Systemic Blockade of ACVR2B Ligands Protects Myocardium from Acute Ischemia-Reperfusion Injury

Activin A and myostatin, members of the transforming growth factor (TGF)-b superfamily of secreted factors, are potent negative regulators of muscle growth, but their contribution to myocardial ischemia-reperfusion (IR) injury is not known. The aim of this study was to investigate if activin 2B (ACVR2B) receptor ligands contribute to myocardial IR injury. Mice were treated with soluble ACVR2B decoy receptor (ACVR2B-Fc) and subjected to myocardial ischemia followed by reperfusion for 6 or 24 h. Systemic blockade of ACVR2B ligands by ACVR2B-Fc was protective against cardiac IR injury, as evidenced by reduced infarcted area, apoptosis, and autophagy and better preserved LV systolic function following IR. ACVR2B-Fc modified cardiac metabolism, LV mitochondrial respiration, as well as cardiac phenotype toward physiological hypertrophy. Similar to its protective role in IR injury in vivo, ACVR2B-Fc antagonized SMAD2 signaling and cell death in cardiomyocytes that were subjected to hypoxic stress. ACVR2B ligand myostatin was found to exacerbate hypoxic stress. In addition to acute cardioprotection in ischemia, ACVR2B-Fc provided beneficial effects on cardiac function in prolonged cardiac stress in cardiotoxicity model. By blocking myostatin, ACVR2B-Fc potentially reduces cardiomyocyte death and modifies cardiomyocyte metabolism for hypoxic conditions to protect the heart from IR injury.Peer reviewe

MiR-185-5p regulates the development of myocardial fibrosis

Background: Cardiac fibrosis stiffens the ventricular wall, predisposes to cardiac arrhythmias and contributes to the development of heart failure. In the present study, our aim was to identify novel miRNAs that regulate the development of cardiac fibrosis and could serve as potential therapeutic targets for myocardial fibrosis. Methods and results: Analysis for cardiac samples from sudden cardiac death victims with extensive myocardial fibrosis as the primary cause of death identified dysregulation of miR-185-5p. Analysis of resident cardiac cells from mice subjected to experimental cardiac fibrosis model showed induction of miR-185-5p expression specifically in cardiac fibroblasts. In vitro, augmenting miR-185-5p induced collagen production and profibrotic activation in cardiac fibroblasts, whereas inhibition of miR-185-5p attenuated collagen production. In vivo, targeting miR-185-5p in mice abolished pressure overload induced cardiac interstitial fibrosis. Mechanistically, miR-185-5p targets apelin receptor and inhibits the anti-fibrotic effects of apelin. Finally, analysis of left ventricular tissue from patients with severe cardiomyopathy showed an increase in miR-185-5p expression together with pro-fibrotic TGF-beta 1 and collagen I. Conclusions: Our data show that miR-185-5p targets apelin receptor and promotes myocardial fibrosis.Peer reviewe

miR-1468-3p promotes aging–related cardiac fibrosis

Abstract Non-coding microRNAs (miRNAs) are powerful regulators of gene expression and critically involved in cardiovascular pathophysiology. The aim of the current study was to identify miRNAs regulating cardiac fibrosis. Cardiac samples of age-matched control subjects and sudden cardiac death (SCD) victims with primary myocardial fibrosis (PMF) were subjected to miRNA profiling. Old SCD victims with PMF and healthy aged human hearts showed increased expression of miR-1468-3p. In vitro studies in human cardiac fibroblasts showed that augmenting miR-1468-3p levels induces collagen deposition and cell metabolic activity and enhances collagen 1, connective tissue growth factor, and periostin expression. In addition, miR-1468-3p promotes cellular senescence with increased senescence-associated β-galactosidase activity and increased expression of p53 and p16. AntimiR-1468-3p antagonized transforming growth factor β1 (TGF-β1)-induced collagen deposition and metabolic activity. Mechanistically, mimic-1468-3p enhanced p38 phosphorylation, while antimiR-1468-3p decreased TGF-β1-induced p38 activation and abolished p38-induced collagen deposition. RNA sequencing analysis, a computational prediction model, and qPCR analysis identified dual-specificity phosphatases (DUSPs) as miR-1468-3p target genes, and regulation of DUSP1 by miR-1468-3p was confirmed with a dual-luciferase reporter assay. In conclusion, miR-1468-3p promotes cardiac fibrosis by enhancing TGF-β1-p38 signaling. Targeting miR-1468-3p in the older population may be of therapeutic interest to reduce cardiac fibrosis

Wnt11 in regulation of physiological and pathological cardiac growth

Abstract Wnt11 regulates early cardiac development and left ventricular compaction in the heart, but it is not known how Wnt11 regulates postnatal cardiac maturation and response to cardiac stress in the adult heart. We studied cell proliferation/maturation in postnatal and adolescent Wnt11 deficient (Wnt11−/−) heart and subjected adult mice with partial (Wnt11+/−) and complete Wnt11 (Wnt11−/−) deficiency to cardiac pressure overload. In addition, we subjected primary cardiomyocytes to recombinant Wnt proteins to study their effect on cardiomyocyte growth. Wnt11 deficiency did not affect cardiomyocyte proliferation or maturation in the postnatal or adolescent heart. However, Wnt11 deficiency led to enlarged heart phenotype that was not accompanied by significant hypertrophy of individual cardiomyocytes. Analysis of stressed adult hearts from wild-type mice showed a progressive decrease in Wnt11 expression in response to pressure overload. When studied in experimental cardiac pressure overload, Wnt11 deficiency did not exacerbate cardiac hypertrophy or remodeling and cardiac function remained identical between the genotypes. When subjecting cardiomyocytes to hypertrophic stimulus, the presence of recombinant Wnt11 together with Wnt5a reduced protein synthesis. In conclusion, Wnt11 deficiency does not affect postnatal cardiomyocyte proliferation but leads to cardiac growth. Interestingly, Wnt11 deficiency alone does not substantially modulate hypertrophic response to pressure overload in vivo. Wnt11 may require cooperation with other noncanonical Wnt proteins to regulate hypertrophic response under stress

Systemic blockade of ACVR2B ligands attenuates muscle wasting in ischemic heart failure without compromising cardiac function

Abstract Signaling through activin receptors regulates skeletal muscle mass and activin receptor 2B (ACVR2B) ligands are also suggested to participate in myocardial infarction (MI) pathology in the heart. In this study, we determined the effect of systemic blockade of ACVR2B ligands on cardiac function in experimental MI, and defined its efficacy to revert muscle wasting in ischemic heart failure (HF). Mice were treated with soluble ACVR2B decoy receptor (ACVR2B‐Fc) to study its effect on post‐MI cardiac remodeling and on later HF. Cardiac function was determined with echocardiography, and myocardium analyzed with histological and biochemical methods for hypertrophy and fibrosis. Pharmacological blockade of ACVR2B ligands did not rescue the heart from ischemic injury or alleviate post‐MI remodeling and ischemic HF. Collectively, ACVR2B‐Fc did not affect cardiomyocyte hypertrophy, fibrosis, angiogenesis, nor factors associated with cardiac regeneration except modification of certain genes involved in metabolism or cell growth/survival. ACVR2B‐Fc, however, was able to reduce skeletal muscle wasting in chronic ischemic HF, accompanied by reduced LC3II as a marker of autophagy and increased mTOR signaling and Cited4 expression as markers of physiological hypertrophy in quadriceps muscle. Our results ascertain pharmacological blockade of ACVR2B ligands as a possible therapy for skeletal muscle wasting in ischemic HF. Pharmacological blockade of ACVR2B ligands preserved myofiber size in ischemic HF, but did not compromise cardiac function nor exacerbate cardiac remodeling after ischemic injury

Activation of the hypoxia response pathway protects against age-induced cardiac hypertrophy

Abstract Aims: We have previously demonstrated protection against obesity, metabolic dysfunction, atherosclerosis and cardiac ischemia in a hypoxia-inducible factor (HIF) prolyl 4-hydroxylase-2 (Hif-p4h-2) deficient mouse line, attributing these protective effects to activation of the hypoxia response pathway in a normoxic environment. We intended here to find out whether the Hif-p4h-2 deficiency affects the cardiac health of these mice upon aging. Methods and results: When the Hif-p4h-2 deficient mice and their wild-type littermates were monitored during normal aging, the Hif-p4h-2 deficient mice had better preserved diastolic function than the wild type at one year of age and less cardiomyocyte hypertrophy at two years. On the mRNA level, downregulation of hypertrophy-associated genes was detected and shown to be associated with upregulation of Notch signaling, and especially of the Notch target gene and transcriptional repressor Hairy and enhancer-of-split-related basic helix-loop-helix (Hey2). Blocking of Notch signaling in cardiomyocytes isolated from Hif-p4h-2 deficient mice with a gamma-secretase inhibitor led to upregulation of the hypertrophy-associated genes. Also, targeting Hey2 in isolated wild-type rat neonatal cardiomyocytes with siRNA led to upregulation of hypertrophic genes and increased leucine incorporation indicative of increased protein synthesis and hypertrophy. Finally, oral treatment of wild-type mice with a small molecule inhibitor of HIF-P4Hs phenocopied the effects of Hif-p4h-2 deficiency with less cardiomyocyte hypertrophy, upregulation of Hey2 and downregulation of the hypertrophy-associated genes. Conclusions: These results indicate that activation of the hypoxia response pathway upregulates Notch signaling and its target Hey2 resulting in transcriptional repression of hypertrophy-associated genes and less cardiomyocyte hypertrophy. This is eventually associated with better preserved cardiac function upon aging. Activation of the hypoxia response pathway thus has therapeutic potential for combating age-induced cardiac hypertrophy

MiR‐185‐5p regulates the development of myocardial fibrosis

Abstract Background: Cardiac fibrosis stiffens the ventricular wall, predisposes to cardiac arrhythmias and contributes to the development of heart failure. In the present study, our aim was to identify novel miRNAs that regulate the development of cardiac fibrosis and could serve as potential therapeutic targets for myocardial fibrosis. Methods and results: Analysis for cardiac samples from sudden cardiac death victims with extensive myocardial fibrosis as the primary cause of death identified dysregulation of miR‐185‐5p. Analysis of resident cardiac cells from mice subjected to experimental cardiac fibrosis model showed induction of miR‐185‐5p expression specifically in cardiac fibroblasts. In vitro, augmenting miR‐185‐5p induced collagen production and profibrotic activation in cardiac fibroblasts, whereas inhibition of miR‐185‐5p attenuated collagen production. In vivo, targeting miR‐185‐5p in mice abolished pressure overload induced cardiac interstitial fibrosis. Mechanistically, miR‐185‐5p targets apelin receptor and inhibits the anti-fibrotic effects of apelin. Finally, analysis of left ventricular tissue from patients with severe cardiomyopathy showed an increase in miR‐185‐5p expression together with pro-fibrotic TGF‐β1 and collagen I. Conclusions: Our data show that miR‐185‐5p targets apelin receptor and promotes myocardial fibrosis