Model Of Human Heart Failure Research Articles

Using a Failing Human Ventricular Cardiomyocyte Model to Re-Evaluate Ca2+ Cycling, Voltage Dependence, and Spark Characteristics

Previous studies have observed alterations in excitation–contraction (EC) coupling during end-stage heart failure that include action potential and calcium (Ca2+) transient prolongation and a reduction of the Ca2+ transient amplitude. Underlying these phenomena are the downregulation of potassium (K+) currents, downregulation of the sarcoplasmic reticulum Ca2+ ATPase (SERCA), increase Ca2+ sensitivity of the ryanodine receptor, and the upregulation of the sodium–calcium (Na=-Ca2+) exchanger. However, in human heart failure (HF), debate continues about the relative contributions of the changes in calcium handling vs. the changes in the membrane currents. To understand the consequences of the above changes, they are incorporated into a computational human ventricular myocyte HF model that can explore the contributions of the spontaneous Ca2+ release from the sarcoplasmic reticulum (SR). The reduction of transient outward K+ current (Ito) is the main membrane current contributor to the decrease in RyR2 open probability and L-type calcium channel (LCC) density which emphasizes its importance to phase 1 of the action potential (AP) shape and duration (APD). During current-clamp conditions, RyR2 hyperphosphorylation exhibits the least amount of Ca2+ release from the SR into the cytosol and SR Ca2+ fractional release during a dynamic slow–rapid–slow (0.5–2.5–0.5 Hz) pacing, but it displays the most abundant and more lasting Ca2+ sparks two-fold longer than a normal cell. On the other hand, under voltage-clamp conditions, HF by decreased SERCA and upregulated INCX show the least SR Ca2+ uptake and EC coupling gain, as compared to HF by hyperphosphorylated RyR2s. Overall, this study demonstrates that the (a) combined effect of SERCA and NCX, and the (b) RyR2 dysfunction, along with the downregulation of the cardiomyocyte’s potassium currents, could substantially contribute to Ca2+ mishandling at the spark level that leads to heart failure.

Empagliflozin prevents heart failure through inhibition of the NHE1-NO pathway, independent of SGLT2

Sodium glucose cotransporter 2 inhibitors (SGLT2i) constitute the only medication class that consistently prevents or attenuates human heart failure (HF) independent of ejection fraction. We have suggested earlier that the protective mechanisms of the SGLT2i Empagliflozin (EMPA) are mediated through reductions in the sodium hydrogen exchanger 1 (NHE1)-nitric oxide (NO) pathway, independent of SGLT2. Here, we examined the role of SGLT2, NHE1 and NO in a murine TAC/DOCA model of HF. SGLT2 knockout mice only showed attenuated systolic dysfunction without having an effect on other signs of HF. EMPA protected against systolic and diastolic dysfunction, hypertrophy, fibrosis, increased Nppa/Nppb mRNA expression and lung/liver edema. In addition, EMPA prevented increases in oxidative stress, sodium calcium exchanger expression and calcium/calmodulin-dependent protein kinase II activation to an equal degree in WT and SGLT2 KO animals. In particular, while NHE1 activity was increased in isolated cardiomyocytes from untreated HF, EMPA treatment prevented this. Since SGLT2 is not required for the protective effects of EMPA, the pathway between NHE1 and NO was further explored in SGLT2 KO animals. In vivo treatment with the specific NHE1-inhibitor Cariporide mimicked the protection by EMPA, without additional protection by EMPA. On the other hand, in vivo inhibition of NOS with L-NAME deteriorated HF and prevented protection by EMPA. In conclusion, the data support that the beneficial effects of EMPA are mediated through the NHE1-NO pathway in TAC/DOCA-induced heart failure and not through SGLT2 inhibition.

Deep learning enables automated volumetric assessments of cardiac function in zebrafish.

ABSTRACTAlthough the zebrafish embryo is a powerful animal model of human heart failure, the methods routinely employed to monitor cardiac function produce rough approximations that are susceptible to bias and inaccuracies. We developed and validated a deep learning-based image-analysis platform for automated extraction of volumetric parameters of cardiac function from dynamic light-sheet fluorescence microscopy (LSFM) images of embryonic zebrafish hearts. This platform, the Cardiac Functional Imaging Network (CFIN), automatically delivers rapid and accurate assessments of cardiac performance with greater sensitivity than current approaches.This article has an associated First Person interview with the first author of the paper.

Induction of cardiac dysfunction in developing and adult zebrafish by chronic isoproterenol stimulation

Zebrafish is a widely used model to evaluate genetic variants and modifiers that can cause heart muscle diseases. Surprisingly, the β-adrenergic receptor (β-AR) pathway in zebrafish is not well characterized, although abnormal β-AR signaling is a major contributor to human heart failure (HF). Chronic β-AR activation in the attempt to normalize heart function in the failing heart results in a reduction of the β-ARs expression and receptor desensitization, largely mediated through G-protein coupled receptor kinase 2 (GRK2) upregulation. This in turn leads to further deterioration of heart function and progression towards HF. This study seeks to systematically characterize the function of the β-AR signaling in developing and adult zebrafish to ultimately assess the ability to induce HF through chronic β-AR activation by isoproterenol (ISO) as established in the mouse model.Larval hearts first responded to ISO by 3dpf, in concordance with robust expression of key components of the β-AR signaling pathway. Although ISO-induced β1-AR and β2-AR isoform upregulation persisted, chronic ISO stimulation for 5d caused systolic cardiac dysfunction concurrently with maximal expression of G-protein-coupled receptor kinase-2 (GRK2). More consistent to mammalians, adult zebrafish developed significant heart failure in concert with β1-AR downregulation, and GRK2 and brain natriuretic peptide (BNP) upregulation in response to prolonged, 14d ISO-stimulation. This was accompanied by significant cell death and inflammation without detectable fibrosis.Our study unveils important characteristics of larvae and adult zebrafish hearts pertaining to β-AR signaling. A lack of β-AR responsiveness and atypical β-AR/GRK2 ratios in larval zebrafish should be considered. Adult zebrafish resembled the mammalian situation on the functional and molecular level more closely, but also revealed differences to dysfunctional mammalian hearts, i.e. lack of fibrosis. Our study establishes the first ISO-inducible HF model in adult zebrafish and present critical characteristics of the zebrafish heart essential to be considered when utilizing the zebrafish as a human disease and future drug discovery model.

Representing variability and transmural differences in a model of human heart failure.

During heart failure (HF) at the cellular level, the electrophysiological properties of single myocytes get remodeled, which can trigger the occurrence of ventricular arrhythmias that could be manifested in many forms such as early afterdepolarizations (EADs) and alternans (ALTs). In this paper, based on experimentally observed human HF data, specific ionic and exchanger current strengths are modified from a recently developed human ventricular cell model: the O'Hara-Virág-Varró-Rudy (OVVR) model. A new transmural HF-OVVR model is developed that incorporates HF changes and variability of the observed remodeling. This new heterogeneous HF-OVVR model is able to replicate many of the failing action potential (AP) properties and the dynamics of both [Ca(2+)]i and [Na(+)]i in accordance with experimental data. Moreover, it is able to generate EADs for different cell types and exhibits ALTs at modest pacing rate for transmural cell types. We have assessed the HF-OVVR model through the examination of the AP duration and the major ionic currents' rate dependence in single myocytes. The evaluation of the model comes from utilizing the steady-state (S-S) and S1-S2 restitution curves and from probing the accommodation of the HF-OVVR model to an abrupt change in cycle length. In addition, we have investigated the effect of chosen currents on the AP properties, such as blocking the slow sodium current to shorten the AP duration and suppress the EADs, and have found good agreement with experimental observations. This study should help elucidate arrhythmogenic mechanisms at the cellular level and predict unseen properties under HF conditions. In addition, this AP cell model might be useful for modeling and simulating HF at the tissue and organ levels.

PKD Regulates Mitochondrial Morphology and Function via Phosphorylation of DLP1 in Cardiac Myocytes.

Regulation of mitochondrial morphology and dynamics is essential for maintaining cardiomyocyte function. Abnormal mitochondrial morphology and mitochondrial dysfunction are frequently observed in both human heart failure (HF) and animal HF models. However, it is still unclear which cardiac signaling pathway(s) regulate(s) mitochondrial morphology/function under pathophysiological conditions. Since Gq‐protein coupled receptor (GqPCR) signaling is critical for the development and progression of HF, we tested the hypothesis that GqPCR stimulation induces pathological changes in mitochondrial morphology/function in cardiomyocytes. We found that protein kinase D (PKD) was activated and translocated to the outer mitochondrial membrane by GqPCR stimulation. GqPCR‐mediated PKD activation induced mitochondrial fragmentation, increased reactive oxygen species generation and mitochondrial permeability transition pore opening followed by caspase‐3 activation. These morphological and functional changes in cardiac mitochondria were mediated via PKD‐dependent phosphorylation of Dynamin‐Like Protein 1 (DLP1) at S637. Importantly, PKD‐dependent DLP1 phosphorylation at S637 concurrent with abnormal mitochondrial morphology and apoptotic signaling were also observed in ventricular tissue from transgenic mice with cardiac‐specific overexpression of constitutively active Gq. In conclusion, we demonstrate that GqPCR stimulation induces mitochondrial fragmentation and dysfunction via PKD‐dependent phosphorylation of DLP1 at S637, which likely contributes to cardiomyocyte dysfunction during HF.

Small-Molecule PKD Inhibitor Prevents Mitochondrial Fragmentation and Dysfunction during Gq-Protein Coupled Receptor Stimulation in Cardiac Cells

Regulation of mitochondrial morphology and dynamics is crucial for the maintenance of various cellular functions in cardiomyocytes. Abnormal mitochondrial morphologies concomitant with mitochondrial dysfunction are frequently observed both in human heart failure (HF) and in animal HF models. However, it is still unclear which cardiac signaling pathways regulate mitochondrial morphology and function under pathophysiological conditions. Recent reports suggest that Gq-protein coupled receptor (GqPCR) signaling pathways are critical for the development and progression of HF. Therefore, we hypothesize that GqPCR stimulation induces mitochondrial fragmentation and dysfunction, which initiates cardiomyocyte death. We found that protein kinase D (PKD) activated by GqPCR signaling was translocated to outer mitochondrial membrane (OMM) observed by Western blot analysis of cytosolic and mitochondria-enriched fractionated proteins and by live cell imaging of fluorescence resonance energy transfer (FRET). We also found that GqPCR-mediated PKD activation induced mitochondrial fragmentation, leading to increased reactive oxygen species (ROS) generation as well as increased mitochondrial permeability transition pore (mPTP) opening, which initiates apoptotic signaling activation and cardiomyocyte death. These morphological and functional changes in cardiac mitochondria were mediated via PKD-dependent phosphorylation of mitochondrial fission protein, Dynamin-Like Protein 1 (DLP1) at S637. Moreover, pretreatment with a novel potent PKD inhibitor CRT0066101 effectively inhibited GqPCR-mediated PKD translocation to OMM, DLP1 phosphorylation at S637, mitochondrial fragmentation, ROS generation and mPTP activation. In conclusion, we demonstrate that GqPCR stimulation induces mitochondrial fragmentation and dysfunction through PKD-dependent phosphorylation of DLP1 at S637, which likely contributes to cardiomyocyte injury. Thus, small-molecule PKD inhibitor may become a novel and potent therapeutic for preventing cardiac cell injury and death during HF.

Nuclear pore rearrangements and nuclear trafficking in cardiomyocytes from rat and human failing hearts.

AimsDuring cardiac hypertrophy, cardiomyocytes (CMs) increase in the size and expression of cytoskeletal proteins while reactivating a foetal gene programme. The process is proposed to be dependent on increased nuclear export and, since nuclear pore trafficking has limited capacity, a linked decrease in import. Our objective was to investigate the role of nuclear import and export in control of hypertrophy in rat and human heart failure (HF).Methods and resultsIn myocardial tissue and isolated CMs from patients with dilated cardiomyopathy, nuclear size was increased; Nucleoporin p62, cytoplasmic RanBP1, and nuclear translocation of importins (α and β) were decreased while Exportin-1 was increased. CM from a rat HF model 16 weeks after myocardial infarction (MI) reproduced these nuclear changes. Nuclear import, determined by the rate of uptake of nuclear localization sequence (NLS)-tagged fluorescent substrate, was also decreased and this change was observed from 4 weeks after MI, before HF has developed. Treatment of isolated rat CMs with phenylephrine (PE) for 48 h produced similar cell and nuclear size increases, nuclear import and export protein rearrangement, and NLS substrate uptake decrease through p38 MAPK and HDAC-dependent pathways. The change in NLS substrate uptake occurred within 15 min of PE exposure. Inhibition of nuclear export with leptomycin B reversed established nuclear changes in PE-treated rat CMs and decreased NLS substrate uptake and cell/nuclear size in human CMs.ConclusionsNuclear transport changes related to increased export and decreased import are an early event in hypertrophic development. Hypertrophy can be prevented, or even reversed, by targeting import/export, which may open new therapeutic opportunities.

Long-term levosimendan treatment improves systolic function and myocardial relaxation in mice with cardiomyocyte-specific disruption of theSerca2gene

In human heart failure (HF), reduced cardiac function has, at least partly, been ascribed to altered calcium homeostasis in cardiomyocytes. The effects of the calcium sensitizer levosimendan on diastolic dysfunction caused by reduced removal of calcium from cytosol in early diastole are not well known. In this study, we investigated the effect of long-term levosimendan treatment in a murine model of HF where the sarco(endo)plasmatic reticulum ATPase (Serca) gene is specifically disrupted in the cardiomyocytes, leading to reduced removal of cytosolic calcium. After induction of Serca2 gene disruption, these mice develop marked diastolic dysfunction as well as impaired contractility. SERCA2 knockout (SERCA2KO) mice were treated with levosimendan or vehicle from the time of KO induction. At the 7-wk end point, cardiac function was assessed by echocardiography and pressure measurements. Vehicle-treated SERCA2KO mice showed significantly diminished left-ventricular (LV) contractility, as shown by decreased ejection fraction, stroke volume, and cardiac output. LV pressure measurements revealed a marked increase in the time constant (τ) of isovolumetric pressure decay, showing impaired relaxation. Levosimendan treatment significantly improved all three systolic parameters. Moreover, a significant reduction in τ toward normalization indicated improved relaxation. Gene-expression analysis, however, revealed an increase in genes related to production of the ECM in animals treated with levosimendan. In conclusion, long-term levosimendan treatment improves both contractility and relaxation in a heart-failure model with marked diastolic dysfunction due to reduced calcium transients. However, altered gene expression related to fibrosis was observed.

Messenger RNA Expression Levels Predict Cellular Electrophysiologic Remodeling in Failing Human Hearts Using a Population-Based Simulation Study

<h3>Background</h3> Alterations in mRNA expression levels are reported in human heart failure (HF). However, their potential causative link to changes in action potential (AP) and calcium transient (Ca_iT) measured in human HF is still unclear. Our aim is to investigate consequences of HF-related mRNA changes on cellular electrophysiology taking into account interindividual variability and uncertainty in membrane-level effect of changed mRNA expression of channel subunits. <h3>Methods</h3> HF and non-HF human cell model populations were constructed using Monte-Carlo sampling and recent mRNA expression data from HF and nondiseased human hearts. Six AP and Ca_iT biomarkers were calculated for each cell at cycle lengths between 1500 and 300 ms. Regression analysis was used to determine key ionic properties underlying biomarker changes. <h3>Results</h3> Simulations show that HF-related mRNA changes result in increase in AP duration, AP triangulation, Ca_iT duration, delay between AP and Ca_iT upstroke as well as decrease in Ca_iT triangulation and amplitude in HF cells. Changes in AP biomarkers in HF are correlated with reduction in I_Kr, whereas changes in Ca_iT biomarkers are correlated with reduction in I_CaL and SERCA. The role of I_CaL is pacing rate dependent. At fast pacing rates, both HF and non-HF cells develop alternans, but the underlying mechanisms differed. <h3>Conclusions</h3> A population-based simulation study reveals the functional consequences of HR-related mRNA changes on cellular electrophysiology, consistent with experimental measurements in human HF. Remodeling in I_Kr, I_CaL, and SERCA are identified as main determinants of AP and Ca_iT changes in HF. Sample AP traces at 1500ms BCL

Affecting Rhomboid-3 Function Causes a Dilated Heart in Adult Drosophila

Drosophila is a well recognized model of several human diseases, and recent investigations have demonstrated that Drosophila can be used as a model of human heart failure. Previously, we described that optical coherence tomography (OCT) can be used to rapidly examine the cardiac function in adult, awake flies. This technique provides images that are similar to echocardiography in humans, and therefore we postulated that this approach could be combined with the vast resources that are available in the fly community to identify new mutants that have abnormal heart function, a hallmark of certain cardiovascular diseases. Using OCT to examine the cardiac function in adult Drosophila from a set of molecularly-defined genomic deficiencies from the DrosDel and Exelixis collections, we identified an abnormally enlarged cardiac chamber in a series of deficiency mutants spanning the rhomboid 3 locus. Rhomboid 3 is a member of a highly conserved family of intramembrane serine proteases and processes Spitz, an epidermal growth factor (EGF)–like ligand. Using multiple approaches based on the examination of deficiency stocks, a series of mutants in the rhomboid-Spitz–EGF receptor pathway, and cardiac-specific transgenic rescue or dominant-negative repression of EGFR, we demonstrate that rhomboid 3 mediated activation of the EGF receptor pathway is necessary for proper adult cardiac function. The importance of EGF receptor signaling in the adult Drosophila heart underscores the concept that evolutionarily conserved signaling mechanisms are required to maintain normal myocardial function. Interestingly, prior work showing the inhibition of ErbB2, a member of the EGF receptor family, in transgenic knock-out mice or individuals that received herceptin chemotherapy is associated with the development of dilated cardiomyopathy. Our results, in conjunction with the demonstration that altered ErbB2 signaling underlies certain forms of mammalian cardiomyopathy, suggest that an evolutionarily conserved signaling mechanism may be necessary to maintain post-developmental cardiac function.

Role of the Transient Outward Current in Regulating Mechanical Properties of Canine Ventricular Myocytes

The transient outward current (I(to)) is a major repolarizing current in the heart. Reduction of I(to) density is consistently observed in human heart failure (HF) and animal HF models. It has been proposed that I(to), via its influence on phase-1 repolarization of the action potential, facilitates L-type Ca(2+) current (I(Ca-L)) activation and sarcoplasmic reticulum Ca(2+) release, and that its down-regulation may contribute to the impaired contractility in failing heart. We used the dynamic clamp to quantitatively examine the influence of I(to) on the mechanical properties of canine left ventricular myocytes at 34 degrees C. In endocardial myocytes, where the native I(to) is small, simulation of an epicardial-level artificial I(to) accentuated the phase-1 repolarization and significantly suppressed cell shortening. The peak amplitude of Ca(2+) transient was also reduced in the presence of simulated I(to), although the rate of rise of the Ca(2+) transient was increased. Conversely, subtraction, or "blockade" of the native I(to) enhanced contractility in epicardial cells. These results agree with the inverse correlation between I(to) levels and myocyte contractility and Ca(2+) transient amplitude in epicardial and endocardial myocytes. Action potential clamp studies showed that the phase-1 repolarization/I(to) versus I(Ca-L) relationship had an inverted-J shape; small I(to) enhanced peak I(Ca-L) while moderate-to-large I(to) decreased peak I(Ca-L) and markedly reduced early Ca(2+) influx. Our results show that epicardial-level of I(to) acts as a negative, rather than positive regulator of myocyte mechanical properties in canine ventricular myocytes.

Role of the Transient Outward Current In Regulating Mechanical Properties of Canine Ventricular Myocytes

The transient outward current (I(to)) is a major repolarizing current in the heart. Reduction of I(to) density is consistently observed in human heart failure (HF) and animal HF models. It has been proposed that I(to), via its influence on phase-1 repolarization of the action potential, facilitates L-type Ca(2+) current (I(Ca-L)) activation and sarcoplasmic reticulum Ca(2+) release, and that its down-regulation may contribute to the impaired contractility in failing heart.We used the dynamic clamp to quantitatively examine the influence of I(to) on the mechanical properties of canine left ventricular myocytes at 34 degrees C. In endocardial myocytes, where the native I(to) is small, simulation of an epicardial-level artificial I(to) accentuated the phase-1 repolarization and significantly suppressed cell shortening. The peak amplitude of Ca(2+) transient was also reduced in the presence of simulated I(to), although the rate of rise of the Ca(2+) transient was increased. Conversely, subtraction, or "blockade" of the native I(to) enhanced contractility in epicardial cells. These results agree with the inverse correlation between I(to) levels and myocyte contractility and Ca(2+) transient amplitude in epicardial and endocardial myocytes. Action potential clamp studies showed that the phase-1 repolarization/I(to) versus I(Ca-L) relationship had an inverted-J shape; small I(to) enhanced peak I(Ca-L) while moderate-to-large I(to) decreased peak I(Ca-L) and markedly reduced early Ca(2+) influx.Our results show that epicardial-level of I(to) acts as a negative, rather than positive regulator of myocyte mechanical properties in canine ventricular myocytes.

Transcriptomic profiling of the canine tachycardia-induced heart failure model: global comparison to human and murine heart failure

Alterations of cardiac gene expression are central to ventricular dysfunction in human heart failure (HF). The canine tachycardia pacing-induced HF model is known to reproduce the main hemodynamic, echocardiographic and electrophysiological changes observed in human HF. In this study, we use this HF model to compare gene expression profiles in the left and right ventricles (LV, RV) of normal and end-stage failing canine hearts and compare the transcription profiles to those in human and murine models of HF. In end-stage HF, the LV exhibits down regulation of genes involved in energy production, cardiac contraction, and modulation of excitation–contraction coupling as compared with normal LV. The majority of transcriptomic changes between normal and end-stage canine HF were shared by the RV and LV. Genes down regulated only in the LV included those involved in aerobic energy production pathways, regulation of actin filament length, and enzyme-linked receptor protein signaling pathways. In normal canine hearts, genes encoding specific components of the contractile apparatus exhibit LV–RV asymmetric expression patterns; in failing hearts, cardiac fetal transcription factors MEF2 and MITF and the stress-responsive transcription factor ATF4 showed interventricular differences in expression. The comparison among the canine tachypacing, mouse transgenic, and human HF reveals that human disease involves down regulation of genes in a broad range of biological processes while experimentally induced HF is associated with down regulation of energy pathways, and that human ischemic HF and canine HF share a similar over representation of transcriptional pathways in the up regulated genes. This study provides insights into the molecular pathways leading to end-stage tachycardia-induced HF, and into global transcriptomic differences between the animal HF models and human HF.

The aging SHR as a model of human heart failure

The aging SHR as a model of human heart failure

The ageing SHR as a model of human heart failure: Responses to low-dose perindopril treatment

The ageing SHR as a model of human heart failure: Responses to low-dose perindopril treatment

The ageing SHR as a model of human heart failure: Responses to L-arginine treatment

The ageing SHR as a model of human heart failure: Responses to L-arginine treatment

Arrhythmogenesis and Contractile Dysfunction in Heart Failure

Ventricular arrhythmias and contractile dysfunction are the main causes of death in human heart failure (HF). In a rabbit HF model reproducing these same aspects of human HF, we demonstrate that a 2-fold functional upregulation of Na(+)-Ca(2+) exchange (NaCaX) unloads sarcoplasmic reticulum (SR) Ca(2+) stores, reducing Ca(2+) transients and contractile function. Whereas beta-adrenergic receptors (beta-ARs) are progressively downregulated in HF, residual beta-AR responsiveness at this critical HF stage allows SR Ca(2+) load to increase, causing spontaneous SR Ca(2+) release and transient inward current carried by NaCaX. A given Ca(2+) release produces greater arrhythmogenic inward current in HF (as a result of NaCaX upregulation), and approximately 50% less Ca(2+) release is required to trigger an action potential in HF. The inward rectifier potassium current (I(K1)) is reduced by 49% in HF, and this allows greater depolarization for a given NaCaX current. Partially blocking I(K1) in control cells with barium mimics the greater depolarization for a given current injection seen in HF. Thus, we present data to support a novel paradigm in which changes in NaCaX and I(K1), and residual beta-AR responsiveness, conspire to greatly increase the propensity for triggered arrhythmias in HF. In addition, NaCaX upregulation appears to be a critical link between contractile dysfunction and arrhythmogenesis.