bluuebunny/crossref_metadata_2025 huggingface dataset:
Impact of referral diagnosis on diagnostic yield in 1-day vs 7-day and 14-day extended wear holter
Authors: Parikh P , Grigoriadis C J , Dunn A , Norlock V , Gillstrom G , Wadhwa M
Published: November 2023

Background

Use of Extended Wear Holter (EWH) with monitoring durations up to 14 days has become common in clinical practice. When ordering a monitor, a referral diagnosis (RefDx) is provided to indicate the reason for the study. We previously presented data showing EWH monitors, irrespective of RefDx, provided a 2x diagnostic yield (DY) at 7d and 2.5x DY at 14d when compared to 1d Holter.

Purpose

To determine the impact of RefDx on DY for clinically significant arrythmias (CSA) using ePatch EWH monitoring for 1d, 7d, and 14d.

Methods

From October - December 2021, retrospective data was analyzed for RefDx (ICD-10 diagnosis code) and DY of CSA for ePatch with 1d, 7d, and 14d of analyzable data. RefDx were categorized as Common and Uncommon. Common RefDx included ICD-10 codes which represented at least 1.5% of total referrals on 1d monitors, while all RefDx with <1.5% were grouped together as Uncommon. CSA was defined as Bradycardia < 40 bpm, Pause ≥ 3 seconds, ≥ 2nd Degree AV Block, AF > 30 sec, SVT > 160 bpm for ≥ 3 beats, and VT > 100 bpm for ≥ 3 beats. DY was calculated as the percentage of patients with at least one CSA over the monitoring duration. Patients with Chronic AF were excluded from this analysis.

Results

RefDx for patients who wore ePatch for 1d (n=21,505), 7d (n=9,252), and 14d (n=11,269) are shown in the Table. Irrespective of monitoring duration, the most frequent RefDx was Palpitations (>35%).

For each RefDx, the DY for any CSA at 1d vs 7d, 1d vs 14d, and 7d vs 14d was statistically significant (see Figure). Across all RefDx, DY increased 1.7x - 2.8x when comparing 1d to 7d, 1.9x - 4.2x when comparing 1d to 14d, and 1.1x – 1.5x when comparing 7d to 14d. For 1d, the lowest DY was for RefDx Chest Pain (13%) while the highest was for RefDx Bradycardia (43%). The DY benefit of extending monitoring from 1d to 7d was normally distributed across RefDX, while the DY benefit of extending monitoring from 7d to 14d was greatest (1.5x) for Chest Pain, and the DY benefit of extending from 1d to 14d was greatest (≥3.4x) for Chest Pain, Tachy Unspecified, Palpitations, Syncope/Collapse, and Dizziness/Giddiness. Notably, the most common RefDx of Palpitations demonstrated a DY of only 17% at 1d that increased 2.5x at 7d and 3.4x at 14d.

Conclusion

The impact of extending monitoring duration from 1d to either 7d or 14d was that a disparity in additive DY was observed when analyzing by RefDx. The 5 RefDx with the lowest DY for 1d of monitoring showed increases of ≥2.5x DY at 7d and ≥3.4x Dy at 14d of monitoring. Conversely, the 3 RefDx with the highest DY for 1d of monitoring showed <2X DY at 7d and <2.2x DY at 14d. Our data suggests that the RefDx can potentially be used to guide an optimal duration of telemetry monitoring.
Massive expansion of functional human iPSC-derived cardiomyocytes by concomitant glycogen synthase kinase-3 beta inhibition and removal of cell-cell contact
Authors: Buikema J W , Lee S , Maas R G J , Van Der Velden J , Sluijter J P G , Wu S M
Published: November 2023

Background

Adult mammalian heart has limited capacity for regeneration. Cardiac injuries such as myocardial infarction lead to significant cardiomyocyte (CM) loss and subsequent heart failure with significant morbidity and mortality. Cell-based therapeutic approaches such as injection of stem cell-derived cells or transplantation of engineered cardiac tissue patches have shown promise to re-muscularize the damaged myocardium and enhance cardiac functions.

Rationale: Modulating signaling pathways including Wnt and Hippo can induce cardiomyocyte proliferation in vivo. Applying these signaling modulators to human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) in vitro can expand CMs modestly (< 5-fold).

Results

Here, we demonstrate massive expansion of hiPSC-CMs in vitro (i.e. 100 to 250-fold) by glycogen synthase kinase-3β (GSK-3β) inhibition using CHIR99021 and concurrent removal of cell-cell contact. We show GSK-3β inhibition suppresses CM maturation while contact removal prevents CMs from cell cycle exit. Remarkably, contact removal enabled 10- to 25-times greater expansion beyond GSK-3β inhibition alone. Mechanistically, persistent CM proliferation required both LEF/TCF activity and AKT phosphorylation but was independent from Yes associated protein (YAP) signaling. Engineered heart tissues from expanded hiPSC-CMs showed comparable contractility to those from unexpanded hiPSC-CMs, demonstrating uncompromised cellular functionality after expansion.

Conclusion

In summary, we uncovered a molecular interplay that enables massive hiPSC-CMs expansion for large-scale drug screening and tissue engineering applications.
Stromal cell models of arrhythmogenic cardiomyopathy
Authors: Iengo L , Maione A S , Meraviglia V , Rabino M , Chiesa M , Catto V , Tondo C , Pompilio G , Bellin M , Sommariva E
Published: November 2023

Background

Arrhythmogenic cardiomyopathy (ACM) is an inherited heart disease mainly associated with mutations in genes encoding cardiac desmosomes, most frequently Plakophilin-2 (PKP2). Primary cardiac mesenchymal stromal cells (CMSCs) were shown to contribute to the aberrant ACM cardiac remodeling based on their ability to differentiate into adipocytes and myofibroblasts, thus representing a useful in vitro model for the disease. The limitations associated with access to biopsies and finite passages in culture have been overcome by the advancement of stem cell technology, especially human induced pluripotent stem cells (hiPSCs) and their ability to differentiate towards cardiac stromal population. Specifically, CMSCs from hiPSCs provide a virtually unlimited cell sources that are genetically identical to patient cells of origin.

Purpose

The study compares hiPSC-derived stromal cells (hiPSC-D) and primary stromal cells (PRs), to explore the feasibility to use iPSC-D as an alternative in vitro model to study ACM.

Methods

PRs were isolated from ACM patients and healthy donors (HC). hiPSCs from a PKP2 mutated ACM patient (c.2013delC) and a sex/aged matched HC were used to produce iPSC-D. Flow cytometry analysis and specific differentiation induction (osteogenic, chondrogenic, adipogenic and pro-fibrotic) were performed to confirm mesenchymal lineage and multipotency. qRT-PCR and western blot analysis were performed to evaluate the expression of desmosomes, both gene and protein levels. Illumina transcriptomic profiling of both PRs and iPSC-D were compared and enriched pathways were identified by GSEA.

Results

Both PR and iPSC-D cells expressed mesenchymal surface markers and were able to differentiate towards osteogenic, chondrogenic, adipogenic and myofibroblast lineages. Desmosomal genes and proteins were expressed by both populations, showing a general trend toward lower expression of desmosomal markers in ACM compared to HC cells. Overall, higher expression of desmosomes was observed in iPSC-D stromal cells compared to their primary counterparts, independently on the genotype. As occurs for PRs, ACM iPSC-D exhibited an increased propensity to accumulate lipid droplets and collagen compared to HC cells. Moreover, both cell populations shared a highly similar transcriptomic profile, showing a significant linear correlation (R=0.85). Interestingly, PRs were enriched for genes associated with the immune response and differentiation processes, hinting to the hypothesis that an inflammatory stimulus received from the cardiac tissue might activate the innate immunity and drive the cell differentiation fates in the PRs.

Conclusion(s)

Both PR and iPSC-D cardiac stromal cells obtained from ACM patients can be used to model the disease, where the choice for the most suitable cell populations can be determined by the experimental strategy and/or the availability of human primary material.
Human epicardial adipose tissue-secreted miR-92a-3p regulates myocardial redox state via paracrine signalling: implications for cardiovascular clinical outcomes
Authors: Badi I , Carena M C , Polkinghorne M , Akoumianakis I , Psarros C , Kotanidis C P , Akawi N , Antonopoulos A S , Sayeed R , Krasopoulos G , Srivastava V , Douglas G , Channon K M , Casadei B , Antoniades C
Published: November 2023

Background

Visceral obesity is directly linked to increased cardiovascular risk, including heart failure.

Purpose

We explored the ability of human epicardial adipose tissue (EAT)-derived microRNAs (miRNAs) to regulate the myocardial redox state and clinical outcomes.

Methods

The 5 study arms included 466 patients undergoing cardiac surgery to perform: 1) the discovery phase in which we screened for 351 miRNAs expressed and released from human EAT; 2) correlation analyses between EAT microRNAs and myocardial expression of their targets or myocardial superoxide production in paired EAT/atrial biopsies; 3) genome-wide association screening for miR-92a-3p expression in EAT to test causality/directionality; 4) ex vivo experiments to investigate the underlying mechanisms that were also studied in vitro and in vivo; 5) an 8-year follow-up study to test the prognostic value of the discovered miRNA. Superoxide (O2.-) generation was measured by lucigenin chemiluminescence with NADPH 100μM stimulation as indicator of NADPH-oxidases activity. Vas2870 400 μM (a specific pan-NADPH oxidase inhibitor) was used to obtain the Vas2870-inhibitable O2.- signal which constitutes a more specific index of NADPH oxidase activity. Activation of Rac1, a key NADPH-oxidases subunit, was evaluated by a commercially available kit. Differentiated H9c2 cells were used as an in vitro model of cardiomyocytes. Doxycycline-inducible Wnt5a-overexpressing mice were used for in vivo experiments.

Results

The EAT secretome profiling on study 1 patients led us to identify 3 microRNAs both expressed and released by EAT whose levels in EAT correlated with oxidative stress in human myocardium. Among these miRNAs only miR-92a-3p reduced NADPH-oxidase-derived superoxide (O2.-) in cardiomyocytes (A). A genetic screening identified 7 SNPs that were associated with high miR-92a-3p levels in EAT (EAT-miR-92a-3p SNPs) and were related to lower myocardial superoxide production (B). miR-92a-3p decreased both activation of Rac1 (not shown) and Wnt5a protein levels in vitro (C). Patients with high EAT miR-92a-3p levels had lower WNT5A levels in the myocardium (not shown). Ex vivo, in vivo and in vitro experiments showed opposite effects of Wnt5a on Rac1-mediated NADPH-oxidases activity (D and not shown). Finally, we found an association of miR-92a-3p levels in EAT with lower relative risk of adverse cardiovascular events (E).

Conclusions

EAT-derived miRNAs exert paracrine effects on the human heart. Indeed miR-92a-3p suppresses the Wnt5a/Rac1/NADPH oxidase axis and improves myocardial redox state. EAT-derived miR-92a-3p is related with improved clinical outcomes and is a rational therapeutic target for the prevention and treatment of obesity-related heart disease.
Analytics of long-term premature ventricular complexes using an insertable cardiac monitor
Authors: Chu A F , Rajagopal G , Sarkar S , Landman S , Koehler J , Pothineni N V K
Published: November 2023

Background

Premature ventricular complexes (PVC) are associated with the development of cardiomyopathy, heart failure and pro-arrhythmia. PVC detection using insertable cardiac monitor (ICM) signal processing has recently been introduced. An ICM device capable of monitoring PVC burden over a longer period can be a useful tool for evaluating long-term PVC burden trends.

Purpose

This study evaluated the various long-term PVC burden trends from patients with implanted ICM capable of continuous PVC burden monitoring.

Methods

The ICM implemented PVC detection algorithm uses various combinations of long-short-long RR interval sequence and similarity and differences in R-wave morphology for three consecutive beats to detect single PVC events. For this study, long-term daily PVC burden trends from real-world patients implanted with ICM which has the capability of PVC detection using the implemented device PVC algorithm were analyzed from the de-identified Electronic Health Record (EHR) database linked with ICM device data warehouse. Examples of long-term PVC burden trends showing dynamic burden changes were evaluated and other PVC burden metrics such as maximum PVC burden, mean, and median PVC burden are analyzed from these ICM patients.

Results

PVC burden data from 1919 unique patients with implanted ICM were analyzed. 1814 patients with at least 90 days of available PVC burden were chosen for evaluation. There were 396,033 days with daily PVC burden data points that were evaluated with an average of 218 (SD=72) days with available PVC burden data per patient. The mean PVC burden which was defined as total PVC count divided by total days with ICM PVC detection enabled was 0.8 (SD=2.3) and median PVC burden was 0.02 (IQR 0-0.6). There were 6945 days (1.8%) with PVC burden values ≥10% in 168 unique patients (9.3%). 786 patients (43%) had a maximum daily PVC burden of ≤5% and 155 patients (9%) had maximum daily PVC burden between 6-10%. 107 patients (6%) had a maximum daily PVC burden between 11-20% and 40 patients (2%) had a maximum daily PVC burden >20%. The clinical history comparison of 1814 ICM patients based on a cutoff of a maximum daily PVC burden of 10% is shown in figure 1. Figure 2 shows examples of long-term daily PVC burden(%) trends from four different patients from this dataset showing dynamic PVC burden variabilities that only long-term monitoring of PVCs could detect.

Conclusion

PVC burden monitoring performed utilizing a unique PVC detection algorithm can provide longitudinal and long-term dynamic PVC burden trends in patients implanted with ICM. Dynamic changes in PVC burden may be associated with cardiac pro-arrhythmia or hospitalizations and requires further investigation.
Performance comparisons between two artificial intelligence based premature ventricular complex detection models using insertable cardiac monitor
Authors: Pothineni N V K , Rajagopal G , Sarkar S , Chu A F
Published: November 2023

Background

Premature ventricular complex (PVC) detection using computationally simple techniques has been recently introduced in insertable cardiac monitors (ICMs). An ICM device capable of monitoring PVC burden over a longer period can be a useful tool for evaluating temporal dynamics of PVC burden leading to appropriate and timely clinical interventions in patients with high PVC burden.

Objective

The study evaluated the performances of two artificial intelligence (AI) based PVC detection models to improve PVC detection accuracy in ICMs.

Methods

We developed and evaluated two deep learning convolution neural network (CNN) based PVC detection models which were trained and validated on ICM ECG data. The models were developed and validated using ECG strips stored by ICM implanted in real world patients and collected in the deidentified ICM device data warehouse. All ECG strips were manually annotated for PVCs beat-by-beat. For the first model, only the ICM ECG of three consecutive beats was provided as an input feature whereas for the second model, additional features including QRS morphology differences, RR interval differences, QRS amplitude and slope differences between adjacent beats were extracted and provided as input. The trained AI models were tested on a validation dataset and performance metrics including accuracy, sensitivity and specificity metrics were computed and compared between both models.

Results

The CNN ResNet-18 models with different input features were trained and validated on a dataset of over 90 patient activated ICM ECG episodes which provided a total of 14,354 beats with over 2,200 PVC beats. This dataset provided PVC beats with different morphologies including monomorphic, polymorphic, bigeminy and trigeminy PVCs. The validation dataset consisted of over 4,300 beats including 664 true PVC beats with various morphologies. On the validation dataset, the first model with only ECG as input obtained an accuracy, sensitivity and specificity of 98%, 92% and 99.1% respectively. Whereas for the second model with ECG plus additional features as input, an accuracy, sensitivity and specificity of 98.9%, 97.3% and 99.2% were obtained respectively (Figure 1). Error analysis of both the AI models revealed that the main reason for misidentifying PVC beats was due to the presence of noise around the QRS complex or the T-wave of the beat in the ECG.

Conclusion

Detection of PVCs utilizing a novel AI algorithm is both feasible and capable of exceeding sensitivity and specificity performance metrics of non-AI based algorithms and AI models with manually extracted ECG features provide better performance for targeted PVC detection in ICMs.
EVs derived from physiologically loaded myocardial slices affect remodelling of overloaded myocardial slices
Authors: Nicastro L , Lal A , Kholia S , Chokron D , Downing B , Emanueli C , Terracciano C
Published: November 2023

Introduction

Mechanical load is an important determinant of cardiac output, and excessive mechanical load can lead to heart remodelling, and ultimately failure. Cardiac intercellular communication is partially mediated by extracellular vesicles (EVs), which are 30-200 nm sized nanoparticles acting as shuttles of many bioactive cargoes that modulate cell behaviour, such as microRNAs. Living myocardial slices are an ideal platform to study how mechanical load affect intercellular communication through EVs, as they can be manipulated to obtain different degrees of preload and are an organotypic multicellular platform that retains the electrophysiology and extracellular matrix of the adult myocardial tissue.

Purpose

The aim of the study is to investigate how EVs derived from physiologically-loaded slices modulate the myocardial remodelling of overloaded slices.

Methods

300 µm slices were obtained from the left ventricle of 300-350 g male Sprague Dawley rats. Mechanical load was applied by stretching the slices to either 2.2 or 2.4 µm of their sarcomere length, to recapitulate physiological and overloaded condition, respectively. Following 48-hours of biomimetic culture, force-stretch relationship of the slices was assessed, the tissue collected for RNA and protein extraction and the media of physiological slices was harvested for EVs isolation by size exclusion chromatography. Physiological EVs were then applied onto overloaded slices (1x108 EVs/slice) and cultured for 48-hours to assess force-stretch relationship, and protein and mRNA expression associated with cardiac remodelling. Physiological and overloaded EVs content was analysed through a microRNA array.

Results

Physiological slices had a significantly higher contractility (P < 0.05), higher blood vessel density (P < 0.0001) and lower apoptosis (P<0.01) compared to overloaded slices. Application of physiological EVs on overloaded slices restored their contractility (P<0.05), attenuated apoptosis by reducing protein PARP-1 expression (P<0.05) and reduced blood vessels density loss (P< 0.01), as shown by Isolectin B4 staining and increased mRNA levels of VEGF. microRNAs analysis showed significant downregulation of miR-346 and miR-16-5p and upregulation of miR-143-3p miR-378a-3p in physiological EVs, compared to overloaded. The upregulation or downregulation of these microRNAs was positively associated with cardiac apoptosis and migration of vascular smooth muscle cells.

Conclusions

Physiological EVs affect remodelling in overloaded slices. This is likely due to an effect on apoptosis or to an indirect effect on endothelial cells. Further validation and mechanistic studies with miRNAs selected candidates may reveal interesting therapeutic targets for heart failure.
RTN3 interacted with FABP5 and promoted lipid droplet biogenesis in cardiomyocytes
Authors: Guo D , Zhang M M , Qi B C , Peng T W , Liu M C , Fu F E N G , Wang Y I N G , Hu L A N G , Li Y A N
Published: November 2023

Background

Lipid droplet (LD) accumulation is a notable feature of obesity-induced cardiomyopathy; however, detailed processes governing cardiac LD content remains poorly understood. Reticulon 3 (RTN3), a tubular endoplasmic reticulum (ER) localized protein, was recently identified as a key modulator of lipid metabolism in adipocytes. But its role in cardiac lipid metabolism and the specific mechanism of RTN3 regulating lipid content are currently unknown.

Purpose

To investigate the role of RTN3 in cardiac LD accumulation and the mechanism by which RTN3 affects LD metabolism.

Methods

We labelled LD, free fatty acids (FFA), and nucleus in cardiomyocytes with fluorescent markers and upregulated/downregulated RTN3 expression with adenovirus. The effect of RTN3 on cardiac LD content and biogenesis was detected with confocal microscopy. Then we investigated the interaction partner of RTN3 with co-immunoprecipitation (Co-IP), liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis, and immunofluorescence staining. The function of the complex which RTN3 formed with its interaction protein was then explored.

Results

RTN3 overexpression significantly increased LD content in cardiomyocytes, which phenocopied the effects of palmitate stress. In contrast, palmitate-induced LD accumulation was blocked by RTN3 knockdown. Tubular ER, where RTN3 residents, is the primary organelle regulating LD biogenesis. Based on that, we hypothesized that the effect of RTN3 on LD content could be attributed to its direct effect on LD biogenesis in the ER. Further FFA tracing results demonstrated that RTN3 overexpression and palmitate could markedly increase FFA-LD colocalization, suggesting the LD biogenesis was activated, but this increase in colocalization was absent in cardiomyocytes infected with Ad sh-RTN3. Live cell imaging also revealed that upregulation of RTN3 significantly accelerated cardiac LD biogenesis and downregulation of RTN3 suppressed palmitate-induce LD biogenesis. Then we investigated the interaction partner of RTN3 and found that RTN3 formed a complex with fatty acids binding protein 5 (FABP5), which is an intracellular carrier for long-chain FFAs. Mechanistically, RTN3 directly bound with FABP5, which facilitated the directed transport of FFA to ER and thus promoted LD biogenesis. FABP5 knockdown blocked RTN3-mediated LD biogenesis. Moreover, we discovered that the 1st-194th amino acids of RTN3 was responsible for interacting with FABP5 and promoting LD biogenesis.

Conclusions

Our study reveals a novel mechanism contributing to LD biogenesis in cardiomyocytes. Based on our results, manipulating LD biogenesis by modulating RTN3 may be a potential strategy for treating LD accumulation in obese patients.RTN3 promoted lipid droplet biogenesisRTN3 formed a protein complex with FABP5
Clinical value of insertable loop recorder: single centre experience
Authors: Ahmed M , Morris J P , Bommireddi S M , Cotici A
Published: November 2023

Background

The aetiology of syncope can be challenging to undermine because of its sporadic nature. Implantable loop recorders (ILR) are recommended to be used early in undetermined syncope to establish a symptom – rhythm correlation if cardiac syncope is suspected. Real world diagnostic yield of ILR is not well established.

Purpose

To evaluate the diagnostic yield of ILR devices in patients with undetermined syncope.

Methods

This single centre, retrospective study, reviewed the electronic medical records of all patients, who underwent ILR insertion for syncope from January 2017 to June 2022. Abnormal rhythm strips on ILR were reviewed by a cardiac physiologist and cardiology consultant to determine its significance.

Primary outcome was to determine the total number of cardiac implantable electronic devices (CIEDs) inserted due to ILR findings. Secondary outcomes included (1) a baseline characteristics comparison between the group of patients who had an arrhythmia detected on ILR and patients who had detected no arrhythmia, and (2) the total number of clinical significant events (CSEs), defined as a symptomatic arrhythmia, an asymptomatic arrhythmia deemed clinically related to the patients’ history of syncope, and the absence of an arrhythmia when the patient experienced syncope. Complications of ILR insertion were recorded.

Given that normality was not assumed, a non-parametric analysis was utilised to compare baseline characteristics.

Results

A total of 215 patients were included in this study, of which 43 (20%) had a CIED inserted due to ILR findings. Baseline characteristics were similar between the groups except age (Table 1).

103 (47.9%) of the patients had a CSE (Figure 1). Symptomatic arrhythmias were the most common clinically significant events, out of which a sinus pause of greater than 6 seconds was most prevalent arrhythmia. There were no complications that required early explant of the ILR in this cohort.

Conclusion

ILR detected a clinically significant arrhythmia and the absence of an arrhythmia in 42.7% and 5.1% of patients with undetermined syncope, respectively. ILR use is a valuable diagnostic tool in this cohort of patients. Further studies looking at cost-effectiveness would be beneficial.Baseline characteristicsCSE and arrhythmias
Recruited macrophages incorporate into the resident cardiac macrophage pool and converge into a common phenotype in the healing infarct, but not in the failing heart exposed to pressure overload
Authors: Vico T , Taglinger M , Haacke V , Zehender C , Heinz B , Von Ehr A , Dederichs T , Ehlert C , Dufner B , Westermann D , Hilgendorf I
Published: November 2023

Introduction

Resident macrophages account for 5% of the cells in the healthy heart. In response to cardiac injury, monocytes infiltrate and differentiate into recruited macrophages which function distinctly from the resident subset of primarily embryonic origin. In this work, we aim to identify the extent to which macrophage origin, tissue location and type of cardiac injury determine macrophage phenotypes in ischemic and non-ischemic cardiac injuries over time.

Results and methods

We use a tamoxifen inducible CX3CR1 Yfp CreER/+ :R26tdT/+ mouse line to visualize and quantify fluxes of resident and recruited macrophages, and their respective localizations within the heart following ischemia and reperfusion (I/R) injury and pressure overload after transversal aortic constriction (TAC), respectively.

Interestingly, macrophage numbers peaked during the first week post-surgery in both the local/acute and the global/continuous injury models. Initially, recruited macrophages outnumbered the resident macrophage pool within the infarct area but ultimately reached a 1:1 equilibrium at 4 weeks post I/R injury and at 8 weeks post TAC. In I/R injury, the 1:1 equilibrium established already during the first week in the remote myocardium. Gene expression profiles changed alongside shifts in macrophage ratios. For instance, in the infarct, recruited macrophages showed upregulation of cytokine production and cell chemotaxis pathways at early stage. By 4 weeks post MI their gene expression profiles have converged with those of resident macrophages. Likewise, in the remote myocardium, recruited and resident macrophages assimilated post MI, overexpressing genes associated adaptative immune response activation and cellular homeostasis.

Similar to the I/R injury setting, a 1:1 equilibrium of resident and recruited macrophages established in the hypertrophied and dilated hearts within 4 to 8 weeks post TAC surgery. However, transcriptional profiles remained differentially regulated between recruited and resident cardiac macrophages at all time points tested, and distinct from those observed post I/R injury: recruited macrophages behavior changed from a profibrotic into a proliferative phenotype, meanwhile resident macrophages featured prominently in tissue homeostasis.

Conclusion

Ischemic and non-ischemic, focal and global, acute and chronic cardiac injuries induce a transient peak of monocyte recruitment and macrophage differentiation in and around cardiac lesions which lead to a permanent integration of recruited monocyte-derived macrophages into the pool of tissue resident macrophages – albeit at different paces. The type of injury and macrophage localization within the remodeling tissue determine partial or complete override of ontogenic cell programs.
Platelet-derived microRNAs play a pivotal role in cardiac remodeling after myocardial ischemia and reperfusion
Authors: Schuette J , Manke M C , Hemmen K , Schoerg B , Ramos G , Pogoda M , Kollotzek F , Muenzer P , Pichler B , Lang F , Nieswandt B , Heinze K G , Gawaz M , Casadei N , Borst O
Published: November 2023

Background

Platelets can infiltrate ischemic myocardium and are increasingly recognized as critical regulators of inflammatory processes during myocardial ischemia and reperfusion (I/R). Platelets contain a broad repertoire of miRNAs, which, under certain conditions such as myocardial ischemia, may be transferred to surrounding cells or released into the microenvironment. Recent studies could demonstrate, that platelets contribute substantially to the circulating miRNA pool holding the potential for so far undiscovered regulatory functions.

Purpose

The present study aimed to determine the role of platelet-derived miRNAs in myocardial injury and repair following myocardial I/R.

Methods and Results

Combining an in vivo model of myocardial I/R, multimodal in vivo and ex vivo imaging approaches (LSFM, PET&MRI, speckle-tracking echocardiography), in vitro immune cell migration assays and Next-generation deep sequencing analysis of platelet miRNA expression pattern in mice with a MK/platelet-specific knockout of pre-miRNA processing ribonuclease Dicer, the present study discloses a key role of platelet-derived miRNAs in the tightly regulated cellular processes orchestrating LV remodeling after myocardial I/R following transient LCA ligation. Disruption of the miRNA processing machinery in platelets by deletion of Dicer resulted in increased myocardial inflammation, impaired angiogenesis, and accelerated development of cardiac fibrosis, culminating in an increased infarct size by d7 that persisted through d28 of myocardial ischemia and reperfusion. Worsened cardiac remodeling after myocardial infarction (MI) in mice with a platelet-specific Dicer deletion (Dicer Pf4Δ/PfΔ4) resulted in an increased fibrotic scar formation and distinguishably increased perfusion defect of the apical and anterolateral wall at d28 post-MI. Altogether, these observations culminated in an impaired LV function and hampered long-term cardiac recovery after experimental MI and reperfusion therapy. Treatment with the P2Y12 antagonist ticagrelor completely reversed increased myocardial damage and adverse cardiac remodeling observed in Dicer Pf4Δ/Pf4Δ mice.

Conclusions

The present study discloses a critical role of platelet-derived miRNA in myocardial inflammation and structural remodeling processes following myocardial ischemia and reperfusion.
Time-course of cardiac gene profiles post-acute lung injury
Authors: Wang H , Steven J Lisco S J L , Irving H Zucker I H Z
Published: November 2023

Patients suffering from acute lung injury (ALI) are at high risk of developing cardiac arrhythmias. In a recent study we demonstrated that bleomycin (Bleo)-induced ALI in rats exhibited significantly more spontaneous premature ventricular contractions (PVCs) from 1 to 3 weeks post induction. Although spontaneous PVCs disappeared at 4 weeks post ALI, PVCs were evoked again by electrical stimulation of the decentralized stellate ganglia in Bleo-exposed rats but not in normal rats. The molecular mechanisms underlying cardiac arrhythmia both post ALI and during the recovery of ALI remains unclear. In the current study, we aimed to utilize non-biased RNA-Seq analysis to discover the time-course changes in cardiac gene profiles both post ALI and during the recovery from ALI. We created an ALI rat model using a single tracheal instillation of bleomycin (2.5 mg/kg) with saline as a vehicle (Veh) control. Blood Troponin level was measured and compared between Veh- and Bleo-treated rats varied between W1 to Week 4 post Veh/Bleo. Cardiac tissues were harvested at W1 or W4 post Veh/Bleo for RNA-seq analysis and validation by real time RT-PCR. One week post ALI, cardiac Troponin was significantly increased in Bleo-treated rats whereas it was undetectable in Veh-treated rats. Cardiac Troponin was largely restored at W2 and was undetectable at W4 post ALI. RNA-seq analysis of the left ventricle (LV) indicated 85 downregulated and 147 upregulated genes at 1 W post ALI vs. Veh. Gene Ontology (GO) enrichment analysis demonstrated that genes related to chronic inflammatory signaling, cell adhesion signaling, and extracellular matrix (ECM) signaling pathways were upregulated at W1 post ALI, suggesting cardiac inflammation. Furthermore, genes related to oxygen carrier activity, oxygen binding and oxygen transport signaling were also significantly altered, which may be due to acute hypoxia caused by ALI. In contrast to W1, RNA-seq analysis at W4 post ALI detected a smaller number of altered genes, including 11 downregulated and 13 upregulated genes between ALI and Veh groups. Comparing these two RNA-Seq databases, 4 genes (i.e., Phactr3, Tgm1, Drd4, Selplg) were identified as changed in both the post ALI state and during the recovery from ALI. Following real time RT-PCR validation confirmed significant changes in these four gene mRNA expressions in both post ALI and during the recovery from ALI. Phactr3 was downregulated while Tgm1, Drd4 and Selplg genes were upregulated in cardiac tissues at w1 and w4 post ALI. Phactr3 gene encodes a member of the phosphatase and actin regulator protein family. Tgm1 plays an important role in stabilizing intercellular junctions in myocardial microvascular endothelial cells, by modulating β-actin filaments. Drd4 gene encodes the D4 subtype of the dopamine receptor. Selplg gene encodes a glycoprotein that regulates T lymphocytes. Future studies are needed to examine the functional roles of these genes in cardiac injury post ALI.
Implantable loop recorder monitoring in inherited cardiomyopathies: yield of clinically relevant arrhythmias
Authors: Trancuccio A , Kukavica D , Zvielli G , Marino M , Gambelli P , Memmi M , Bloise R , Napolitano C , Mazzanti A , Priori S G
Published: November 2023

Background

Implantable loop recorders (ILRs) are a minimally invasive tool for the diagnosis of arrhythmias in patients with unexplained syncope or asymptomatic arrhythmias. Their use in patients with inherited arrhythmia syndromes has been recommended in the recent 2022 ESC Guidelines (1). However, the clinical utility of ILRs in patients with inherited cardiomyopathies remains unexplored.

Purpose

The purpose of this study was to evaluate the yield of ILR monitoring in detecting clinically relevant events in a single cohort of patients with inherited cardiomyopathies.

Methods

We prospectively collected demographic, clinical, and follow-up data from patients with inherited cardiomyopathies (Arrhythmogenic Cardiomyopathy, ACM; Hypertrophic Cardiomyopathy, HCM; Dilated Cardiomyopathy, DCM; Left Ventricular Non-Compaction, LVNC) who were implanted with an ILR and were followed with telemedicine. The primary endpoint was a clinically significant event recorded with the ILR that required an update to an implantable cardioverter defibrillator (ICD).

Results

The study cohort included 53 consecutive patients (55% males; mean age at ILR implantation 35±14 years) with inherited cardiomyopathies: 33/53 (62%) with ACM, 9/53 (17%) with HCM, 7/53 (13%) LVNC, 4/33 (8%) DCM. None of the patients implanted had previously experienced life-threatening arrhythmias. During a median follow-up of 2.6 years (IQR 1.4-3.7 years) after ILR implantation, clinically relevant arrhythmic events were documented in 9/53 (17%) of patients. Specifically, sustained or non-sustained ventricular tachycardia episodes were detected in 8 patients (5 with ACM, 2 with LVNC, 1 with LMNA-related DCM), whereas one patient with HCM showed symptomatic pauses > 3 seconds due to sick sinus syndrome. In all cases, the detection of such episodes led to the upgrade to ICD implantation.

Conclusions

ILR monitoring of patients with inherited cardiomyopathies identified clinically relevant events in almost one-fifth of patients, suggesting that the use of ILR may substantially improve the management of these patients.

Figure 1. Representative example of clinically actionable event detected by implantable loop recorder (ILR) in a patient with inherited cardiomyopathy. The patient, a 19-year-old male with a definitive diagnosis of Arrhythmogenic Right Ventricular Cardiomyopathy (ARVC) and a pathogenic PKP2 truncating variant, presented with palpitations and vertigo during physical exertion one year after the implantation of an ILR. ILR monitoring documented a rapid tachycardia with a mean frequency of 250 bpm lasting 3 minutes, prompting an indication for an implantable cardioverter defibrillator (ICD).Figure 1.
Cardiac extracellular matrix at microvascular obstruction area after reperfused myocardial infarction: a morphometric study
Authors: Rios Navarro C , Molina-Garcia T , Ortega M , Gavara J , De Dios E , Perez-Sole N , Marcos-Garces V , Merenciano-Gonzalez H , Chorro F J , Bodi V , Ruiz-Sauri A
Published: November 2023

Background

Extracellular matrix (ECM), a structural and biomechanical support to cardiomyocytes, suffers substantial alterations after myocardial infarction (MI). Despite a complete reopening at epicardial level, hypoperfusion within the infarcted myocardium, known as microvascular obstruction (MVO), occurs. Extensive clinical studies and meta-analysis reported the association between the magnitude of MVO and the appearance of adverse remodeling; thus, scrutinizing ECM composition within the infarcted areas with impaired tissue-level reperfusion could provide fundamental insight regarding MVO pathophysiology.

Purpose

To histologically compare ECM composition within the infarct zone regarding the presence or absence of MVO using an experimental swine model of reperfused MI.

Methods

MI was induced in swine (n=10) by transitory 90-min coronary occlusion followed by 7 days of reperfusion. Prior to euthanasia, intracoronary thioflavin-S was infused. Within the infarcted myocardium, regions displaying MVO (thioflavin-S-) or no MVO (thioflavin-S+) were isolated and stained to morphometrically compare ECM composition.

Results

Regarding cellularity, areas with MVO displayed an enlarged presence of neutrophils and lymphocytes, whilst no differences in the amount of macrophages and myofibroblasts were detected in comparison to no MVO-myocardium. Those regions with macroscopic MVO showed lower capillary density than those without MVO. According to the fibers and ground substance, a significant reduction in the extension of type I collagen, laminin, and fibronectin together with an augmentation of glycosaminoglycan were noted in areas showing MVO compared to those without microvascular injury. No changes in the amount of type III collagen was observed.

Conclusion

The ECM composition in infarcted regions with MVO displays a higher presence of inflammatory infiltrate and glycosaminoglycans as well as a reduced amount of microvessels and fibers compared to those areas without microvascular hypoperfusion. These characteristics might underlie the development of adverse ventricular remodeling in MI patients with extensive MVO.
The selective Rho Kinase 2 inhibitor Chroman-1, is a potent vasodilator but does not compromise left ventricular function in-vivo
Authors: Pearce L , Yusof L , He D , Davidson S M , Yellon D M
Published: November 2023

Background

In the field of cardioprotection, focus is turning to limiting microvascular obstruction (MVO) post myocardial infarction. The development of MVO carries a poor prognosis, which is independent of infarct burden [1]. Commonly used vasodilators in the treatment of MVO e.g., Ca2+ blockers, beta-blockers can induce negatively inotropic effects. This is a serious problem as hypotension and shock during acute MI are harmful. Rho kinase (ROCK) inhibitors induce vasodilation by inhibiting the phosphorylation of myosin light chain kinase (MLC2) in vascular smooth muscle (VSMC) [2]. ROCK1/2 inhibitors are known to cause transient negative inotropic effects [3]. However, the potential side-effects of the selective ROCK2i, Chroman-1 (Ch1) are unknown.

Purpose

We aimed to i) Confirm the expression of ROCK2 mRNA in aorta and coronary vasculature, ii) To investigate the vasodilatory properties of Ch1 iii) To confirm that Ch1 does not acutely impair left ventricular ejection fraction (LVEF%) and blood pressure in-vivo.

Methods

RNA scope in-situ hybridisation was performed with a fluorescent, multiplex assay for ROCK1/2 & VSMC mRNA in whole hearts, coronary vasculature and aorta from male SD rats. Thoracic aorta was dissected and 3mm segments suspended into a tissue bath. Vascular rings were constricted with 1µM PE and treated with [10-12]-[10-5]mM Ch1. In a separate set of in-vivo experiments, male SD rats 250-300g were anaesthetised with isoflurane, intubated and ventilated. LVEF% was measured in a closed chest model, following the addition of Ch1 (30-300µg/kg) or DMSO vehicle, administered i.p at 15-minute intervals. Mean arterial pressure (MAP, mmHg) and heart rate (BPM) were measured throughout.

Results

ROCK2 mRNA was localised to myocardium, aorta and coronary circulation VSMC. Figure 1 demonstrates that Ch1 induced dose dependent vasodilation of PE constricted aortic rings vs control (EC50 Ch1 95% CI [5.1e-008, 2.2e-007] n=6). In-vivo, Ch1 at all doses (30-300µg/kg) did not significantly affect LVEF% (p>0.05, n=5) nor average MAP at doses 30-100µg/kg (p>0.05, n=5). At the highest dose, i.e 300µg/kg, Ch1 decreased average MAP (mmHg) compared to control +/-SEM (76+/-7.1 vs 92+/-2.8, p=0.02, n=5), but not below 70mmHg threshold. Ch1 did not significantly affect mean heart rate at any dose (p>0.05, n=5).

Conclusion

The selective ROCK2 inhibitor Ch1, induces potent arterial vasodilation, without compromising LVEF% or haemodynamics. It is therefore, a suitable drug for further preclinical studies of MVO in ischaemia/reperfusion (I/R).Figure 1:Ch1 induces dose dependent vasodilation of aortic rings, **p<0.005, ***p<0.001
S1P content as a characteristic of the cardioprotective properties of HDL in coronary heart disease and diabetes mellitus
Authors: Polzin A M I N , Benkhoff M A R C E L , Dannenberg L I S A , Barcik M A I K E , Schroeder N A T A L I , Weske S A R A H , Vogt J E N S , Wolnitzke P H I L I P , Mourikis P H I L I P , Keul P E T R A , Sarabhai T H E R E S , Zeus T O B I A S , Roden M I C H A E , Kelm M A L T E , Levkau B O D O
Published: November 2023

Background

Many basic science studies have demonstrated the cardioprotective effect of high-density lipoprotein (HDL). At the same time, several clinical studies in which plasma levels of HDL were pharmacologically increased showed no benefit in cardiovascular risk patients. Accordingly, HDL quantity seems to be less important than HDL quality. Many beneficial effects of HDL are attributed to the bioactive sphingolipid S1P in HDL (HDL-S1P).

Purpose

In this study, we are now investigating a) the cardioprotective properties of HDL of different patient groups and b) the possibility of loading S1P to restore cardioprotection that may be lacking.

Methods

HDL was extracted from plasma of diabetic (dmHDL), as well as CHD patients (CHD-HDL) and healthy subjects (healthy HDL) of the same age by ultracentrifugation and injected i.v. into the mouse five minutes before 30 minutes of ischemia (43 mg/KG). This was followed by 24 hours of reperfusion. Systolic cardiac function was determined by ultrasound, infarct size by TTC (2,3,5-triphenyltetrazolium chloride) staining, and S1P levels by mass spectrometry. The level of apolipoprotein M in HDL was determined by ELISA.

Results

Mice treated with healthy HDL showed smaller infarct size and improved ejection fraction (EF) after I/R than control mice (IS: control 42.95±7.40% [n=26] vs. healthy HDL 33.15±4.68% [n=16], p=0.0001; EF: control 33.68±4.49% vs. healthy HDL 40.00±5.74%, p=0.0016). Treatment with CHD-HDL or dmHDL showed no cardioprotective effects compared to control mice (IS: CHD-HDL 42.22±6.53% [n=16], p=0.9863; dmHDL 43.67±7.81% [n=12], p=0.9899). In both cases, the lack of cardioprotection is associated with lowered HDL-S1P levels (total HDL 210.6±32.6 pmol/mg; CHD-HDL 157.6±49.0 pmol/mg, p=0.0073; dmHDL 171.1±64.7 pmol/mg, p=0.0176). Loading the "sick" HDL with S1P restores cardioprotection only in CHD-HDL (IS: CHD-HDL 42.22±6.53% [n=16] vs. CHD-HDL loaded 32.03±6.45% [n=10], p=0.0023), but not in dmHDL (IS: dmHDL 43.67±7.81% [n=12] vs. dmHDL loaded 38.44±5.98% [n=7], p=0.5110). The very low level of apolipoprotein M (ApoM), responsible for S1P binding, could be a reason for this (total HDL 23.45±6.23 µg/mg [n=15] vs. dmHDL 13.79±4.72 µg/mg [n=14], p=0.0007).

Conclusion

We highlighted worsened cardioprotection as another characteristic of HDL dysfunction in CHD and in diabetes. This characteristic was based on a lower HDL-S1P content and could be therapeutically corrected by loading S1P in the case of CHD-HDL. A low content of ApoM in dmHDL may prevent successful restoration of cardioprotection by S1P loading.
New therapeutic strategy targeting regulation of the SERCA2 complex protects from myocardial ischemia-reperfusion injury
Authors: Calejo A , Ostensen E , Aronsen M , Lygren B , Skogestad J , Klaveness J , Sjaastad I , Tasken K
Published: November 2023

7 million people suffer every year from myocardial infarction. There is significant unmet medical need for cardioprotection during MI treatment. The beta-adrenergic receptor/cyclic AMP/protein kinase A (PKA) signalling pathway regulates heart rate and contractility. Activation of cAMP-dependent PKA causes phospholamban (PLB) phosphorylation and its ability to inhibit sarcoplasmic reticulum calcium ATPase (SERCA) is lost. Discrete control of PLB phosphorylation is facilitated by the A-kinase anchoring protein (AKAP) 18g/d, which holds PKA and PLB in close proximity. We are developing a novel drug candidate, named 13M, with a highly selective mechanism of action in cardioprotection currently in pre-clinical phase. Our drug candidate disrupts the AKAP18g/d-PLB protein-protein interaction (PPI) as this may protect from ischemia reperfusion injury (IRI). We used AlphaScreen assay to screen a library of 79,000 small molecular compounds for compounds that disrupted the AKAP18δ-PLB PPI. Following secondary and tertiary screens of sub-libraries, the resulting hits were further characterized and similarity searches carried out to delineate a compound structure for further derivatization. Newly synthesized compounds were further tested in a cell-line based assay to determine compound toxicity. A combination of biochemistry, electrophysiology and heart function experiments demonstrated that the compounds exclusively block the adrenergic effect on SERCA2 without affecting its basal activity. A 40% reduction in infarction size and improved long term cardiac function was observed in an animal disease model for ischemia-reperfusion injury.