Abstract Introduction Mast cells reside in cardiac tissue and release inflammatory mediators upon IgE-mediated activation. While mast cells have been implicated in cardiac remodeling, the relationship between IgE and arrhythmias remains unexplored. We hypothesized that elevated IgE is associated with increased atrial fibrillation (AF) risk and that IgE-dependent mast cell activation promotes cardiac inflammation and sympathetic remodeling. Methods We performed a propensity-matched cohort study using TriNetX comparing patients with elevated IgE (>1000 IU/mL) versus normal IgE (< 100 IU/mL), matched on 29 covariates. The primary outcome was incident AF/flutter over 5 years. In parallel, C57BL/6J mice were sensitized to peanut allergen and fed peanut for 4 weeks. Hearts were analyzed for mast cell density, inflammatory cytokines, tyrosine hydroxylase (TH) as a sympathetic nerve marker, and sinoatrial node (SAN) macrophage infiltration. Results After matching, 54,007 patients per cohort were analyzed (mean age 29). Elevated IgE was associated with increased AF risk (1.54% vs 1.25%; OR 1.24, 95% CI 1.12-1.37, p < 0.0001). In mice, sensitization increased cardiac mast cells 3-fold (p < 0.01), an effect absent in IgE-deficient mice. Notably, much of the increase in mast cells was observed in or near cardiac ganglia and conducting system, key locations in the development or arrhythmia. Sensitized mice demonstrated elevated cardiac IL-1α, IL-6, IL-13, and TNFα. TH staining revealed 7-fold increased right atrial sympathetic nerve density (1.5% vs 0.2% area, p < 0.001). CD68 immunostaining showed increased macrophage infiltration within the SAN of sensitized animals. Conclusion Elevated IgE is associated with increased AF risk in young adults. IgE-dependent mast cell activation drives cardiac inflammation, sympathetic hyperinnervation, and SAN inflammation. These findings identify allergic sensitization as a novel arrhythmia risk factor with therapeutic implications. Funding Source n/a Topic Categories Neuroimmunology (NEUR)

Interpreting bulk RNA sequencing from heterogeneous tissues like the post-myocardial infarction (MI) heart is confounded by dynamic changes in cell-type composition. To address this, we developed a computational approach using single-nucleus RNA sequencing (snRNA-seq) references to estimate and correct for cell-type abundance shifts in bulk transcriptomic data. We applied this method to analyze infarct border zone transcriptomes from wild-type (WT) and cardiomyocyte-specific α1A-adrenergic receptor knockout (cmAKO) mice subjected to MI via left coronary artery ligation or sham surgery. Our analysis revealed exaggerated cardiomyocyte loss and fibroblast gain in cmAKO mice post-MI compared to WT, implicating α1A-ARs in maintaining cellular homeostasis. We then demonstrate the confounding effect of composition changes though simulations: a modest 10% change in the major cell type's abundance caused over 20% of transcripts to appear as differentially expressed genes (DEGs) when composition was ignored. Applying our correction method refined the interpretation of MI-induced transcriptomic changes, attributing many apparent DEGs, particularly those related to metabolism and inflammation, to shifts in cell abundance rather than direct transcriptional regulation. Importantly, the correction also unveiled previously masked biological processes associated with the cmAKO-specific response to MI, including pathways related to cell adhesion, cell cycle regulation, and stress response, highlighting potential intrinsic mechanisms of α1A-AR cardioprotection. RNAscope validation supported the composition-aware findings for key genes. This work presents a robust method for dissecting bulk RNA-seq data from complex tissues and provides refined insights into the cellular and molecular roles of cardiomyocyte α1A-ARs during cardiac injury and remodeling.

Neutrophils, the most abundant leukocytes in human blood, have long been recognized as critical first responders in the innate immune system's defense against pathogens. Some of the more notable innate antimicrobial properties of neutrophils include generation of superoxide free radicals like myeloperoxidase, production of proteases that reshape the extracellular matrix allowing for easier access to infected tissues, and release of neutrophil extracellular traps, extruded pieces of DNA that ensnare bacterial and fungi. These mechanisms developed to provide neutrophils with a vast array of specialized functions to provide the host defense against infection in an acute setting. However, emerging evidence over the past few decades has revealed a far more complex and nuanced role for these neutrophil-driven processes in various chronic conditions, particularly in cardiovascular diseases. The pathophysiology of cardiac diseases involves a complex interplay of hemodynamic, neurohumoral, and inflammatory factors. Neutrophils, as key mediators of inflammation, contribute significantly to this intricate network. Their involvement extends far beyond their classical role in pathogen clearance, encompassing diverse functions that can both exacerbate tissue damage and contribute to repair processes. Here, we consider the contributions of neutrophils to myocardial infarction, heart failure, cardiac arrhythmias, and nonischemic cardiomyopathies. Understanding these complex interactions is crucial for developing novel therapeutic strategies aimed at modulating neutrophil functions in these highly morbid cardiac diseases.

Trametinib (Trm) is a highly selective MEK inhibitor that potently and persistently abrogates ERK1/2 activation. Trm initially was used to treat BRAF V600E-mutated melanoma but its FDA-approved indications are expanding rapidly. Trm generally is well tolerated but it can cause dose-limiting cardiomyopathy and heart failure. Here we characterize a mouse model of Trm cardiotoxicity using complementary in vitro approaches to show that Trm induces mitochondrial dysfunction in cardiomyocytes and some cancer cell types. In vivo , Trm caused contractile dysfunction within 3 days and heart failure within 2 weeks. High resolution respirometry using isolated cardiac mitochondria revealed that Trm compromises oxidative metabolism, in part through blunted activity of Electron Transport System Complexes. Trm-mediated mitochondrial injury led to the release of mitochondrial Damage-Associated Molecular Patterns including mitochondrial DNA in both mice and humans, triggering activation of canonical innate immune pathways including cGAS-STING. In multiple rodent and human cardiomyocyte platforms, Trm diminished mitochondrial respiratory capacity at nanomolar concentrations but this lesion was reversed by expression of a phosphomimetic STAT3-S727 construct. We also found that Trm induced mitochondrial dysfunction in some but not all cancer cell lines, identifying a previously unrecognized effect that could contribute to Trm’s anti-cancer efficacy.

Every year, more than a million people in the United States undergo chemotherapy or radiation therapy for cancer, as estimated by the CDC. While chemotherapy has been an instrumental tool for treating cancer, it also causes severe adverse effects. The more commonly acknowledged adverse effects include hair loss, fatigue, and nausea, but a more severe and longer lasting side effect is cardiotoxicity. Cardiotoxicity, or heart damage, is a common complication of cancer treatments. It can range from mild to severe, and it can affect some patients temporarily or others permanently, even after they are cured of cancer. Dexrazoxane is the only FDA-approved drug for treating anthracycline induced cardiotoxicity, but it also has drawbacks and adverse effects. There is no other type of chemotherapy induced cardiotoxicity that has an approved treatment option. In this review, we discuss the pathophysiology of chemotherapeutic-induced cardiotoxicity, methods and guidelines of diagnosis, methods of treatment and mitigation, and current drug delivery approaches in therapeutic development.

Mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) are a promising treatment for myocardial infarction (MI), but their therapeutic efficacy is limited by inefficient accumulation at the target site. A minimally invasive MSC EV therapy that enhances EV accumulation at the disease site and extends EV retention could significantly improve post-infarct cardiac regeneration. Here, we show that EVs decorated with the next-generation of high-affinity (HiA) heterodimerizing leucine zippers, termed HiA Zippersomes, amplify targetable surface areas through in situ crosslinking and exhibited ~7-fold enhanced accumulation within the infarcted myocardium in mice after 3 days and continued to be retained up to Day 21, surpassing the performance of unmodified EVs. After MI in mice, HiA Zippersomes increase the ejection fraction by 53% and 100% compared with unmodified EVs and phosphate-buffered saline (PBS), respectively. This notable improvement in cardiac function played a crucial role in restoring healthy heart performance. HiA Zippersomes also robustly decrease infarct size by 52% and 60% compared with unmodified EVs and PBS, respectively, thus representing a promising platform for minimally invasive vesicle delivery to the infarcted heart compared to intramyocardial injections.

Background:Recent advances in single cell sequencing have led to an increased focus on the role of cell-type composition in phenotypic presentation and disease progression. Cell-type composition research in the heart is challenging due to large, frequently multinucleated cardiomyocytes that preclude most single cell approaches from obtaining accurate measurements of cell composition. Our in silico studies reveal that ignoring cell type composition when calculating differentially expressed genes (DEGs) can have significant consequences. For example, a relatively small change in cell abundance of only 10% can result in over 25% of DEGs being false positives. Methods:We have implemented an algorithmic approach that uses snRNAseq datasets as a reference to accurately calculate cell type compositions from bulk RNAseq datasets through robust data cleaning, gene selection, and multi-sample cross-subject and cross-cell-type deconvolution. We applied our approach to cardiomyocyte-specific α1A adrenergic receptor (CM-α1A-AR) knockout mice. 8-12 week-old mice (either WT or CM-α1A-KO) were subjected to permanent left coronary artery (LCA) ligation or sham surgery (n=4 per group). Transcriptomes from the infarct border zones were collected 3 days later and analyzed using our algorithm to determine cell-type abundances, corrected differential expression calculations using DESeq2, and validated these findings using RNAscope. Results:Uncorrected DEGs for the CM-α1A-KO X LCA interaction term featured many cell-type specific genes such as Timp4 (fibroblasts) and Aplnr (cardiomyocytes) and overall GO enrichment for terms pertaining to cardiomyocyte differentiation (P=3.1E-4). Using our algorithm, we observe a striking loss of cardiomyocytes and gain in fibroblasts in the α1A-KO + LCA mice that was not recapitulated in WT + LCA animals, although we did observe a similar increase in macrophage abundance in both conditions. This recapitulates prior results that showed a much more severe heart failure phenotype in CM-α1A-KO + LCA mice. Following correction for cell-type, our DEGs now highlight a novel set of genes enriched for GO terms such as cardiac contraction (P=3.7E-5) and actin filament organization (P=6.3E-5). Conclusions:Our algorithm identifies and corrects for cell-type abundance in bulk RNAseq datasets opening new avenues for research on novel genes and pathways as well as an improved understanding of the role of cardiac cell types in cardiovascular disease.

AbstractAimsThe objective of this study was to examine associations between elevated depressive symptoms and increased risk of adverse clinical events patients with heart failure and reduced ejection fraction (HFrEF), as well as the potential contribution of health behaviours.Methods and resultsOne hundred forty‐two men and women with HFrEF were enrolled through heart failure (HF) clinics and followed over time. At baseline and 6 months, depressive symptoms were assessed by the Beck Depression Inventory‐II (BDI‐II) and HFrEF disease activity by B‐type natriuretic peptide (BNP). The Self‐Care of Heart Failure Index (SCHFI) was used to assess HF self‐care behaviours. Proportional hazards regression models assessed the contribution of depressive symptoms and HFrEF disease biomarkers on death or cardiovascular hospitalization. Over a median follow‐up period of 4 years, 42 patients (30%) died, and 84 (60%) had cardiovascular hospitalizations. A 10‐point higher baseline BDI‐II score was associated with a 35% greater risk of death or cardiovascular hospitalization. Higher baseline BDI‐II scores were associated with poorer HF self‐care maintenance behaviours (R = −0.30, P < 0.001) and fewer daily steps (R = −0.19, P = 0.04), suggesting that elevated depressive symptoms may diminish important health behaviours. Increases in plasma BNP over 6 months were associated with worse outcomes. Changes in BDI‐II and plasma BNP over 6 months were positively related (R = 0.25, P = 0.004).ConclusionsThis study confirms that elevated depressive symptoms are associated with an increased likelihood of adverse clinical outcomes in patients with HFrEF. Poor health behaviours may contribute to the adverse association of elevated depressive symptoms with the increased hazard of adverse clinical outcomes.

Background: Venipuncture is the standard technique for blood sample acquisition but requires expertise, larger volumes of blood, and can be challenging in special populations. Capillary sampling may provide an alternative, but its reliability is unclear for assessing hematological parameters, especially following acute exercise. Objectives: The purpose of the study was to compare the agreement and accuracy of complete blood counts (CBC) from capillary and venous blood sampled before, immediately after a single bout of aerobic exercise, and into recovery. An exploratory purpose was to examine potential biological sex differences within the CBC between blood sampling sites. Methods: Recreationally active healthy adults (N = 13 male and N = 13 female) completed three visits including familiarization, graded exercise testing, and a 40-minute exercise bout at 90–98% of ventilatory threshold. Venous and capillary blood samples were collected simultaneously at rest, immediately post-exercise (0 h) and 30-minutes into recovery (0.5 h). Results: White blood cells (WBC), lymphocytes (LYM), and neutrophils (NEUT) were strongly correlated at all timepoints (r > 0.8, p < 0.05) with low bias, and moderate level of agreement (LOA). Mixed cells (MXD), hemoglobin (HGB), and hematocrit (HCT) were strongly correlated at baseline (r > 0.7, p < 0.05), but showed weaker correlations following exercise. There were no differences in mobilization or egress of CBC outcomes between sites by sex except for lymphocyte egress. Males had lower lymphocyte egress in venous versus capillary sampling whereas females had higher lymphocyte egress in venous versus capillary sampling. Discussion: These data indicate capillary sampling is an accurate alternative to venous sampling following acute exercise based on LOA and reliability data.

AIMS:The sympathetic nervous system regulates numerous critical aspects of mitochondrial function in the heart through activation of adrenergic receptors (ARs) on cardiomyocytes. Mounting evidence suggests that α1-ARs, particularly the α1A subtype, are cardioprotective and may mitigate the deleterious effects of chronic β-AR activation by shared ligands. The mechanisms underlying these adaptive effects remain unclear. Here, we tested the hypothesis that α1A-ARs adaptively regulate cardiomyocyte oxidative metabolism in both the uninjured and infarcted heart.METHODS:We used high resolution respirometry, fatty acid oxidation (FAO) enzyme assays, substrate-specific electron transport chain (ETC) enzyme assays, transmission electron microscopy (TEM) and proteomics to characterize mitochondrial function comprehensively in the uninjured hearts of wild type and α1A-AR knockout mice and defined the effects of chronic β-AR activation and myocardial infarction on selected mitochondrial functions.RESULTS:We found that isolated cardiac mitochondria from α1A-KO mice had deficits in fatty acid-dependent respiration, FAO, and ETC enzyme activity. TEM revealed abnormalities of mitochondrial morphology characteristic of these functional deficits. The selective α1A-AR agonist A61603 enhanced fatty-acid dependent respiration, fatty acid oxidation, and ETC enzyme activity in isolated cardiac mitochondria. The β-AR agonist isoproterenol enhanced oxidative stress in vitro and this adverse effect was mitigated by A61603. A61603 enhanced ETC Complex I activity and protected contractile function following myocardial infarction.CONCLUSIONS:Collectively, these novel findings position α1A-ARs as critical regulators of cardiomyocyte metabolism in the basal state and suggest that metabolic mechanisms may underlie the protective effects of α1A-AR activation in the failing heart.

Introduction: Trametinib (Trm) is a highly selective inhibitor of MEK1/2, downstream targets of the RAS signaling pathway that has been used widely for the treatment of BRAF V600E/K-mutant melanoma. Multiple clinical trials are also underway investigating its use in colorectal, prostate cancer, leukemia, and triple negative breast cancer. Trm is generally well tolerated but can be associated with potentially serious cardiotoxicity in 5-20% of treated patients. The specific mechanisms driving Trm-induced cardiomyopathy (TIC) are largely unexplored. The goal of this project is to investigate the role that resident and infiltrating inflammatory cells play in TIC. Hypothesis: Our data suggest that Trm provokes cardiomyocyte immune responses including increased production of inflammatory cytokines, NF-KB signaling, and macrophage infiltration. We hypothesize that this activation occurs through DAMP-driven activation of the TLR and cGAS-STING pathway. Methods: C57BL/6 mice were gavaged with Trm 3mg/kg/day for 3 days prior to sacrifice. Heart tissue was analyzed via RNAseq, confocal microscopy, and flow cytometry. In vitro assays were completed using neonatal rat ventricular myocytes (NRVM). Results: RNAseq data of whole heart tissue from Trm treated mice surprisingly demonstrated increased expression of transcripts in inflammatory pathways. In vitro stimulation of NRVMs with Trm demonstrated increased production of inflammatory cytokines including IFNy and increased NF-KB signaling. Confocal microscopy of Trm treated heart tissue demonstrated increased CD68+ macrophage recruitment compared to control. None of these findings would have been predicted by our previous knowledge of cardiomyocyte MEK biology. Conclusions: Trm treatment is associated with cardiomyopathy in up to 20% of patients. Understanding the mechanisms driving this disease pathogenesis may help reduce likelihood of adverse cardiac events associated with future MEK inhibitor therapy used alone or in combination with other agents. Our novel findings suggest that Trm causes immune dysregulation and inflammation in the heart. Future studies will focus on further characterization of immune cell subsets involved and mechanisms of innate immune activation in TIC.

Clonal hematopoiesis (CH) refers to the expansion of hematopoietic stem cell clones and their cellular progeny due to somatic mutations, mosaic chromosomal alterations (mCAs), or copy number variants which naturally accumulate with age. CH has been linked to increased risk of blood cancers, but CH has also been linked to adverse cardiovascular outcomes. A combination of clinical outcome studies and mouse models have offered strong evidence that CH mutations either correlate with or cause atherosclerosis, diabetes mellitus, chronic kidney disease, heart failure, pulmonary hypertension, aortic aneurysm, myocardial infarction, stroke, aortic stenosis, poor outcomes following transcatheter aortic valve replacement (TAVR) or orthotopic heart transplant, death or need of renal replacement therapy secondary to cardiogenic shock, death from cardiovascular causes at large, and enhance anthracycline cardiac toxicity. Mechanistically, some adverse outcomes are caused by macrophage secretion of IL-1β and IL-6, neutrophil invasion of injured myocardium, and T-cell skewing towards inflammatory phenotypes. CH mutations lead to harmful inflammation and arterial wall invasion by bone marrow-derived cells resulting in poor cardiovascular health and outcomes. Blockade of IL-1β or JAK2 signaling are potential avenues for preventing CH-caused cardiovascular morbidity and mortality.