available CVD risk predictors with interaction terms. We calculated and compared individuals’ 5-year CVD risk from the age-only and full models and assessed model performance using Harrell’s C-statistics and calibration plots.

Results

Risk estimates for individuals using the two models showed substantial differences. A 65-year-old woman had predicted 5-year risks of 4.9% with the age-only model but 16.1% with the full model; a 74-year-old man had predicted risks of 15.2% and 9.2%, respectively, with age-only and full models (Figure 1). However, the models had similar discrimination and calibration (Figure 2). Age-only models had good discrimination with C-statistics of 0.773 (95% CI: 0.771–0.775) in women and 0.740 (0.738–0.742) in men, and calibrated well in both sexes. The full models had slightly better discrimination (C-statistics: 0.814 [0.812–0.816] in women and 0.769 [0.767–0.771] in men), and very similar calibration to the age-only models in both sexes.

Conclusions

Age-only models showed good discrimination and were well calibrated, but predicted very different risks in individual case studies compared to models with multiple predictors. This study calls into question the use of discrimination and calibration metrics as the main measures for comparing the performance of different CVD risk prediction models. We recommend that examples of predicted risk for individuals and an assessment of the magnitude and prevalence of cardiovascular predictors should also be considered when comparing the performance of prognostic models.
Identification of cell adhesion molecules for the maturation of engineered heart tissue
Authors: Fujiwara Y , Naka Y , Sasalo M , Imahashi K , Yoshida Y
Published: October 2024

Background

Engineered cardiac tissue (ECT) utilizing human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CMs) is anticipated to function as a translational in vitro model. However, hiPSC-CMs display phenotypes indicative of immaturity, and the structural organization of ECT composed of hiPSC-CMs is insuficient when compared to that of mature adult hearts. Although the mechanisms for hiPSC-CMs maturation have been reported, little is known about the mechanisms regulating the formation of in vitro tissue such as ECT. Cell adhesion molecules play important roles in heart formation in vivo through cell-cell interactions. However, specific cell adhesion molecules responsible for tissue structure formation in in vitro tissue models, such as ECTs, have not been identified. We hypothesize that cell adhesion molecules, which regulate cell-cell attachment, are crucial for in vitro tissue formation.

Purpose

This study aims to identify the key cell adhesion molecules involved in cell-cell interaction and assess the effects of their expression on hiPSC-CMs and EHT functions.

Methods and Results

We developed a cell attachment assay to evaluate the attachment ability of hiPCS-CMs and discovered that hiPSC-CMs produced via embryoid body method (EB-hiPSC-CMs) exhibited a higher attachment ability than those produced by monolayer method (Mono-hiPSC-CMs). Further analysis revealed no significant difference in anoikis sensitivity between EB-hiPSC-CMs and Mono-hiPSC-CMs, suggesting that the observed difference in attachment ability were due to differencies in cell adhesion efficiency between hiPSC-CMs generated by two methods. Moreover, RNA-seq analysis identified nine cell adhesion molecule genes that were highly expressed in EB-hiPSC-CMs. We identified the cell adhesion molecules responsible for the attachment ability of hiPSC-CMs by knockdown and over-expression experiments. Moreover, over-expression experiments using adeno-associated virus (AAV) demonstrate that hiPSC-CMs with overexpression of a cell adhesion molecule improved calcium handling and increased propagation speed, and ECTs with overexpression of a cell adhesion molecule showed enhanced sarcomere alignment and sarcomere length compared to controls.

Conclusion(s)

We observed differences in attachment ability between hiPSC-CMs generated by two differentiation methods. Transcriptome analysis of these hiPSC-CMs identified key cell adhesion molecules that regulate cell-cell attachment ability of hiPSC-CMs. Furthermore, increment of attachment ability of hiPSC-CMs by overexpression of a cell adhesion molecule enhanced hiPSC-CMs and ECT functions.
SMAD3 silencing induces Epithelial-to-Mesenchymal Transition (EMT) towards cardiac pericyte progenitors with proangiogenic potential in hiPSC-derived epicardial cells
Authors: Miyoshi Y , Lecena-Cacace A , Yoshida Y
Published: October 2024

Background

The epicardium is only a single mesothelial layer and "quiescent" in the adult heart. However, once the heart is injured, the epicardium reactivates and acts as a key for cardiac regeneration through epithelium-to-mesenchymal transition (EMT). While the canonical TGF-β-SMAD pathway plays a central role in regulating EMT, SMAD3 functions independent of this pathway in epicardium remain unknown.

Purpose

The purpose of this study is to investigate the effects of SMAD3 downregulation on EMT in epicardial cells.

Methods

First, we differentiated human iPSC-derived epicardial cells (EPI cells) by mimicking the process of cardiac mesoderm development. Next, siRNA targeting SMAD3 was transfected into EPI cells to silence SMAD3. We performed qRT-PCR, western blotting, immunocytochemistry, flow cytometry, transcriptional analysis, endothelial tube formation assay, and paracrine experiments using FUCCI (fluorescent ubiquitination-based cell cycle indicator) system to evaluate the phenotype of EPI cells with silenced SMAD3.

Results

We found that SMAD3 silencing in EPI cells resulted in loss of epicardial identity and upregulation of multiple EMT markers. Additionally, NG2 and ENG (CD105), both cardiac pericyte markers, were highly expressed in SMAD3-silenced EPI cells. RNA-sequence also revealed that SMAD3 silencing in EPI cells induced EMT facilitated towards cardiac pericyte progenitors with proangiogenic potential. In endothelial tube formation assay, SMAD3-silenced EPI cells demonstrated enhanced tube formation compared to the control (1.48-fold increase in the number of junctions, n=3; p<0.0001), which indicated their angiogenic potential. At last, we revealed the paracrine effect of SMAD3-silenced EPI cells to enhance the proliferative reactivation in iPSC-derived cardiomyocytes by showing upregulation in CDK6/CCND1 axis and FUCCI-green.

Conclusion

Our findings demonstrate a new role of SMAD3 biology in the human epicardium, specifically in the generation of cardiac pericyte progenitor cells that contribute to better responses in angiogenesis of the heart microvasculature enabling the possibility to continue exploiting epicardial biology for effective cardiac regeneration.
The BET protein inhibitor apabetalone (RVX-208) prevents doxorubicin-induced endothelial senescence
Authors: Mohammed S A , Telesca M , Gorica E , Panteloglou G , Mongelli A , Ruschitzka F , Costantino S , Paneni F
Published: October 2024

Background

Accumulation of senescent endothelial cells (ECs) plays a pivotal role in cardiometabolic disorders and lifespan reduction(1). Yet, only a few senolytics are known to date, partly due to the poor grasp of the molecular mechanisms that control the senescence survival programme. Bromo-and extraterminal domain (BET) proteins, known epigenetic reader proteins, recognize acetylated histone tails and regulate gene transcription. RVX-208, an FDA-approved BET inhibitor, has shown to modulate transcriptional programs involved in inflammation, cancer, and renal disease(2). Yet, its potential role in EC senescence and cardio-oncology remains elusive.

Purpose

Our study aims to investigate whether pharmacological modulation of BET proteins by RVX-208 can mitigate doxorubicin-induced endothelial senescence.

Methods

Human aortic endothelial cells (HAECs) were exposed to different concentrations of Doxo (50nM-500nM) for 48 hours. Cell morphology changes, cell cycle arrest, senescence associated secreted phenotype (SASP) release, upregulation of SA-b gal activity, DNA damage response (gH2AX) and expression of p21, p16 and p53 were used as molecular markers of cellular senescence. Conditioned media was collected from Doxo-treated ECs and processed for molecular analyses or used for treatment of healthy, non-senescent ECs. To test whether RVX-208 exerts any senolytic or senomorphic effect, HAECs were treated with Doxo (100 nM) in presence or absence of RVX-208 (5mM-20mM) for 48 hrs. RNA-seq and proteomic profiling were performed in vehicle and Doxo-treated ECs.

Results

Doxo-treated ECs showed a dose-dependent increase in senescence markers (p16, p21 and p53) and DNA-damage response (gH2AX). FACS analysis revealed an accumulation of cells in G2/M phase in Doxo-treated ECs (100 nM). Of interest, treatment with RVX-208 (15mM) prevented the upregulation of senescent markers (p16, p21 and p53) while rescuing alterations in cell morphology and reducing the number of SA-b gal positive cells. RVX-208 prevented Doxo-induced upregulation of genes involved in inflammation, oxidative stress and mitochondrial dysfunction as shown by RNA-seq and proteomic analysis. Conditioned media from Doxo-treated ECs showed an enrichment of SASP-associated markers (IL6, CCL2, IL8, TIMP2 and PDGF-b) as shown by dot-blot arrays. Exposure of healthy ECs to conditioned media from senescent ECs recapitulated aging features. Notably, RVX-208 blunted the release of SASP markers (IL6, CCL2 and IL8) from Doxo-treated ECs. Mechanistically, we found enhanced BET protein occupancy (BRD4) with concomitant enrichment of activating chromatin marks (H3K27Ac and H3K4me) on the promoter of senescence genes COL1A2, CDKN1C and SESN3.

Conclusion

Targeting BET proteins with RVX-208 may offer a promising therapeutic strategy to prevent endothelial aging in cancer patients undergoing cardiotoxic therapies.
Heritable heart failure traits in mice undergoing early life stress
Authors: Costantino S , Mohammed S A , Gorica E , Manuella F , Telesca M , Ambrosini S , Boscardin C , Ruschitzka F , Mansuy I , Paneni F
Published: October 2024

Introduction

Adverse childhood experiences, known as early life stress (ELS), are associated with increased cardiovascular disease risk in later life, yet the underlying mechanisms remain elusive. Recent evidence indicates that parental life experiences can be transmitted to the offspring. Whether ELS exerts transgenerational effects on the heart is unknown.

Purpose

To investigate the effects of ELS on cardiac structure and function in exposed parents and in their offspring, across 3 generations.

Methods

We used a mouse model of ELS (MSUS) that mimics the exposure to traumatic experiences during childhood in humans. Mouse pups (F1) were separated from their mother (F0) unpredictably each day for 3 hours from postnatal day 1 (PND1) to PND14. During separation, dams were exposed to an additional unpredictable stressor by being subjected to either, a forced swim (18°C water for 5 minutes) or a 20-minute physical tube restraint, anytime (unpredictably) during the 3 hours. From PND15, mice are left undisturbed with their mother until PND21, are then raised normally until adulthood. Control litters were raised normally. A high-resolution Micro-Ultrasound System (Vevo 3100, Visualsonics) was used for the echocardiographic assessment at 6, 12 and 18 months both in the exposed animals (F0) as well as in their offspring (F1) and grandoffspring (F2). Both male and female mice were studied. Heart weight/tibia length and left ventricle weight/tibia length were used to assess cardiac mass while myocardial fibrosis was assessed by Masson's Trichrome and Picriosirius Red stainings. Lung congestion was assessed as lung wet/dry weight ratio. Single cell RNA sequencing (scRNAseq) was used in LV specimens from MSUS mice to unveil cell-specific transcriptional changes. Finally, MSUS mice underwent environmental enrichment (cages containing running wheels, maze) to investigate any rescuing effect on cardiac function.

Results

F1 mice displayed increased LV mass, impaired diastolic function, myocardial fibrosis and lung congestion. A time-dependent worsening of cardiac performance was observed from 6 to 18 months, both in males and females. ScRNAseq unveiled dysregulation of transcriptional programs underlying inflammation and lipotoxicity in the cardiomyocyte and endothelial cell clusters. The MSUS offspring did not show changes of cardiac function at 6 months, however diastolic dysfunction and lung congestion were observed at 12 and 18 months. A similar impairment of cardiac function was observed in the MSUS grandoffspring (F3). Of interest, 6-week exposure to an environmental enrichment protocol was able to improve LV mass, diastolic function and lung congestion in 12 months-old MSUS mice.

Conclusions

ELS induces a transgenerational transmission of cardiac phenotypic alterations which can be rescued by EE. Our results shed light on the potential role of ELS on heart failure development and potential mitigation strategies.
Exome sequencing identifies a novel FBN1 variant in a Chinese family with Marfan syndrome that includes aberrant right subclavian artery
Authors: Huang S , Lei L M , Zhang R Y
Published: October 2024

Background

Marfan syndrome (MFS) is an inherited autosomal dominant disorder that affects connective tissue with an incidence of about 1 in 5,000 to 10,000 people. Ninety percent of MFS is caused by mutations in the fibrillin-1 (fbn1) gene. We recruited a family with MFS phenotype in South China and identified a novel variant in fbn1 by whole exome sequencing (WES). The zebrafish share high genetic similarity to humans. This study aimed to generate this novel mutation in the fbn1 gene of zebrafish using the CRISPR/Cas9 technology and researched whether this variant is pathogenic.

Methods

A three-generation consanguineous family was recruited in this study, which was visited and interviewed for the clinical data and family history. Peripheral blood samples were collected from family members for DNA isolation and WES. The 3D structure of the protein was predicted by Alphafold. CRISPR/Cas9 was applied to generate an fbn1 nonsense mutation (fbn1+/−) in zebrafish. Morphological abnormalities were assessed in F2 fbn1+/− zebrafish by comparing with the wild-type (WT) controls at different development stages.

Results

Our study recruited a family with MFS phenotype in South China and identified a novel variant [NM_000138.5; c.7764C>G: p.(Y2588*)] in fbn1. Through Sanger sequencing, we found that family members Ⅲ-1 (son) and Ⅲ-2 (daughter) had the fbn1 variant segregated with MFS in the family. The p.Y2588* mutation resulted in the absence of 283 amino acids, thereby leading to the deficiency of 17 β-folded structures, 4 α-helix structures, and various loop structures. Compared with WT zebrafish, F2 fbn1+/− zebrafish exhibited aortic arches bleeding, abnormal angiogenesis, decreased cardiac volume, cardiac function defects, curly tail, and cartilage malformations.

Conclusion

A novel variant [NM_000138.5; c.7764C>G: p.(Y2588*)] was identified in fbn1 gene, which has not been reported in previous literature. The variant caused loss of function through premature protein truncation or nonsense-mediated mRNA decay. In zebrafish, we identified functional evidence of the detected nonsense mutation, which aligns with the clinical significance, suggesting a pathogenic variant of fbn1. Zebrafish as a novel, efficient tool to allow for the quick classification of unknown variants in fbn1 and improve the clinical diagnosis and treatment of MFS. It has brought the implementation of personal and precision medicine in the clinic within reach and made it possible to find patient-specific treatments.Marfanoid symptoms of the patient.Phenotypic Spectrum of fbn1+/− Zebrafish
Transcriptome-wide association study discovers functional long non-coding RNAs in coronary artery disease
Authors: Li S , Li L , Diagel A , Chen Z , Schunkert H
Published: October 2024

Genetically regulated long non-coding RNAs (lncRNAs) in coronary artery disease (CAD) have not been investigated. To discover CAD-relevant lncRNAs, we performed the transcriptome-wide association study (TWAS) using a subset genomic data of CARDIoGRAMplusC4D Consortium and transcriptomic data of nine CAD-relevant tissue types from Genotyped Tissue Expression (GTEx) and Stockholm-Tartu Atherosclerosis Reverse Network Engineering Task (STARNET) projects. We identified 20 lncRNAs associated with CAD in a tissue-specific fashion. The eQTL-GWAS colocalization analysis associated 15 lncRNAs with cardiometabolic traits which could bridge the lncRNA to CAD risk. The co-expression analysis identified co-expressed protein-coding genes of 16 lncRNAs and suggested their involved biological pathways. Four antisense lncRNAs from the same locus, CDH13-AS2, CDH13-AS1, CEDORA, and CTD-3253I12.1, not only shared cardiometabolic traits but also exclusively co-expressed with the host gene, CDH13. Our biological experiments using dRfxCas13d-based RNA immunoprecipitation, CRISPR/Cas9 gene knockdown, and dCas9-VP64-based transcriptional activation in endothelial cells, elucidated CDH13-AS2 as a stabilizer of CDH13 mRNA and suggested therapeutic potential of CDH13-AS2 overexpression.
Combined class I and class III antiarrhythmic effects of sotagliflozin - insights from heterologous expressions systems and atrial cardiomyocytes
Authors: Weirauch L W , Stelzle M S , Paasche A P , Prueser M P , Kraft M K , Wiedmann F W , Zaradzki M Z , Soethoff J S , Arif R A , Karck M K , Frey N F , Schmidt C
Published: October 2024

Background

The demand for more effective treatment options for atrial fibrillation (AF), the most common sustained arrhythmia, is driven by its direct impact on morbidity and mortality. Current therapies, such as antiarrhythmic drugs and catheter ablation, are limited by suboptimal efficacy and adverse effects or periinterventional complications. Therefore, safe and effective treatment of AF remains a major unmet clinical need. Sodium-glucose cotransporter (SGLT) inhibitors have shown promise in reducing the incidence of AF in large cardiovascular endpoint trials. However, the exact mechanisms of action remain unclear. While only two SGLT2 inhibitors are approved and clinically used in Europe, sotagliflozin, which inhibits both SGLT1 and SGLT2, might also have potential for the treatment of AF.

Purpose

This study aims to elucidate the effects of sotagliflozin on cardiac action potential formation. This understanding could optimise its clinical use, identify new targets for the treatment of atrial fibrillation and ultimately improve the quality of life of patients suffering from AF.

Methods

The effects of sotagliflozin on cardiac ionic currents were assessed using two-electrode voltage clamp measurements of several sodium and potassium channels heterologously expressed in Xenopus laevis oocytes. Subsequently, ascending concentrations of sotagliflozin were administered to isolated human atrial cardiomyocytes during patch-clamp measurements to investigate its effect on atrial action potential morphology.

Results

At a concentration of 100 µM, sotagliflozin showed significant inhibitory effects on three out of ten ion channels tested: NaV1.5 sodium channel current was reduced by 22,3 % %, hERG and KV4.3 potassium currents were reduced by 30,4 % and 22,5 % respectively. Patch-clamp experiments, performed on isolated human atrial cardiomyocytes showed a dose-dependent reduction in upstroke velocity and action potential amplitude by 20,8 % and 13,8 % at a concentration of 20 µM, reflecting class-I-antiarrhythmic effects. Furthermore, a prolongation of AP duration measured at 50 % (APD50) and 90 % (APD90) of repolarization by 57,7 % and 27,9 % at a sotagliflozin concentration of 20 µM was noted, implying additional class-III-antiarrhythmic action.

Conclusions

Sotagliflozin exerts direct inhibitory effects on heterologously expressed NaV1.5, hERG and KV4.3 channels, resulting in a reduction in upstroke velocity and action potential amplitude, as well as a prolongation of APD50 and APD90 levels in isolated human atrial cardiomyocytes. These results extend the pharmacological profile of sotagliflozin and suggest a potential role for the clinical use of combined SGLT1/SGLT2 inhibitors in the treatment of AF.
Dual inhibition of SGLT 1 and 2 alleviates intracellular sodium overload in uraemic cardiomyopathy
Authors: Young M , Miller J J J J , Shattock M J , Eykyn T R , Cullen F , Yaqoob M M , Aksentijevic D
Published: October 2024

Background

Chronic kidney disease (CKD) is a leading cause of mortality with the risk of developing cardiovascular disease higher than progression to kidney failure. The aetiology of uraemic cardiomyopathy is multifactorial and includes metabolic derangement. It was previously shown that elevated intracellular sodium (Nai) is causally linked to cardiac metabolic impairment and that targeting sodium homeostasis can reprogramme cardiac metabolism. Dual inhibition of sodium-glucose co-transporters (SGLT) 1 and 2 is an emerging therapy for CKD, however the impact on intracellular sodium and cardiac metabolism is yet to be elucidated.

Purpose

This study aimed to assess whether dual SGLT1/2 inhibitor sotagliflozin impacts cardiometabolic phenotype in CKD.

Methods

CKD was surgically induced by 5/6 nephrectomy in Wistar rats (n=46) for a period of 4 weeks, with chronic sotagliflozin treatment (5mg/kg) given for the last 3 weeks. Upon termination, hearts were Langendorff-perfused and studied using 31P and 23Na NMR spectroscopy (Sham n=7, CKD n=6, CKD+sotagliflozin n=5). To assess the cardiac-specific effect of the drug, acute sotagliflozin treatment (30mM) was delivered in the Langendorff perfusate to an additional group of CKD hearts (n=5). Furthermore, the effect of chronic sotagliflozin treatment on in vivo cardiac function (echocardiography n=23) and metabolic profile (1H NMR spectroscopy n=15) was assessed.

Results

CKD animals were characterized by significant renal dysfunction (2-fold increase in urea and creatinine vs sham, p<0.05), and HFpEF (EF(%) sham 82±2 vs CKD 80±1 p>0.05; diastolic dysfunction (E/A) sham 1.39±0.03 vs CKD 1.11±0.01, p=0.0018). Myocardial Nai was significantly elevated in CKD animals compared to sham controls (TQF 23Na 3.04±0.18 vs 2.01±0.32 arb units, p=0.03). Chronic sotagliflozin treatment didn’t impact renal or cardiac dysfunction. Neither chronic nor acute drug treatment impacted baseline CKD cardiac energetics (PCr/ATP sham 1.53±0.22, CKD 1.25±0.13, chronic 1.16±0.14, acute 1.18±0.13, p>0.05; ΔGATP (kJ/mol) sham -64±4, CKD -66±8, chronic -56±1, acute -59±3, p>0.05). However, both chronic and acute sotagliflozin treatment normalised the increased myocardial Nai (TQF 23Na chronic 2.17±0.12, acute 2.23±0.14 arb units, p>0.05 vs sham). Chronic drug dosing altered the systemic metabolic profile of CKD animals reducing the levels of circulating triglycerides and FFA (p<0.05) as well as altering metabolomic profile of liver, muscle and kidneys.

Conclusion(s)

We have shown that uraemic cardiomyopathy is characterised by HFpEF, elevated myocardial Nai and altered systemic metabolic profile. Both chronic and acute treatment with dual SGLT1/2 inhibitor sotagliflozin normalised myocardial Nai. The alleviation of Nai load could lead to improved cardiometabolic outcomes in CKD.
Reward system activation improves recovery from acute myocardial infarction
Authors: Haykin H , Haykin H , Avishai E A , Krot M K , Ghiringhelli M G , Debby-Azulay H D , Rolls A R , Gepstein L G
Published: October 2024

Introduction

Psychological processes play a crucial role in the recovery from acute myocardial infarction (AMI), yet the underlying mechanisms of these effects remain elusive. Here, we focused on the reward system, which mediates positively valenced emotions, motivation, and expectation and is based mainly on dopaminergic neurotransmission from the ventral tegmental area (VTA).

Purpose

The study examines the impact of activating dopaminergic neurons in the VTA following AMI in mice, aiming to understand the extent and mechanisms through which motivational brain network contribute to cardiac recovery process.

Methods

We used Designer Receptor Exclusively Activated by Designer Drugs (DREADDs) to target neuronal activity. We performed an AMI model and assessed cardiac function using cardiac magnetic resonance (CMR). We used histology, immunohistochemistry, proteomic analysis, ELISA, reverse transcription-PCR, endothelial tube formation assay, and pseudorabies-virus neuronal tracing to investigate the mechanism.

Results

Chemogenetic activation of dopaminergic neurons in the VTA significantly modified the remodeling processes by reducing scar tissue (Fig. 1a) and increased vascularization (Fig. 1b) 15 days following AMI, leading to improved left-ventricular performance compared to control (Fig. 1c). These effects were mediated through several physiological mechanisms, including alterations in immune activity, and reduced adrenergic inputs to the liver. We further demonstrate an anatomical connection between the VTA and the liver that follows AMI manifested by altered transcription of complement component 3 (C3). We established the essential role of C3 in endothelial tube formation in vitro and its necessity in the beneficial effects of VTA activation on the recovery from AMI in vivo (Fig.2).

Conclusion

These findings establish a causal connection between a motivational brain network and recovery from AMI, introducing potential new therapeutic avenues for intervention.Effects of VTA activation on cardiac recMechanism of VTA effects
sRAGE attenuates myocardial ischemia-reperfusion injury by recruiting Tregs through chemokine CCL4
Authors: Zhang W , Jiang X , Liu J , Yu P , Zeng X , Guo C
Published: October 2024

Background

Immune response has recently been shown to play a crucial role in myocardial ischemia-reperfusion (MIR). Our previous studies have confirmed that soluble receptors for advanced glycation end-products (sRAGE) could reduce myocardial ischemia-reperfusion injury (MIRI) by increasing the number of regulatory T cells (Tregs). However, the precise mechanism is still incompletely understood.

Purpose

The present study aimed to determine which kind of chemokines play a major role in recruiting Tregs during MIRI. The potential mechanism of sRAGE on the chemokine production was further explored.

Methods

We modeled mice with cardiac-specific overexpression of sRAGE by crossing the floxed sRAGE mouse model with the cardiomyocyte-specific Cre transgenic mouse. Experimental models were established utilizing left anterior descending coronary artery ligation in mice and oxygen-glucose deprivation/reoxygenation (OGD/R) in vitro. Transcriptomics, quantitative real-time polymerase chain reaction, immunohistochemistry, flow cytometry, and western blotting were used to determine the chemokines and related pathways involved in the recruitment of Tregs during MIRI.

Results

Using transcriptomics, quantitative real-time polymerase chain reaction, and western blotting approach, we identify the chemokine CCL4 plays a crucial role during regulatory T cell recruitment in the heart after MIRI. In vivo studies show that depletion of CCL4 reduces Tregs infiltration into the ischemic myocardium, subsequently deteriorating cardiac injury post-MIR. Mechanistically, sRAGE promotes cardiomyocytes to express and secrete CCL4 via the JNK/AP-1 pathway in vitro. Selective inhibition of the JNK/AP-1 pathway could decrease CCL4 production in sRAGE overexpression cardiomyocytes.

Conclusions

sRAGE induces Tregs recruitment and alleviates MIRI via the JNK/AP-1/CCL4 pathway, preserving myocardial function and impeding inflammation and apoptosis, providing new perspectives for immunotherapeutic targets in MIRI treatment.
Macrophage-expressed coagulation factor 7 promotes adverse cardiac remodeling
Authors: Garlapati V , Luo Q , Posma J , Aluia M , Nguyen T S , Molitor M , Finger S , Harms G , Bopp T , Wolfram R , Wenzel P
Published: October 2024

Background

Excess fibrotic remodeling leads to cardiac dysfunction in ischemic heart disease and is driven by MAP kinase-dependent transforming growth factor-ß1 (TGF-ß1) activation by coagulation signaling of myeloid cells. How coagulation-inflammatory circuits can be specifically targeted to achieve beneficial macrophage reprogramming after myocardial infarction (MI) is incompletely understood.

Methods

Mice with permanent ligation of the proximal left anterior descending artery (LAD) were used to model ischemic heart disease and analyzed by single cell RNA sequencing, protein expression changes, confocal microscopy, and longitudinal monitoring of recovery. We probed the role of the tissue factor (TF)-factor 7 (F7)-integrin ß1-protease activated receptor (PAR) 2 signaling complex by utilizing genetic mouse models and pharmacological intervention.

Results

Cleavage-insensitive PAR2R38E and myeloid cell integrin ß1-deficient mice had improved cardiac function after MI compared to controls. Proximity ligation assays of monocytic cells in the infarcted myocardium demonstrated that colocalization of F7 with integrin ß1 was diminished in monocyte/macrophage F7-deficient mice. F7fl/fl CX3CR1Cre relative to littermate control mice showed reduced TGF-ß1 and MAP kinase activation, as well as cardiac dysfunction after MI, despite unaltered overall recruitment of myeloid cells into the infarct zone. Single cell mRNA sequencing of CD45+ cells 3 and 7 days after MI uncovered a trajectory from ischemic myocardium-recruited monocytes to inflammatory TF+/F7+/TREM1+ macrophages. As early as 7 days after MI, macrophage F7-deletion led to an expansion of Olfml3+ macrophages with a reparative phenotype and, conversely, to a reduction of TF+/F7+/TREM1+ macrophages, which were also attenuated in activation-resistant PAR2R38E mice.

To demonstrate the therapeutic potential of inhibiting the TF-F7-PAR2 signaling complex, we injected a specific monoclonal anti-TF antibody that lacks anticoagulant activity. Short-term treatment from day 1-5 after non-reperfused MI improved cardiac dysfunction, decreased excess fibrosis, attenuated vascular endothelial dysfunction, and increased survival 28 days after MI.

Conclusion

Extravascular TF-F7-PAR2 complex signaling drives inflammatory macrophage polarization in ischemic heart disease. Targeting this signaling complex for specific therapeutic macrophage reprogramming following MI attenuates cardiac fibrosis and improves cardiovascular function.
Collagen VI and endotrophin: potential extracellular matrix targets to attenuate adverse cardiac remodeling post-infarction
Authors: Garcia-Arguinzonis M , Vilahur G , De Santisteban V , Badimon L , Padro T
Published: October 2024

Background

The extracellular matrix (ECM) is a key component of cardiac repair after myocardial infarction (post-MI). After tissue damage following myocardial ischaemia, the ECM becomes enriched in proteases that are known to induce partial enzymatic cleavage and fragmentation of native structural and fibrillar macromolecules such as collagens. While some of the resulting protein fragments (matrikines) may have beneficial effects by mediating an active scar formation, other such as the collagen VI (COL6) proteolytic component "endothrophin" has been related to deleterious evolution and fibrosis, particularly in the presence of cardiovascular risk factors.

Purpose

We aimed to investigate the collagen profile in the structural fraction of ECM (enriched in fibrillar and structural proteins) of post-MI hearts at late evolution stages after MI.

Methods

The ECM protein profile was investigated in the ischemic (ISCH) and non-ischemic (NISCH) zones of the post-MI myocardium in pigs. MI was induced by closed-chest balloon occlusion of the LAD for 90 minutes. Studies were performed in normo- and hypercholesterolemic (diet induced) animals. Myocardial tissue was obtained 30d after MI. ECM proteins were selectively extracted, identified by LC/MS-MS and validated by Western Blot. Plasma levels of endothrophin were quantified by ELISA. Myocardial tissue from the same areas was characterized by immunohistochemistry.

Results

The ECM in the ISCH zone showed a differential signature of 39 proteins (14 upregulated / 25 downregulated) 30 days after MI compared to the NISCH area of the same animals. In the ISCH region, significant changes in collagen referring to COL6A3 and COL6A1 (increase) and COL6A2 and COL6A6 (decrease) were detected. COL6A3 protein content was 1.8fold higher (p=0.01) in the ISCH-ECM. A similar increase was found in normo- and hypercholesterolemic animals. A higher trend in COL6A3 levels was found in the ISCH- vs, the NISCH-zone in the myocardial cellular fraction suggesting that the COL6 ECM-related increase in ISCH is produced at a transcript level. Importantly, the COL6A3 found in the ECM did not contain the protein peptide (C5 domain) corresponding to endotrophin matrikine, suggesting its cleavage and release. Using a specific ELISA, hypercholesterolemic animals showed significantly higher endothrophin plasma levels than normolipemic controls (p=0.003) even in the absence of MI.

Conclusion

Our results evidence an increase in COL6A3 content in the ECM of the ischemic myocardium at 30 days post-MI. COL6A3 is depleted in a peptide of the C5 domain corresponding to endotrophin, found to be enhanced in hypercholesterolemia. These findings support a role for collagen type VI in adverse cardiac remodeling post-MI.
Interleukin-22 inhibits right ventricular remodelling by a unique mechanism in a pulmonary artery constriction model
Authors: Shibao K , Yasukawa H , Akagaki D , Okabe K , Shima K , Fukumoto Y
Published: October 2024

Background

Fibrosis and remodeling of right ventricle (RV) are associated with prognosis in patients with RV failure. Interleukin-22 (IL-22) is a member of the IL-10 cytokine family, play roles in tissue protection and wound repair. We have previously shown that IL-22 plays an important role in cardiac remodeling after acute myocardial infarction; however, the role of IL-22 in RV fibrosis and remodeling remains elusive.

Purpose

We investigated the role of IL-22 in the pathogenesis of RV remodeling during pressure overload.

Methods

Pulmonary artery banding (PAB) is performed by placing a 6-0 suture around the pulmonary artery over a 24 G needle in wild type mice (C57BL/6J) and IL-22 knockout mice (IL-22 KO). Only mice with moderate pulmonary artery stenosis (peak pressure gradient across the pulmonary band: 25–40 mmHg at 1 week after surgery by echo Doppler) were included in the study protocol. Four weeks after PAB, RV function was measured by echocardiography and real-time PCR analysis was performed to measure Col1a1, Col3a1, BNP, and transcriptome analysis was performed. Survival confirmation of mice with moderate pulmonary artery stenosis was also performed up to 56 days.

Results

IL-22 KO mice had significantly higher 56-day mortality rate compared with WT mice after PAC (p<0.01). Four weeks after PAB, IL-22 KO mice showed markedly increased RV weight (IL-22 KO mice vs. wild-type mice; RV/LV+IVS: 0.48±0.05 vs 0.40±0.02, P=0.002). IL-22 KO mice exhibited significant RV enlargement and RV dysfunction 4 weeks after PAB. (IL-22 KO mice vs wild type mice; RVEDV: 10±2.3mm2 vs 12.9±3.1mm2, P=0.005, TAPSE: 10.8±1.8mm vs 9.5±1.7mm, P=0.04, FAC: 35.2±7.8% vs 23±6.0%, P<0.001). The expressions of Col1a1 and Col3a1 were no difference between in IL22-KO mice and Wild mice. However, BNP, a cardioprotective marker, tended to be lower in IL-22KO mice. The transcriptome analysis also showed that the group of fibrosis-related genes that are elevated in PAC in WT mice were less elevated in IL-22 KO mice.

Conclusion

These results suggest that IL-22 is involved in RV remodelling during RV pressure loading without a general mechanism.
Incidence of acute cellular and antibody mediated rejection in heart transplantation from a tertiary care center
Authors: Mallick S K , Arava S , Singh S P , Vij A , Hote M P , Seth S , Ray R
Published: October 2024

Background

Heart transplant greatly increases the survival in patients with end stage heart failure. However, cardiologists and cardiothoracic surgeons face difficulty in detecting rejection at times. Post-cardiac transplant surveillance biopsy remains the gold standard for diagnosing rejection and therapeutic intervention.

Purpose

The objective is to report our experience of monitoring rejection by endomyocardial biopsy (EMB).

Methods

The index study is an ambispective one where we evaluated all the patients who underwent heart transplant at our center from 1994 to 2021.

Results

Heart transplant were carried out in 72 patients within a period of 28 years, with majority being done after 2016. The patients mean age was 31.6 years (10-59 years). The commonest indication was dilated cardiomyopathy (67%) followed by ischemic cardiomyopathy (2.5%), congenital heart disease (7%) and other miscellaneous causes (5%). The mean age of donors was 31.8 years (14-55 years). Eight of the 72 patients died without any post cardiac transplant endomyocardial biopsy. The remaining 64 patients underwent cumulatively a total of 271 endomyocardial biopsy procedures. Majority of endomyocaridal biopsies contained three cores (range 1-7). First EMB procedure was performed in 17 (26.6%) patients within first seven days, 35(53.1%) within the second week and 12(20.3%) after 14 days. Histologically rejection was identified in 42 (66%) patients. Thirty-nine (61%) had only acute cellular rejection (ACR), one (1.6%) had only antibody mediated rejection (AMR) and two (3.1%) had mixed ACR and AMR. These 42 patients with rejection had a total of 90 (33.2%) episodes of ACR detected in 271 endomyocardial biopsies. Out of 90 ACR, 44 (48.9%) occurred in less than six months; seventeen (18.9% of 90 ACR) episodes occurred between six and 12 months and 29 (32.2%) after one year. The mean of presentation of first ACR episode was 36 weeks (range 5 days to 3.6 years). There were 172 endomyocardial procedures within one year of transplantation of which 61 (35%) showed rejection. Ninety-nine endomyocardial procedures were performed one year of transplantation of which 29 (29%) showed rejection. Most common grade in both the time period was ACR grade 1R (International Society for Heart and Lung Transplantation (ISHLT) 2004) (72.1% in ≤1 year and 65.5% in >1 year) followed by 2R and 3R. Thirty-eight (90.4%) of 42 patients expired in less than five years since transplantation. Two (4.8%) patients survived between five to ten years post transplantation and two (4.8%) survived more than ten years. The longest cardiac transplant recipient had completed 23 years since cardiac transplant and is alive till date.

Conclusions

EMB being the gold standard for cardiac allograft rejection, the cardiac pathologists need to be aware of the various grades of rejection including mixed rejection.Acute cellular rejection (ISHLT 2004)
Comparative effectiveness of pulsed field ablation and cryoballoon ablation for atrial fibrillation: a systematic review and meta-analysis
Authors: Freitas M , Lucena L , Guida C , Bulhoes E , Vieira R , Donadon I , Barbalho M , Vilela M , Cazeiro D , Brito D , Dantas J
Published: October 2024

Background

Cryoballoon ablation (CBA) is a well-established technique for treating atrial fibrillation (AF), with a low complication rate related to cryoenergy-induced lesions. Pulsed Field Ablation (PFA) has emerged as a non-thermal alternative, with promising potential to further reduce complications.

Purpose

We aim to conduct a systematic review and meta-analysis comparing the efficacy and safety of PFA versus CBA in patients with AF.

Methods

We conducted a systematic search across PubMed, Embase, and the Cochrane Central Register of Controlled Trials to identify randomized controlled trials (RCTs) or observational studies published between 2013 and 2024 that compared PFA and CBA ablation techniques in patients with AF. Statistical analysis was carried out utilizing RevMan 5.1.7. Heterogeneity was assessed with I² statistics; p-values inferior to 0.05 and I²>25% were considered as significant heterogeneity.

Results

A total of 840 studies were screened in the initial search, and 26 studies were selected for full-text review based on inclusion criteria. Out of these, 8 observational studies involving 3,113 patients were included. Among these, 1,088 (34.9%) underwent PFA, while 2,025 (65.0%) underwent CBA. Periprocedural complications were significantly lower in patients who underwent PFA compared with the CBA group (RR 0.46; 95% CI 0.29 to 0.74; p=0.001; I²=1%; Fig. 1A). Furthermore, there was a significant reduction in procedure time in the PFA group (MD -8.21 min; 95% CI -13.74 to -2.69; p=0.004; I²=96%; Fig. 1B). However, no statistically significant difference was observed among groups regarding fluoroscopy time (MD 1.63 min; 95% CI -0.22 to 3.48; p=0.08; I²=92%; Fig. 2A), or recurrence of atrial arrhythmias after the blanking period (RR 0.84; 95% CI 0.68 to 1.04; p=0.12; I²=6%; Fig. 2B). Additionally, a subgroup analysis involving only propensity score-matched cohorts was conducted but didn’t show any statistical difference for any outcomes evaluated.

Conclusion

In this systematic review and meta-analysis, PFA was associated with a lower risk of periprocedural complications and a shorter procedure duration, compared to CBA in patients with AF. Additionally, it didn’t show a potential for reducing the recurrence of atrial arrhythmias, and the data regarding fluoroscopy time were inconclusive.Primary outcomesSecondary outcomes
SGLT2 inhibitor empagliflozin prevents anthracycline-induced cardiotoxicity in a large-animal model
Authors: Medina-Hernandez D , Galan-Arriola C , Cadiz-Barrera L , Higuero-Verdejo M , Lopez-Martin G , Sanchez-Gonzalez J , Ibanez B
Published: October 2024

Background

While the development of cancer therapies has significantly improved the survival of cancer patients, more than 35% of cancer survivors treated with anthracyclines will develop cardiotoxicity. There is currently a lack of preventative therapies against anthracycline-induced cardiotoxicity (AIC). AIC is characterised by a disruption in myocardial metabolism. Sodium-glucose co-transporter (SLGT2) inhibitors represent a potential therapy as they have been shown to modulate myocardial substrate utilisation in other heart-failure contexts.

Purpose

We developed a highly translational large-animal model to test the ability of SGLT2 inhibitor empagliflozin to prevent AIC at different doses.

Methods

Female Large-White pigs were randomised (2:1:1) to no treatment (AIC-control group) or empagliflozin at a daily oral dose of 10 mg or 20 mg, beginning at the start of the doxorubicin regimen and continuing to 21 weeks. All pigs received 6 doses of doxorubicin intravenous infusion (25 mg/m2/every 3 weeks) with 2 additional off-dose visits. Pigs were assessed by cardiac magnetic resonance (CMR) and MR spectroscopy (MRS) at baseline and every 3 weeks for 21 weeks. At the end of 21-week follow-up, the LV was collected and processed for analysis by transmission electron microscopy (TEM) and mitochondrial respirometry.

Results

The primary outcome, final LVEF, was significantly higher in pigs receiving 20 mg empagliflozin than in the AIC-control group (57.5% (IQR=55.5, 60.3) vs 47.0% (40.8, 47.8); p=0.027). Final LVEF in pigs receiving 10 mg empagliflozin was 51% (46.5, 55.5) (p=0.020 vs 20 mg empagliflozin). The 20 mg empagliflozin dose prevented the doxorubicin-induced change in cardiac metabolic substrate utilisation and was associated with preservation of myocardial energetics (ATP/PCr ratio on MRS). TEM revealed disrupted mitochondrial structure and cristae density in AIC-control pigs, while mitochondria respirometry detected deteriorated respiratory function. Both mitochondrial structure and function were preserved in empagliflozin-treated pigs, with the 20 mg dose having the strongest cardioprotective effect.

Conclusion

The results from our translational large-animal model provide strong evidence that SGLT2 inhibition with empagliflozin prevents AIC by preserving LV systolic function and energetics as well as mitochondrial structural integrity and function. Our results establish the basis for clinical trial testing SGLT2i therapy in patients at high risk of AIC.
Mammalian Diaphanous-related formin 1 (mDia1) inducing diabetic cardiomyopathy by regulating metabolic reprogramming
Authors: Dang M , Jin J , Wang Y
Published: October 2024

Background

Mammalian Diaphanous-related formin 1 (mDia1) is one of the formin family proteins and acts as a rho-effector molecule which is involved in actin organization. Previous studies showed that mDia1 aggravated compensatory cardiac hypertrophy in response to pressure overload. However, the role of mDia1 in diabetic cardiomyopathy and its mechanisms remain elusive.

Objective

To clarify the mechanisms of mDia1 in DCM through mediating metabolic reprogramming.

Methods

5-week-old of mDia1 ko mice were used to induce DCM by feeding high-fat diet (HFD) for 16 weeks. The blood glucose and body weight of the mice were measured regularly. Echocardiography was used to evaluate cardiac function. HE and WGA staining were used to evaluate hypertrophy of cardiomyocytes. Transmission electron microscopy and mitotracker staining were used to observe the morphology of mitochondrial in heart. Proteomics and metabolomics were used to explore the metabolites in hearts of mice. Primary neonatal cardiomyocytes were isolated and stimulated with 35 mM glucose and 250 μM palmitate in vitro. Adenovirus expressing shRNA of mDia1 was used to knockdown mDia1 in cardiomyocytes. Mitochondria were isolated from primary cardiomyocytes to verify the translocation of mDia1. Co-Immunoprecipitation was used to confirm the interaction between mDia1 with succinate dehydrogenase subunits A (SDHA).

Results

mDia1 had no effects to blood glucose and body weight after 16-week HFD feeding. Loss of mDia1 ameliorated diastolic dysfunction in DCM mice with shorter isovolumic relaxation time (IVRT) and increased E/A ratio in left ventricular. In addition, cardiac hypertrophy was significantly improved in mDia1 ko mice. mDia1 deficiency tended to maintain the normal mitochondrial structure. Results of proteomics showed that mDia1 was involved in mitochondrial function, ATP synthesis and metabolic pathways. Metabolomics showed that HFD induced accumulation of α-ketoglutaric acid in wild type mice compared with mDia1 ko mice. However, mDia1 ko significantly increased fumaric acid but did not affect succinyl-CoA and succinic acid compared with wild type mice. Furthermore，we found that mDia1 translocated into mitochondrial with high glucose and palmitate stimuli. Results of CoIP showed mDia1 interacted with SDHA both in cardiomyocytes and HEK293 cells.

Conclusions

mDia1 deteriorated DCM through translocating into mitochondrial and interacting with SDHA, a process dependent on metabolic pathways.Figure of Mechanism