Publications by authors named "Roger J Hajjar"

360 Publications

Molecules linked to Ras signaling as therapeutic targets in cardiac pathologies.

Biol Res 2021 Aug 3;54(1):23. Epub 2021 Aug 3.

Division of Nephrology, Department of Medicine, Jacobs School of Medicine and Biomedical Sciences, State University of New York at Buffalo, Clinical and Translational Research Center, 875 Ellicott Street, Suite 8030B, Buffalo, NY, 14203, USA.

The Ras family of small Guanosine Triphosphate (GTP)-binding proteins (G proteins) represents one of the main components of intracellular signal transduction required for normal cardiac growth, but is also critically involved in the development of cardiac hypertrophy and heart failure. The present review provides an update on the role of the H-, K- and N-Ras genes and their related pathways in cardiac diseases. We focus on cardiac hypertrophy and heart failure, where Ras has been studied the most. We also review other cardiac diseases, like genetic disorders related to Ras. The scope of the review extends from fundamental concepts to therapeutic applications. Although the three Ras genes have a nearly identical primary structure, there are important functional differences between them: H-Ras mainly regulates cardiomyocyte size, whereas K-Ras regulates cardiomyocyte proliferation. N-Ras is the least studied in cardiac cells and is less associated to cardiac defects. Clinically, oncogenic H-Ras causes Costello syndrome and facio-cutaneous-skeletal syndromes with hypertrophic cardiomyopathy and arrhythmias. On the other hand, oncogenic K-Ras and alterations of other genes of the Ras-Mitogen-Activated Protein Kinase (MAPK) pathway, like Raf, cause Noonan syndrome and cardio-facio-cutaneous syndromes characterized by cardiac hypertrophy and septal defects. We further review the modulation by Ras of key signaling pathways in the cardiomyocyte, including: (i) the classical Ras-Raf-MAPK pathway, which leads to a more physiological form of cardiac hypertrophy; as well as other pathways associated with pathological cardiac hypertrophy, like (ii) The SAPK (stress activated protein kinase) pathways p38 and JNK; and (iii) The alternative pathway Raf-Calcineurin-Nuclear Factor of Activated T cells (NFAT). Genetic alterations of Ras isoforms or of genes in the Ras-MAPK pathway result in Ras-opathies, conditions frequently associated with cardiac hypertrophy or septal defects among other cardiac diseases. Several studies underline the potential role of H- and K-Ras as a hinge between physiological and pathological cardiac hypertrophy, and as potential therapeutic targets in cardiac hypertrophy and failure.
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http://dx.doi.org/10.1186/s40659-021-00342-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8330049PMC
August 2021

Direct Reprogramming Induces Vascular Regeneration Post Muscle Ischemic Injury.

Mol Ther 2021 Jul 28. Epub 2021 Jul 28.

Cardiovascular Research Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA, 10029; Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA, 10029; Black Family Stem Cell Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA, 10029. Electronic address:

Reprogramming non-cardiomyocytes (non-CMs) into cardiomyocyte (CM)-like cells is a promising strategy for cardiac regeneration in conditions such as ischemic heart disease. Here, we used a modified mRNA (modRNA) gene delivery platform to deliver a cocktail of four cardiac-reprogramming genes (Gata4 (G), Mef2c (M), Tbx5 (T) and Hand2 (H)) together with three reprogramming-helper genes (Dominant Negative (DN)-TGFβ, DN-Wnt8a and Acid ceramidase (AC)), termed 7G-modRNA, to induce CM-like cells. We showed that 7G-modRNA reprogrammed 57% of CM-like cells in vitro. Through a lineage-tracing model, we determined that delivering the 7G-modRNA cocktail at the time of myocardial infarction reprogrammed ∼25% of CM-like cells in the scar area and significantly improved cardiac function, scar size, long-term survival and capillary density. Mechanistically, we determined that while 7G-modRNA cannot create de-novo beating CMs in vitro or in vivo, it can significantly upregulate pro-angiogenic mesenchymal stromal cells markers and transcription factors. We also demonstrated that our 7G-modRNA cocktail leads to neovascularization in ischemic-limb injury, indicating CM-like cells importance in other organs besides the heart. modRNA is currently being used around the globe for vaccination against COVID-19, and this study proves this is a safe, highly efficient gene delivery approach with therapeutic potential to treat ischemic diseases.
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http://dx.doi.org/10.1016/j.ymthe.2021.07.014DOI Listing
July 2021

Global genome analysis reveals a vast and dynamic anellovirus landscape within the human virome.

Cell Host Microbe 2021 Aug 27;29(8):1305-1315.e6. Epub 2021 Jul 27.

Ring Therapeutics, Cambridge, MA 02139, USA. Electronic address:

Anelloviruses are a ubiquitous component of healthy human viromes and remain highly prevalent after being acquired early in life. The full extent of "anellome" diversity and its evolutionary dynamics remain unexplored. We employed in-depth sequencing of blood-transfusion donor(s)-recipient pairs coupled with public genomic resources for a large-scale assembly of anellovirus genomes and used the data to characterize global and personal anellovirus diversity through time. The breadth of the anellome is much greater than previously appreciated, and individuals harbor unique anellomes and transmit lineages that can persist for several months within a diverse milieu of endemic host lineages. Anellovirus sequence diversity is shaped by extensive recombination at all levels of divergence, hindering traditional phylogenetic analyses. Our findings illuminate the transmission dynamics and vast diversity of anelloviruses and set the foundation for future studies to characterize their biology.
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http://dx.doi.org/10.1016/j.chom.2021.07.001DOI Listing
August 2021

Impaired Right Ventricular Calcium Cycling Is an Early Risk Factor in R14del-Phospholamban Arrhythmias.

J Pers Med 2021 Jun 3;11(6). Epub 2021 Jun 3.

Department of Pharmacology and Systems Physiology, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA.

The inherited mutation (R14del) in the calcium regulatory protein phospholamban (PLN) is linked to malignant ventricular arrhythmia with poor prognosis starting at adolescence. However, the underlying early mechanisms that may serve as prognostic factors remain elusive. This study generated humanized mice in which the endogenous gene was replaced with either human wild type or R14del-PLN and addressed the early molecular and cellular pathogenic mechanisms. R14del-PLN mice exhibited stress-induced impairment of atrioventricular conduction, and prolongation of both ventricular activation and repolarization times in association with ventricular tachyarrhythmia, originating from the right ventricle (RV). Most of these distinct electrocardiographic features were remarkably similar to those in R14del-PLN patients. Studies in isolated cardiomyocytes revealed RV-specific calcium defects, including prolonged action potential duration, depressed calcium kinetics and contractile parameters, and elevated diastolic Ca-levels. Ca-sparks were also higher although SR Ca-load was reduced. Accordingly, stress conditions induced after contractions, and inclusion of the CaMKII inhibitor KN93 reversed this proarrhythmic parameter. Compensatory responses included altered expression of key genes associated with Ca-cycling. These data suggest that R14del-PLN cardiomyopathy originates with RV-specific impairment of Ca-cycling and point to the urgent need to improve risk stratification in asymptomatic carriers to prevent fatal arrhythmias and delay cardiomyopathy onset.
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http://dx.doi.org/10.3390/jpm11060502DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8226909PMC
June 2021

Arrhythmia Mechanism and Dynamics in a Humanized Mouse Model of Inherited Cardiomyopathy Caused by Phospholamban R14del Mutation.

Circulation 2021 Aug 24;144(6):441-454. Epub 2021 May 24.

Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, New York, NY (N.R., M.C., D.J., Z.I., D.C., E.K., F.S., F.G.A.).

Background: Arginine (Arg) 14 deletion (R14del) in the calcium regulatory protein phospholamban (hPLN) has been identified as a disease-causing mutation in patients with an inherited cardiomyopathy. Mechanisms underlying the early arrhythmogenic phenotype that predisposes carriers of this mutation to sudden death with no apparent structural remodeling remain unclear.

Methods: To address this, we performed high spatiotemporal resolution optical mapping of intact hearts from adult knock-in mice harboring the human PLN (wildtype [WT], n=12) or the heterozygous human PLN mutation (R14del, n=12) before and after ex vivo challenge with isoproterenol and rapid pacing.

Results: Adverse electrophysiological remodeling was evident in the absence of significant structural or hemodynamic changes. R14del hearts exhibited increased arrhythmia susceptibility compared with wildtype. Underlying this susceptibility was preferential right ventricular action potential prolongation that was unresponsive to β-adrenergic stimulation. A steep repolarization gradient at the left ventricular/right ventricular interface provided the substrate for interventricular activation delays and ultimately local conduction block during rapid pacing. This was followed by the initiation of macroreentrant circuits supporting the onset of ventricular tachycardia. Once sustained, these circuits evolved into high-frequency rotors, which in their majority were pinned to the right ventricle. These rotors exhibited unique spatiotemporal dynamics that promoted their increased stability in R14del compared with wildtype hearts.

Conclusions: Our findings highlight the crucial role of primary electric remodeling caused by the hPLN mutation. These inherently arrhythmogenic features form the substrate for adrenergic-mediated VT at early stages of PLN induced cardiomyopathy.
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http://dx.doi.org/10.1161/CIRCULATIONAHA.119.043502DOI Listing
August 2021

Adverse effects of hydroxychloroquine and azithromycin on contractility and arrhythmogenicity revealed by human engineered cardiac tissues.

J Mol Cell Cardiol 2021 04 27;153:106-110. Epub 2020 Dec 27.

Sardocor, Boston, Massachusetts, United States. Electronic address:

The coronavirus disease 2019 (COVID-19) outbreak caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a global pandemic as declared by World Health Organization (WHO). In the absence of an effective treatment, different drugs with unknown effectiveness, including antimalarial hydroxychloroquine (HCQ), with or without concurrent administration with azithromycin (AZM), have been tested for treating COVID-19 patients with developed pneumonia. However, the efficacy and safety of HCQ and/or AZM have been questioned by recent clinical reports. Direct effects of these drugs on the human heart remain very poorly defined. To better understand the mechanisms of action of HCQ +/- AZM, we employed bioengineered human ventricular cardiac tissue strip (hvCTS) and anisotropic sheet (hvCAS) assays, made with human pluripotent stem cell (hPSC)-derived ventricular cardiomyocytes (hvCMs), which have been designed for measuring cardiac contractility and electrophysiology, respectively. Our hvCTS experiments showed that AZM induced a dose-dependent negative inotropic effect which could be aggravated by HCQ; electrophysiologically, as revealed by the hvCAS platform, AZM prolonged action potentials and induced spiral wave formations. Collectively, our data were consistent with reported clinical risks of HCQ and AZM on QTc prolongation/ventricular arrhythmias and development of heart failure. In conclusion, our study exposed the risks of HCQ/AZM administration while providing mechanistic insights for their toxicity. Our bioengineered human cardiac tissue constructs therefore provide a useful platform for screening cardiac safety and efficacy when developing therapeutics against COVID-19.
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http://dx.doi.org/10.1016/j.yjmcc.2020.12.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7765761PMC
April 2021

Specific Modified mRNA Translation System.

Circulation 2020 Dec 21;142(25):2485-2488. Epub 2020 Dec 21.

Cardiovascular Research Center (A.M., A.A.K., E.C., Y.S., L.Z.), Icahn School of Medicine at Mount Sinai, New York.

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http://dx.doi.org/10.1161/CIRCULATIONAHA.120.047211DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7768930PMC
December 2020

The matricellular protein CCN5 prevents adverse atrial structural and electrical remodelling.

J Cell Mol Med 2020 10 4;24(20):11768-11778. Epub 2020 Sep 4.

College of Life Sciences, Gwangju Institute of Science and Technology, Gwangju, Korea.

Atrial structural remodelling including atrial hypertrophy and fibrosis is a key mediator of atrial fibrillation (AF). We previously demonstrated that the matricellular protein CCN5 elicits anti-fibrotic and anti-hypertrophic effects in left ventricles under pressure overload. We here determined the utility of CCN5 in ameliorating adverse atrial remodelling and arrhythmias in a murine model of angiotensin II (AngII) infusion. Advanced atrial structural remodelling was induced by AngII infusion in control mice and mice overexpressing CCN5 either through transgenesis (CCN5 Tg) or AAV9-mediated gene transfer (AAV9-CCN5). The mRNA levels of pro-fibrotic and pro-inflammatory genes were markedly up-regulated by AngII infusion, which was significantly normalized by CCN5 overexpression. In vitro studies in isolated atrial fibroblasts demonstrated a marked reduction in AngII-induced fibroblast trans-differentiation in CCN5-treated atria. Moreover, while AngII increased the expression of phosphorylated CaMKII and ryanodine receptor 2 levels in HL-1 cells, these molecular features of AF were prevented by CCN5. Electrophysiological studies in ex vivo perfused hearts revealed a blunted susceptibility of the AAV9-CCN5-treated hearts to rapid atrial pacing-induced arrhythmias and concomitant reversal in AngII-induced atrial action potential prolongation. These data demonstrate the utility of a gene transfer approach targeting CCN5 for reversal of adverse atrial structural and electrophysiological remodelling.
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http://dx.doi.org/10.1111/jcmm.15789DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7579720PMC
October 2020

Induction and Characterization of Pulmonary Hypertension in Mice using the Hypoxia/SU5416 Model.

J Vis Exp 2020 06 3(160). Epub 2020 Jun 3.

Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai;

Pulmonary Hypertension (PH) is a pathophysiological condition, defined by a mean pulmonary arterial pressure exceeding 25 mm Hg at rest, as assessed by right heart catheterization. A broad spectrum of diseases can lead to PH, differing in their etiology, histopathology, clinical presentation, prognosis, and response to treatment. Despite significant progress in the last years, PH remains an uncured disease. Understanding the underlying mechanisms can pave the way for the development of new therapies. Animal models are important research tools to achieve this goal. Currently, there are several models available for recapitulating PH. This protocol describes a two-hit mouse PH model. The stimuli for PH development are hypoxia and the injection of SU5416, a vascular endothelial growth factor (VEGF) receptor antagonist. Three weeks after initiation of Hypoxia/SU5416, animals develop pulmonary vascular remodeling imitating the histopathological changes observed in human PH (predominantly Group 1). Vascular remodeling in the pulmonary circulation results in the remodeling of the right ventricle (RV). The procedures for measuring RV pressures (using the open chest method), the morphometrical analyses of the RV (by dissecting and weighing both cardiac ventricles) and the histological assessments of the remodeling (both pulmonary by assessing vascular remodeling and cardiac by assessing RV cardiomyocyte hypertrophy and fibrosis) are described in detail. The advantages of this protocol are the possibility of the application both in wild type and in genetically modified mice, the relatively easy and low-cost implementation, and the quick development of the disease of interest (3 weeks). Limitations of this method are that mice do not develop a severe phenotype and PH is reversible upon return to normoxia. Prevention, as well as therapy studies, can easily be implemented in this model, without the necessity of advanced skills (as opposed to surgical rodent models).
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http://dx.doi.org/10.3791/59252DOI Listing
June 2020

Pkm2 Regulates Cardiomyocyte Cell Cycle and Promotes Cardiac Regeneration.

Circulation 2020 04 11;141(15):1249-1265. Epub 2020 Feb 11.

Cardiovascular Research Center (A.M, N.S., A.A.K., I.M., T.M. K.B., M.T.K.S., E.C., Y.S., J.G.O., P.L, A.G.-S., C.K., M.M., L.Z.), Icahn School of Medicine at Mount Sinai, New York.

Background: The adult mammalian heart has limited regenerative capacity, mostly attributable to postnatal cardiomyocyte cell cycle arrest. In the last 2 decades, numerous studies have explored cardiomyocyte cell cycle regulatory mechanisms to enhance myocardial regeneration after myocardial infarction. Pkm2 (Pyruvate kinase muscle isoenzyme 2) is an isoenzyme of the glycolytic enzyme pyruvate kinase. The role of Pkm2 in cardiomyocyte proliferation, heart development, and cardiac regeneration is unknown.

Methods: We investigated the effect of Pkm2 in cardiomyocytes through models of loss (cardiomyocyte-specific Pkm2 deletion during cardiac development) or gain using cardiomyocyte-specific Pkm2 modified mRNA to evaluate Pkm2 function and regenerative affects after acute or chronic myocardial infarction in mice.

Results: Here, we identify Pkm2 as an important regulator of the cardiomyocyte cell cycle. We show that Pkm2 is expressed in cardiomyocytes during development and immediately after birth but not during adulthood. Loss of function studies show that cardiomyocyte-specific Pkm2 deletion during cardiac development resulted in significantly reduced cardiomyocyte cell cycle, cardiomyocyte numbers, and myocardial size. In addition, using cardiomyocyte-specific Pkm2 modified RNA, our novel cardiomyocyte-targeted strategy, after acute or chronic myocardial infarction, resulted in increased cardiomyocyte cell division, enhanced cardiac function, and improved long-term survival. We mechanistically show that Pkm2 regulates the cardiomyocyte cell cycle and reduces oxidative stress damage through anabolic pathways and β-catenin.

Conclusions: We demonstrate that Pkm2 is an important intrinsic regulator of the cardiomyocyte cell cycle and oxidative stress, and highlight its therapeutic potential using cardiomyocyte-specific Pkm2 modified RNA as a gene delivery platform.
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http://dx.doi.org/10.1161/CIRCULATIONAHA.119.043067DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7241614PMC
April 2020

Single SERCA2a Therapy Ameliorated Dilated Cardiomyopathy for 18 Months in a Mouse Model of Duchenne Muscular Dystrophy.

Mol Ther 2020 03 10;28(3):845-854. Epub 2020 Jan 10.

Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, MO 65212, USA; Department of Neurology, School of Medicine, University of Missouri, Columbia, MO 65212, USA; Department of Biomedical, Biological & Chemical Engineering, College of Engineering, University of Missouri, Columbia, MO 65212, USA; Department of Biomedical Sciences, College of Veterinary Medicine, University of Missouri, Columbia, MO 65212, USA. Electronic address:

Loss of dystrophin leads to Duchenne muscular dystrophy (DMD). A pathogenic feature of DMD is the significant elevation of cytosolic calcium. Supraphysiological calcium triggers protein degradation, membrane damage, and eventually muscle death and dysfunction. Sarcoplasmic/endoplasmic reticulum (SR) calcium ATPase (SERCA) is a calcium pump that transports cytosolic calcium to the SR during excitation-contraction coupling. We hypothesize that a single systemic delivery of SERCA2a with adeno-associated virus (AAV) may improve calcium recycling and provide long-lasting benefits in DMD. To test this, we injected an AAV9 human SERCA2a vector (6 × 10 viral genome particles/mouse) intravenously to 3-month-old mdx mice, the most commonly used DMD model. Immunostaining and western blot showed robust human SERCA2a expression in the heart and skeletal muscle for 18 months. Concomitantly, SR calcium uptake was significantly improved in these tissues. SERCA2a therapy significantly enhanced grip force and treadmill performance, completely prevented myocardial fibrosis, and normalized electrocardiograms (ECGs). Cardiac catheterization showed normalization of multiple systolic and diastolic hemodynamic parameters in treated mice. Importantly, chamber dilation was completely prevented, and ejection fraction was restored to the wild-type level. Our results suggest that a single systemic AAV9 SERCA2a therapy has the potential to provide long-lasting benefits for DMD.
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http://dx.doi.org/10.1016/j.ymthe.2019.12.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7054741PMC
March 2020

AAV1.SERCA2a Gene Therapy Reverses Pulmonary Fibrosis by Blocking the STAT3/FOXM1 Pathway and Promoting the SNON/SKI Axis.

Mol Ther 2020 02 6;28(2):394-410. Epub 2019 Dec 6.

Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA. Electronic address:

Inhibition of pulmonary fibrosis (PF) by restoring sarco/endoplasmic reticulum calcium ATPase 2a isoform (SERCA2a) expression using targeted gene therapy may be a potentially powerful new treatment approach for PF. Here, we found that SERCA2a expression was significantly decreased in lung samples from patients with PF and in the bleomycin (BLM) mouse model of PF. In the BLM-induced PF model, intratracheal aerosolized adeno-associated virus serotype 1 (AAV1) encoding for human SERCA2a (AAV1.hSERCA2a) reduces lung fibrosis and associated vascular remodeling. SERCA2a gene therapy also decreases right ventricular pressure and hypertrophy in both prevention and curative protocols. In vitro, we observed that SERCA2a overexpression inhibits fibroblast proliferation, migration, and fibroblast-to-myofibroblast transition induced by transforming growth factor β (TGF-β1). Thus, pro-fibrotic gene expression is prevented by blocking nuclear factor κB (NF-κB)/interleukin-6 (IL-6)-induced signal transducer and activator of transcription 3 (STAT3) activation. This effect is signaled toward an inhibitory mechanism of small mother against decapentaplegic (SMAD)/TGF-β signaling through the repression of OTU deubiquitinase, ubiquitin aldehyde binding 1 (OTUB1) and Forkhead box M1 (FOXM1). Interestingly, this cross-inhibition leads to an increase of SKI and SnoN expression, an auto-inhibitory feedback loop of TGF-β signaling. Collectively, our results demonstrate that SERCA2a gene transfer attenuates bleomycin (BLM)-induced PF by blocking the STAT3/FOXM1 pathway and promoting the SNON/SKI Axis. Thus, SERCA2a gene therapy may be a potential therapeutic target for PF.
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http://dx.doi.org/10.1016/j.ymthe.2019.11.027DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7001085PMC
February 2020

Echocardiographic Left Ventricular Mass Estimation: Two-Dimensional Area-Length Method is Superior to M-Mode Linear Method in Swine Models of Cardiac Diseases.

J Cardiovasc Transl Res 2020 08 11;13(4):648-658. Epub 2019 Dec 11.

Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, New York, NY, USA.

Echocardiography offers rapid and cost-effective estimations of left ventricular (LV) mass, but its accuracy in patients with cardiac disease remains unclear. LV mass was measured by M-mode-based linear method and two-dimensional echocardiography (2DE)-based area-length method in pig models and correlation with actual LV weight was assessed. Twenty-six normal, 195 ischemic heart disease (IHD), and 33 non-IHD HF pigs were included. A strong positive linear relationship to the actual LV weight was found with 2DE-based area-length method (r = 0.82, p < 0.001), whereas a moderate relationship was found with M-mode method in the overall population (r = 0.68, p < 0.001). Two correlation coefficients were significantly different (p < 0.001), and were driven mainly by incremental overestimation of LV mass in heavier hearts using the M-mode method. IHD and LV dilation were the factors contributing to overestimation using M-mode method. 2DE-based area-length method provides a better estimation of LV weight in swine models of HF, particularly in those with IHD.
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http://dx.doi.org/10.1007/s12265-019-09937-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7286771PMC
August 2020

Adult human cardiac stem cell supplementation effectively increases contractile function and maturation in human engineered cardiac tissues.

Stem Cell Res Ther 2019 12 4;10(1):373. Epub 2019 Dec 4.

Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, One Gustave L. Levy Pl, Box 1030, New York, NY, 10029, USA.

Background: Delivery of stem cells to the failing heart is a promising therapeutic strategy. However, the improvement in cardiac function in animal studies has not fully translated to humans. To help bridge the gap between species, we investigated the effects of adult human cardiac stem cells (hCSCs) on contractile function of human engineered cardiac tissues (hECTs) as a species-specific model of the human myocardium.

Methods: Human induced pluripotent stem cell-derived cardiomyoctes (hCMs) were mixed with Collagen/Matrigel to fabricate control hECTs, with an experimental group of hCSC-supplemented hECT fabricated using a 9:1 ratio of hCM to hCSC. Functional testing was performed starting on culture day 6, under spontaneous conditions and also during electrical pacing from 0.25 to 1.0 Hz, measurements repeated at days 8 and 10. hECTs were then frozen and processed for gene analysis using a Nanostring assay with a cardiac targeted custom panel.

Results: The hCSC-supplemented hECTs displayed a twofold higher developed force vs. hCM-only controls by day 6, with approximately threefold higher developed stress and maximum rates of contraction and relaxation during pacing at 0.75 Hz. The spontaneous beat rate characteristics were similar between groups, and hCSC supplementation did not adversely impact beat rate variability. The increased contractility persisted through days 8 and 10, albeit with some decrease in the magnitude of the difference of the force by day 10, but with developed stress still significantly higher in hCSC-supplemented hECT; these findings were confirmed with multiple hCSC and hCM cell lines. The force-frequency relationship, while negative for both, control (- 0.687 Hz; p = 0.013 vs. zero) and hCSC-supplemented (- 0.233 Hz;p = 0.067 vs. zero) hECTs, showed a significant rectification in the regression slope in hCSC-supplemented hECT (p = 0.011 vs. control). Targeted gene exploration (59 genes) identified a total of 14 differentially expressed genes, with increases in the ratios of MYH7/MHY6, MYL2/MYL7, and TNNI3/TNNI1 in hCSC-supplemented hECT versus controls.

Conclusions: For the first time, hCSC supplementation was shown to significantly improve human cardiac tissue contractility in vitro, without evidence of proarrhythmic effects, and was associated with increased expression of markers of cardiac maturation. These findings provide new insights about adult cardiac stem cells as contributors to functional improvement of human myocardium.
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http://dx.doi.org/10.1186/s13287-019-1486-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6894319PMC
December 2019

A novel secreted-cAMP pathway inhibits pulmonary hypertension via a feed-forward mechanism.

Cardiovasc Res 2020 07;116(8):1500-1513

Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, One Gustave L. Levy Place, Box 1030, New York, NY 10029, USA.

Aims: Cyclic adenosine monophosphate (cAMP) is the predominant intracellular second messenger that transduces signals from Gs-coupled receptors. Intriguingly, there is evidence from various cell types that an extracellular cAMP pathway is active in the extracellular space. Herein, we investigated the role of extracellular cAMP in the lung and examined whether it may act on pulmonary vascular cell proliferation and pulmonary vasculature remodelling in the pathogenesis of pulmonary hypertension (PH).

Methods And Results: The expression of cyclic AMP-metabolizing enzymes was increased in lungs from patients with PH as well as in rats treated with monocrotaline and mice exposed to Sugen/hypoxia. We report that inhibition of the endogenous extracellular cAMP pathway exacerbated Sugen/hypoxia-induced lung remodelling. We found that application of extracellular cAMP induced an increase in intracellular cAMP levels and inhibited proliferation and migration of pulmonary vascular cells in vitro. Extracellular cAMP infusion in two in vivo PH models prevented and reversed pulmonary and cardiac remodelling associated with PH. Using protein expression analysis along with luciferase assays, we found that extracellular cAMP acts via the A2R/PKA/CREB/p53/Cyclin D1 pathway.

Conclusions: Taken together, our data reveal the presence of an extracellular cAMP pathway in pulmonary arteries that attempts to protect the lung during PH, and suggest targeting of the extracellular cAMP signalling pathway to limit pulmonary vascular remodelling and PH.
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http://dx.doi.org/10.1093/cvr/cvz244DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7314640PMC
July 2020

Sarco/endoplasmic reticulum Ca-ATPase is a more effective calcium remover than sodium-calcium exchanger in human embryonic stem cell-derived cardiomyocytes.

Am J Physiol Heart Circ Physiol 2019 11 26;317(5):H1105-H1115. Epub 2019 Jul 26.

Dr. Li Dak-Sum Research Centre, The University of Hong Kong, Pokfulam, Hong Kong.

Human pluripotent stem cell (hPSCs)-derived ventricular (V) cardiomyocytes (CMs) display immature Ca-handing properties with smaller transient amplitudes and slower kinetics due to such differences in crucial Ca-handling proteins as the poor sarco/endoplasmic reticulum Ca-ATPase (SERCA) pump but robust Na-Ca exchanger (NCX) activities in human embryonic stem cell (ESC)-derived VCMs compared with adult. Despite their fundamental importance in excitation-contraction coupling, the relative contribution of SERCA and NCX to Ca-handling of hPSC-VCMs remains unexplored. We systematically altered the activities of SERCA and NCX in human embryonic stem cell-derived ventricular cardiomyocytes (hESC-VCMs) and their engineered microtissues, followed by examining the resultant phenotypic consequences. SERCA overexpression in hESC-VCMs shortened the decay of Ca transient at low frequencies (0.5 Hz) without affecting the amplitude, SR Ca content and Ca baseline. Interestingly, short hairpin RNA-based NCX suppression did not prolong the transient decay, indicating a compensatory response for Ca removal. Although hESC-VCMs and their derived microtissues exhibited negative frequency-transient/force responses, SERCA overexpression rendered them less negative at high frequencies (>2 Hz) by accelerating Ca sequestration. We conclude that for hESC-VCMs and their microtissues, SERCA, rather than NCX, is the main Ca remover during diastole; poor SERCA expression is the leading cause for immature negative-frequency/force responses, which can be partially reverted by forced expression. Combinatorial approach to mature calcium handling in hESC-VCMs may help shed further mechanistic insights. In this study of human pluripotent stem cell-derived cardiomyocytes, we studied the role of sarco/endoplasmic reticulum Ca-ATPase (SERCA) and Na-Ca exchanger (NCX) in Ca handling. Our data support the notion that SERCA is more effective in cytosolic calcium removal than the NCX.
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http://dx.doi.org/10.1152/ajpheart.00540.2018DOI Listing
November 2019

Allele-Specific Silencing Ameliorates Restrictive Cardiomyopathy Attributable to a Human Myosin Regulatory Light Chain Mutation.

Circulation 2019 08 18;140(9):765-778. Epub 2019 Jul 18.

Division of Cardiovascular Medicine (K.Z.-R., A.D., P.C., G.R., C.S., J.L., T.F., W.N.P., S.S., K.S., N.S., N.J., Y.H., M.T.W., E.A.A.), Stanford University School of Medicine, CA.

Background: Restrictive cardiomyopathy is a rare heart disease associated with mutations in sarcomeric genes and with phenotypic overlap with hypertrophic cardiomyopathy. There is no approved therapy directed at the underlying cause. Here, we explore the potential of an interfering RNA (RNAi) therapeutic for a human sarcomeric mutation in MYL2 causative of restrictive cardiomyopathy in a mouse model.

Methods: A short hairpin RNA (M7.8L) was selected from a pool for specificity and efficacy. Two groups of myosin regulatory light chain N47K transgenic mice were injected with M7.8L packaged in adeno-associated virus 9 at 3 days of age and 60 days of age. Mice were subjected to treadmill exercise and echocardiography after treatment to determine maximal oxygen uptake and left ventricular mass. At the end of treatment, heart, lung, liver, and kidney tissue was harvested to determine viral tropism and for transcriptomic and proteomic analysis. Cardiomyocytes were isolated for single-cell studies.

Results: A one-time injection of AAV9-M7.8L RNAi in 3-day-old humanized regulatory light chain mutant transgenic mice silenced the mutated allele (RLC-47K) with minimal effects on the normal allele (RLC-47N) assayed at 16 weeks postinjection. AAV9-M7.8L RNAi suppressed the expression of hypertrophic biomarkers, reduced heart weight, and attenuated a pathological increase in left ventricular mass. Single adult cardiac myocytes from mice treated with AAV9-M7.8L showed partial restoration of contraction, relaxation, and calcium kinetics. In addition, cardiac stress protein biomarkers, such as calmodulin-dependent protein kinase II and the transcription activator Brg1 were reduced, suggesting recovery toward a healthy myocardium. Transcriptome analyses further revealed no significant changes of argonaute (AGO1, AGO2) and endoribonuclease dicer (DICER1) transcripts, and endogenous microRNAs were preserved, suggesting that the RNAi pathway was not saturated.

Conclusions: Our results show the feasibility, efficacy, and safety of RNAi therapeutics directed towards human restrictive cardiomyopathy. This is a promising step toward targeted therapy for a prevalent human disease.
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http://dx.doi.org/10.1161/CIRCULATIONAHA.118.036965DOI Listing
August 2019

Modulation of chromatin remodeling proteins SMYD1 and SMARCD1 promotes contractile function of human pluripotent stem cell-derived ventricular cardiomyocyte in 3D-engineered cardiac tissues.

Sci Rep 2019 05 16;9(1):7502. Epub 2019 May 16.

Stem Cell and Regenerative Medicine Consortium, The University of Hong Kong, Pok Fu Lam, Hong Kong.

Human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) have the ability of differentiating into functional cardiomyocytes (CMs) for cell replacement therapy, tissue engineering, drug discovery and toxicity screening. From a scale-free, co-expression network analysis of transcriptomic data that distinguished gene expression profiles of undifferentiated hESC, hESC-, fetal- and adult-ventricular(V) CM, two candidate chromatin remodeling proteins, SMYD1 and SMARCD1 were found to be differentially expressed. Using lentiviral transduction, SMYD1 and SMARCD1 were over-expressed and suppressed, respectively, in single hESC-VCMs as well as the 3D constructs Cardiac Micro Tissues (CMT) and Tissue Strips (CTS) to mirror the endogenous patterns, followed by dissection of their roles in controlling cardiac gene expression, contractility, Ca-handling, electrophysiological functions and in vitro maturation. Interestingly, compared to independent single transductions, simultaneous SMYD1 overexpression and SMARCD1 suppression in hESC-VCMs synergistically interacted to increase the contractile forces of CMTs and CTSs with up-regulated transcripts for cardiac contractile, Ca-handing, and ion channel proteins. Certain effects that were not detected at the single-cell level could be unleashed under 3D environments. The two chromatin remodelers SMYD1 and SMARCD1 play distinct roles in cardiac development and maturation, consistent with the notion that epigenetic priming requires triggering signals such as 3D environmental cues for pro-maturation effects.
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http://dx.doi.org/10.1038/s41598-019-42953-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6522495PMC
May 2019

Animal model of left atrial thrombus in congestive heart failure in rats.

Am J Physiol Heart Circ Physiol 2019 07 10;317(1):H63-H72. Epub 2019 May 10.

Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai , New York, New York.

The aim of the present study was to develop and study a new model of left atrial thrombus (LAT) in rat with congestive heart failure (CHF). CHF was induced by aortic banding for 2 mo, followed by ischemia-reperfusion (I/R) and subsequent aortic debanding for 1 mo. Cardiac function and the presence of LAT were assessed by echocardiography. Masson's staining was performed for histological analysis. All CHF rats presented with significantly decreased cardiac function, fibrosis in remote myocardium, and pulmonary edema. The incidence rate of LAT was 18.8% in the rats. LAT was associated with severity of aortic constriction, aortic pressure gradient, aortic blood flow velocity, and pulmonary edema but not myocardial infarction or a degree of left ventricular depression. The progressive process of thrombogenesis was characterized by myocyte hypertrophy, fibrosis, and inflammation in the left atrial wall. Fibrin adhesion and clot formation were observed, whereas most LAT presented as a relatively hard "mass," likely attributable to significant fibrosis in the middle and outer layers. Some LAT mass showed focal necrosis as well as fibrin bulging. Most LAT occurred at the upper anterior wall of the left atrial appendage. Aortic debanding had no significant impact on large LATs (>5 mm) that had formed, whereas small LATs (<5 mm) regressed 1 mo after aortic release. LAT is found in a rat model of aortic banding plus I/R followed by aortic debanding. The model provides a platform to study molecular mechanisms and potential new pathways for LAT treatment. It is critically important to have a rodent model to study the molecular mechanism of thrombogenesis in the left atrium. Left atrial thrombus (LAT) is not a simple fibrin clot like those seen in peripheral veins or arteries. Rather, LAT is a cellular mass that likely develops in conjunction with blood clotting. Studying this phenomenon will help us understand congestive heart failure and promote new therapies for LAT.
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http://dx.doi.org/10.1152/ajpheart.00086.2019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6692738PMC
July 2019

CXCR4 Cardiac Specific Knockout Mice Develop a Progressive Cardiomyopathy.

Int J Mol Sci 2019 May 8;20(9). Epub 2019 May 8.

Department of Physiology and Biophysics, College of Medicine, Howard University, Washington, DC 20060, USA.

Activation of multiple pathways is associated with cardiac hypertrophy and heart failure. We previously published that CXCR4 negatively regulates β-adrenergic receptor (β-AR) signaling and ultimately limits β-adrenergic diastolic (Ca) accumulation in cardiac myocytes. In isolated adult rat cardiac myocytes; CXCL12 treatment prevented isoproterenol-induced hypertrophy and interrupted the calcineurin/NFAT pathway. Moreover; cardiac specific CXCR4 knockout mice show significant hypertrophy and develop cardiac dysfunction in response to chronic catecholamine exposure in an isoproterenol-induced (ISO) heart failure model. We set this study to determine the structural and functional consequences of CXCR4 myocardial knockout in the absence of exogenous stress. Cardiac phenotype and function were examined using (1) gated cardiac magnetic resonance imaging (MRI); (2) terminal cardiac catheterization with in vivo hemodynamics; (3) histological analysis of left ventricular (LV) cardiomyocyte dimension; fibrosis; and; (4) transition electron microscopy at 2-; 6- and 12-months of age to determine the regulatory role of CXCR4 in cardiomyopathy. Cardiomyocyte specific-CXCR4 knockout (CXCR4 cKO) mice demonstrate a progressive cardiac dysfunction leading to cardiac failure by 12-months of age. Histological assessments of CXCR4 cKO at 6-months of age revealed significant tissue fibrosis in knockout mice versus wild-type. The expression of atrial naturietic factor (ANF); a marker of cardiac hypertrophy; was also increased with a subsequent increase in gross heart weights. Furthermore, there were derangements in both the number and the size of the mitochondria within CXCR4 cKO hearts. Moreover, CXCR4 cKO mice were more sensitive to catocholamines, their response to β-AR agonist challenge via acute isoproterenol (ISO) infusion demonstrated a greater increase in ejection fraction, dp/dt, and contractility index. Interestingly, prior to ISO infusion, there were significant differences in baseline hemodynamics between the CXCR4 cKO compared to littermate controls. However, upon administering ISO, the CXCR4 cKO responded in a robust manner overcoming the baseline hemodynamic deficits reaching WT values supporting our previous data that CXCR4 negatively regulates β-AR signaling. This further supports that, in the absence of the physiologic negative modulation, there is an overactivation of down-stream pathways, which contribute to the development and progression of contractile dysfunction. Our results demonstrated that CXCR4 plays a non-developmental role in regulating cardiac function and that CXCR4 cKO mice develop a progressive cardiomyopathy leading to clinical heart failure.
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http://dx.doi.org/10.3390/ijms20092267DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6539363PMC
May 2019

Child Health Promotion in Underserved Communities: The FAMILIA Trial.

J Am Coll Cardiol 2019 04;73(16):2011-2021

Zena and Michael A. Wiener Cardiovascular Institute, Icahn School of Medicine at Mount Sinai, New York, New York; Centro Nacional de Investigaciones Cardiovasculares, Madrid, Spain. Electronic address:

Background: Preschool-based interventions offer promise to instill healthy behaviors in children, which can be a strategy to reduce the burden of cardiovascular disease later. However, their efficacy in underserved communities is not well established.

Objectives: The purpose of this study was to assess the impact of a preschool-based health promotion educational intervention in an underserved community.

Methods: This cluster-randomized controlled study involved 15 Head Start preschools in Harlem, New York. Schools and their children were randomized 3:2 to receive either a 4-month (50 h) educational intervention to instill healthy behaviors in relation to diet, physical activity, body/heart awareness, and emotion management; or their standard curriculum (control). The primary outcome was the change from baseline in the overall knowledge, attitudes, and habits (KAH) score of the children at 5 months. As secondary outcomes, we evaluated the changes in KAH subcomponents and emotion comprehension. Linear mixed-effects models were used to test for intervention effects.

Results: The authors enrolled 562 preschool children age 3 to 5 years, 51% female, 54% Hispanic/Latino, and 37% African-American. Compared with the control group, the mean relative change from baseline in the overall KAH score was ∼2.2 fold higher in the intervention group (average absolute difference of 2.86 points; 95% confidence interval: 0.58 to 5.14; p = 0.014). The maximal effect was observed in children who received >75% of the curriculum. Physical activity and body/heart awareness components, and knowledge and attitudes domains, were the main drivers of the effect (p values <0.05). Changes in emotion comprehension trended toward favoring intervened children.

Conclusions: This multidimensional school-based educational intervention may be an effective strategy for establishing healthy behaviors among preschoolers from a diverse and socioeconomically disadvantaged community. Early primordial prevention strategies may contribute to reducing the global burden of cardiovascular disease. (Family-Based Approach in a Minority Community Integrating Systems-Biology for Promotion of Health [FAMILIA]; NCT02343341).
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http://dx.doi.org/10.1016/j.jacc.2019.01.057DOI Listing
April 2019

MicroRNA-25 upregulation protects spinal cords, yet is bad for the heart: The dark side of noncoding RNAS.

J Thorac Cardiovasc Surg 2019 09 16;158(3):e87-e88. Epub 2019 Apr 16.

Department of Cardiology, Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, New York, NY.

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http://dx.doi.org/10.1016/j.jtcvs.2019.03.054DOI Listing
September 2019

Empagliflozin Ameliorates Adverse Left Ventricular Remodeling in Nondiabetic Heart Failure by Enhancing Myocardial Energetics.

J Am Coll Cardiol 2019 04;73(15):1931-1944

AtheroThrombosis Research Unit, Cardiovascular Institute, Icahn School of Medicine at Mount Sinai, New York, New York. Electronic address:

Background: Empagliflozin cardiac benefits in the EMPA-REG OUTCOME (Empagliflozin Cardiovascular Outcome Event Trial in Type 2 Diabetes Mellitus Patients) trial cannot be explained exclusively by its antihyperglycemic activity.

Objectives: The hypothesis was that empagliflozin's cardiac benefits are mediated by switching myocardial fuel metabolism away from glucose toward ketone bodies (KB), which improves myocardial energy production.

Methods: Heart failure was induced in nondiabetic pigs (n = 14) by 2-h balloon occlusion of the proximal left anterior descending artery. Animals were randomized to empagliflozin or placebo for 2 months. Animals were evaluated with cardiac magnetic resonance imaging and 3-dimensional echocardiography. Myocardial metabolite consumption was analyzed by simultaneous blood sampling from coronary artery and coronary sinus. Myocardial samples were obtained for molecular evaluation. Nonmyocardial infarction animals served as comparison.

Results: Despite similar initial ischemic myocardial injury in both groups, the empagliflozin group showed amelioration of adverse remodeling at 2 months (lower left ventricular [LV] mass, reduced LV dilatation, less LV sphericity) versus the control group. LV systolic function (LV ejection fraction and echocardiography-derived strains) was improved, as was neurohormonal activation. Compared with nonmyocardial infarction, control animals increased myocardial glucose consumption mainly through anaerobic glycolysis while reducing utilization of free fatty acid (FFA) and branched-chain amino acid (BCAA). Empagliflozin-treated pigs did not consume glucose (reduction in myocardial glucose uptake, and glucose-related enzymes) but instead switched toward utilization of KB, FFA, and BCAA (increased myocardial uptake of these 3 metabolites, and enhanced expression/activity of the enzymes implicated in the metabolism of KB/FFA/BCAA). Empagliflozin increased myocardial ATP content and enhanced myocardial work efficiency.

Conclusions: Empagliflozin ameliorates adverse cardiac remodeling and heart failure in a nondiabetic porcine model. Empagliflozin switches myocardial fuel utilization away from glucose toward KB, FFA, and BCAA, thereby improving myocardial energetics, enhancing LV systolic function, and ameliorating adverse LV remodeling.
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http://dx.doi.org/10.1016/j.jacc.2019.01.056DOI Listing
April 2019

The Left Pneumonectomy Combined with Monocrotaline or Sugen as a Model of Pulmonary Hypertension in Rats.

J Vis Exp 2019 03 8(145). Epub 2019 Mar 8.

Cardiovascular Research Center, Department of Cardiology, Icahn School of Medicine at Mount Sinai;

In this protocol, we detail the correct procedural steps and necessary precautions to successfully perform a left pneumonectomy and induce PAH in rats with the additional administration of monocrotaline (MCT) or SU5416 (Sugen). We also compare these two models to other PAH models commonly used in research. In the last few years, the focus of animal PAH models has moved towards studying the mechanism of angioproliferation of plexiform lesions, in which the role of increased pulmonary blood flow is considered as an important trigger in the development of severe pulmonary vascular remodeling. One of the most promising rodent models of increased pulmonary flow is the unilateral left pneumonectomy combined with a "second hit" of MCT or Sugen. The removal of the left lung leads to increased and turbulent pulmonary blood flow and vascular remodeling. Currently, there is no detailed procedure of the pneumonectomy surgery in rats. This article details a step-by-step protocol of the pneumonectomy surgical procedure and post-operative care in male Sprague-Dawley rats. Briefly, the animal is anesthetized and the chest is opened. Once the left pulmonary artery, pulmonary vein, and bronchus are visualized, they are ligated and the left lung is removed. The chest then closed and the animal recovered. Blood is forced to circulate only on the right lung. This increased vascular pressure leads to a progressive remodeling and occlusion of small pulmonary arteries. The second hit of MCT or Sugen is used one week post-surgery to induce endothelial dysfunction. The combination of increased blood flow in the lung and endothelial dysfunction produces severe PAH. The primary limitation of this procedure is that it requires general surgical skills.
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http://dx.doi.org/10.3791/59050DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7992111PMC
March 2019

Surgical and physiological challenges in the development of left and right heart failure in rat models.

Heart Fail Rev 2019 09;24(5):759-777

Cardiovascular Research Center, Department of Cardiology, Icahn School of Medicine at Mount Sinai, 1470 Madison Ave., Box 1030, New York, NY, 10029-6574, USA.

Rodent surgical animal models of heart failure (HF) are critically important for understanding the proof of principle of the cellular alterations underlying the development of the disease as well as evaluating therapeutics. Robust, reproducible rodent models are a prerequisite to the development of pharmacological and molecular strategies for the treatment of HF in patients. Due to the absence of standardized guidelines regarding surgical technique and clear criteria for HF progression in rats, objectivity is compromised. Scientific publications in rats rarely fully disclose the actual surgical details, and technical and physiological challenges. This lack of reporting is one of the main reasons that the outcomes specified in similar studies are highly variable and associated with unnecessary loss of animals, compromising scientific assessment. This review details rat circulatory and coronary arteries anatomy, the surgical details of rat models that recreate the HF phenotype of myocardial infarction, ischemia/reperfusion, left and right ventricular pressure, and volume overload states, and summarizes the technical and physiological challenges of creating HF. The purpose of this article is to help investigators understand the underlying issues of current HF models in order to reduce variable results and ensure successful, reproducible models of HF.
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http://dx.doi.org/10.1007/s10741-019-09783-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6698228PMC
September 2019

Role of SIRT1 in Modulating Acetylation of the Sarco-Endoplasmic Reticulum Ca-ATPase in Heart Failure.

Circ Res 2019 04;124(9):e63-e80

From the Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, New York (P.A.G., D.J., A.L., P.L., J.G.O., V.C., R.J.H., C.K.).

Rationale: SERCA2a, sarco-endoplasmic reticulum Ca-ATPase, is a critical determinant of cardiac function. Reduced level and activity of SERCA2a are major features of heart failure. Accordingly, intensive efforts have been made to develop efficient modalities for SERCA2a activation. We showed that the activity of SERCA2a is enhanced by post-translational modification with SUMO1 (small ubiquitin-like modifier 1). However, the roles of other post-translational modifications on SERCA2a are still unknown.

Objective: In this study, we aim to assess the role of lysine acetylation on SERCA2a function and determine whether inhibition of lysine acetylation can improve cardiac function in the setting of heart failure.

Methods And Results: The acetylation of SERCA2a was significantly increased in failing hearts of humans, mice, and pigs, which is associated with the reduced level of SIRT1 (sirtuin 1), a class III histone deacetylase. Downregulation of SIRT1 increased the SERCA2a acetylation, which in turn led to SERCA2a dysfunction and cardiac defects at baseline. In contrast, pharmacological activation of SIRT1 reduced the SERCA2a acetylation, which was accompanied by recovery of SERCA2a function and cardiac defects in failing hearts. Lysine 492 (K492) was of critical importance for the regulation of SERCA2a activity via acetylation. Acetylation at K492 significantly reduced the SERCA2a activity, presumably through interfering with the binding of ATP to SERCA2a. In failing hearts, acetylation at K492 appeared to be mediated by p300 (histone acetyltransferase p300), a histone acetyltransferase.

Conclusions: These results indicate that acetylation/deacetylation at K492, which is regulated by SIRT1 and p300, is critical for the regulation of SERCA2a activity in hearts. Pharmacological activation of SIRT1 can restore SERCA2a activity through deacetylation at K492. These findings might provide a novel strategy for the treatment of heart failure.
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http://dx.doi.org/10.1161/CIRCRESAHA.118.313865DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6483854PMC
April 2019

Targeted Gene Delivery through the Respiratory System: Rationale for Intratracheal Gene Transfer.

J Cardiovasc Dev Dis 2019 Feb 15;6(1). Epub 2019 Feb 15.

Cardiovascular Research Center, Department of Cardiology, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA.

Advances in DNA- and RNA-based technologies have made gene therapy suitable for many lung diseases, especially those that are hereditary. The main objective of gene therapy is to deliver an adequate amount of gene construct to the intended target cell, achieve stable transduction in target cells, and to produce a clinically therapeutic effect. This review focuses on the cellular organization in the normal lung and how gene therapy targets the specific cell types that are affected by pulmonary disorders caused by genetic mutations. Furthermore, it examines the pulmonary barriers that can compromise the absorption and transduction of viral vectors and genetic agents by the lung. Finally, it discusses the advantages and limitations of direct intra-tracheal gene delivery with different viral vectors in small and large animal models and in clinical trials.
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http://dx.doi.org/10.3390/jcdd6010008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6462990PMC
February 2019

Role of the PRC2-Six1-miR-25 signaling axis in heart failure.

J Mol Cell Cardiol 2019 04 13;129:58-68. Epub 2019 Feb 13.

Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, New York, NY, USA. Electronic address:

The reduced expression of cardiac sarco-endoplasmic reticulum Ca ATPase (SERCA2a) is a hallmark of heart failure. We previously showed that miR-25 is a crucial transcriptional regulator of SERCA2a in the heart. However, the precise mechanism of cardiac miR-25 regulation is largely unknown. Literatures suggested that miR-25 is regulated by the transcriptional co-factor, sine oculis homeobox homolog 1 (Six1), which in turn is epigenetically regulated by polycomb repressive complex 2 (PRC 2) in cardiac progenitor cells. Therefore, we aimed to investigate whether Six1 and PRC2 are indeed involved in the regulation of the miR-25 level in the setting of heart failure. Six1 was up-regulated in the failing hearts of humans and mice. Overexpression of Six1 led to adverse cardiac remodeling, whereas knock-down of Six1 attenuated pressure overload-induced cardiac dysfunction. The adverse effects of Six1 were ameliorated by knock-down of miR-25. The epigenetic repression on the Six1 promoter by PRC2 was significantly reduced in failing hearts. Epigenetic repression of Six1 is relieved through a reduction of PRC2 activity in heart failure. Six1 up-regulates miR-25, which is followed by reduction of cardiac SERCA2a expression. Collectively, these data showed that the PRC2-Six1-miR-25 signaling axis is involved in heart failure. Our finding introduces new insight into potential treatments of heart failure.
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http://dx.doi.org/10.1016/j.yjmcc.2019.01.017DOI Listing
April 2019

Mitochondrial Morphology, Dynamics, and Function in Human Pressure Overload or Ischemic Heart Disease With Preserved or Reduced Ejection Fraction.

Circ Heart Fail 2019 02;12(2):e005131

Department of Cardiovascular Diseases, (A.H.C., M.M.R.), Mayo Clinic, Rochester, MN.

Background: The FOXO3a (forkhead box O3a)-BNIP3 (B-cell lymphoma 2/adenovirus E1B 19kDa interacting protein 3) pathway modulates mitochondrial dynamics and function and contributes to myocardial remodeling in rodent models of heart failure. We sought to investigate the expression of this pathway along with the expression of mitochondrial biogenesis (PGC-1α [peroxisome proliferator-activated receptor-γ coactivator-1α]), dynamics (DRP-1 [dynamin-related protein 1], OPA-1 [optic atrophy 1], and MFN 2 [mitofusin 2]), and oxidative phosphorylation (citrate synthase and electron transport chain complexes) markers and COX IV (cytochrome C oxidase) activity in myocardium from patients with valvular or ischemic heart disease and heart failure with preserved ejection fraction (HFpEF) or heart failure with reduced ejection fraction (HFrEF).

Methods And Results: Subepicardial left ventricular biopsies (10×1×1 mm) were obtained at aortic valve replacement (HFpEF, n=5; and HFrEF, n=4), coronary artery bypass grafting (HFpEF, n=5; and HFrEF, n=5), or left ventricular assist device implantation (HFrEF, n=4). Subepicardial biopsies from patients with normal left ventricular function (n=2) and from donor hearts (n=3) served as controls (normal). Relative to normal, mitochondrial fragmentation and cristae destruction were evident, and mitochondrial area was decreased in HFpEF; 1.00±0.09 versus 0.71±0.08; P=0.016. These mitochondrial morphological changes were more pronounced in HFrEF (0.54±0.06); P=0.002 HFpEF versus HFrEF. BNIP3 (monomer+dimer) expression was increased in HFpEF (3.99±2.44) and in HFrEF (5.19±1.70) relative to normal; P=0.004 and P<0.001, respectively. However, BNIP3 monomer was increased in HFrEF (4.32±1.43) compared with normal (0.99±0.06) and HFpEF (1.97±0.90); P=0.001 and 0.004, respectively. The HFrEF group uniquely showed increase in DRP-1 expression (1.94±0.38) and decreases in PGC-1α expression (0.61±0.07) and COX IV activity (0.70±0.10) relative to normal; P=0.013, P<0.001, and P<0.001, respectively, with no significant change in electron transport chain complexes expression.

Conclusions: These findings in human myocardium confirm studies in rodents where contractile dysfunction is associated with activation of the FOXO3a-BNIP3 pathway and altered mitochondrial dynamics, biogenesis, and function.
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http://dx.doi.org/10.1161/CIRCHEARTFAILURE.118.005131DOI Listing
February 2019

Lentiviral-Mediated Interleukin-10 Gene Therapy for Lung Transplantation.

J Thorac Cardiovasc Surg 2019 02;157(2):817-818

Department of Cardiology, Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, New York, NY.

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http://dx.doi.org/10.1016/j.jtcvs.2018.07.052DOI Listing
February 2019
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