Cardiac Growth Research Articles

Abstract 4134900: Cellular Crosstalk in Endocardial Fibroelastosis: NRG Signaling Dysregulation Contributes to Fibrosis and Myocardial Dysfunction in Hypoplastic Left Heart Complex

Background: Endocardial fibroelastosis (EFE) is marked by a thickened fibro-elastic layer derived by endothelial-to-mesenchymal transition (EndMT), leading to restrictive myocardial function and poor cardiac growth, complicating biventricular repair in hypoplastic left heart complex (HLHC) patients. Neuregulin (NRG) signaling, crucial in EndMT and myocardial development, is implicated in various cardiomyopathies and heart failure, and its dysregulation may drive EFE pathogenesis. Hypothesis: NRG signaling dysregulation influences the cellular and molecular mechanisms of EFE. Methods: Single nucleus RNA sequencing (snRNA-seq) was conducted on human left ventricular (LV) tissue resected from 7 EFE patients (0.4-15 years) and 4 non-congenital heart disease (CHD) postmortem human controls (0.02-16 years). Differential gene expression was analyzed using the Wilcoxon Rank Sum test. We used CellChat, a computational tool for analyzing intercellular communication networks, to identify signaling changes via quantitative contrasts and joint manifold learning. Results: We identified 11 cell types in 64,963 nuclei. In EFE samples compared to non-CHD samples, NRG1-expressing fibroblasts increased from 3.4% to 21.2% (P<0.0001), and NRG2-expressing cardiomyocytes from 11.9% to 45.9% (P<0.001). CellChat analysis revealed a denser NRG signaling network in EFE, with enhanced interactions among cardiomyocytes, endocardial cells, fibroblasts, mast cells, and smooth muscle cells (Fig. 1A-B). Cardiomyocytes emerged as significant senders of NRG signaling in EFE, while endocardial cells, though still key senders, showed reduced influence. Fibroblasts became strong mediators, receivers, and influencers, smooth muscle cells emerged as prominent receivers, and mast cells gained roles as mediators and influencers (Fig. 1 C-D). These changes indicate a complex NRG signaling network in EFE, highlighting altered cellular interactions driving disease progression. Conclusion: Dysregulated NRG signaling in EFE contributes to pathological cellular crosstalk, fibrosis, and myocardial dysfunction, potentially hindering successful biventricular repair in HLHC patients. Targeting NRG pathways could represent a novel therapeutic approach. These findings offer promising implications for affected patients, potentially improving outcomes. Future research should explore additional molecular pathways involved in EFE and assess the therapeutic efficacy of targeting NRG signaling.

Sex-specific regulation of the cardiac transcriptome by the protein phosphatase 2A regulatory subunit B55α

Protein phosphatase 2A (PP2A) regulatory subunit B55α has been implicated in the transcriptional regulation of cardiac growth and fibrosis by suppressing HDAC5/MEF2 signalling in cardiomyocytes. We created and characterised two mouse models with global or cardiomyocyte-specific disruption of the gene encoding B55α (Ppp2r2a) to conduct the first detailed exploration of B55α in the heart. Global homozygous B55α knockout mice died in utero, while heterozygous mice had thinner left ventricular walls at 12 months, an effect more pronounced in males. At 10–12 weeks of age, cardiomyocyte-specific B55α knockout mice displayed normal cardiac morphology with increased left ventricular collagen deposition, identifying B55α as a negative regulator of cardiac fibrosis. Gene expression analyses demonstrated extensive remodelling of the cardiac transcriptome in male but not female mice, revealing a sexually dimorphic role for B55α in cardiac transcriptional regulation. These findings provide a basis for future work investigating B55α in cardiac stress settings.

Embryonic and Fetal Heart Development Before 12 Weeks of Gestation.

To assess embryonic and fetal cardiac growth and development using transvaginal 2-dimensional sonography before 12 weeks of gestation. Transvaginal scans for first-trimester dating were performed for 131 normal fetuses at 8-11 + 6 weeks of gestation. The basal-apical length (BAL), transverse length (TL), cardiac circumference (ECC), embryonic cardiac area (ECA), global sphericity index (GSI), and cardio-thoracic area ratio (CTAR) were able to be obtained in 105 normal embryos and fetuses. Nomograms for several cardiac parameters including BAL, TL, ECC, ECA, GSI, and CTAR were constructed. BAL, TL, ECC, and ECA increased curvilinearly with advancing gestation (R2 = 0.97406, 0.980396, 0.978359, and 0.920705, respectively, P < .001). GSI (mean, 1.14; SD, 0.10) and CTAR (mean, 15.7%; SD, 3.3%) values were constant at 8-11 + 6 weeks of gestation. There were significant curvilinear correlations between BAL, TL, ECC, and ECA, and crown-rump length (CRL) (R2 = 0.975976, 0.983482, 0.980673, and 0.929936, respectively, P < .001). GSI and CTAR values were not changed with the increase of CRL during this period. Our results provide nomograms for several cardiac parameters which may improve the understanding of embryonic and fetal cardiac growth and development prior to 12 weeks of gestation.

PFKP inhibition protects against pathological cardiac hypertrophy by regulating protein synthesis

Metabolic reprogramming precedes most alterations during pathological cardiac hypertrophy and heart failure (HF). Recent studies have revealed that Phosphofructokinase, platelet (PFKP) has a wealth of metabolic and non-metabolic functions. In this study, we explored the role of PFKP in cardiac hypertrophic growth and HF. The expression level of PFKP was elevated both in pathological cardiac remodeling mouse model challenged by transverse aortic constriction (TAC) surgery and in the neonatal rat cardiomyocytes (NRCMs) stimulated by phenylephrine (PE). In global PFKP knockout (PFKP-KO) mice, cardiac hypertrophy was ameliorated under TAC surgery, while overexpression of PFKP by intravenous injection of adeno-associated virus 9 (AAV9) under the cardiac troponin T (cTnT) promoter worsened myocardial hypertrophy and fibrosis. In NRCMs, small interfering RNA (SiRNA) knockdown or adenovirus (Adv) overexpression of PFKP was employed and the intervention of PFKP showed a similar phenotype. Mechanistically, immunoprecipitation combined with liquid chromatography-tandem mass spectrometry (IP-MS/MS) analysis was used to identify the interacting proteins of PFKP. Eukaryotic translation initiation factor 2 subunit beta (EIF2S2) was identified as the downstream target of PFKP. In the PE-stimulated NRCM hypertrophy model and mouse TAC model, knocking down EIF2S2 after PFKP overexpression reduced the synthesis of new proteins and alleviated the hypertrophy phenotype. Our findings illuminate that PFKP participates in pathological cardiac hypertrophy partly by regulating protein synthesis through EIF2S2, which provides a new clue for the involvement of metabolic intermediates in signal transduction.

Long noncoding RNA AK144717 exacerbates pathological cardiac hypertrophy through modulating the cellular distribution of HMGB1 and subsequent DNA damage response

DNA damage induced by oxidative stress during cardiac hypertrophy activates the ataxia telangiectasia mutated (ATM)-mediated DNA damage response (DDR) signaling, in turn aggravating the pathological cardiomyocyte growth. This study aims to identify the functional associations of long noncoding RNA (lncRNAs) with cardiac hypertrophy and DDR. The altered ventricular lncRNAs in the mice between sham and transverse aortic constriction (TAC) group were identified by microarray analysis, and a novel lncRNA AK144717 was found to gradually upregulate during the development of pathological cardiac hypertrophy induced by TAC surgery or angiotensin II (Ang II) stimulation. Silencing AK144717 had a similar anti-hypertrophic effect to that of ATM inhibitor KU55933 and also suppressed the activated ATM-DDR signaling induced by hypertrophic stimuli. The involvement of AK144717 in DDR and cardiac hypertrophy was closely related to its interaction with HMGB1, as silencing HMGB1 abolished the effects of AK144717 knockdown. The binding of AK144717 to HMGB1 prevented the interaction between HMGB1 and SIRT1, contributing to the increased acetylation and then cytosolic translocation of HMGB1. Overall, our study highlights the role of AK144717 in the hypertrophic response by interacting with HMGB1 and regulating DDR, hinting that AK144717 is a promising therapeutic target for pathological cardiac growth.

Molecular regulation of reversible cardiac remodeling: lessons from species with extreme physiological adaptations.

Some vertebrates evolved to have a remarkable capacity for anatomical and physiological plasticity in response to environmental challenges. One example of such plasticity can be found in the ambush-hunting snakes of the genus Python, which exhibit reversible cardiac growth with feeding. The predation strategy employed by pythons is associated with months-long fasts that are arrested by ingestion of large prey. Consequently, digestion compels a dramatic increase in metabolic rate and hypertrophy of multiple organs, including the heart. In this Review, we summarize the post-prandial cardiac adaptations in pythons at the whole-heart, cellular and molecular scales. We highlight circulating factors and cellular signaling pathways that are altered during digestion to affect cardiac form and function and propose possible mechanisms that may drive the post-digestion regression of cardiac mass. Adaptive physiological cardiac hypertrophy has also been observed in other vertebrates, including in fish acclimated to cold water, birds flying at high altitudes and exercising mammals. To reveal potential evolutionarily conserved features, we summarize the molecular signatures of reversible cardiac remodeling identified in these species and compare them with those of pythons. Finally, we offer a perspective on the potential of biomimetics targeting the natural biology of pythons as therapeutics for human heart disease.

Abstract 35: The Dual Endothelin-1 Antagonist Aprocitentan Attenuates Mitochondrial Oxidative Stress in Human Cardiac Fibroblasts

Endothelin-1 (ET-1) is a peptide hormone primarily produced by endothelial cells that functions as a potent vasoconstrictor through two receptors, ETA and ETB. While most research on ET-1 has focused on endothelial dysfunction and hypertension, increasing evidence shows that ET-1 is also involved in other physiological processes such as cell proliferation, inflammation, and tissue remodeling. Dysregulation of ET-1 has been implicated in the pathogenesis of various diseases, including heart failure, atherosclerosis, fibrosis, systemic sclerosis, and cancer. Aprocitentan (also known as ACT-132577) is a relatively new dual ET-1 receptor antagonist that prevents ET-1 from binding to both endothelin receptors and has been shown to significantly reduce blood pressure in patients with resistant hypertension. Our hypothesis was that aprocitentan could reduce the proliferation rate and mitochondrial ROS production induced by TGF-β in human cardiac fibroblasts (HCFs). HCFs were isolated from hearts from fully de-identified cadaveric donors and cultured in cardiac fibroblast growth medium. HCFs were treated with aprocitentan (4 μM) and/or TGF-β1 (10 ng/ml) for 72 hours (doses and times identified in preliminary dose-response and time-course assays). Aprocitentan reduced the TGF-β1-induced proliferation rate in HCFs by approximately 23% ( Figure 1A ). Our study also aimed to determine if aprocitentan could mitigate mitochondrial dysfunction. Since the mitochondrion is a major source of reactive oxygen species (ROS), which can promote myofibroblast differentiation, mitochondrial properties in HCFs were assessed using MitoTracker Green and MitoSOX staining. TGF-β1 treatment increased mitochondrial ROS production, which was significantly reduced by aprocitentan ( Figure 1BC ); furthermore, aprocitentan reduced mitochondrial fragmentation induced by TGF-β1 treatment ( Figure 1D ). To the best of our knowledge, we are the first to demonstrate that aprocitentan can reduce the increased production of mitochondrial ROS, mitochondrial fragmentation, and cell proliferation rate induced by TGF-β1 in HCFs. This discovery could pave the way for new research and therapeutic strategies for treating fibrosis in cardiovascular disorders.

Anabolic deficits and divergent unfolded protein response underlie skeletal and cardiac muscle growth impairments in the Yoshida hepatoma tumor model of cancer cachexia.

Cancer cachexia manifests as whole body wasting, however, the precise mechanisms governing the alterations in skeletal muscle and cardiac anabolism have yet to be fully elucidated. In this study, we explored changes in anabolic processes in both skeletal and cardiac muscles in the Yoshida AH-130 ascites hepatoma model of cancer cachexia. AH-130 tumor-bearing rats experienced significant losses in body weight, skeletal muscle, and heart mass. Skeletal and cardiac muscle loss was associated with decreased ribosomal (r)RNA, and hypophosphorylation of the eukaryotic factor 4E binding protein 1. Endoplasmic reticulum stress was evident by higher activating transcription factor mRNA in skeletal muscle and growth arrest and DNA damage-inducible protein (GADD)34 mRNA in both skeletal and cardiac muscles. Tumors provoked an increase in tissue expression of interferon-γ in the heart, while an increase in interleukin-1β mRNA was apparent in both skeletal and cardiac muscles. We conclude that compromised skeletal muscle and heart mass in the Yoshida AH-130 ascites hepatoma model involves a marked reduction translational capacity and efficiency. Furthermore, our observations suggest that endoplasmic reticulum stress and tissue production of pro-inflammatory factors may play a role in the development of skeletal and cardiac muscle wasting.

Chronic cold exposure causes left ventricular hypertrophy that appears to be physiological.

Exposure to winter cold causes an increase in energy demands to meet the challenge of thermoregulation. In small rodents, this increase in cardiac output leads to a profound cardiac hypertrophy, 2-3x that typically seen with exercise training. The nature of this hypertrophy and its relevance to winter mortality remains unclear. Our goal was to characterize cold-induced cardiac hypertrophy and to assess its similarity to either exercise-induced (physiological) hypertrophy or the pathological hypertrophy of hypertension. We hypothesized that cold-induced hypertrophy will most closely resemble exercise-induced hypertrophy, but be another unique pathway for physiological cardiac growth. We found that cold-induced hypertrophy was largely reversed after return to warm temperatures. Further, metabolic rates were elevated while gene expression and mitochondrial enzyme activities indicative of pathology were absent. A gene expression panel comparing hearts of exercised and cold exposed mice further suggests that these activities are similar, although not identical. In conclusion, we found that chronic cold led to a phenotype that most closely resembled physiological hypertrophy, with enhanced metabolic rate, without induction of fetal genes , but with decreased expression of genes associated with fatty acid oxidation, suggesting that heart failure is not a cause of winter mortality in small rodents and identifying a novel approach for the study of cardiac growth.

Opa1 processing is dispensable in mouse development but is protective in mitochondrial cardiomyopathy.

Mitochondrial fusion and fission accompany adaptive responses to stress and altered metabolic demands. Inner membrane fusion and cristae morphogenesis depends on optic atrophy 1 (Opa1), which is expressed in different isoforms and is cleaved from a membrane-bound, long to a soluble, short form. Here, we have analyzed the physiological role of Opa1 isoforms and Opa1 processing by generating mouse lines expressing only one cleavable Opa1 isoform or a non-cleavable variant thereof. Our results show that expression of a single cleavable or non-cleavable Opa1 isoform preserves embryonic development and the health of adult mice. Opa1 processing is dispensable under metabolic and thermal stress but prolongs life span and protects against mitochondrial cardiomyopathy in OXPHOS-deficient Cox10-/- mice. Mechanistically, loss of Opa1 processing disturbs the balance between mitochondrial biogenesis and mitophagy, suppressing cardiac hypertrophic growth in Cox10-/- hearts. Our results highlight the critical regulatory role of Opa1 processing, mitochondrial dynamics, and metabolism for cardiac hypertrophy.

Sex-Dependent Cardiac Growth Responses to Adrenergic Stimulation in Mice Lacking the Protein Phosphatase Regulator B55α

Sex-Dependent Cardiac Growth Responses to Adrenergic Stimulation in Mice Lacking the Protein Phosphatase Regulator B55α

CITED4 gene therapy protects against maladaptive cardiac remodeling after ischemia/reperfusion injury in mice

Cardiac signaling pathways functionally important in the heart's response to exercise often protect the heart against pathological stress, potentially providing novel therapeutic targets. However, it is important to determine which of these pathways can be feasibly targeted invivo. Transgenic overexpression of exercise-induced CITED4 has been shown to protect against adverse remodeling after ischemia/reperfusion injury (IRI). Here we investigated whether somatic gene transfer of CITED4 in a clinically relevant time frame could promote recovery after IRI. Cardiac CITED4 gene delivery via intravenous AAV9 injections in wild type mice led to an approximately 3-fold increase in cardiac CITED4 expression. After 4weeks, CITED4-treated animals developed physiological cardiac hypertrophy without adverse remodeling. In IRI, delivery of AAV9-CITED4 after reperfusion resulted in a 6-fold increase in CITED4 expression 1week after surgery, as well as decreased apoptosis, fibrosis, and inflammatory markers, culminating in a smaller scar and improved cardiac function 8weeks after IRI, compared with control mice receiving AAV9-GFP. Somatic gene transfer of CITED4 induced a phenotype suggestive of physiological cardiac growth and mitigated adverse remodeling after ischemic injury. These studies support the feasibility of CITED4 gene therapy delivered in a clinically relevant time frame to mitigate adverse ventricular remodeling after ischemic injury.

Synergistic Biophysics and Machine Learning Modeling to Rapidly Predict Cardiac Growth Probability.

Computational models that can predict growth and remodeling of the heart could have important clinical applications. However, the time it takes to calibrate and run current models while considering data uncertainty and variability makes them impractical for routine clinical use. This study aims to address this need by creating a computational framework to efficiently predict cardiac growth probability. We utilized a biophysics model to rapidly simulate cardiac growth following mitral valve regurgitation (MVR). Here we developed a two-tiered Bayesian History Matching approach augmented with Gaussian process emulators for efficient calibration of model parameters to align with growth outcomes within a 95% confidence interval. We first generated a synthetic data set to assess the accuracy of our framework, and the effect of changes in data uncertainty on growth predictions. We then calibrated our model to match baseline and chronic canine MVR data and used an independent data set to successfully validate the ability of our calibrated model to accurately predict cardiac growth probability. The combined biophysics and machine learning modeling framework we proposed in this study can be easily translated to predict patient-specific cardiac growth.

Functional-Molecular Mechanisms of Sympathetic-Parasympathetic Dysfunction in PVC-Induced Cardiomyopathy Revealed by Dual Stressor PVC-Exercise Challenge

BackgroundThe significance of autonomic dysfunction in premature ventricular contraction–induced cardiomyopathy (PVC-CM) remain unknown. ObjectivesUtilizing a novel “dual stressor” provocative challenge combining exercise with premature ventricular contraction (PVCs), the authors characterized the functional and molecular mechanisms of cardiac autonomic (cardiac autonomic nervous system) remodeling in a PVC-CM animal model. MethodsIn 15 canines (8 experimental, 7 sham), we implanted pacemakers and neurotelemetry devices and subjected animals to 12 weeks of bigeminal PVCs to induce PVC-CM. Sympathetic nerve activity (SNA), vagal nerve activity (VNA), and heart rate were continuously recorded before, during, and after treadmill exercise challenge with and without PVCs, at baseline and after development of PVC-CM. Western blot and enzyme-linked immunosorbent assay were used to evaluate molecular markers of neural remodeling. ResultsExercise triggered an increase in both SNA and VNA followed by late VNA withdrawal. With PVCs, the degree of exercise-induced SNA augmentation was magnified, whereas late VNA withdrawal became blunted. After PVC-CM development, SNA was increased at rest but failed to adequately augment during exercise, especially with PVCs, coupled with impaired VNA and heart rate recovery after exercise. In the remodeled cardiac autonomic nervous system, there was widespread sympathetic hyperinnervation and elevated transcardiac norepinephrine levels but unchanged parasympathetic innervation, indicating sympathetic overload. However, cardiac nerve growth factor was paradoxically downregulated, suggesting an antineurotrophic counteradaptive response to PVC-triggered sympathetic overload. ConclusionsSympathetic overload, sympathetic dysfunction, and parasympathetic dysfunction in PVC-CM are unmasked by combined exercise and PVC challenge. Reduced cardiac neurotrophic factor might underlie the mechanisms of this dysfunction. Neuromodulation therapies to restore autonomic function could constitute a novel therapeutic approach for PVC-CM.

Kv beta complex facilitates exercise-induced augmentation of myocardial perfusion and cardiac growth.

The oxygen sensitivity of voltage-gated potassium (Kv) channels regulates cardiovascular physiology. Members of the Kv1 family interact with intracellular Kvβ proteins, which exhibit aldo-keto reductase (AKR) activity and confer redox sensitivity to Kv channel gating. The Kvβ proteins contribute to vasoregulation by controlling outward K+ currents in smooth muscle upon changes in tissue oxygen consumption and demand. Considering exercise as a primary physiological stimulus of heightened oxygen demand, the current study tested the role of Kvβ proteins in exercise performance, exercise-induced adaptations in myocardial perfusion, and physiological cardiac growth. Our findings reveal that genetic ablation of Kvβ2 proteins diminishes baseline exercise capacity in mice and attenuates the enhancement in exercise performance observed after long-term training. Moreover, we demonstrate that Kvβ2 proteins are critical for exercise-mediated enhancement in myocardial perfusion during cardiac stress as well as adaptive changes in cardiac structure. Our results underscore the importance of Kvβ proteins in metabolic vasoregulation, highlighting their role in modulating both exercise capacity and cardiovascular benefits associated with training. Furthermore, our study sheds light on a novel molecular target for enhancing exercise performance and improving the health benefits associated with exercise training in patients with limited capacity for physical activity.

Circadian control of histone turnover during cardiac development and growth

During postnatal cardiac hypertrophy, cardiomyocytes undergo mitotic exit, relying on DNA replication-independent mechanisms of histone turnover to maintain chromatin organization and gene transcription. In other tissues, circadian oscillations in nucleosome occupancy influence clock-controlled gene expression, suggesting a role for the circadian clock in temporal control of histone turnover and coordinated cardiomyocyte gene expression. To elucidate roles for the master circadian transcription factor, Bmal1, in histone turnover, chromatin organization, and myocyte-specific gene expression and cell growth in the neonatal period. Bmal1 knockdown in neonatal rat ventricular myocytes (NRVM) decreased myocyte size, total cellular protein synthesis, and transcription of the fetal hypertrophic gene Nppb following treatment with serum or the α-adrenergic agonist phenylephrine (PE). Depletion of Bmal1 decreased expression of clock-controlled genes Per2 and Tcap, as well as Sik1, a Bmal1 target upregulated in adult versus embryonic hearts. Bmal1 knockdown impaired Per2 and Sik1 promoter accessibility as measured by MNase-qPCR and impaired histone turnover as measured by metabolic labeling of acid-soluble chromatin fractions. Sik1 knockdown in turn decreased myocyte size, while simultaneously inhibiting Nppb transcription and activating Per2 transcription. Linking these changes to chromatin remodeling, depletion of the replication-independent histone variant H3.3a inhibited myocyte hypertrophy and prevented PE-induced changes in clock-controlled gene transcription. Bmal1 is required for neonatal myocyte growth, replication-independent histone turnover, and chromatin organization at the Sik1 promoter. Sik1 represents a novel clock-controlled gene that coordinates myocyte growth with hypertrophic and clock-controlled gene transcription. Replication-independent histone turnover is required for transcriptional remodeling of clock-controlled genes in cardiac myocytes in response to growth stimuli.

Revealing functional-molecular parasympathetic dysfunction in PVC-induced cardiomyopathy via dual stressor PVC and exercise

Abstract Background Exercise intolerance is a common presentation of premature ventricular contractions-induced cardiomyopathy (PVC-CM). In a canine PVC-CM model, previous studies have shown cardiac autonomic nervous system remodeling, resulting in persistent sympathetic excess. Nevertheless, the molecular mechanisms and functional implications of this neural remodeling, as well as parasympathetic alterations, remain unclear. Purpose We aim to evaluate the functional-molecular mechanisms of parasympathetic remodeling and its potential contribution to exercise intolerance in an animal model of PVC-CM. Methods We studied 15 canines (8 experimental vs 7 sham controls) chronically instrumented with pacemakers and telemetry devices to deliver 12-weeks of bigeminal PVCs (200ms coupling) to induce PVC-CM and to record sympathetic (SNA), vagal nerve activity (VNA) and heart rate (HR) over this period. Exercise challenge was conducted in sinus rhythm (SR) and in the presence of bigeminal PVCs ("dual stressor") at baseline and after development of PVC-CM. Sympatho-vagal neural and HR responses were assessed during and after exercise at baseline and after PVC-CM development, with and without PVCs, thus comparing baseline SR, baseline with PVCs, CM SR and CM with PVCs. Western Blot was performed on intrinsic and extrinsic cardiac neural tissues to evaluate molecular mechanisms of neural remodeling. Results Compared to baseline, SNA was significantly higher in PVC-CM in all phases of exercise (rest, exercise, recovery, p=0.03). In contrast, VNA was significantly lower in PVC-CM than baseline during exercise recovery (P=0.014). After exercise, HR recovery (HRR) was impaired (p=0.001), VNA lower (p=0.014) and SNA (p=0.03) higher in PVC-CM compared with baseline. Tyrosine hydroxylase (TH) immunoreactivity in the left ventricle (LV, p=0.001) and stellate ganglia (SG, p=0.03) and trans-cardiac plasma NE levels (p=0.03) were higher in PVC-CM compared with sham. Choline Acetyltransferase (CHAT) was not significantly different in either vagus (p=0.33) or LV (p=0.8) between PVC-CM vs sham. Cardiac (LV) Nerve Growth Factor (NGF), Growth Associated Protein (GAP-43) and beta 1-adrenoreceptor levels were lower (p=0.042, p=0.003, p=0.002 respectively) whereas beta 2-adreno- and muscarinic M2-cholinergic receptors were higher (p=0.02, p&amp;lt;0.01 respectively) in PVC-CM compared with sham. Conclusion Neural remodeling in PVC-CM results in an imbalanced sympathetic excess and sympathetic-parasympathetic dysfunction characterized by a loss of sympathetic functional reserve during exercise and impaired parasympathetic activity and HRR after exercise. Adaptive downregulation of NGF might explain this imbalance and dysfunction. These data open the intriguing possibility of neural-targeted therapies to reverse remodeling and restore autonomic function disrupted by chronic PVCs.GraphsMolecular

YOUNG WOMAN WITH LEFT VENTRICULAR DYSFUNCTION AND MITRAL LAMBL EXCRESCENCE: WHAT TO DO WHEN ISCHEMIC STROKE IS RECURRENT?

Abstract Lambl excrescence describing rare cardiac growths that develop at the valvular coaptation sites of the heart which are seen as a thin, hypermobile and filiform strand on an echocardiogram. These filiform strands can be noted in association with embolic stroke. Also heart failure (HF) and stroke frequently coexist because of an overlap of shared risk factors and subsequent mechanisms and a higher risk of stroke is present also in HF patients in sinus rythm (SR). The clinical case concerns a 54–year–old woman. In March 2020 she was admitted for ischemic stroke by occlusion of middle cerebral artery subjected to thrombolysis and mechanical thromboctomy. Electrocardiogram showed SR and left branch block. Echocardiogram and echocardiogram transesophageal showed left ventricular (LV) dysfunction (EF 40%) withouth clear presence of emboligen sources. Thrombophilic screening and neurological checks were negative. Single antiaggregant therapy has been started. New hospitalization in October 2023 for recurrent ischemic stroke. Echocardiogram showed worsening of LV dysfunction (EF 20%). Transesophageal echocardiogram showed the presence of a filiform filament on the mitral leaflets on the atrial side (Fig 1) without the presence of intracavitary thrombi. Coronary TC angiography (Fig 2) was negative. Cardiac magnetic resonance (Fig 3) confirmed reduced ejection fraction and showed subepicardial late gadolinium enhancement in lateral wall. There are no clear evidence–based guidelines for the treatment of Lambl excrescences. When associated with stroke, an exhaustive stroke workup to identify the potential cause of stroke should always be undertaken. If the workup remains negative without any identifiable cause, then the patients can be treated with antiplatelet agents such as aspirin and clopidogrel or anticoagulation with warfarin. Also the indication to antithrombotic strategies in patients with HF, SR and stroke is controversial. There are no data to support a routine strategy of anticoagulation in patients with HF with LV dysfuncion in SR who do not have history of paroxysmal atrial fibrillation (ESC 2022 HF guidelines). So which therapy to start in young patient, with recurrent strokes, heart failure /severe LV dysfunction and mitral lambl excrescence? Oral anticoagulant? Double platelet antiaggregation? Oral anticoagulant and single platelet antiaggregation? Definitely the choice is not simple but we decided to start warfarin and continue follow–up.

Biological sex, sex steroids and sex chromosomes contribute to mouse cardiac aging.

After menopause, the incidence of cardiovascular disease rapidly rises in women. The disappearing protection provided by sex steroids is a consequence of the development of many risk factors. Preclinical studies are necessary to understand better the effects of ovarian hormones loss cardiac aging. To mimic menopause in mice and study its consequences, we delayed ovariectomy at 12 months and followed animals for 12 months. Using RNA sequencing, we investigated changes in the myocardial exome with aging. In addition, with four-core genotypes (FCG) transgenic mice, we studied sex chromosome effects on cardiac aging. Heart weight increased from 3 to 24 months (males + 35%, females + 29%). In males, 75% of this increase had occurred at 12 months; in females, only 30%. Gonadectomy of mice at 12 months blocked cardiac hypertrophy in both sexes during the second year of life. The dosage of the X chromosomes did not influence cardiac growth in young and older mice. We performed an RNA sequencing study in young and old mice. We identified new highly expressed genes modulated during aging (Bdh, Myot, Cpxm2, and Slc38a1). The myocardial exome in older animals displayed few differences related to the animal's sex or the presence or absence of sex steroids for a year. We show that the morphological evolution of the heart depends on the biological sex via gonadal sex hormone actions. The myocardial exome of old male and female mice is relatively similar. Our study emphasizes the need to consider sex steroid effects in studying cardiac aging.

Triiodo-L-thyronine (T3) downregulates Npr1 gene (coding for natriuretic peptide receptor-A) transcription in H9c2 cells: involvement of β-AR-ROS signaling.

Natriuretic peptide receptor-A (NPR-A) signaling system is considered as an intrinsic productive mechanism of the heart that opposes abnormal cardiac remodeling and hypertrophic growth. NPR-A is coded by Npr1 gene, and its expression is downregulated in the hypertrophied heart. We sought to examine the levels of Npr1 gene transcription in triiodo-L-thyronine (T3) treated hypertrophied cardiomyocyte (H9c2) cells, in vitro, and also the involvement of β-adrenergic receptor (β-AR) - Reactive oxygen species (ROS) signaling system in the down-regulation of Npr1 transcription also studied. Anti-hypertrophic Npr1 gene transcription was monitored in control and T3-treated (dose and time dependent) H9c2 cells, using a real time PCR method. Further, cell size, intracellular cGMP, ROS, hypertrophy markers (ANP, BNP, α-sk, α-MHC and β-MHC), β-AR, and protein kinase cGMP-dependent 1 (PKG-I) genes expression were also determined. The intracellular cGMP and ROS levels were determined by ELISA and DCF dye method, respectively. In addition, to neutralize T3 mediated ROS generation, H9c2 cells were treated with T3 in the presence and absence of antioxidants [curcumin (CU) or N-acetyl-L-cysteine (NAC)]. A dose dependent (10 pM, 100 pM, 1 nM and 10 nM) and time dependent (12 h, 24 h and 48 h) down-regulation of Npr1 gene transcription (20, 39, 60, and 74% respectively; 18, 55, and 85%, respectively) were observed in T3-treated H9c2 cells as compared with control cells. Immunofluorescence analysis also revealed that a marked down regulation of NPR- A protein in T3-treated cells as compared with control cells. Further, a parallel downregulation of cGMP and PKG-I (2.4 fold) were noticed in the T3-treated cells. In contrast, a time dependent increased expression of β-AR (60, 72, and 80% respectively) and ROS (26, 48, and 74%, respectively) levels were noticed in T3-treated H9c2 cells as compared with control cells. Interestingly, antioxidants, CU or NAC co-treated T3 cells displayed a significant reduction in ROS (69 and 81%, respectively) generation and to increased Npr1 gene transcription (81 and 88%, respectively) as compared with T3 alone treated cells. Our result suggest that down regulation of Npr1 gene transcription is critically involved in T3- induced hypertrophic growth in H9c2 cells, and identifies the cross-talk between T3-β-AR-ROS and NPR-A signaling.