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 in vivo. 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 4 weeks, 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 1 week after surgery, as well as decreased apoptosis, fibrosis, and inflammatory markers, culminating in a smaller scar and improved cardiac function 8 weeks 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.

ShuffleAnalyzer: A Comprehensive Tool to Visualize DNA Shuffling.

DNA shuffling is a powerful technique for generating synthetic DNA via recombination of homologous parental sequences. Resulting chimeras are often incorporated into complex libraries for functionality screenings that identify novel variants with improved characteristics. To survey shuffling efficiency, subsequences of chimeras can be computationally assigned to their corresponding parental counterpart, yielding insight into frequency of recombination events, diversity of shuffling libraries and actual composition of final variants. Whereas tools for parental assignment exist, they do not provide direct visualization of the results, making the analysis time-consuming and cumbersome. Here we present ShuffleAnalyzer, a comprehensive, user-friendly, Python-based analysis tool that directly generates graphical outputs of parental assignments and is freely available under a BSD-3 license (https://github.com/joerg-swg/ShuffleAnalyzer/releases). Besides DNA shuffling, peptide insertions can be simultaneously analyzed and visualized, which makes ShuffleAnalyzer a highly valuable tool for integrated approaches often used in synthetic biology, such as AAV capsid engineering in gene therapy applications.

S100A1's single cysteine is an indispensable redox switch for the protection against diastolic calcium waves in cardiomyocytes.

The EF-hand calcium (Ca2+) sensor protein S100A1 combines inotropic with antiarrhythmic potency in cardiomyocytes (CMs). Oxidative posttranslational modification (ox-PTM) of S100A1's conserved, single-cysteine residue (C85) via reactive nitrogen species (i.e., S-nitrosylation or S-glutathionylation) has been proposed to modulate conformational flexibility of intrinsically disordered sequence fragments and to increase the molecule's affinity toward Ca2+. Considering the unknown biological functional consequence, we aimed to determine the impact of the C85 moiety of S100A1 as a potential redox switch. We first uncovered that S100A1 is endogenously glutathionylated in the adult heart in vivo. To prevent glutathionylation of S100A1, we generated S100A1 variants that were unresponsive to ox-PTMs. Overexpression of wild-type (WT) and C85-deficient S100A1 protein variants in isolated CM demonstrated equal inotropic potency, as shown by equally augmented Ca2+ transient amplitudes under basal conditions and ß-adrenergic receptor (ßAR) stimulation. However, in contrast, ox-PTM defective S100A1 variants failed to protect against arrhythmogenic diastolic sarcoplasmic reticulum (SR) Ca2+ waves and ryanodine receptor 2 (RyR2) hypernitrosylation during ßAR stimulation. Despite diastolic performance failure, C85-deficient S100A1 protein variants exerted similar Ca2+-dependent interaction with the RyR2 than WT-S100A1. Dissecting S100A1's molecular structure-function relationship, our data indicate for the first time that the conserved C85 residue potentially acts as a redox switch that is indispensable for S100A1's antiarrhythmic but not its inotropic potency in CMs. We, therefore, propose a model where C85's ox-PTM determines S100A1's ability to beneficially control diastolic but not systolic RyR2 activity.NEW & NOTEWORTHY S100A1 is an emerging candidate for future gene-therapy treatment of human chronic heart failure. We aimed to study the significance of the conserved single-cysteine 85 (C85) residue in cardiomyocytes. We show that S100A1 is endogenously glutathionylated in the heart and demonstrate that this is dispensable to increase systolic Ca2+ transients, but indispensable for mediating S100A1's protection against sarcoplasmic reticulum (SR) Ca2+ waves, which was dependent on the ryanodine receptor 2 (RyR2) nitrosylation status.

miR-222 inhibits pathological cardiac hypertrophy and heart failure.

AIMS: Physiological cardiac hypertrophy occurs in response to exercise and can protect against pathological stress. In contrast, pathological hypertrophy occurs in disease and often precedes heart failure. The cardiac pathways activated in physiological and pathological hypertrophy are largely distinct. Our prior work demonstrated that miR-222 increases in exercised hearts and is required for exercise-induced cardiac hypertrophy and cardiomyogenesis. Here, we sought to define the role of miR-222 in pathological hypertrophy. METHODS AND RESULTS: We found that miR-222 also increased in pathological hypertrophy induced by pressure overload. To assess its functional significance in this setting, we generated a miR-222 gain-of-function model through cardiac-specific constitutive transgenic miR-222 expression (TgC-miR-222) and used locked nucleic acid anti-miR specific for miR-222 to inhibit its effects. Both gain- and loss-of-function models manifested normal cardiac structure and function at baseline. However, after transverse aortic constriction (TAC), miR-222 inhibition accelerated the development of pathological hypertrophy, cardiac dysfunction, and heart failure. Conversely, miR-222-overexpressing mice had less pathological hypertrophy after TAC, as well as better cardiac function and survival. We identified p53-up-regulated modulator of apoptosis, a pro-apoptotic Bcl-2 family member, and the transcription factors, Hmbox1 and nuclear factor of activated T-cells 3, as direct miR-222 targets contributing to its roles in this context. CONCLUSION: While miR-222 is necessary for physiological cardiac growth, it inhibits cardiac growth in response to pressure overload and reduces adverse remodelling and cardiac dysfunction. These findings support the model that physiological and pathological hypertrophy are fundamentally different. Further, they suggest that miR-222 may hold promise as a therapeutic target in pathological cardiac hypertrophy and heart failure.

Current understanding of thermo(dys)regulation in severe burn injury and the pathophysiological influence of hypermetabolism, adrenergic stress and hypothalamic regulation-a systematic review.

Background: In this systematic review, we summarize the aetiology as well as the current knowledge regarding thermo(dys)regulation and hypothermia after severe burn trauma and aim to present key concepts of pathophysiology and treatment options. Severe burn injuries with >20% total body surface area (TBSA) affected commonly leave the patient requiring several surgical procedures, prolonged hospital stays and cause substantial changes to body composition and metabolism in the acute and long-term phase. Particularly in severely burned patients, the loss of intact skin and the dysregulation of peripheral and central thermoregulatory processes may lead to substantial complications. Methods: A systematic and protocol-based search for suitable publications was conducted following the PRISMA guidelines. Articles were screened and included if deemed eligible. This encompasses animal-based in vivo studies as well as clinical studies examining the control-loops of thermoregulation and metabolic stability within burn patients. Results: Both experimental animal studies and clinical studies examining thermoregulation and metabolic functions within burn patients have produced a general understanding of core concepts which are, nonetheless, lacking in detail. We describe the wide range of pathophysiological alterations observed after severe burn trauma and highlight the association between thermoregulation and hypermetabolism as well as the interactions between nearly all organ systems. Lastly, the current clinical standards of mitigating the negative effects of thermodysregulation and hypothermia are summarized, as a comprehensive understanding and implementation of the key concepts is critical for patient survival and long-term well-being. Conclusions: The available in vivo animal models have provided many insights into the interwoven pathophysiology of severe burn injury, especially concerning thermoregulation. We offer an outlook on concepts of altered central thermoregulation from non-burn research as potential areas of future research interest and aim to provide an overview of the clinical implications of temperature management in burn patients.

Long-term sequelae of critical illness in sepsis, trauma and burns: A systematic review and meta-analysis.

BACKGROUND: Sepsis, major trauma, and severe burn injury are life-threatening critical illnesses that remain significant contributors to worldwide morbidity and mortality. The three underlying etiologies share pathophysiological similarities: hyperinflammation, hypermetabolism, and acute immunomodulation. The aims of this study were to assess the current state of long-term outcome research and to identify key outcome parameters between the three forms of critical illness. METHODS: This systematic review and meta-analysis (MA) were conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-analyses guidelines. PubMed was searched from January 1, 1975, to December 31, 2019. Studies were assessed for eligibility by independent reviewers. Inclusion criteria were studies reporting at least a 6-month follow-up of health-related quality of life and organ-specific sequelae within the three etiologies: severe burn injury, sepsis, and major trauma. RESULTS: In total, 125 articles could be included in the systematic review and 74 in the MA. The mean follow-up time was significantly longer in burn studies, compared with sepsis and trauma studies. The majority of patients were from the sepsis group, followed by burns, and major trauma studies. In the overall health-related quality of life, as assessed by Short Form 36 and European Quality-of-Life Index, the three different etiologies were comparable with one another. CONCLUSION: The effects of critical illness on survivors persist for years after hospitalization. Well-reported and reliable data on the long-term outcomes are imperative, as they can be used to determine the treatment choice of physicians and to guide the expectations of patients, improving the overall quality of care of three significant patient cohorts. LEVEL OF EVIDENCE: Systematic review and MA, level III.

Differential regulation of KCa 2.1 (KCNN1) K+ channel expression by histone deacetylases in atrial fibrillation with concomitant heart failure.

Atrial fibrillation (AF) with concomitant heart failure (HF) poses a significant therapeutic challenge. Mechanism-based approaches may optimize AF therapy. Small-conductance, calcium-activated K+ (KCa , KCNN) channels contribute to cardiac action potential repolarization. KCNN1 exhibits predominant atrial expression and is downregulated in chronic AF patients with preserved cardiac function. Epigenetic regulation is suggested by AF suppression following histone deacetylase (HDAC) inhibition. We hypothesized that HDAC-dependent KCNN1 remodeling contributes to arrhythmogenesis in AF complicated by HF. The aim of this study was to assess KCNN1 and HDAC1-7 and 9 transcript levels in AF/HF patients and in a pig model of atrial tachypacing-induced AF with reduced left ventricular function. In HL-1 atrial myocytes, tachypacing and anti-Hdac siRNAs were employed to investigate effects on Kcnn1 mRNA levels. KCNN1 expression displayed side-specific remodeling in AF/HF patients with upregulation in left and suppression in right atrium. In pigs, KCNN1 remodeling showed intermediate phenotypes. HDAC levels were differentially altered in humans and pigs, reflecting highly variable epigenetic regulation. Tachypacing recapitulated downregulation of Hdacs 1, 3, 4, 6, and 7 with a tendency towards reduced Kcnn1 levels in vitro, indicating that atrial high rates induce remodeling. Finally, Kcnn1 expression was decreased by knockdown of Hdacs 2, 3, 6, and 7 and enhanced by genetic Hdac9 inactivation, while anti-Hdac 1, 4, and 5 siRNAs did not affect Kcnn1 transcript levels. In conclusion, KCNN1 and HDAC expression is differentially remodeled in AF complicated by HF. Direct regulation of KCNN1 by HDACs in atrial myocytes provides a basis for mechanism-based antiarrhythmic therapy.

Trigger-Specific Remodeling of KCa2 Potassium Channels in Models of Atrial Fibrillation.

AIM: Effective antiarrhythmic treatment of atrial fibrillation (AF) constitutes a major challenge, in particular, when concomitant heart failure (HF) is present. HF-associated atrial arrhythmogenesis is distinctly characterized by prolonged atrial refractoriness. Small-conductance, calcium-activated K+ (KCa, SK, KCNN) channels contribute to cardiac action potential repolarization and are implicated in AF susceptibility and therapy. The mechanistic impact of AF/HF-related triggers on atrial KCa channels is not known. We hypothesized that tachycardia, stretch, ß-adrenergic stimulation, and hypoxia differentially determine KCa2.1-2.3 channel remodeling in atrial cells. METHODS: KCNN1-3 transcript levels were assessed in AF/HF patients and in a pig model of atrial tachypacing-induced AF with reduced left ventricular function. HL-1 atrial myocytes were subjected to proarrhythmic triggers to investigate the effects on Kcnn mRNA and KCa channel protein. RESULTS: Atrial KCNN1-3 expression was reduced in AF/HF patients. KCNN2 and KCNN3 suppression was recapitulated in the corresponding pig model. In contrast to human AF, KCNN1 remained unchanged in pigs. Channel- and stressor-specific remodeling was revealed in vitro. Lower expression levels of KCNN1/KCa2.1 were linked to stretch and ß-adrenergic stimulation. Furthermore, KCNN3/KCa2.3 expression was suppressed upon tachypacing and hypoxia. Finally, KCNN2/KCa2.2 abundance was specifically enhanced by hypoxia. CONCLUSION: Reduction of KCa2.1-2.3 channel expression might contribute to the action potential prolongation in AF complicated by HF. Subtype-specific KCa2 channel remodeling induced by tachypacing, stretch, ß-adrenergic stimulation, or hypoxia is expected to differentially determine atrial remodeling, depending on patient-specific activation of each triggering factor. Stressor-dependent KCa2 regulation in atrial myocytes provides a starting point for mechanism-based antiarrhythmic therapy.

Ligand-activated RXFP1 gene therapy ameliorates pressure overload-induced cardiac dysfunction.

Recurrent episodes of decompensated heart failure (HF) represent an emerging cause of hospitalizations in developed countries with an urgent need for effective therapies. Recently, the pregnancy-related hormone relaxin (RLN) was found to mediate cardio-protective effects and act as a positive inotrope in the cardiovascular system. RLN binds to the RLN family peptide receptor 1 (RXFP1), which is predominantly expressed in atrial cardiomyocytes. We therefore hypothesized that ventricular RXFP1 expression might exert potential therapeutic effects in an in vivo model of cardiac dysfunction. Thus, mice were exposed to pressure overload by transverse aortic constriction and treated with AAV9 to ectopically express RXFP1. To activate RXFP1 signaling, RLN was supplemented subcutaneously. Ventricular RXFP1 expression was well tolerated. Additional RLN administration not only abrogated HF progression but restored left ventricular systolic function. In accordance, upregulation of fetal genes and pathological remodeling markers were significantly reduced. In vitro, RLN stimulation of RXFP1-expressing cardiomyocytes induced downstream signaling, resulting in protein kinase A (PKA)-specific phosphorylation of phospholamban (PLB), which was distinguishable from ß-adrenergic activation. PLB phosphorylation corresponded to increased calcium amplitude and contractility. In conclusion, our results demonstrate that ligand-activated cardiac RXFP1 gene therapy represents a therapeutic approach to attenuate HF with the potential to adjust therapy by exogenous RLN supplementation.

Consensus Transcriptional Landscape of Human End-Stage Heart Failure.

Background Transcriptomic studies have contributed to fundamental knowledge of myocardial remodeling in human heart failure (HF). However, the key HF genes reported are often inconsistent between studies, and systematic efforts to integrate evidence from multiple patient cohorts are lacking. Here, we aimed to provide a framework for comprehensive comparison and analysis of publicly available data sets resulting in an unbiased consensus transcriptional signature of human end-stage HF. Methods and Results We curated and uniformly processed 16 public transcriptomic studies of left ventricular samples from 263 healthy and 653 failing human hearts. First, we evaluated the degree of consistency between studies by using linear classifiers and overrepresentation analysis. Then, we meta-analyzed the deregulation of 14 041 genes to extract a consensus signature of HF. Finally, to functionally characterize this signature, we estimated the activities of 343 transcription factors, 14 signaling pathways, and 182 micro RNAs, as well as the enrichment of 5998 biological processes. Machine learning approaches revealed conserved disease patterns across all studies independent of technical differences. These consistent molecular changes were prioritized with a meta-analysis, functionally characterized and validated on external data. We provide all results in a free public resource (https://saezlab.shinyapps.io/reheat/) and exemplified usage by deciphering fetal gene reprogramming and tracing the potential myocardial origin of the plasma proteome markers in patients with HF. Conclusions Even though technical and sampling variability confound the identification of differentially expressed genes in individual studies, we demonstrated that coordinated molecular responses during end-stage HF are conserved. The presented resource is crucial to complement findings in independent studies and decipher fundamental changes in failing myocardium.

Development of an AAV9-RNAi-mediated silencing strategy to abrogate TRPM4 expression in the adult heart.

The cation channel transient receptor potential melastatin 4 (TRPM4) is a calcium-activated non-selective cation channel and acts in cardiomyocytes as a negative modulator of the L-type Ca2+ influx. Global deletion of TRPM4 in the mouse led to increased cardiac contractility under ß-adrenergic stimulation. Consequently, cardiomyocyte-specific inactivation of the TRPM4 function appears to be a promising strategy to improve cardiac contractility in heart failure patients. The aim of this study was to develop a gene therapy approach in mice that specifically silences the expression of TRPM4 in cardiomyocytes. First, short hairpin RNAmiR30 (shRNAmiR30) sequences against the TRPM4 mRNA were screened in vitro using lentiviral transduction for a stable expression of the shRNA cassettes. Western blot analysis identified three efficient shRNAmiR30 sequences out of six, which reduced the endogenous TRPM4 protein level by up to 90 ± 6%. Subsequently, the most efficient shRNAmiR30 sequences were delivered into cardiomyocytes of adult mice using adeno-associated virus serotype 9 (AAV9)-mediated gene transfer. Initially, the AAV9 vector particles were administered via the lateral tail vein, which resulted in a downregulation of TRPM4 by 46 ± 2%. Next, various optimization steps were carried out to improve knockdown efficiency in vivo. First, the design of the expression cassette was streamlined for integration in a self-complementary AAV vector backbone for a faster expression. Compared to the application via the lateral tail vein, intravenous application via the retro-orbital sinus has the advantage that the vector solution reaches the heart directly and in a high concentration, and eventually a TRPM4 knockdown efficiency of 90 ± 7% in the heart was accomplished by this approach. By optimization of the shRNAmiR30 constructs and expression cassette as well as the route of AAV9 vector application, a 90% reduction of TRPM4 expression was achieved in the adult mouse heart. In the future, AAV9-RNAi-mediated inactivation of TRPM4 could be a promising strategy to increase cardiac contractility in preclinical animal models of acute and chronic forms of cardiac contractile failure.

Epigenetic regulation of cardiac electrophysiology in atrial fibrillation: HDAC2 determines action potential duration and suppresses NRSF in cardiomyocytes.

Atrial fibrillation (AF) is associated with electrical remodeling, leading to cellular electrophysiological dysfunction and arrhythmia perpetuation. Emerging evidence suggests a key role for epigenetic mechanisms in the regulation of ion channel expression. Histone deacetylases (HDACs) control gene expression through deacetylation of histone proteins. We hypothesized that class I HDACs in complex with neuron-restrictive silencer factor (NRSF) determine atrial K+ channel expression. AF was characterized by reduced atrial HDAC2 mRNA levels and upregulation of NRSF in humans and in a pig model, with regional differences between right and left atrium. In vitro studies revealed inverse regulation of Hdac2 and Nrsf in HL-1 atrial myocytes. A direct association of HDAC2 with active regulatory elements of cardiac K+ channels was revealed by chromatin immunoprecipitation. Specific knock-down of Hdac2 and Nrsf induced alterations of K+ channel expression. Hdac2 knock-down resulted in prolongation of action potential duration (APD) in neonatal rat cardiomyocytes, whereas inactivation of Nrsf induced APD shortening. Potential AF-related triggers were recapitulated by experimental tachypacing and mechanical stretch, respectively, and exerted differential effects on the expression of class I HDACs and K+ channels in cardiomyocytes. In conclusion, HDAC2 and NRSF contribute to AF-associated remodeling of APD and K+ channel expression in cardiomyocytes via direct interaction with regulatory chromatin regions. Specific modulation of these factors may provide a starting point for the development of more individualized treatment options for atrial fibrillation.

Updated and enhanced pig cardiac transcriptome based on long-read RNA sequencing and proteomics.

Clinically translatable large animal models have become indispensable for cardiovascular research, clinically relevant proof of concept studies and for novel therapeutic interventions. In particular, the pig has emerged as an essential cardiovascular disease model, because its heart, circulatory system, and blood supply are anatomically and functionally similar to that of humans. Currently, molecular and omics-based studies in the pig are hampered by the incompleteness of the genome and the lack of diversity of the corresponding transcriptome annotation. Here, we employed Nanopore long-read sequencing and in-depth proteomics on top of Illumina RNA-seq to enhance the pig cardiac transcriptome annotation. We assembled 15,926 transcripts, stratified into coding and non-coding, and validated our results by complementary mass spectrometry. A manual review of several gene loci, which are associated with cardiac function, corroborated the utility of our enhanced annotation. All our data are available for download and are provided as tracks for integration in genome browsers. We deem this resource as highly valuable for molecular research in an increasingly relevant large animal model.

HDAC2-dependent remodeling of KCa2.2 (KCNN2) and KCa2.3 (KCNN3) K+ channels in atrial fibrillation with concomitant heart failure.

AIMS: Atrial fibrillation (AF) with concomitant heart failure (HF) is associated with prolonged atrial refractoriness. Small-conductance, calcium-activated K+ (KCa, KCNN) channels promote action potential (AP) repolarization. KCNN2 and KCNN3 variants are associated with AF risk. In addition, histone deacetylase (HDAC)-related epigenetic mechanisms have been implicated in AP regulation. We hypothesized that HDAC2-dependent remodeling of KCNN2 and KCNN3 expression contributes to atrial arrhythmogenesis in AF complicated by HF. The objectives were to assess HDAC2 and KCNN2/3 transcript levels in AF/HF patients and in a pig model, and to investigate cellular epigenetic effects of HDAC2 inactivation on KCNN expression. MATERIALS AND METHODS: HDAC2 and KCNN2/3 transcript levels were quantified in patients with AF and HF, and in a porcine model of atrial tachypacing-induced AF and reduced left ventricular function. Tachypacing and anti-Hdac2 siRNA treatment were employed in HL-1 atrial myocytes to study effects on KCNN2/3 mRNA and KCa protein abundance. KEY FINDINGS: Atrial KCNN2 and KCNN3 expression was reduced in AF/HF patients and in a corresponding pig model. HDAC2 displayed significant downregulation in humans and a tendency towards reduced expression in right atrial tissue of pigs. Tachypacing recapitulated downregulation of Kcnn2/KCa2.2, Kcnn3/KCa2.3 and Hdac2/HDAC2, indicating that high atrial rates trigger epigenetic remodeling mechanisms. Finally, knock-down of Hdac2 in vitro reduced Kcnn3/KCa2.3 expression. SIGNIFICANCE: KCNN2/3 and HDAC2 expression is suppressed in AF complicated by HF. Hdac2 directly regulates Kcnn3 mRNA levels in atrial cells. The mechanistic and therapeutic significance of epigenetic electrophysiological effects in AF requires further validation.

Deep Characterization of Circular RNAs from Human Cardiovascular Cell Models and Cardiac Tissue.

For decades, cardiovascular disease (CVD) has been the leading cause of death throughout most developed countries. Several studies relate RNA splicing, and more recently also circular RNAs (circRNAs), to CVD. CircRNAs originate from linear transcripts and have been shown to exhibit tissue-specific expression profiles. Here, we present an in-depth analysis of sequence, structure, modification, and cardiac circRNA interactions. We used human induced pluripotent stem cell-derived cardiac myocytes (hiPSC-CMs), human healthy and diseased (ischemic cardiomyopathy, dilated cardiomyopathy) cardiac tissue, and human umbilical vein endothelial cells (HUVECs) to profile circRNAs. We identified shared circRNAs across all samples, as well as model-specific circRNA signatures. Based on these circRNAs, we identified 63 positionally conserved and expressed circRNAs in human, pig, and mouse hearts. Furthermore, we found that the sequence of circRNAs can deviate from the sequence derived from the genome sequence, an important factor in assessing potential functions. Integration of additional data yielded evidence for m6A-methylation of circRNAs, potentially linked to translation, as well as, circRNAs overlapping with potential Argonaute 2 binding sites, indicating potential association with the RISC complex. Moreover, we describe, for the first time in cardiac model systems, a sub class of circRNAs containing the start codon of their primary transcript (AUG circRNAs) and observe an enrichment for m6A-methylation for AUG circRNAs.

From traditional pharmacological towards nucleic acid-based therapies for cardiovascular diseases.

Nucleic acid-based therapeutics are currently developed at large scale for prevention and management of cardiovascular diseases (CVDs), since: (i) genetic studies have highlighted novel therapeutic targets suggested to be causal for CVD; (ii) there is a substantial recent progress in delivery, efficacy, and safety of nucleic acid-based therapies; (iii) they enable effective modulation of therapeutic targets that cannot be sufficiently or optimally addressed using traditional small molecule drugs or antibodies. Nucleic acid-based therapeutics include (i) RNA-targeted therapeutics for gene silencing; (ii) microRNA-modulating and epigenetic therapies; (iii) gene therapies; and (iv) genome-editing approaches (e.g. CRISPR-Cas-based): (i) RNA-targeted therapeutics: several large-scale clinical development programmes, using antisense oligonucleotides (ASO) or short interfering RNA (siRNA) therapeutics for prevention and management of CVD have been initiated. These include ASO and/or siRNA molecules to lower apolipoprotein (a) [apo(a)], proprotein convertase subtilisin/kexin type 9 (PCSK9), apoCIII, ANGPTL3, or transthyretin (TTR) for prevention and treatment of patients with atherosclerotic CVD or TTR amyloidosis. (ii) MicroRNA-modulating and epigenetic therapies: novel potential therapeutic targets are continually arising from human non-coding genome and epigenetic research. First microRNA-based therapeutics or therapies targeting epigenetic regulatory pathways are in clinical studies. (iii) Gene therapies: EMA/FDA have approved gene therapies for non-cardiac monogenic diseases and LDL receptor gene therapy is currently being examined in patients with homozygous hypercholesterolaemia. In experimental studies, gene therapy has significantly improved cardiac function in heart failure animal models. (iv) Genome editing approaches: these technologies, such as using CRISPR-Cas, have proven powerful in stem cells, however, important challenges are remaining, e.g. low rates of homology-directed repair in somatic cells such as cardiomyocytes. In summary, RNA-targeted therapies (e.g. apo(a)-ASO and PCSK9-siRNA) are now in large-scale clinical outcome trials and will most likely become a novel effective and safe therapeutic option for CVD in the near future. MicroRNA-modulating, epigenetic, and gene therapies are tested in early clinical studies for CVD. CRISPR-Cas-mediated genome editing is highly effective in stem cells, but major challenges are remaining in somatic cells, however, this field is rapidly advancing.

Cardiac Dysfunction in Severely Burned Patients: Current Understanding of Etiology, Pathophysiology, and Treatment.

Patients who experience severe burn injuries face a massive inflammatory response resulting in hemodynamic and cardiovascular complications. Even after immediate and appropriate resuscitation, removal of burn eschar and covering of open areas, burn patients remain at high risk for serious morbidity and mortality. As a result of the massive fluid shifts following the initial injury, along with large volume fluid resuscitation, the cardiovascular system is critically affected. Further, increased inflammation, catecholamine surge, and hypermetabolic syndrome impact cardiac dysfunction, which worsens outcomes of burn patients. This review aimed to summarize the current knowledge about the effect of burns on the cardiovascular system.A comprehensive search of the PubMed and Embase databases and manual review of articles involving effects of burns on the cardiovascular system was conducted.Many burn units use multimodal monitors (e.g., transpulmonary thermodilution) to assess hemodynamics and optimize cardiovascular function. Echocardiography is often used for additional evaluations of hemodynamically unstable patients to assess systolic and diastolic function. Due to its noninvasive character, echocardiography can be repeated easily, which allows us to follow patients longitudinally.The use of anabolic and anticatabolic agents has been shown to be beneficial for short- and long-term outcomes of burn survivors. Administration of propranolol (non-selective ß-receptor antagonist) or oxandrolone (synthetic testosterone) for up to 12 months post-burn counteracts hypermetabolism during hospital stay and improves cardiac function.A comprehensive understanding of how burns lead to cardiac dysfunction and new therapeutic options could contribute to better outcomes in this patient population.

The lipid droplet-associated protein ABHD5 protects the heart through proteolysis of HDAC4.

Catecholamines stimulate the first step of lipolysis by PKA-dependent release of the lipid droplet-associated protein ABHD5 from perilipin to co-activate the lipase ATGL. Here, we unmask a yet unrecognized proteolytic and cardioprotective function of ABHD5. ABHD5 acts in vivo and in vitro as a serine protease cleaving HDAC4. Through the production of an N-terminal polypeptide of HDAC4 (HDAC4-NT), ABHD5 inhibits MEF2-dependent gene expression and thereby controls glucose handling. ABHD5-deficiency leads to neutral lipid storage disease in mice. Cardiac-specific gene therapy of HDAC4-NT does not protect from intra-cardiomyocyte lipid accumulation but strikingly from heart failure, thereby challenging the concept of lipotoxicity-induced heart failure. ABHD5 levels are reduced in failing human hearts and murine transgenic ABHD5 expression protects from pressure-overload induced heart failure. These findings represent a conceptual advance by connecting lipid with glucose metabolism through HDAC4 proteolysis and enable new translational approaches to treat cardiometabolic disease.

Comprehensive cardiac phenotyping in large animals: comparison of pressure-volume analysis and cardiac magnetic resonance imaging in pig post-myocardial infarction systolic heart failure.

Large animal ischemic cardiomyopathy models are widely used for preclinical testing of promising novel therapeutic approaches. Pressure volume (PV) loop analysis and cardiac magnetic resonance imaging (CMRI) allow functional and morphological phenotyping. In this study we performed a comparative analysis of both methods highlighting the strength of each and their synergistic potential. Myocardial infarction (MI) was created in German farm pigs (German Landrace) by 2 h LCX occlusion (n = 11) and subsequent reperfusion. Cardiac function was assessed by PV-loops and CMRI 56 and 112 days post-MI. Two hours occlusion of the LCX led to mid-size left ventricular (LV) MI represented by high-sensitive troponin T (hsTnT) 3 days post-MI, correlating well with cardiac CMRI late enhancement. CMRI determined end-diastolic and end-systolic volumes significantly increased post-MI, while ejection fraction was reduced in infarcted animals compared to the sham group (n = 6). PV-loop derived preload-insensitive parameters of systolic and diastolic function were diminished post-MI compared to sham animals while preload-dependent parameters only deteriorated in advanced HF. PV-loop analysis significantly correlates with CMRI analysis of cardiac function in pig post-MI ischemic cardiomyopathy. PV-Loop analysis accurately quantifies LV volumetry and function in post-MI HF, and thus eccentric LV morphology. PV-loop analysis correlates well to cardiac MRI. Preload-insensitive parameters show high sensitivity to quantify HF while preload-sensitive parameters are not able to quantify early-stages of LV HF.