Topic: Basic Research

Abstract

STING, a cytosolic DNA sensor, plays a critical role in atherogenesis: a link between innate immunity and chronic inflammation caused by lifestyle-related diseases.

Pham PT, Fukuda D, Nishimoto S, Kim-Kaneyama JR, ... Barber GN, Sata M
Aims 
Lifestyle-related diseases promote atherosclerosis, a chronic inflammatory disease; however, the molecular mechanism remains largely unknown. Endogenous DNA fragments released under over-nutrient condition provoke sterile inflammation through the recognition by DNA sensors. Here, we investigated the role of stimulator of interferon genes (STING), a cytosolic DNA sensor, in atherogenesis.
Methods and results 
Apolipoprotein E-deficient (Apoe-/-) mice fed a western-type diet (WTD), a hypercholesterolaemic mouse model, showed higher STING expression and markers for DNA damage such as γH2AX, p53, and single-stranded DNA (ssDNA) accumulation in macrophages in the aorta compared with wild-type (WT) mice. The level of cGAMP, a STING agonist, in the aorta was higher in Apoe-/- mice. Genetic deletion of Sting in Apoe-/- mice reduced atherosclerotic lesions in the aortic arch, lipid, and macrophage accumulation in plaques, and inflammatory molecule expression in the aorta compared with the control. Pharmacological blockade of STING using a specific inhibitor, C-176, ameliorated atherogenesis in Apoe-/- mice. In contrast, bone marrow-specific STING expression in Apoe-/- mice stimulated atherogenesis. Expression or deletion of STING did not affect metabolic parameters and blood pressure. In vitro studies revealed that STING activation by cGAMP or mitochondrial DNA accelerated inflammatory molecule expression (e.g. TNF-α or IFN-β) in mouse and human macrophages. Activation of nuclear factor-κB and TANK binding kinase 1 was involved in STING-associated vascular inflammation and macrophage activation. Furthermore, human atherosclerotic lesions in the carotid arteries expressed STING and cGAMP.
Conclusion 
Stimulator of interferon genes stimulates pro-inflammatory activation of macrophages, leading to the development of atherosclerosis. Stimulator of interferon genes signalling may serve as a potential therapeutic target for atherosclerosis.

Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2021. For permissions, please email: [email protected]

Eur Heart J: 05 Jul 2021; epub ahead of print
Pham PT, Fukuda D, Nishimoto S, Kim-Kaneyama JR, ... Barber GN, Sata M
Eur Heart J: 05 Jul 2021; epub ahead of print | PMID: 34226923
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Abstract

Regulation of the Methylation and Expression Levels of the BMPR2 Gene by SIN3a as a Novel Therapeutic Mechanism in Pulmonary Arterial Hypertension.

Bisserier M, Mathiyalagan P, Zhang S, Elmastour F, ... Sahoo S, Hadri L
Background
Epigenetic mechanisms are critical in the pathogenesis of pulmonary arterial hypertension (PAH). Previous studies have suggested that hypermethylation of the BMPR2 (bone morphogenetic protein receptor type 2) promoter is associated with BMPR2 downregulation and progression of PAH. Here, we investigated for the first time the role of SIN3a (switch-independent 3a), a transcriptional regulator, in the epigenetic mechanisms underlying hypermethylation of BMPR2 in the pathogenesis of PAH.
Methods
We used lung samples from PAH patients and non-PAH controls, preclinical mouse and rat PAH models, and human pulmonary arterial smooth muscle cells. Expression of SIN3a was modulated using a lentiviral vector or a siRNA in vitro and a specific adeno-associated virus serotype 1 or a lentivirus encoding for human SIN3a in vivo.
Results
SIN3a is a known transcriptional regulator; however, its role in cardiovascular diseases, especially PAH, is unknown. It is interesting that we detected a dysregulation of SIN3 expression in patients and in rodent models, which is strongly associated with decreased BMPR2 expression. SIN3a is known to regulate epigenetic changes. Therefore, we tested its role in the regulation of BMPR2 and found that BMPR2 is regulated by SIN3a. It is interesting that SIN3a overexpression inhibited human pulmonary arterial smooth muscle cells proliferation and upregulated BMPR2 expression by preventing the methylation of the BMPR2 promoter region. RNA-sequencing analysis suggested that SIN3a downregulated the expression of DNA and histone methyltransferases such as DNMT1 (DNA methyltransferase 1) and EZH2 (enhancer of zeste 2 polycomb repressive complex 2) while promoting the expression of the DNA demethylase TET1 (ten-eleven translocation methylcytosine dioxygenase 1). Mechanistically, SIN3a promoted BMPR2 expression by decreasing CTCF (CCCTC-binding factor) binding to the BMPR2 promoter. Last, we identified intratracheal delivery of adeno-associated virus serotype human SIN3a to be a beneficial therapeutic approach in PAH by attenuating pulmonary vascular and right ventricle remodeling, decreasing right ventricle systolic pressure and mean pulmonary arterial pressure, and restoring BMPR2 expression in rodent models of PAH.
Conclusions
All together, our study unveiled the protective and beneficial role of SIN3a in pulmonary hypertension. We also identified a novel and distinct molecular mechanism by which SIN3a regulates BMPR2 in human pulmonary arterial smooth muscle cells. Our study also identified lung-targeted SIN3a gene therapy using adeno-associated virus serotype 1 as a new promising therapeutic strategy for treating patients with PAH.



Circulation: 05 Jul 2021; 144:52-73
Bisserier M, Mathiyalagan P, Zhang S, Elmastour F, ... Sahoo S, Hadri L
Circulation: 05 Jul 2021; 144:52-73 | PMID: 34078089
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Abstract

Cooperative Binding of ETS2 and NFAT Links Erk1/2 and Calcineurin Signaling in the Pathogenesis of Cardiac Hypertrophy.

Luo Y, Jiang N, May HI, Luo X, ... Gillette TG, Hill JA
Background
Cardiac hypertrophy is an independent risk factor for heart failure, a leading cause of morbidity and mortality globally. The calcineurin/NFAT (nuclear factor of activated T cells) pathway and the MAPK (mitogen-activated protein kinase)/Erk (extracellular signal-regulated kinase) pathway contribute to the pathogenesis of cardiac hypertrophy as an interdependent network of signaling cascades. How these pathways interact remains unclear and few direct targets responsible for the prohypertrophic role of NFAT have been described.
Methods
By engineering cardiomyocyte-specific ETS2 (a member of the E26 transformation-specific sequence [ETS] domain family) knockout mice, we investigated the role of ETS2 in cardiac hypertrophy. Primary cardiomyocytes were used to evaluate ETS2 function in cell growth.
Results
ETS2 is phosphorylated and activated by Erk1/2 on hypertrophic stimulation in both mouse (n=3) and human heart samples (n=8 to 19). Conditional deletion of ETS2 in mouse cardiomyocytes protects against pressure overload-induced cardiac hypertrophy (n=6 to 11). Silencing of ETS2 in the hearts of calcineurin transgenic mice significantly attenuates hypertrophic growth and contractile dysfunction (n=8). As a transcription factor, ETS2 is capable of binding to the promoters of hypertrophic marker genes, such as ANP, BNP, and Rcan1.4 (n=4). We report that ETS2 forms a complex with NFAT to stimulate transcriptional activity through increased NFAT binding to the promoters of at least 2 hypertrophy-stimulated genes: Rcan1.4 and microRNA-223 (=n4 to 6). Suppression of microRNA-223 in cardiomyocytes inhibits calcineurin-mediated cardiac hypertrophy (n=6), revealing microRNA-223 as a novel prohypertrophic target of the calcineurin/NFAT and Erk1/2-ETS2 pathways.
Conclusions
Our findings point to a critical role for ETS2 in calcineurin/NFAT pathway-driven cardiac hypertrophy and unveil a previously unknown molecular connection between the Erk1/2 activation of ETS2 and expression of NFAT/ETS2 target genes.



Circulation: 05 Jul 2021; 144:34-51
Luo Y, Jiang N, May HI, Luo X, ... Gillette TG, Hill JA
Circulation: 05 Jul 2021; 144:34-51 | PMID: 33821668
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Abstract

Fibroblast-Specific Proteotranscriptomes Reveal Distinct Fibrotic Signatures of Human Sinoatrial Node in Nonfailing and Failing Hearts.

Kalyanasundaram A, Li N, Gardner ML, Artiga EJ, ... Mohler PJ, Fedorov VV
Background
Up to 50% of the adult human sinoatrial node (SAN) is composed of dense connective tissue. Cardiac diseases including heart failure (HF) may increase fibrosis within the SAN pacemaker complex, leading to impaired automaticity and conduction of electric activity to the atria. Unlike the role of cardiac fibroblasts in pathologic fibrotic remodeling and tissue repair, nothing is known about fibroblasts that maintain the inherently fibrotic SAN environment.
Methods
Intact SAN pacemaker complex was dissected from cardioplegically arrested explanted nonfailing hearts (non-HF; n=22; 48.7±3.1 years of age) and human failing hearts (n=16; 54.9±2.6 years of age). Connective tissue content was quantified from Masson trichrome-stained head-center and center-tail SAN sections. Expression of extracellular matrix proteins, including collagens 1 and 3A1, CILP1 (cartilage intermediate layer protein 1), and POSTN (periostin), and fibroblast and myofibroblast numbers were quantified by in situ and in vitro immunolabeling. Fibroblasts from the central intramural SAN pacemaker compartment (≈10×5×2 mm3) and right atria were isolated, cultured, passaged once, and treated ± transforming growth factor β1 and subjected to comprehensive high-throughput next-generation sequencing of whole transcriptome, microRNA, and proteomic analyses.
Results
Intranodal fibrotic content was significantly higher in SAN pacemaker complex from HF versus non-HF hearts (57.7±2.6% versus 44.0±1.2%; P<0.0001). Proliferating phosphorylated histone 3+/vimentin+/CD31- (cluster of differentiation 31) fibroblasts were higher in HF SAN. Vimentin+/α-smooth muscle actin+/CD31- myofibroblasts along with increased interstitial POSTN expression were found only in HF SAN. RNA sequencing and proteomic analyses identified unique differences in mRNA, long noncoding RNA, microRNA, and proteomic profiles between non-HF and HF SAN and right atria fibroblasts and transforming growth factor β1-induced myofibroblasts. Specifically, proteins and signaling pathways associated with extracellular matrix flexibility, stiffness, focal adhesion, and metabolism were altered in HF SAN fibroblasts compared with non-HF SAN.
Conclusions
This study revealed increased SAN-specific fibrosis with presence of myofibroblasts, CILP1, and POSTN-positive interstitial fibrosis only in HF versus non-HF human hearts. Comprehensive proteotranscriptomic profiles of SAN fibroblasts identified upregulation of genes and proteins promoting stiffer SAN extracellular matrix in HF hearts. Fibroblast-specific profiles generated by our proteotranscriptomic analyses of the human SAN provide a comprehensive framework for future studies to investigate the role of SAN-specific fibrosis in cardiac rhythm regulation and arrhythmias.



Circulation: 12 Jul 2021; 144:126-143
Kalyanasundaram A, Li N, Gardner ML, Artiga EJ, ... Mohler PJ, Fedorov VV
Circulation: 12 Jul 2021; 144:126-143 | PMID: 33874740
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Abstract

Salt Transiently Inhibits Mitochondrial Energetics in Mononuclear Phagocytes.

Geisberger S, Bartolomaeus H, Neubert P, Willebrand R, ... Kempa S, Müller DN
Background
Dietary high salt (HS) is a leading risk factor for mortality and morbidity. Serum sodium transiently increases postprandially but can also accumulate at sites of inflammation affecting differentiation and function of innate and adaptive immune cells. Here, we focus on how changes in extracellular sodium, mimicking alterations in the circulation and tissues, affect the early metabolic, transcriptional, and functional adaption of human and murine mononuclear phagocytes.
Methods
Using Seahorse technology, pulsed stable isotope-resolved metabolomics, and enzyme activity assays, we characterize the central carbon metabolism and mitochondrial function of human and murine mononuclear phagocytes under HS in vitro. HS as well as pharmacological uncoupling of the electron transport chain under normal salt is used to analyze mitochondrial function on immune cell activation and function (as determined by Escherichia coli killing and CD4+ T cell migration capacity). In 2 independent clinical studies, we analyze the effect of a HS diet during 2 weeks (URL: http://www.clinicaltrials.gov. Unique identifier: NCT02509962) and short-term salt challenge by a single meal (URL: http://www.clinicaltrials.gov. Unique identifier: NCT04175249) on mitochondrial function of human monocytes in vivo.
Results
Extracellular sodium was taken up into the intracellular compartment, followed by the inhibition of mitochondrial respiration in murine and human macrophages. Mechanistically, HS reduces mitochondrial membrane potential, electron transport chain complex II activity, oxygen consumption, and ATP production independently of the polarization status of macrophages. Subsequently, cell activation is altered with improved bactericidal function in HS-treated M1-like macrophages and diminished CD4+ T cell migration in HS-treated M2-like macrophages. Pharmacological uncoupling of the electron transport chain under normal salt phenocopies HS-induced transcriptional changes and bactericidal function of human and murine mononuclear phagocytes. Clinically, also in vivo, rise in plasma sodium concentration within the physiological range reversibly reduces mitochondrial function in human monocytes. In both a 14-day and single meal HS challenge, healthy volunteers displayed a plasma sodium increase of [Formula: see text] and [Formula: see text] respectively, that correlated with decreased monocytic mitochondrial oxygen consumption.
Conclusions
Our data identify the disturbance of mitochondrial respiration as the initial step by which HS mechanistically influences immune cell function. Although these functional changes might help to resolve bacterial infections, a shift toward proinflammation could accelerate inflammatory cardiovascular disease.



Circulation: 12 Jul 2021; 144:144-158
Geisberger S, Bartolomaeus H, Neubert P, Willebrand R, ... Kempa S, Müller DN
Circulation: 12 Jul 2021; 144:144-158 | PMID: 33906377
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Abstract

Hyperglycaemia Induces Trained Immunity in Macrophages and Their Precursors and Promotes Atherosclerosis.

Edgar L, Akbar N, Braithwaite AT, Krausgruber T, ... Carnicer R, Choudhury RP
Background: Cardiovascular risk in diabetes remains elevated despite glucose lowering therapies. We hypothesised that hyperglycaemia induces trained immunity in macrophages, promoting persistent pro-atherogenic characteristics.
Methods:
Bone marrow derived macrophages from control and mice with diabetes were grown in physiological glucose (5 mM) and subject to RNA-sequencing (n=6), ATAC-sequencing (n=6) and ChIP-sequencing (n=6) for determination of hyperglycaemia-induced trained immunity. Bone marrow transplantation from mice with (n=9) or without (n=6) diabetes into [normoglycaemic] Ldlr -/- mice was used to assess its functional significance in vivo. Evidence of hyperglycaemia-induced trained immunity was sought in human peripheral blood mononuclear cells (PBMCs) from patients with diabetes (n=8) compared with case controls (n=16) and in human atherosclerotic plaque macrophages excised by laser capture microdissection.
Results:
In macrophages, high extracellular glucose promoted pro-inflammatory gene expression and pro-atherogenic functional characteristics, through glycolysis-dependent mechanisms. Bone marrow-derived macrophages (BMDM) from diabetic mice, retained these characteristics, even when cultured in physiological glucose, indicating hyperglycaemia-induced trained immunity. Bone marrow transplantation from diabetic mice into [normoglycaemic] Ldlr -/- mice increased aortic root atherosclerosis, confirming a disease-relevant and persistent form of trained innate immunity. Integrated ATAC-seq, ChIP-seq and RNA-seq analyses of haematopoietic stem cells and BMDM revealed a pro-inflammatory \"priming effect\" in diabetes. The pattern of open chromatin implicated transcription factor, RUNX1, while transcriptomes of atherosclerotic plaque macrophages and peripheral leukocytes in patients with type 2 diabetes were enriched for RUNX1 targets, consistent with a potential role in human disease. Pharmacological inhibition of RUNX1 in vitro inhibited the trained phenotype. Conclusions: Hyperglycaemia-induced trained immunity may explain why targeting elevated glucose is ineffective in reducing macrovascular risk in diabetes and suggests new targets for disease prevention and therapy.




Circulation: 12 Jul 2021; epub ahead of print
Edgar L, Akbar N, Braithwaite AT, Krausgruber T, ... Carnicer R, Choudhury RP
Circulation: 12 Jul 2021; epub ahead of print | PMID: 34255973
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Abstract

Epigenetic Regulation by Suv4-20h1 in Cardiopulmonary Progenitor Cells is Required to Prevent Pulmonary Hypertension and COPD.

Qi H, Liu H, Pullamsetti SS, Günther S, ... Yuan X, Braun T
Background: The etiology of life-threatening cardiopulmonary diseases such as Pulmonary Hypertension (PH) and Chronic Obstructive Pulmonary Disease (COPD) originates from a complex interplay of environmental factors and genetic predispositions, which is not fully understood. Likewise, little is known about developmental abnormalities or epigenetic dysregulations that might predispose for PH or COPD in adult individuals.
Methods:
To identify pathology-associated epigenetic alteration in diseased lung tissues, we screened a cohort of human PH and COPD patients for changes of histone modifications by immunofluorescence staining. To analyze the function of H4K20me2/3 in lung pathogenesis, we developed a series of Suv4-20h1 knockout mouse lines targeting cardiopulmonary progenitor cells (CPPs) and different heart and lung cell types, followed by hemodynamic studies and morphometric assessment of tissue samples. Molecular, cellular and biochemical techniques were applied to analyze the function of Suv4-20h1-dependent epigenetic processes in cardiopulmonary progenitor cells and their derivatives.
Results:
We discovered a strong reduction of the histone modifications H4K20me2/3 in human COPD but not PH patients, which depend on the activity of the H4K20 di-methyltransferase SUV4-20H1. Loss of Suv4-20h1 in CPPs caused a COPD-like/PH phenotype in mice including formation of perivascular tertiary lymphoid tissue and goblet cell hyperplasia, hyper-proliferation of smooth muscle cells/myofibroblasts, impaired alveolarization and maturation defects of the microvasculature leading to massive right ventricular dilatation and premature death. Mechanistically, SUV4-20H1 binds directly to the 5\'-upstream regulatory element of superoxide dismutase 3 (Sod3) gene to repress its expression. Increased levels of the extracellular SOD3 enzyme in Suv4-20h1 mutants increases hydrogen peroxide (H2O2) concentrations, causing vascular defects and impairing alveolarization. Conclusions: Our findings reveal a pivotal role of the histone modifier SUV4-20H1 in cardiopulmonary co-development and uncover developmental origins of cardiopulmonary diseases. We assume that the study will facilitate the understanding of pathogenic events causing PH and COPD, and aid the development of epigenetic drugs for treatment of cardiopulmonary diseases.




Circulation: 11 Jul 2021; epub ahead of print
Qi H, Liu H, Pullamsetti SS, Günther S, ... Yuan X, Braun T
Circulation: 11 Jul 2021; epub ahead of print | PMID: 34247492
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Abstract

MicroRNA-21 Controls Circadian Regulation of Apoptosis in Atherosclerotic Lesions.

Schober A, Blay RM, Saboor Maleki S, Zahedi F, ... Weber C, Nazari-Jahantigh M
Background: The necrotic core partly formed by ineffective efferocytosis increases the risk of an atherosclerotic plaque rupture. microRNAs contribute to necrotic core formation by regulating efferocytosis and macrophage apoptosis. Atherosclerotic plaque rupture occurs at increased frequency in the early morning, indicating diurnal changes in plaque vulnerability. Although circadian rhythms play a role in atherosclerosis, the molecular clock output pathways that control plaque composition and rupture susceptibility are unclear.
Methods:
Circadian gene expression, necrotic core size, and apoptosis and efferocytosis in aortic lesions were investigated at different times of the day in Apoe-/-Mir21+/+ mice and Apoe-/- Mir21-/- mice after consumption of a high-fat diet for 12 weeks feeding. Genome-wide gene expression and lesion formation were analyzed in bone marrow (BM)-transplanted mice. Diurnal changes in apoptosis and clock gene expression were determined in human atherosclerotic lesions.
Results:
The expression of molecular clock genes, lesional apoptosis, and necrotic core size were diurnally regulated in Apoe-/- mice. Efferocytosis did not match the diurnal increase in apoptosis at the beginning of the active phase. However, in parallel with apoptosis, expression levels of oscillating Mir21 strands decreased in the mouse atherosclerotic aorta. Mir21 knockout abolished circadian regulation of apoptosis and reduced necrotic core size, but did not affect core clock gene expression. Further, Mir21 knockout upregulated expression of pro-apoptotic XIAP associated factor 1 (Xaf1) in the atherosclerotic aorta, which abolished circadian expression of Xaf1. The anti-apoptotic effect of Mir21 was mediated by non-canonical targeting of Xaf1 through both Mir21 strands. Mir21 knockout in BM cells also reduced atherosclerosis and necrotic core size. Circadian regulation of clock gene expression was confirmed in human atherosclerotic lesions. Apoptosis oscillated diurnally in phase with XAF1 expression, demonstrating an early morning peak anti-phase to that of the Mir21 strands. Conclusions: Our findings suggest that the molecular clock in atherosclerotic lesions induces a diurnal rhythm of apoptosis regulated by circadian Mir21 expression in macrophages that is not matched by efferocytosis, thus increasing the size of the necrotic core.




Circulation: 07 Jul 2021; epub ahead of print
Schober A, Blay RM, Saboor Maleki S, Zahedi F, ... Weber C, Nazari-Jahantigh M
Circulation: 07 Jul 2021; epub ahead of print | PMID: 34233454
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Abstract

Red Blood Cell and Endothelial eNOS Independently Regulate Circulating Nitric Oxide Metabolites and Blood Pressure.

Leo F, Suvorava T, Heuser SK, Li J, ... Kelm M, Cortese-Krott MM
Background: Current paradigms suggest that nitric oxide (NO) produced by endothelial cells (ECs) via endothelial nitric oxide synthase (eNOS) in the vessel wall is the primary regulator of blood flow and blood pressure. However, red blood cells (RBCs) also carry a catalytically active eNOS, but its role is controversial and remains undefined. This study aimed to elucidate the functional significance of red cell eNOS compared to EC eNOS for vascular hemodynamics and NO metabolism.
Methods:
We generated tissue-specific \"loss-\" and \"gain-of-function\" models for eNOS by using cell-specific Cre-induced gene inactivation or reactivation. We created two founder lines carrying a floxed eNOS (eNOSflox/flox) for Cre-inducible knock out (KO), as well as gene construct with an inactivated floxed/inverted exon (eNOSinv/inv) for a Cre-inducible knock in (KI), which respectively allow targeted deletion or reactivation of eNOS in erythroid cells (RBC eNOS KO or RBC eNOS KI mice) or endothelial cells (EC eNOS KO or EC eNOS KI mice). Vascular function, hemodynamics, and NO metabolism were compared ex vivo and in vivo.
Results:
The EC eNOS KOs exhibited significantly impaired aortic dilatory responses to acetylcholine, loss of flow-mediated dilation (FMD), and increased systolic and diastolic blood pressure. RBC eNOS KO mice showed no alterations in acetylcholine-mediated dilation or FMD but were hypertensive. Treatment with the NOS inhibitor L-NAME further increased blood pressure in RBC eNOS KOs, demonstrating that eNOS in both ECs and RBCs contributes to blood pressure regulation. While both EC eNOS KOs and RBC eNOS KOs had lower plasma nitrite and nitrate concentrations, the levels of bound NO in RBCs were lower in RBC eNOS KOs as compared to EC eNOS KOs. Crucially, reactivation of eNOS in ECs or RBCs rescues the hypertensive phenotype of the eNOSinv/inv mice, while the levels of bound NO were restored only in RBC eNOS KI mice. Conclusions:These data reveal that eNOS in ECs and RBCs contribute independently to blood pressure homeostasis.




Circulation: 06 Jul 2021; epub ahead of print
Leo F, Suvorava T, Heuser SK, Li J, ... Kelm M, Cortese-Krott MM
Circulation: 06 Jul 2021; epub ahead of print | PMID: 34229449
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Abstract

Interleukin-1α (IL-1α) is a Central Regulator of Leukocyte-Endothelial Adhesion in Myocardial Infarction and in Chronic Kidney Disease.

Schunk SJ, Triem S, Schmit D, Zewinger S, ... Ampofo E, Speer T
Background: Cardiovascular diseases (CVD) and chronic kidney disease (CKD) are highly prevalent, aggravate each other, and account for substantial mortality. Both conditions are characterized by activation of the innate immune system. The alarmin IL-1α is expressed in a variety of cell types promoting (sterile) systemic inflammation. The aim of the present study was to examine the role of IL-1α in mediating inflammation in the setting of acute myocardial infarction (AMI) and CKD.
Methods:
We assessed the expression of IL-1α on the surface of monocytes from patients with AMI and patients with CKD and determined its association with atherosclerotic CVD events during follow-up in an explorative clinical study. Furthermore, we assessed the inflammatory effects of IL-1α in several organ injury models in Il1a-/- and Il1b-/- mice and investigated the underlying mechanisms in vitro in monocytes and endothelial cells.
Results:
IL-1α is strongly expressed on the surface of monocytes from patients with AMI and CKD compared to healthy controls. Higher IL-1α surface expression on monocytes from patients with AMI and CKD was associated with a higher risk for atherosclerotic CVD events, which underlines the clinical relevance of IL-1α. In mice, IL-1α, but not IL-1β, mediates leukocyte-endothelial adhesion as determined by intravital microscopy. IL-1α promotes accumulation of macrophages and neutrophils in inflamed tissue in vivo. Furthermore, IL-1α on monocytes stimulates their homing at sites of vascular injury. A variety of stimuli such as free fatty acids or oxalate crystals induce IL-1α surface expression and release by monocytes, which then mediates their adhesion to the endothelium via IL-1 receptor-1. Besides, IL-1α promotes expression of the vascular cell adhesion molecule-1 (VCAM-1) on endothelial cells thereby fostering the adhesion of circulating leukocytes. IL-1α induces inflammatory injury after experimental AMI and abrogation of IL-1α prevents the development of CKD in oxalate or adenine-fed mice. Conclusions: IL-1α represents a key mediator of leukocyte-endothelial adhesion and inflammation in AMI and CKD. Inhibition of IL-1α may serve as a novel anti-inflammatory treatment strategy.




Circulation: 30 Jun 2021; epub ahead of print
Schunk SJ, Triem S, Schmit D, Zewinger S, ... Ampofo E, Speer T
Circulation: 30 Jun 2021; epub ahead of print | PMID: 34192892
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Abstract

Defective Flow-Migration Coupling Causes Arteriovenous Malformations in Hereditary Hemorrhagic Telangiectasia.

Park H, Furtado J, Poulet M, Chung M, ... Schwartz MA, Eichmann A
Background: Activin receptor-like kinase 1 (ALK1) is an endothelial transmembrane serine threonine kinase receptor for BMP family ligands that plays a critical role in cardiovascular development and pathology. Loss-of-function mutations in the ALK1 gene cause type 2 hereditary hemorrhagic telangiectasia (HHT), a devastating disorder that leads to arteriovenous malformations (AVMs). Here we show that ALK1 controls endothelial cell polarization against the direction of blood flow and flow-induced endothelial migration from veins through capillaries into arterioles.
Methods:
Using Cre lines that recombine in different subsets of arterial, capillary-venous or endothelial tip cells, we showed that capillary-venous Alk1 deletion was sufficient to induce AVM formation in the postnatal retina.
Results:
ALK1 deletion impaired capillary-venous endothelial cell polarization against the direction of blood flow in vivo and in vitro. Mechanistically, ALK1 deficient cells exhibited increased integrin signaling interaction with VEGFR2, which enhanced downstream YAP/TAZ nuclear translocation. Pharmacological inhibition of integrin or YAP/TAZ signaling rescued flow migration coupling and prevented vascular malformations in Alk1 deficient mice. Conclusions: Our study reveals ALK1 as an essential driver of flow-induced endothelial cell migration and identifies loss of flow-migration coupling as a driver of AVM formation in HHT disease. Integrin-YAP/TAZ signaling blockers are new potential targets to prevent vascular malformations in HHT patients.




Circulation: 28 Jun 2021; epub ahead of print
Park H, Furtado J, Poulet M, Chung M, ... Schwartz MA, Eichmann A
Circulation: 28 Jun 2021; epub ahead of print | PMID: 34182767
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Abstract

Cyclin D2 Overexpression Enhances the Efficacy of Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes for Myocardial Repair in a Swine Model of Myocardial Infarction.

Zhao M, Nakada Y, Wei Y, Bian W, ... Walcott GP, Zhang J
Background
Human induced pluripotent stem cells with normal (wild-type) or upregulated (overexpressed) levels of CCND2 (cyclin D2) expression were differentiated into cardiomyocytes (CCND2WTCMs or CCND2OECMs, respectively) and injected into infarcted pig hearts.
Methods
Acute myocardial infarction was induced by a 60-minute occlusion of the left anterior descending coronary artery. Immediately after reperfusion, CCND2WTCMs or CCND2OECMs (3×107 cells each) or an equivalent volume of the delivery vehicle was injected around the infarct border zone area.
Results
The number of the engrafted CCND2OECMs exceeded that of the engrafted CCND2WTCMs from 6- to 8-fold, rising from 1 week to 4 weeks after implantation. In contrast to the treatment with the CCND2WTCMs or the delivery vehicle, the administration of CCND2OECM was associated with significantly improved left ventricular function, as revealed by magnetic resonance imaging. This correlated with reduction of infarct size, fibrosis, ventricular hypertrophy, and cardiomyocyte apoptosis, and increase of vascular density and arterial density, as per histologic analysis of the treated hearts. Expression of cell proliferation markers (eg, Ki67, phosphorylated histone 3, and Aurora B kinase) was also significantly upregulated in the recipient cardiomyocytes from the CCND2OECM-treated than from the CCND2WTCM-treated pigs. The cell proliferation rate and the hypoxia tolerance measured in cultured human induced pluripotent stem cell cardiomyocytes were significantly greater after treatment with exosomes isolated from the CCND2OECMs (CCND2OEExos) than from the CCND2WTCMs (CCND2WTExos). As demonstrated by our study, CCND2OEExos can also promote the proliferation activity of postnatal rat and adult mouse cardiomyocytes. A bulk miRNA sequencing analysis of CCND2OEExos versus CCND2WTExos identified 206 and 91 miRNAs that were significantly upregulated and downregulated, respectively. Gene ontology enrichment analysis identified significant differences in the expression profiles of miRNAs from various functional categories and pathways, including miRNAs implicated in cell-cycle checkpoints (G2/M and G1/S transitions), or the mechanism of cytokinesis.
Conclusions
We demonstrated that enhanced potency of CCND2OECMs promoted myocyte proliferation in both grafts and recipient tissue in a large mammal acute myocardial infarction model. These results suggest that CCND2OECMs transplantation may be a potential therapeutic strategy for the repair of infarcted hearts.



Circulation: 19 Jul 2021; 144:210-228
Zhao M, Nakada Y, Wei Y, Bian W, ... Walcott GP, Zhang J
Circulation: 19 Jul 2021; 144:210-228 | PMID: 33951921
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Abstract

Novel Role of GPR35 (G-Protein-Coupled Receptor 35) in the Regulation of Endothelial Cell Function and Blood Pressure.

Li H, Nguyen H, Meda Venkata SP, Koh JY, ... Chen W, Wang JM
GPR35 (G-protein-coupled receptor 35) is a poorly characterized receptor that has garnered increased interest as a therapeutic target through its implications in a range of inflammatory and cardiovascular diseases, but its biological functions stay largely unknown. The current study evaluated the effect of GPR35 on endothelial cell (EC) functions and hemodynamic homeostasis. In primary human aortic ECs, the expression of GPR35 was manipulated by transfections of adenovirus carrying either GPR35 cDNA or shRNA against GPR35, using adenovirus carrying β-gal as control. Mouse aortic ECs were isolated and cultured from GPR35 knockout and wild-type control mice. Our results indicated that genetic inhibition of GPR35 in human and mouse ECs significantly promoted cell proliferation, migration, and tube formation in vitro. The GCH1 (guanosine triphosphate cyclohydrolase I)-mediated biosynthesis of tetrahydrobiopterin was enhanced, reducing intracellular superoxide. Knocking down GCH1 or eNOS (endothelial nitric oxide synthase) significantly blunted the robust angiogenesis induced by GPR35 suppression. Male GPR35 knockout mice demonstrated reduced basal arterial blood pressure and an attenuated onset of hypertension in deoxycorticosterone acetate-salt induced hypertensive model compared with male GPR35 wild-type control mice in vivo, with concomitant improved endothelium-dependent vasodilation and decreased superoxide in isolated aortas. The difference in arterial blood pressure was absent between female GPR35 wild-type control and female GPR35 knockout mice. Our study provides novel insights into the roles of GPR35 in endothelial function and vascular tone modulation that critically contribute to the pathophysiology of blood pressure elevation. Antagonizing GPR35 activity might represent a potentially effective therapeutic approach to restore EC function and hemodynamic homeostasis.



Hypertension: 18 Jul 2021:HYPERTENSIONAHA12015423; epub ahead of print
Li H, Nguyen H, Meda Venkata SP, Koh JY, ... Chen W, Wang JM
Hypertension: 18 Jul 2021:HYPERTENSIONAHA12015423; epub ahead of print | PMID: 34275335
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Abstract

In Vivo Characterization of Endogenous Cardiovascular Extracellular Vesicles in Larval and Adult Zebrafish.

Scott A, Sueiro Ballesteros L, Bradshaw M, Tsuji C, ... Emanueli C, Richardson RJ
Objective
Extracellular vesicles (EVs) facilitate molecular transport across extracellular space, allowing local and systemic signaling during homeostasis and in disease. Extensive studies have described functional roles for EV populations, including during cardiovascular disease, but the in vivo characterization of endogenously produced EVs is still in its infancy. Because of their genetic tractability and live imaging amenability, zebrafish represent an ideal but under-used model to investigate endogenous EVs. We aimed to establish a transgenic zebrafish model to allow the in vivo identification, tracking, and extraction of endogenous EVs produced by different cell types. Approach and
Results:
Using a membrane-tethered fluorophore reporter system, we show that EVs can be fluorescently labeled in larval and adult zebrafish and demonstrate that multiple cell types including endothelial cells and cardiomyocytes actively produce EVs in vivo. Cell-type specific EVs can be tracked by high spatiotemporal resolution light-sheet live imaging and modified flow cytometry methods allow these EVs to be further evaluated. Additionally, cryo electron microscopy reveals the full morphological diversity of larval and adult EVs. Importantly, we demonstrate the utility of this model by showing that different cell types exchange EVs in the adult heart and that ischemic injury models dynamically alter EV production.
Conclusions
We describe a powerful in vivo zebrafish model for the investigation of endogenous EVs in all aspects of cardiovascular biology and pathology. A cell membrane fluorophore labeling approach allows cell-type specific tracing of EV origin without bias toward the expression of individual protein markers and will allow detailed future examination of their function.



Arterioscler Thromb Vasc Biol: 14 Jul 2021:ATVBAHA121316539; epub ahead of print
Scott A, Sueiro Ballesteros L, Bradshaw M, Tsuji C, ... Emanueli C, Richardson RJ
Arterioscler Thromb Vasc Biol: 14 Jul 2021:ATVBAHA121316539; epub ahead of print | PMID: 34261327
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Impact:
Abstract

Fibrous Caps in Atherosclerosis Form by Notch-Dependent Mechanisms Common to Arterial Media Development.

Martos-Rodríguez CJ, Albarrán-Juárez J, Morales-Cano D, Caballero A, ... Carramolino L, Bentzon JF
Objective
Atheromatous fibrous caps are produced by smooth muscle cells (SMCs) that are recruited to the subendothelial space. We tested whether the recruitment mechanisms are the same as in embryonic artery development, which relies prominently on Notch signaling to form the subendothelial medial SMC layers. Approach and
Results:
Notch elements were expressed in regions of fibrous cap in human and mouse plaques. To assess the causal role of Notch signaling in cap formation, we studied atherosclerosis in mice where the Notch pathway was inactivated in SMCs by conditional knockout of the essential effector transcription factor RBPJ. The recruitment of cap SMCs was significantly reduced without major effects on plaque size. Lineage tracing revealed the accumulation of SMC-derived plaque cells in the cap region was unaltered but that Notch-defective cells failed to re-acquire the SMC phenotype in the cap. Conversely, to analyze whether the loss of Notch signaling is required for SMC-derived cells to accumulate in atherogenesis, we studied atherosclerosis in mice with constitutive activation of Notch signaling in SMCs achieved by conditional expression of the Notch intracellular domain. Forced Notch signaling inhibited the ability of medial SMCs to contribute to plaque cells, including both cap SMCs and osteochondrogenic cells, and significantly reduced atherosclerosis development.
Conclusions
Sequential loss and gain of Notch signaling is needed to build the cap SMC population. The shared mechanisms with embryonic arterial media assembly suggest that the cap forms as a neo-media that restores the connection between endothelium and subendothelial SMCs, transiently disrupted in early atherogenesis.



Arterioscler Thromb Vasc Biol: 14 Jul 2021:ATVBAHA120315627; epub ahead of print
Martos-Rodríguez CJ, Albarrán-Juárez J, Morales-Cano D, Caballero A, ... Carramolino L, Bentzon JF
Arterioscler Thromb Vasc Biol: 14 Jul 2021:ATVBAHA120315627; epub ahead of print | PMID: 34261328
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Impact:
Abstract

Genome-Wide Association Study Identifies a Functional Variant Associated With HDL-C (High-Density Lipoprotein Cholesterol) Levels and Premature Coronary Artery Disease.

León-Mimila P, Villamil-Ramírez H, Macias-Kauffer LR, Jacobo-Albavera L, ... Villarreal-Molina T, Canizales-Quinteros S
Objective
Low HDL-C (high-density lipoprotein cholesterol) is the most frequent dyslipidemia in Mexicans, but few studies have examined the underlying genetic basis. Our purpose was to identify genetic variants associated with HDL-C levels and cardiovascular risk in the Mexican population. Approach and
Results:
A genome-wide association studies for HDL-C levels in 2335 Mexicans, identified four loci associated with genome-wide significance: CETP, ABCA1, LIPC, and SIDT2. The SIDT2 missense Val636Ile variant was associated with HDL-C levels and was replicated in 3 independent cohorts (P=5.9×10-18 in the conjoint analysis). The SIDT2/Val636Ile variant is more frequent in Native American and derived populations than in other ethnic groups. This variant was also associated with increased ApoA1 and glycerophospholipid serum levels, decreased LDL-C (low-density lipoprotein cholesterol) and ApoB levels, and a lower risk of premature CAD. Because SIDT2 was previously identified as a protein involved in sterol transport, we tested whether the SIDT2/Ile636 protein affected this function using an in vitro site-directed mutagenesis approach. The SIDT2/Ile636 protein showed increased uptake of the cholesterol analog dehydroergosterol, suggesting this variant affects function. Finally, liver transcriptome data from humans and the Hybrid Mouse Diversity Panel are consistent with the involvement of SIDT2 in lipid and lipoprotein metabolism.
Conclusions
This is the first genome-wide association study for HDL-C levels seeking associations with coronary artery disease in the Mexican population. Our findings provide new insight into the genetic architecture of HDL-C and highlight SIDT2 as a new player in cholesterol and lipoprotein metabolism in humans.



Arterioscler Thromb Vasc Biol: 07 Jul 2021:ATVBAHA120315391; epub ahead of print
León-Mimila P, Villamil-Ramírez H, Macias-Kauffer LR, Jacobo-Albavera L, ... Villarreal-Molina T, Canizales-Quinteros S
Arterioscler Thromb Vasc Biol: 07 Jul 2021:ATVBAHA120315391; epub ahead of print | PMID: 34233476
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Impact:
Abstract

Endothelial-Mesenchymal Transition in Cardiovascular Disease.

Alvandi Z, Bischoff J
Endothelial-to-mesenchymal transition is a dynamic process in which endothelial cells suppress constituent endothelial properties and take on mesenchymal cell behaviors. To begin the process, endothelial cells loosen their cell-cell junctions, degrade the basement membrane, and migrate out into the perivascular surroundings. These initial endothelial behaviors reflect a transient modulation of cellular phenotype, that is, a phenotypic modulation, that is sometimes referred to as partial endothelial-to-mesenchymal transition. Loosening of endothelial junctions and migration are also seen in inflammatory and angiogenic settings such that endothelial cells initiating endothelial-to-mesenchymal transition have overlapping behaviors and gene expression with endothelial cells responding to inflammatory signals or sprouting to form new blood vessels. Reduced endothelial junctions increase permeability, which facilitates leukocyte trafficking, whereas endothelial migration precedes angiogenic sprouting and neovascularization; both endothelial barriers and quiescence are restored as inflammatory and angiogenic stimuli subside. Complete endothelial-to-mesenchymal transition proceeds beyond phenotypic modulation such that mesenchymal characteristics become prominent and endothelial functions diminish. In proadaptive, regenerative settings the new mesenchymal cells produce extracellular matrix and contribute to tissue integrity whereas in maladaptive, pathologic settings the new mesenchymal cells become fibrotic, overproducing matrix to cause tissue stiffness, which eventually impacts function. Here we will review what is known about how TGF (transforming growth factor) β influences this continuum from junctional loosening to cellular migration and its relevance to cardiovascular diseases.



Arterioscler Thromb Vasc Biol: 30 Jun 2021:ATVBAHA121313788; epub ahead of print
Alvandi Z, Bischoff J
Arterioscler Thromb Vasc Biol: 30 Jun 2021:ATVBAHA121313788; epub ahead of print | PMID: 34196216
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Impact:
Abstract

Salt-Inducible Kinase 3 Promotes Vascular Smooth Muscle Cell Proliferation and Arterial Restenosis by Regulating AKT and PKA-CREB Signaling.

Cai Y, Wang XL, Lu J, Lin X, Dong J, Guzman RJ
Objective
Arterial restenosis is the pathological narrowing of arteries after endovascular procedures, and it is an adverse event that causes patients to experience recurrent occlusive symptoms. Following angioplasty, vascular smooth muscle cells (SMCs) change their phenotype, migrate, and proliferate, resulting in neointima formation, a hallmark of arterial restenosis. SIKs (salt-inducible kinases) are a subfamily of the AMP-activated protein kinase family that play a critical role in metabolic diseases including hepatic lipogenesis and glucose metabolism. Their role in vascular pathological remodeling, however, has not been explored. In this study, we aimed to understand the role and regulation of SIK3 in vascular SMC migration, proliferation, and neointima formation. Approach and
Results:
We observed that SIK3 expression was low in contractile aortic SMCs but high in proliferating SMCs. It was also highly induced by growth medium in vitro and in neointimal lesions in vivo. Inactivation of SIKs significantly attenuated vascular SMC proliferation and up-regulated p21CIP1 and p27KIP1. SIK inhibition also suppressed SMC migration and modulated actin polymerization. Importantly, we found that inhibition of SIKs reduced neointima formation and vascular inflammation in a femoral artery wire injury model. In mechanistic studies, we demonstrated that inactivation of SIKs mainly suppressed SMC proliferation by down-regulating AKT (protein kinase B) and PKA (protein kinase A)-CREB (cAMP response element-binding protein) signaling. CRTC3 signaling likely contributed to SIK inactivation-mediated antiproliferative effects.
Conclusions
These findings suggest that SIK3 may play a critical role in regulating SMC proliferation, migration, and arterial restenosis. This study provides insights into SIK inhibition as a potential therapeutic strategy for treating restenosis in patients with PAD.



Arterioscler Thromb Vasc Biol: 30 Jun 2021:ATVBAHA121316219; epub ahead of print
Cai Y, Wang XL, Lu J, Lin X, Dong J, Guzman RJ
Arterioscler Thromb Vasc Biol: 30 Jun 2021:ATVBAHA121316219; epub ahead of print | PMID: 34196217
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Impact:
Abstract

Smooth Muscle Cell-Enriched Long Noncoding RNA Regulates Cell Plasticity and Atherosclerosis by Interacting With Serum Response Factor.

Ni H, Haemmig S, Deng Y, Chen J, ... Dai Q, Feinberg MW
Objective
Vascular smooth muscle cell (VSMC) plasticity plays a critical role in the development of atherosclerosis. Long noncoding RNAs (lncRNAs) are emerging as important regulators in the vessel wall and impact cellular function through diverse interactors. However, the role of lncRNAs in regulating VSMCs plasticity and atherosclerosis remains unclear. Approach and
Results:
We identified a VSMC-enriched lncRNA cardiac mesoderm enhancer-associated noncoding RNA (CARMN) that is dynamically regulated with progression of atherosclerosis. In both mouse and human atherosclerotic plaques, CARMN colocalized with VSMCs and was expressed in the nucleus. Knockdown of CARMN using antisense oligonucleotides in Ldlr-/- mice significantly reduced atherosclerotic lesion formation by 38% and suppressed VSMCs proliferation by 45% without affecting apoptosis. In vitro CARMN gain- and loss-of-function studies verified effects on VSMC proliferation, migration, and differentiation. TGF-β1 (transforming growth factor-beta) induced CARMN expression in a Smad2/3-dependent manner. CARMN regulated VSMC plasticity independent of the miR143/145 cluster, which is located in close proximity to the CARMN locus. Mechanistically, lncRNA pulldown in combination with mass spectrometry analysis showed that the nuclear-localized CARMN interacted with SRF (serum response factor) through a specific 600-1197 nucleotide domain. CARMN enhanced SRF occupancy on the promoter regions of its downstream VSMC targets. Finally, knockdown of SRF abolished the regulatory role of CARMN in VSMC plasticity.
Conclusions
The lncRNA CARMN is a critical regulator of VSMC plasticity and atherosclerosis. These findings highlight the role of a lncRNA in SRF-dependent signaling and provide implications for a range of chronic vascular occlusive disease states.



Arterioscler Thromb Vasc Biol: 21 Jul 2021:ATVBAHA120315911; epub ahead of print
Ni H, Haemmig S, Deng Y, Chen J, ... Dai Q, Feinberg MW
Arterioscler Thromb Vasc Biol: 21 Jul 2021:ATVBAHA120315911; epub ahead of print | PMID: 34289702
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Impact:
Abstract

Analysis of F-Sodium Fluoride Positron Emission Tomography Signal Sources in Atherosclerotic Minipigs Shows Specific Binding of F-Sodium Fluoride to Plaque Calcifications.

Nogales P, Velasco C, Mota-Cobián A, González-Cintado L, ... Mateo J, Bentzon JF
Objective
18F-sodium fluoride (18F-NaF) positron emission tomography (PET) imaging is thought to visualize active atherosclerotic plaque calcification. This is supported by the binding of 18F-NaF to plaque calcification ex vivo, but no prior studies have examined binding of 18F-NaF to human-like plaque in vivo. Our aim was to validate the specificity of 18F-NaF PET for plaque calcifications in atherosclerotic minipigs. Approach and
Results:
Gain-of-function PCSK9D374Y (proprotein convertase/subtilisin kexin type 9) transgenic Yucatan minipigs (n=4) were fed high-fat diet for 2.5 years to develop atherosclerosis and then subjected to 18F-NaF PET/computed tomography imaging. The heart, aorta, and iliac arteries were immediately re-scanned ex vivo after surgical extraction. Lesions from the abdominal aorta, iliac arteries, and coronary arteries were cryo-sectioned for autoradiography. Histological plaque characteristics, PET/computed tomography signal, and autoradiography were linked through regression and co-localization analysis. Arterial 18F-NaF PET signal had intensities comparable to clinical scans and colocalized moderately with calcification detected by computed tomography. Histological analysis showed calcification spanning from microcalcifications near lipid pools and necrotic core to more homogenous macrocalcifications. Comparison with arteries from autopsy cases confirmed the resemblance in localization and appearance with early human plaque calcification. Regression analysis in the abdominal aorta showed correlations with calcified plaque but could not rule out contributions from noncalcified plaque. This was resolved by autoradiography, which showed specific accumulation in plaque calcifications in all examined arteries. In the context of porcine abdominal aorta, 18F-NaF PET imaging was, however, less accurate than computed tomography for detecting small calcifications.
Conclusions
18F-NaF accumulates specifically in calcifications of atherosclerotic plaques in vivo.



Arterioscler Thromb Vasc Biol: 21 Jul 2021:ATVBAHA121316075; epub ahead of print
Nogales P, Velasco C, Mota-Cobián A, González-Cintado L, ... Mateo J, Bentzon JF
Arterioscler Thromb Vasc Biol: 21 Jul 2021:ATVBAHA121316075; epub ahead of print | PMID: 34289703
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Abstract

High spatial endothelial shear stress gradient independently predicts site of acute coronary plaque rupture and erosion.

Thondapu V, Mamon C, Poon EKW, Kurihara O, ... Barlis P, Jang IK
Aims
To investigate local haemodynamics in the setting of acute coronary plaque rupture and erosion.
Methods and results
Intracoronary optical coherence tomography performed in 37 patients with acute coronary syndromes caused by plaque rupture (n = 19) or plaque erosion (n = 18) was used for three-dimensional reconstruction and computational fluid dynamics simulation. Endothelial shear stress (ESS), spatial ESS gradient (ESSG), and oscillatory shear index (OSI) were compared between plaque rupture and erosion through mixed-effects logistic regression. Lipid, calcium, macrophages, layered plaque, and cholesterol crystals were also analysed. By multivariable analysis, only high ESSG [odds ratio (OR) 5.29, 95% confidence interval (CI) 2.57-10.89, P < 0.001], lipid (OR 12.98, 95% CI 6.57-25.67, P < 0.001), and layered plaque (OR 3.17, 95% CI 1.82-5.50, P < 0.001) were independently associated with plaque rupture. High ESSG (OR 13.28, 95% CI 6.88-25.64, P < 0.001), ESS (OR 2.70, 95% CI 1.34-5.42, P = 0.005), and OSI (OR 2.18, 95% CI 1.33-3.54, P = 0.002) independently associated with plaque erosion. ESSG was higher at rupture sites than erosion sites [median (interquartile range): 5.78 (2.47-21.15) vs. 2.62 (1.44-6.18) Pa/mm, P = 0.009], OSI was higher at erosion sites than rupture sites [1.04 × 10-2 (2.3 × 10-3-4.74 × 10-2) vs. 1.29 × 10-3 (9.39 × 10-5-3.0 × 10-2), P < 0.001], but ESS was similar (P = 0.29).
Conclusions
High ESSG is independently associated with plaque rupture while high ESSG, ESS, and OSI associate with plaque erosion. While ESSG is higher at rupture sites than erosion sites, OSI is higher at erosion sites and ESS was similar. These results suggest that ESSG and OSI may play critical roles in acute plaque rupture and erosion, respectively.

Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2020. For permissions, please email: [email protected]

Cardiovasc Res: 06 Jul 2021; 117:1974-1985
Thondapu V, Mamon C, Poon EKW, Kurihara O, ... Barlis P, Jang IK
Cardiovasc Res: 06 Jul 2021; 117:1974-1985 | PMID: 32832991
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Abstract

Interleukin-7 and interleukin-15 drive CD4+CD28null T lymphocyte expansion and function in patients with acute coronary syndrome.

Bullenkamp J, Mengoni V, Kaur S, Chhetri I, ... Kaski JC, Dumitriu IE
Aims
Inflammation has important roles in atherosclerosis. CD4+CD28null (CD28null) T cells are a specialized T lymphocyte subset that produce inflammatory cytokines and cytotoxic molecules. CD28null T cells expand preferentially in patients with acute coronary syndrome (ACS) rather than stable angina and are barely detectable in healthy subjects. Importantly, ACS patients with CD28null T-cell expansion have increased risk for recurrent acute coronary events and poor prognosis, compared to ACS patients in whom this cell subset does not expand. The mechanisms regulating CD28null T-cell expansion in ACS remain elusive. We therefore investigated the role of cytokines in CD28null T-cell expansion in ACS.
Methods and results
High-purity sorted CD4+ T cells from ACS patients were treated with a panel of cytokines (TNF-α, IL-1β, IL-6, IL-7, and IL-15), and effects on the number, phenotype, and function of CD28null T cells were analysed and compared to the control counterpart CD28+ T-cell subset. IL-7- and IL-15-induced expansion of CD28null T cells from ACS patients, while inflammatory cytokines TNF-α, IL-1β, and IL-6 did not. The mechanisms underlying CD28null T-cell expansion by IL-7/IL-15 were preferential activation and proliferation of CD28null T cells compared to control CD28+ T cells. Additionally, IL-7/IL-15 markedly augmented CD28null T-cell cytotoxic function and interferon-γ production. Further mechanistic analyses revealed differences in baseline expression of component chains of IL-7/IL-15 receptors (CD127 and CD122) and increased baseline STAT5 phosphorylation in CD28null T cells from ACS patients compared to the control CD28+ T-cell subset. Notably, we demonstrate that CD28null T-cell expansion was significantly inhibited by Tofacitinib, a selective JAK1/JAK3 inhibitor that blocks IL-7/IL-15 signalling.
Conclusion
Our novel data show that IL-7 and IL-15 drive the expansion and function of CD28null T cells from ACS patients suggesting that IL-7/IL-15 blockade may prevent expansion of these cells and improve patient outcomes.

© The Author(s) 2020. Published by Oxford University Press on behalf of the European Society of Cardiology.

Cardiovasc Res: 06 Jul 2021; 117:1935-1948
Bullenkamp J, Mengoni V, Kaur S, Chhetri I, ... Kaski JC, Dumitriu IE
Cardiovasc Res: 06 Jul 2021; 117:1935-1948 | PMID: 32647892
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Impact:
Abstract

Preclinical evidence for the therapeutic value of TBX5 normalization in arrhythmia control.

Rathjens FS, Blenkle A, Iyer LM, Renger A, ... Zelarayan LC, Zafeiriou MP
Aims 
Arrhythmias and sudden cardiac death (SCD) occur commonly in patients with heart failure. We found T-box 5 (TBX5) dysregulated in ventricular myocardium from heart failure patients and thus we hypothesized that TBX5 reduction contributes to arrhythmia development in these patients. To understand the underlying mechanisms, we aimed to reveal the ventricular TBX5-dependent transcriptional network and further test the therapeutic potential of TBX5 level normalization in mice with documented arrhythmias.
Methods and results 
We used a mouse model of TBX5 conditional deletion in ventricular cardiomyocytes. Ventricular (v) TBX5 loss in mice resulted in mild cardiac dysfunction and arrhythmias and was associated with a high mortality rate (60%) due to SCD. Upon angiotensin stimulation, vTbx5KO mice showed exacerbated cardiac remodelling and dysfunction suggesting a cardioprotective role of TBX5. RNA-sequencing of a ventricular-specific TBX5KO mouse and TBX5 chromatin immunoprecipitation was used to dissect TBX5 transcriptional network in cardiac ventricular tissue. Overall, we identified 47 transcripts expressed under the control of TBX5, which may have contributed to the fatal arrhythmias in vTbx5KO mice. These included transcripts encoding for proteins implicated in cardiac conduction and contraction (Gja1, Kcnj5, Kcng2, Cacna1g, Chrm2), in cytoskeleton organization (Fstl4, Pdlim4, Emilin2, Cmya5), and cardiac protection upon stress (Fhl2, Gpr22, Fgf16). Interestingly, after TBX5 loss and arrhythmia development in vTbx5KO mice, TBX5 protein-level normalization by systemic adeno-associated-virus (AAV) 9 application, re-established TBX5-dependent transcriptome. Consequently, cardiac dysfunction was ameliorated and the propensity of arrhythmia occurrence was reduced.
Conclusions 
This study uncovers a novel cardioprotective role of TBX5 in the adult heart and provides preclinical evidence for the therapeutic value of TBX5 protein normalization in the control of arrhythmia.

© The Author(s) 2020. Published by Oxford University Press on behalf of the European Society of Cardiology.

Cardiovasc Res: 06 Jul 2021; 117:1908-1922
Rathjens FS, Blenkle A, Iyer LM, Renger A, ... Zelarayan LC, Zafeiriou MP
Cardiovasc Res: 06 Jul 2021; 117:1908-1922 | PMID: 32777030
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Impact:
Abstract

Impaired cytoplasmic domain interactions cause co-assembly defect and loss of function in the p.Glu293Lys KNCJ2 variant isolated from an Andersen-Tawil syndrome patient.

Déri S, Borbás J, Hartai T, Hategan L, ... Sepp R, Ördög B
Aims
Subunit interactions at the cytoplasmic domain interface (CD-I) have recently been shown to control gating in inward rectifier potassium channels. Here we report the novel KCNJ2 variant p.Glu293Lys that has been found in a patient with Andersen-Tawil syndrome type 1 (ATS1), causing amino acid substitution at the CD-I of the inward rectifier potassium channel subunit Kir2.1. Neither has the role of Glu293 in gating control been investigated nor has a pathogenic variant been described at this position. This study aimed to assess the involvement of Glu293 in CD-I subunit interactions and to establish the pathogenic role of the p.Glu293Lys variant in ATS1.
Methods and results
The p.Glu293Lys variant produced no current in homomeric form and showed dominant-negative effect over wild-type (WT) subunits. Immunocytochemical labelling showed the p.Glu293Lys subunits to distribute in the subsarcolemmal space. Salt bridge prediction indicated the presence of an intersubunit salt bridge network at the CD-I of Kir2.1, with the involvement of Glu293. Subunit interactions were studied by the NanoLuc® Binary Technology (NanoBiT) split reporter assay. Reporter constructs carrying NanoBiT tags on the intracellular termini produced no bioluminescent signal above background with the p.Glu293Lys variant in homomeric configuration and significantly reduced signals in cells co-expressing WT and p.Glu293Lys subunits simultaneously. Extracellularly presented reporter tags, however, generated comparable bioluminescent signals with heteromeric WT and p.Glu293Lys subunits and with homomeric WT channels.
Conclusions
Loss of function and dominant-negative effect confirm the causative role of p.Glu293Lys in ATS1. Co-assembly of Kir2.1 subunits is impaired in homomeric channels consisting of p.Glu293Lys subunits and is partially rescued in heteromeric complexes of WT and p.Glu293Lys Kir2.1 variants. These data point to an important role of Glu293 in mediating subunit assembly, as well as in gating of Kir2.1 channels.

Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2020. For permissions, please email: [email protected]

Cardiovasc Res: 06 Jul 2021; 117:1923-1934
Déri S, Borbás J, Hartai T, Hategan L, ... Sepp R, Ördög B
Cardiovasc Res: 06 Jul 2021; 117:1923-1934 | PMID: 32810216
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Impact:
Abstract

The physiological and pathological functions of VEGFR3 in cardiac and lymphatic development and related diseases.

Monaghan RM, Page DJ, Ostergaard P, Keavney BD
Vascular endothelial growth factor receptors (VEGFRs) are part of the evolutionarily conserved VEGF signalling pathways that regulate the development and maintenance of the body\'s cardiovascular and lymphovascular systems. VEGFR3, encoded by the FLT4 gene, has an indispensable and well-characterized function in development and establishment of the lymphatic system. Autosomal dominant VEGFR3 mutations, that prevent the receptor functioning as a homodimer, cause one of the major forms of hereditary primary lymphoedema; Milroy disease. Recently, we and others have shown that FLT4 variants, distinct to those observed in Milroy disease cases, predispose individuals to Tetralogy of Fallot, the most common cyanotic congenital heart disease, demonstrating a novel function for VEGFR3 in early cardiac development. Here, we examine the familiar and emerging roles of VEGFR3 in the development of both lymphovascular and cardiovascular systems, respectively, compare how distinct genetic variants in FLT4 lead to two disparate human conditions, and highlight the research still required to fully understand this multifaceted receptor.

© The Author(s) 2020. Published by Oxford University Press on behalf of the European Society of Cardiology.

Cardiovasc Res: 06 Jul 2021; 117:1877-1890
Monaghan RM, Page DJ, Ostergaard P, Keavney BD
Cardiovasc Res: 06 Jul 2021; 117:1877-1890 | PMID: 33067626
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Impact:
Abstract

Adult zebrafish ventricular electrical gradients as tissue mechanisms of ECG patterns under baseline vs. oxidative stress.

Zhao Y, James NA, Beshay AR, Chang EE, ... Nguyen B, Nguyen TP
Aims
In mammalian ventricles, electrical gradients establish electrical heterogeneities as essential tissue mechanisms to optimize mechanical efficiency and safeguard electrical stability. Electrical gradients shape mammalian electrocardiographic patterns; disturbance of electrical gradients is proarrhythmic. The zebrafish heart is a popular surrogate model for human cardiac electrophysiology thanks to its remarkable recapitulation of human electrocardiogram and ventricular action potential features. Yet, zebrafish ventricular electrical gradients are largely unexplored. The goal of this study is to define the zebrafish ventricular electrical gradients that shape the QRS complex and T wave patterns at baseline and under oxidative stress.
Methods and results
We performed in vivo electrocardiography and ex vivo voltage-sensitive fluorescent epicardial and transmural optical mapping of adult zebrafish hearts at baseline and during acute H2O2 exposure. At baseline, apicobasal activation and basoapical repolarization gradients accounted for the polarity concordance between the QRS complex and T wave. During H2O2 exposure, differential regional impairment of activation and repolarization at the apex and base disrupted prior to baseline electrical gradients, resulting in either reversal or loss of polarity concordance between the QRS complex and T wave. KN-93, a specific calcium/calmodulin-dependent protein kinase II inhibitor (CaMKII), protected zebrafish hearts from H2O2 disruption of electrical gradients. The protection was complete if administered prior to oxidative stress exposure.
Conclusions
Despite remarkable apparent similarities, zebrafish and human ventricular electrocardiographic patterns are mirror images supported by opposite electrical gradients. Like mammalian ventricles, zebrafish ventricles are also susceptible to H2O2 proarrhythmic perturbation via CaMKII activation. Our findings suggest that the adult zebrafish heart may constitute a clinically relevant model to investigate ventricular arrhythmias induced by oxidative stress. However, the fundamental ventricular activation and repolarization differences between the two species that we demonstrated in this study highlight the potential limitations when extrapolating results from zebrafish experiments to human cardiac electrophysiology, arrhythmias, and drug toxicities.

Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2020. For permissions, please email: [email protected]

Cardiovasc Res: 06 Jul 2021; 117:1891-1907
Zhao Y, James NA, Beshay AR, Chang EE, ... Nguyen B, Nguyen TP
Cardiovasc Res: 06 Jul 2021; 117:1891-1907 | PMID: 32735330
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Impact:
Abstract

Obesity, kidney dysfunction, and inflammation: interactions in hypertension.

Hall JE, Mouton AJ, da Silva AA, Omoto ACM, ... Li X, do Carmo JM
Obesity contributes 65-75% of the risk for human primary (essential) hypertension (HT) which is a major driver of cardiovascular and kidney diseases. Kidney dysfunction, associated with increased renal sodium reabsorption and compensatory glomerular hyperfiltration, plays a key role in initiating obesity-HT and target organ injury. Mediators of kidney dysfunction and increased blood pressure include (i) elevated renal sympathetic nerve activity (RSNA); (ii) increased antinatriuretic hormones such as angiotensin II and aldosterone; (iii) relative deficiency of natriuretic hormones; (iv) renal compression by fat in and around the kidneys; and (v) activation of innate and adaptive immune cells that invade tissues throughout the body, producing inflammatory cytokines/chemokines that contribute to vascular and target organ injury, and exacerbate HT. These neurohormonal, renal, and inflammatory mechanisms of obesity-HT are interdependent. For example, excess adiposity increases the adipocyte-derived cytokine leptin which increases RSNA by stimulating the central nervous system proopiomelanocortin-melanocortin 4 receptor pathway. Excess visceral, perirenal and renal sinus fat compress the kidneys which, along with increased RSNA, contribute to renin-angiotensin-aldosterone system activation, although obesity may also activate mineralocorticoid receptors independent of aldosterone. Prolonged obesity, HT, metabolic abnormalities, and inflammation cause progressive renal injury, making HT more resistant to therapy and often requiring multiple antihypertensive drugs and concurrent treatment of dyslipidaemia, insulin resistance, diabetes, and inflammation. More effective anti-obesity drugs are needed to prevent the cascade of cardiorenal, metabolic, and immune disorders that threaten to overwhelm health care systems as obesity prevalence continues to increase.

Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2020. For permissions, please email: [email protected]

Cardiovasc Res: 06 Jul 2021; 117:1859-1876
Hall JE, Mouton AJ, da Silva AA, Omoto ACM, ... Li X, do Carmo JM
Cardiovasc Res: 06 Jul 2021; 117:1859-1876 | PMID: 33258945
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Impact:
Abstract

Update on apolipoprotein B.

Sniderman A, Langlois M, Cobbaert C
Purpose of review
The 2019 European Society of Cardiology/European Atherosclerosis Society Guidelines concluded that apolipoprotein B (apoB) was a more accurate measure of cardiovascular risk and a better guide to the adequacy of lipid lowering than low-density lipoprotein cholesterol (LDL-C) or non-high-density lipoprotein cholesterol (HDL-C). Also, they stated that apoB can be measured more accurately than LDL-C or non-HDL-C. This strong endorsement of the central role of apoB contrasts with the limited endorsement of apoB by the 2018 American College of Cardiology/American Heart Association Multisociety Guidelines. Nevertheless, both retained LDL-C as the primary metric to guide statin/ezetimibe/Proprotein convertase subtilisin/kexin type 9 (PCSK9) therapy.
Recent findings
This essay will review the most important recent advances in knowledge about apoB with particular emphasis on the results of Mendelian randomization studies and a new discordance analysis in subjects on statin therapy. We will also lay out why using LDL-C to guide the adequacy of lipid lowering therapy represents an interpretive error of the results of the statin/ezetimibe/PCSK9 inhibitor randomized clinical trials and therefore why apoB should be the primary metric to guide statin/ezetimibe/PCSK9 therapy.
Summary
There is now a robust body of evidence demonstrating the superiority of apoB over LDL-C and non-HDL-C as a clinical marker of cardiovascular risk. LDL-C is not the appropriate marker to assess the benefits of statin/ezetimibe/PCSK9 therapy.

Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.

Curr Opin Lipidol: 31 Jul 2021; 32:226-230
Sniderman A, Langlois M, Cobbaert C
Curr Opin Lipidol: 31 Jul 2021; 32:226-230 | PMID: 33870931
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Abstract

Inhibition of angiopoietin-like 3 for the management of severe hypercholesterolemia.

Mohamed F, Botha TC, Raal FJ
Purpose for review
Despite the therapeutic advances for patients with severe hypercholesterolemia, particularly those with homozygous familial hypercholesterolemia (HoFH), most patients are unable to achieve target low-density lipoprotein cholesterol (LDL-C) levels with the current available standard lipid-lowering therapy (LLT). We review the role of angiopoietin-like 3 (ANGPTL3) inhibition as an additional therapeutic option for severe hypercholesterolemia, particularly HoFH.
Recent findings
Evinacumab is a monoclonal antibody against ANGPTL3, and reduces LDL-C independent of LDL-receptor activity. ANGPTL3 inhibitors are effective in lowering LDL-C in patients with FH, with a 50% reduction in LDL-C in those with HoFH. Longer-term efficacy and safety have been demonstrated with reductions in LDL-C maintained following 48 weeks of therapy. Gene silencing strategies directed against ANGPTL3 include antisense oligonucleotide and small-interfering ribonucleic acid (siRNA). ARO-ANG3 is a siRNA directed against ANGPTL3 messenger ribonucleic acid and is associated with up to a 42% reduction in LDL-C.
Summary
With the promise of these emerging novel therapeutics directed against ANGPTL3 on the horizon, achieving acceptable target LDL-C levels in HoFH without the need for lipoprotein apheresis may finally be a realistic goal and we can anticipate a decrease in cardiovascular morbidity and mortality in these difficult to treat patients.

Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.

Curr Opin Lipidol: 31 Jul 2021; 32:213-218
Mohamed F, Botha TC, Raal FJ
Curr Opin Lipidol: 31 Jul 2021; 32:213-218 | PMID: 33883446
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Abstract

Lipids and peripheral neuropathy.

Iqbal Z, Bashir B, Ferdousi M, Kalteniece A, ... Malik RA, Soran H
Purpose of review
Hyperlipidaemia is associated with the development of neuropathy. Indeed, a mechanistic link between altered lipid metabolism and peripheral nerve dysfunction has been demonstrated in a number of experimental and clinical studies. Furthermore, post hoc analyses of clinical trials of cholesterol and triglyceride-lowering pharmacotherapy have shown reduced rates of progression of diabetic neuropathy. Given, there are currently no FDA approved disease-modifying therapies for diabetic neuropathy, modulation of lipids may represent a key therapeutic target for the treatment of diabetic nerve damage. This review summarizes the current evidence base on the role of hyperlipidaemia and lipid lowering therapy on the development and progression of peripheral neuropathy.
Recent findings
A body of literature supports a detrimental effect of dyslipidaemia on nerve fibres resulting in somatic and autonomic neuropathy. The case for an important modulating role of hypertriglyceridemia is stronger than for low-density lipoprotein cholesterol (LDL-C) in relation to peripheral neuropathy. This is reflected in the outcomes of clinical trials with the different therapeutic agents targeting hyperlipidaemia reporting beneficial or neutral effects with statins and fibrates. The potential concern with the association between proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitor therapy and cognitive decline raised the possibility that extreme LDL-C lowering may result in neurodegeneration. However, studies in murine models and data from small observational studies indicate an association between increased circulating PCSK9 levels and small nerve fibre damage with a protective effect of PCSK9i therapy against small fibre neuropathy. Additionally, weight loss with bariatric surgery leads to an improvement in peripheral neuropathy and regeneration of small nerve fibres measured with corneal confocal microscopy in people with obesity with or without type 2 diabetes. These improvements correlate inversely with changes in triglyceride levels.
Summary
Hyperlipidaemia, particularly hypertriglyceridemia, is associated with the development and progression of neuropathy. Lipid modifying agents may represent a potential therapeutic option for peripheral neuropathy. Post hoc analyses indicate that lipid-lowering therapies may halt the progression of neuropathy or even lead to regeneration of nerve fibres. Well designed randomized controlled trials are needed to establish if intensive targeted lipid lowering therapy as a part of holistic metabolic control leads to nerve fibre regeneration and improvement in neuropathy symptoms.

Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.

Curr Opin Lipidol: 31 Jul 2021; 32:249-257
Iqbal Z, Bashir B, Ferdousi M, Kalteniece A, ... Malik RA, Soran H
Curr Opin Lipidol: 31 Jul 2021; 32:249-257 | PMID: 34101657
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Abstract

PDE5 Inhibition Suppresses Ventricular Arrhythmias by Reducing SR Ca Content.

Hutchings DC, Pearman CM, Madders GW, Woods LS, ... Dibb KM, Trafford AW
Rationale: Phosphodiesterase-5 (PDE5) inhibition reduces the occurrence of ventricular arrhythmias following myocardial ischemia. However, the mechanisms of the anti-arrhythmic effects of PDE5 inhibition are unknown. Diastolic calcium (Ca2+) waves lead to arrhythmias by inducing delayed after-depolarizations. Ca2+ waves are initiated when sarcoplasmic reticulum (SR) Ca2+ content reaches a threshold level and the SR releases Ca2+ spontaneously and generates a depolarizing inward sodium-calcium exchange (NCX) current. Objective: To determine the effects of PDE5 inhibition on the propensity for ventricular arrhythmias in a pro-arrhythmic large animal model and establish the role of alterations of intracellular Ca2+ cycling / SR Ca2+ content.
Methods and results:
Arrhythmia burden, monophasic action potentials and beat-to-beat variability of repolarization were measured in a sheep model using the IKr inhibitor dofetilide to induce QT prolongation and arrhythmia. Ca2+ transients, Ca2+ waves and SR Ca2+ content were measured in isolated ventricular myocytes. PDE5 inhibition was achieved using acute application of sildenafil and protein kinase G (PKG) was inhibited with KT5823. PDE5 inhibition reduced beat-to-beat variability of repolarization and suppressed after-depolarizations, premature ventricular complexes, and torsade de pointes in vivo. In single cells, dofetilide-induced DADs and triggered action potentials were suppressed by PDE5 inhibition. PDE5 inhibition decreased Ca2+ wave frequency in all cells and abolished waves in 12/22 cells. A decrease in SR Ca2+ uptake, increased trans-sarcolemmal Ca2+ efflux and reduced trans-sarcolemmal Ca2+ influx led to a reduction of SR Ca2+ content, and Ca2+ wave abolition. These effects were dependent on PKG activation. Conclusions: PDE5 inhibition acutely suppresses triggered ventricular arrhythmias in vivo and cellular data suggests this occurs via suppression of cellular Ca2+ waves. These novel anti-arrhythmic properties of PDE5 inhibition are mediated by a reduction of SR Ca2+ content and are PKG-dependent.




Circ Res: 11 Jul 2021; epub ahead of print
Hutchings DC, Pearman CM, Madders GW, Woods LS, ... Dibb KM, Trafford AW
Circ Res: 11 Jul 2021; epub ahead of print | PMID: 34247494
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Abstract

Natriuretic peptide receptor B maintains heart rate and sinoatrial node function via cyclic GMP-mediated signaling.

Dorey TW, Mackasey M, Jansen HJ, McRae MD, ... Atkinson L, Rose RA
Aims
Heart rate (HR) is a critical indicator of cardiac performance that is determined by sinoatrial node (SAN) function and regulation. Natriuretic peptides, including C-type NP (CNP) have been shown to modulate ion channel function in the SAN when applied exogenously. CNP is the only NP that acts as a ligand for natriuretic peptide receptor-B (NPR-B). Despite these properties, the ability of CNP and NPR-B to regulate HR and intrinsic SAN automaticity in vivo, and the mechanisms by which it does so, are incompletely understood. Thus, the objective of this study was to determine the role of NPR-B signaling in regulating HR and SAN function.
Methods and results
We have used NPR-B deficient mice (NPR-B+/-) to study HR regulation and SAN function using telemetry in conscious mice, intracardiac electrophysiology in anesthetized mice, high resolution optical mapping in isolated SAN preparations, patch-clamping in isolated SAN myocytes, and molecular biology in isolated SAN tissue. These studies demonstrate that NPR-B+/- mice exhibit slow HR, increased corrected SAN recovery time, and slowed SAN conduction. Spontaneous AP firing frequency in isolated SAN myocytes was impaired in NPR-B+/- mice due to reductions in the hyperpolarization activated current (If) and L-type Ca2+ current (ICa,L). If and ICa,L were reduced due to lower cGMP levels and increased hydrolysis of cAMP by phosphodiesterase 3 (PDE3) in the SAN. Inhibiting PDE3 or restoring cGMP signaling via application of 8-Br-cGMP abolished the reductions in cAMP, AP firing, If, and ICa,L, and normalized SAN conduction, in the SAN in NPR-B+/- mice. NPR-B+/- mice did not exhibit changes in SAN fibrosis and showed no evidence of cardiac hypertrophy or changes in ventricular function.
Conclusions
NPR-B plays an essential physiological role in maintaining normal HR and SAN function by modulating ion channel function in SAN myocytes via a cGMP/PDE3/cAMP signaling mechanism.

Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2021. For permissions please email: [email protected]

Cardiovasc Res: 16 Jul 2021; epub ahead of print
Dorey TW, Mackasey M, Jansen HJ, McRae MD, ... Atkinson L, Rose RA
Cardiovasc Res: 16 Jul 2021; epub ahead of print | PMID: 34273155
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Abstract

Nicotine promotes vascular calcification via intracellular Ca2+-mediated, Nox5-induced oxidative stress and extracellular vesicle release in vascular smooth muscle cells.

Petsophonsakul P, Burgmaier M, Willems B, Heeneman S, ... Furmanik M, Schurgers L
Aims
Smokers are at increased risk of cardiovascular events. However, the exact mechanisms through which smoking influences cardiovascular disease resulting in accelerated atherosclerosis and vascular calcification are unknown. The aim of this study was to investigate effects of nicotine on initiation of vascular smooth muscle cell (VSMC) calcification and to elucidate underlying mechanisms.
Methods and results
We assessed vascular calcification of 62 carotid lesions of both smoking and non-smoking patients using ex vivo micro-computed-tomography (µCT)-scanning. Calcification was present more often in carotid plaques of smokers (n = 22 of 30, 73.3%) compared to non-smokers (n = 11 of 32, 34.3%; p < 0.001), confirming higher atherosclerotic burden. The difference was particularly profound for microcalcifications, which was 17-fold higher in smokers compared to non-smokers. In vitro, nicotine induced human primary VSMC calcification, increased osteogenic gene expression (Runx2, Osx, BSP and OPN), and extracellular vesicle (EV) secretion. The pro-calcifying effects of nicotine were mediated by Ca2+-dependent Nox5. SiRNA knock-down of Nox5 inhibited nicotine-induced EV release and calcification. Moreover, pre-treatment of hVSMCs with vitamin K2 ameliorated nicotine-induced intracellular oxidative stress, EV secretion, and calcification. Using nicotinic acetylcholine receptor (nAChR) blockers α-bungarotoxin and hexamethonium bromide we found that the effects of nicotine on intracellular Ca2+ and oxidative stress were mediated by α7 and α3 nAChR. Finally, we showed that Nox5 expression was higher in carotid arteries of smokers and correlated with calcification levels in these vessels.
Conclusion
In this study we provide evidence that nicotine induces Nox5-mediated pro-calcific processes as novel mechanism of increased atherosclerotic calcification. We identified that activation of α7 and α3 nAChR by nicotine increases intracellular Ca2+ and initiates calcification of hVSMCs through increased Nox5 activity, leading to oxidative stress-mediated EV release. Identifying the role of Nox5-induced oxidative stress opens novel avenues for diagnosis and treatment of smoking-induced cardiovascular disease.

© The Author(s) 2021. Published by Oxford University Press on behalf of the European Society of Cardiology.

Cardiovasc Res: 16 Jul 2021; epub ahead of print
Petsophonsakul P, Burgmaier M, Willems B, Heeneman S, ... Furmanik M, Schurgers L
Cardiovasc Res: 16 Jul 2021; epub ahead of print | PMID: 34273166
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Abstract

Bcar1/p130Cas is essential for ventricular development and neural crest cell remodelling of the cardiac outflow tract.

Mahmoud M, Evans I, Wisniewski L, Tam Y, ... Scambler P, Zachary I
Aim
The adapter protein p130Cas, encoded by the Bcar1 gene, is a key regulator of cell movement, adhesion, and cell cycle control in diverse cell types. Bcar1 constitutive knockout mice are embryonic lethal by embryonic days (E) 11.5-12.5, but the role of Bcar1 in embryonic development remains unclear. Here, we investigated the role of Bcar1 specifically in cardiovascular development and defined the cellular and molecular mechanisms disrupted following targeted Bcar1 deletions.
Methods and results
We crossed Bcar1 floxed mice with Cre transgenic lines allowing for cell-specific knockout either in smooth muscle and early cardiac tissues (SM22-Cre), mature smooth muscle cells (smMHC-Cre), endothelial cells (Tie2-Cre), second heart field cells (Mef2c-Cre), or neural crest cells (NCC) (Pax3-Cre) and characterised these conditional knock outs using a combination of histological and molecular biology techniques.Conditional knockout of Bcar1 in SM22-expressing smooth muscle cells and cardiac tissues (Bcar1SM22KO) was embryonically lethal from E14.5-15.5 due to severe cardiovascular defects, including abnormal ventricular development and failure of outflow tract (OFT) septation leading to a single outflow vessel reminiscent of persistent truncus arteriosus. SM22-restricted loss of Bcar1 was associated with failure of OFT cushion cells to undergo differentiation to septal mesenchymal cells positive for SMC-specific α-actin, and disrupted expression of proteins and transcription factors involved in epithelial-to-mesenchymal transformation (EMT). Furthermore, knockout of Bcar1 specifically in NCC (Bcar1PAX3KO) recapitulated part of the OFT septation and aortic sac defects seen in the Bcar1SM22KO mutants, indicating a cell-specific requirement for Bcar1 in NCC essential for OFT septation. In contrast, conditional knockouts of Bcar1 in differentiated smooth muscle, endothelial cells, and second heart field cells survived to term and were phenotypically normal at birth and post-natally.
Conclusions
Our work reveals a cell-specific requirement for Bcar1 in NCC, early myogenic and cardiac cells, essential for OFT septation, myocardialisation and EMT/cell cycle regulation and differentiation to myogenic lineages.
Translational perspective
The molecular pathways coordinating cardiogenesis and the remodelling of the OFT are complex, and dysregulation of these pathways causes human heart defects. Our findings highlight a specific requirement for Bcar1 essential for cardiogenesis. Furthermore, the failure of OFT septation in Bcar1SM22KO mice resembles persistent truncus arteriosus (PTA), a feature of several human congenital heart diseases, including DiGeorge Syndrome. Our findings have implications for the mechanisms underlying the pathogenesis of congenital heart disease, and suggest that mice with conditional Bcar1 deletions may be useful models for dissecting mechanisms involved in the pathogenesis of human heart defects.

© The Author(s) 2021. Published by Oxford University Press on behalf of the European Society of Cardiology.

Cardiovasc Res: 15 Jul 2021; epub ahead of print
Mahmoud M, Evans I, Wisniewski L, Tam Y, ... Scambler P, Zachary I
Cardiovasc Res: 15 Jul 2021; epub ahead of print | PMID: 34270692
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Abstract

HDAC1: an environmental sensor regulating endothelial function.

Dunaway LS, Pollock JS
The histone deacetylases (HDACs) are a family of enzymes that catalyze lysine deacetylation of both histone and non-histone proteins. Here we review, summarize, and provide perspectives on the literature regarding one such HDAC, HDAC1, in endothelial biology. In the endothelium, HDAC1 mediates the effects of external and environmental stimuli by regulating major endothelial functions such as angiogenesis, inflammatory signaling, redox homeostasis, and nitric oxide signaling. Angiogenesis is most often, but not exclusively, repressed by endothelial HDAC1. The regulation of inflammatory signaling is more complex as HDAC1 promotes or suppresses inflammatory signaling depending upon the environmental stimuli. HDAC1 is protective in models of atherosclerosis where loss of HDAC1 results in increased cytokine and cell adhesion molecule abundance. In other models, HDAC1 promotes inflammation by increasing cell adhesion molecules and repressing claudin-5 expression. Consistently, from many investigations, HDAC1 decreases antioxidant enzyme expression and nitric oxide production in the endothelium. HDAC1 decreases antioxidant enzyme expression through the deacetylation of histones and transcription factors, and also regulates nitric oxide production through regulating both the expression and activity of nitric oxide synthase 3. The HDAC1-dependent regulation of endothelial function through the deacetylation of both histone and non-histone proteins ultimately impacts whole animal physiology and health.

Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2021. For permissions, please email: [email protected]

Cardiovasc Res: 14 Jul 2021; epub ahead of print
Dunaway LS, Pollock JS
Cardiovasc Res: 14 Jul 2021; epub ahead of print | PMID: 34264338
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Abstract

A Neuroligin Isoform Translated by circNlgn Contributes to Cardiac Remodeling.

Du WW, Xu J, Yang W, Wu N, ... Maksimovic K, Yang BB
Rationale: Fibrotic cardiac remodeling is a maladaptive response to acute or chronic injury that leads to arrythmia and progressive heart failure. The underlying mechanisms remain unclear.Objective: We performed high-throughput RNA sequencing to analyze circular RNA (circRNA) profile in human cardiac disease and developed transgenic mice to explore the roles of circNlgn.
Methods and results:
Using RNA sequencing, we found that circular neuroligin RNA (circNlgn) was highly upregulated in myocardial tissues of patients with selected congenital heart defects with cardiac overload. Back-splicing of the neuroligin gene led to the translation of a circular RNA-derived peptide (Nlgn173) with a 9-amino-acid nuclear localization motif. Binding of this motif to the structural protein LaminB1 facilitated the nuclear localization of Nlgn173. CHIP analysis demonstrated subsequent binding of Nlgn173 to both ING4 and C8orf44-SGK3 promoters, resulting in aberrant collagen deposition, cardiac fibroblast proliferation, and reduced cardiomyocyte viability. Three-dimensional ultrasound imaging of circNlgn transgenic mice showed impaired left ventricular function, with further impairment when subjected to left ventricular pressure overload compared to wild type mice. Nuclear translocation of Nlgn173, dysregulated expression of ING4 and C8orf44-SGK3, and immunohistochemical markers of cardiac fibrosis were detected in a panel of 145 patient specimens. Phenotypic changes observed in left ventricular pressure overload and transgenic mice were abrogated with silencing of circNlgn or its targets ING4 and SGK3. Conclusions: We show that a circular RNA can be translated into a novel protein isoform. Dysregulation of this process contributes to fibrosis and heart failure in cardiac overload-induced remodeling. This mechanism may hold therapeutic implications for cardiac disease.




Circ Res: 14 Jul 2021; epub ahead of print
Du WW, Xu J, Yang W, Wu N, ... Maksimovic K, Yang BB
Circ Res: 14 Jul 2021; epub ahead of print | PMID: 34261347
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Abstract

Interleukin-5-induced eosinophil population improves cardiac function after myocardial infarction.

Xu JY, Xiong YY, Tang RJ, Jiang WY, ... Li XD, Yang YJ
Aims
Interleukin (IL)-5 mediates the development of eosinophils (EOS) that are essential for tissue post-injury repair. It remains unknown whether IL-5 plays a role in heart repair after myocardial infarction (MI). This study aims to test whether IL-5-induced EOS population promotes the healing and repair process post-MI and to reveal the underlying mechanisms.
Method and results
MI was induced by permanent ligation of the left anterior descending coronary artery in wild-type C57BL/6 mice. Western blot and real-time polymerase chain reaction revealed elevated expression of IL-5 in the heart at 5 days post-MI. Immunohistostaining indicated that IL-5 was secreted mainly from macrophages and type 2 innate lymphoid cells in the setting of experimental MI. External supply of recombinant mouse IL-5 (20 min, 1 day, and 2 days after MI surgery) reduced the infarct size and increased ejection fraction and angiogenesis in the border zone. A significant expansion of EOS was detected in both the peripheral blood and infarcted myocardium after IL-5 administration. Pharmacological depletion of EOS by TRFK5 pretreatment muted the beneficial effects of IL-5 in MI mice. Mechanistic studies demonstrated that IL-5 increased the accumulation of CD206+ macrophages in infarcted myocardium at 7 days post-MI. In vitro co-culture experiments showed that EOS shifted bone marrow-derived macrophage polarization towards the CD206+ phenotypes. This activity of EOS was abolished by IL-4 neutralizing antibody, but not IL-10 or IL-13 neutralization. Western blot analyses demonstrated that EOS promoted the macrophage downstream signal transducer and activator of transcription 6 (STAT6) phosphorylation.
Conclusion
IL-5 facilitates the recovery of cardiac dysfunction post-MI by promoting EOS accumulation and subsequent CD206+ macrophage polarization via the IL-4/STAT6 axis.
Translational perspective
Accumulating evidence suggests that modulation of innate and adaptive immune responses is a promising therapeutic strategy for myocardial infarction. In this study, we demonstrate that IL-5 exerts cardioprotective effects on infarcted myocardium by promoting eosinophil accumulation and subsequent CD206+ macrophage polarization via the IL-4/STAT6 axis. Hence, regulation of cardiac IL-5 level or eosinophil count may become a therapeutic approach for post-myocardial infarction cardiac repair in humans.

Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2021. For permissions please email: [email protected]

Cardiovasc Res: 13 Jul 2021; epub ahead of print
Xu JY, Xiong YY, Tang RJ, Jiang WY, ... Li XD, Yang YJ
Cardiovasc Res: 13 Jul 2021; epub ahead of print | PMID: 34259869
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Abstract

Focus on the road to modelling cardiomyopathy in muscular dystrophy.

Canonico F, Chirivi M, Maiullari F, Milan M, ... Bearzi C, D\'Amario D
Alterations in the DMD gene, which codes for the protein dystrophin, cause forms of dystrophinopathies such as Duchenne muscular dystrophy, an X-linked disease. Cardiomyopathy linked to DMD mutations is becoming the leading cause of death in patients with dystrophinopathy. Since phenotypic pathophysiological mechanisms are not fully understood, the improvement and development of new disease models, considering their relative advantages and disadvantages, is essential. The application of genetic engineering approaches on induced pluripotent stem cells, such as gene editing technology, enables the development of physiologically relevant human cell models for in vitro dystrophinopathy studies. The combination of induced pluripotent stem cells-derived cardiovascular cell types and 3 D bioprinting technologies hold great promise for the study of dystrophin-linked cardiomyopathy. This combined approach enables the assessment of responses to physical or chemical stimuli, and the influence of pharmaceutical approaches. The critical objective of in vitro microphysiological systems is to more accurately reproduce the microenvironment observed in vivo. Ground-breaking methodology involving the connection of multiple microphysiological systems comprised of different tissues would represent a move toward precision body-on-chip disease modelling could lead to a critical expansion in what is known about inter-organ responses to disease and novel therapies that have the potential to replace animal models. In this review, we will focus on the generation, development, and application of current cellular, animal and potential for bio-printed models, in the study of the pathophysiological mechanisms underlying dystrophin-linked cardiomyopathy in the direction of personalized medicine.

Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2021. For permissions please email: [email protected]

Cardiovasc Res: 11 Jul 2021; epub ahead of print
Canonico F, Chirivi M, Maiullari F, Milan M, ... Bearzi C, D'Amario D
Cardiovasc Res: 11 Jul 2021; epub ahead of print | PMID: 34254111
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Impact:
Abstract

Disrupted Resolution Mechanisms Favor Altered Phagocyte Responses in Covid-19.

Koenis DS, Beegun I, Jouvene C, Aguirre GA, ... Pfeffer P, Dalli J
Rationale: Resolution mechanisms are central in both the maintenance of homeostasis and the return to catabasis following tissue injury and/or infections. Amongst the pro-resolving mediators, the essential fatty acid-derived specialized pro-resolving lipid mediators (SPM) govern immune responses to limit disease severity. Notably, little is known about the relationship between the expression and activity of SPM pathways, circulating phagocyte function and disease severity in patients infected with novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) leading to coronavirus disease 2019 (COVID-19). Objective: Herein, we investigated the link between circulating SPM concentrations and phagocyte activation status and function in COVID-19 patients (n=39) compared to healthy (n=12) and post-COVID-19 (n=8) volunteers.Methods and Results:Lipid mediator profiling demonstrated that plasma SPM concentrations were upregulated in patients with mild COVID-19 and are downregulated in those with severe disease. SPM concentrations were correlated with both circulating phagocyte activation status and function. Perturbations in plasma SPM concentrations and phagocyte activation were retained after the resolution of COVID-19 clinical symptoms. Treatment of patients with dexamethasone upregulated both the expression of SPM biosynthetic enzymes in circulating phagocytes and plasma concentration of these mediators. Furthermore, incubation of phagocytes from COVID-19 patients with SPM rectified their phenotype and function. This included a downregulation in the expression of activation markers, a decrease in the Tissue Factor and inflammatory cytokine expression, and an upregulation of bacterial phagocytosis.Conclusions: The present findings suggest that downregulation of systemic SPM concentrations is linked with both increased disease severity and dysregulated phagocyte function. They also identify the upregulation of these mediators by dexamethasone as a potential mechanism in host protective activities elicited by this drug in COVID-19 patients. Taken together, our findings elucidate a role for altered resolution mechanisms in the disruption of phagocyte responses and the propagation of systemic inflammation in COVID-19.



Circ Res: 08 Jul 2021; epub ahead of print
Koenis DS, Beegun I, Jouvene C, Aguirre GA, ... Pfeffer P, Dalli J
Circ Res: 08 Jul 2021; epub ahead of print | PMID: 34238021
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Impact:
Abstract

Loss of Hepatic Angiotensinogen Attenuates Sepsis-Induced Myocardial Dysfunction.

Rong J, Tao X, Lin Y, Zheng H, ... Xu Y, Wang J
Rationale: The renin-angiotensin system (RAS) is a complex regulatory network that maintains normal physiological functions. The role of the RAS in sepsis-induced myocardial dysfunction (SIMD) is poorly defined. Angiotensinogen (AGT) is the unique precursor of the RAS and gives rise to all angiotensin peptides. The effects and mechanisms of AGT in development of SIMD have not been defined. Objective: To determine a role of AGT in SIMD and investigate the underlying mechanisms.
Methods and results:
Either intraperitoneal injection of lipopolysaccharide (LPS) or cecal ligation and puncture (CLP) significantly enhanced AGT abundances in liver, heart, and plasma. Deficiency of hepatocyte-derived AGT (hepAGT), rather than cardiomyocyte-derived AGT (carAGT), alleviated septic cardiac dysfunction in mice and prolonged survival time. Further investigations revealed that the effects of hepAGT on SIMD were partially associated with augmented angiotensin II (AngII) production in circulation. In addition, hepAGT was internalized by LDL receptor-related protein 1 (LRP1) in cardiac fibroblasts (CF), and subsequently activated NLRP3 inflammasome via an AngII-independent pathway, ultimately promoting SIMD by suppressing Sarco(endo)plasmic reticulum Ca(2+)-ATPase 2a (SERCA2a) abundances in cardiomyocytes (CM). Conclusions: HepAGT promoted SIMD via both AngII-dependent and AngII-independent pathways. We identified a liver-heart axis by which AGT regulated development of SIMD. Our study may provide a potential novel therapeutic target for SIMD.




Circ Res: 08 Jul 2021; epub ahead of print
Rong J, Tao X, Lin Y, Zheng H, ... Xu Y, Wang J
Circ Res: 08 Jul 2021; epub ahead of print | PMID: 34238019
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Impact:
Abstract

Glucose Derivative Induced Vasculopathy in Children on Chronic Peritoneal Dialysis.

Bartosova M, Zhang C, Schaefer B, Herzog R, ... Kratochwill K, Schmitt CP
Rationale
Patients with chronic kidney disease (CKD) have an exceedingly high cardiovascular risk; which further increases in patients on peritoneal dialysis (PD). The pathophysiological role of reactive metabolites accumulating in CKD such as glucose degradation products (GDP) is uncertain.
Objective
Delineating the impact of GDP present in PD fluids in accelerated vasculopathy development in patients with CKD.
Methods and results
Omental and parietal peritoneal tissues were obtained from 107 children with CKD prior to dialysis, and 90 children on chronic PD with PD fluids containing very low or high concentrations of GDP. Omental arterioles, protected from local PD fluid exposure by surrounding fat, were microdissected for multi-omics analyses. High-GDP exposed omental arterioles exhibited three-fold higher advanced glycation endproduct concentrations and upregulated genes involved in cell death/apoptosis and suppressed genes related to cell viability/survival, cytoskeleton organization and immune response biofunctions. Vasculopathy associated canonical pathways concordantly regulated on gene- and protein level with high-GDP exposure included cell death/proliferation, apoptosis, cytoskeleton organization, metabolism and detoxification, cell junction signaling, and immune response. Parietal peritoneal arterioles of patients exposed to high-GDP fluids exhibited lumen narrowing compared to patients with CKD5 and patients on low-GDP PD, intima thickness was increased. Protein quantification verified increased proapoptotic activity and cytoskeleton disintegration, single-molecule-localization microscopy demonstrated arteriolar endothelial zonula occludens-1 (ZO-1) disruption. Absolute and per endoluminal surface length, arteriolar endothelial cell counts inversely correlated with GDP exposure, caspase-3, TGF-ß induced pSMAD2/3, interleukin-6, ZO-1 abundance and lumen narrowing. In vitro, 3,4-dideoxyglucosone-3-ene reduced lamin-A/C and membrane ZO-1 assembly, increased pSMAD2/3, and ionic and 4- and 10kDa permeability of arterial endothelial cells.
Conclusions
Our findings indicate a fundamental role of GDP in PD associated vasculopathy, exerted by endothelial cell junction and cytoskeleton disruption, and induction of apoptosis. They should redirect the focus of research and intervention on targeting reactive metabolite overload in CKD and PD.



Circ Res: 07 Jul 2021; epub ahead of print
Bartosova M, Zhang C, Schaefer B, Herzog R, ... Kratochwill K, Schmitt CP
Circ Res: 07 Jul 2021; epub ahead of print | PMID: 34233458
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Impact:
Abstract

Role of VLDL receptor in atherogenesis.

Takahashi S
Purpose of review
My group previously discovered and characterized the rabbit and human VLDL receptors. For more than 30 years, I have focused on research regarding the functions of VLDL receptors in the fields of lipoprotein metabolism and atherogenesis. In this review, I introduce the roles of VLDL receptors in lipoprotein metabolism under physiological conditions and in atherogenesis under nonphysiological conditions.
Recent findings
I propose that the VLDL receptor plays key roles in the metabolism of postprandial remnant lipoproteins in concert with lipoprotein lipase (LPL). Furthermore, I propound a new mechanism for macrophage foam cell formation via VLDL receptors by remnant lipoproteins and lipoprotein(a) [Lp(a)] in addition to scavenger receptor pathways.
Summary
The VLDL receptor is a so-called macrophage β-VLDL receptor, which is involved in macrophage foam cell formation by remnant lipoproteins. Furthermore, Lp(a) is a VLDL receptor ligand and is directly taken up through macrophage VLDL receptors for macrophage foam cell formation. Additionally, the roles of VLDL receptors in atherogenesis are canvassed.
Supplementary video abstract
http://links.lww.com/COL/A21.

Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.

Curr Opin Lipidol: 31 Jul 2021; 32:219-225
Takahashi S
Curr Opin Lipidol: 31 Jul 2021; 32:219-225 | PMID: 34227576
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Abstract

ANXA7 Regulates Platelet Lipid Metabolism and Ca Release in Arterial Thrombosis.

Manke MC, Geue S, Coman C, Peng B, ... Ahrends R, Borst O
Rationale: Platelet activation after contact to subendothelial collagen leads to acute arterial thrombosis. Annexin A7 (ANXA7) is a phospholipid-binding protein participating in the regulation of intracellular Ca2+ and exocytosis. Objective: The present study aimed to determine the role of ANXA7 in platelet Ca2+ signaling and lipid metabolism during platelet activation in arterial thrombosis using the ANXA7 inhibitor ABO and gene-targeted mice lacking Anxa7 (Anxa7-/-).
Methods and results:
ANXA7 is strongly expressed in platelets. Functionally, luminescence aggregometry revealed significantly abrogated aggregation and secretion of ABO-treated or Anxa7-/- platelets when compared with untreated or Anxa7+/+ platelets after activation with collagen or the GPVI-specific agonist collagen-related peptide (CRP). Furthermore, while both thrombus formation on collagen-coated surfaces under high arterial shear rates in ABO-treated or Anxa7-deficient whole blood, and thrombotic vascular occlusion after FeCl3-induced injury in vivo in Anxa7-/- bone marrow chimeric mice were significantly diminished, no prolongation of bleeding time was observed in ABO-treated or Anxa7-/- mice. Fura-2-AM spectrofluorimetry unraveled a blunted [Ca2+]i increase in ABO-treated or Anxa7-/- platelets after GPVI stimulation. Due to an abolished PLCy2 phosphorylation, Anxa7-/- platelets displayed abrogated intracellular Ca2+ mobilization following CRP-dependent platelet activation. Quantitative lipidomics analysis further revealed that ANXA7 critically affects platelet oxylipin metabolism following GPVI-dependent platelet activation. Anxa7-/- platelets showed a significantly reduced generation of several bioactive metabolites, particularly TxA2 and 12(S)-HETE. Finally, defective PLCy2 phosphorylation and blunted [Ca2+]i increase in Anxa7-/- platelets could be rescued by exogenous addition of 12(S)-HETE, indicating that ANXA7 is a critical regulator of the platelet 12-lipoxygenase in GPVI-dependent platelet Ca2+ signaling during arterial thrombosis. Conclusions:The present study unravels ANXA7 as a regulator of oxylipin metabolism and Ca2+-dependent platelet activation downstream of GPVI. ANXA7 plays an important role in platelet signaling during arterial thrombosis and thus may reflect a promising target for novel antiplatelet strategies.




Circ Res: 27 Jun 2021; epub ahead of print
Manke MC, Geue S, Coman C, Peng B, ... Ahrends R, Borst O
Circ Res: 27 Jun 2021; epub ahead of print | PMID: 34176316
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Abstract

Cell senescence: basic mechanisms and the need for computational networks in vascular ageing.

Regnault V, Challande P, Pinet F, Li Z, Lacolley P
This review seeks to provide an update of the mechanisms of vascular cell senescence, from newly identified molecules to arterial ageing phenotypes, and finally to present a computational approach to connect these selected proteins in biological networks. We will discuss current key signalling and gene expression pathways by which these focus proteins and networks drive normal and accelerated vascular ageing. We also review the possibility that senolytic drugs, designed to restore normal cell differentiation and function, could effectively treat multiple age-related vascular diseases. Finally, we discuss how cell senescence is both a cause and a consequence of vascular ageing because of the possible feedback controls between identified networks.

Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2020. For permissions, please email: journals.pe[email protected]

Cardiovasc Res: 06 Jul 2021; 117:1841-1858
Regnault V, Challande P, Pinet F, Li Z, Lacolley P
Cardiovasc Res: 06 Jul 2021; 117:1841-1858 | PMID: 33206947
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Abstract

Genetic determinants of LDL cholesterol and risk of intracerebral haemorrhage.

Szejko N, Kirsch E, Falcone GJ
Purpose of review
The role of lipids in spontaneous, nontraumatic intracerebral haemorrhage (ICH) remains controversial, as some studies suggest that lower levels of total and LDL cholesterol could increase the risk of this disease. Because of their random assortment during meiosis, genetic variants known to associate with lipid levels can be used as instruments to evaluate this relationship from a causal perspective. The purpose of this review is to summarize the existing literature related to genetically determined LDL cholesterol levels and risk of ICH.
Recent findings
A number of studies have demonstrated that lower LDL levels are associated with a higher risk of ICH and a higher burden of neuroimaging markers of cerebral small vessel disease, such as microbleeds and white matter hyperintensity volume. As for genetically elevated lipid levels, several studies confirmed an inverse association between LDL levels and ICH. However, a number of observational studies and large meta-analyses of clinical trials of statins have failed to show such association.
Summary
Observational studies and clinical trials of statins have yielded inconsistent results regarding a possible link between LDL levels and the risk of ICH. Genetic studies focused on genetically elevated LDL levels and risk of ICH have, for the most, found an inverse association.

Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.

Curr Opin Lipidol: 31 Jul 2021; 32:244-248
Szejko N, Kirsch E, Falcone GJ
Curr Opin Lipidol: 31 Jul 2021; 32:244-248 | PMID: 34010223
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Impact:
Abstract

New insights in statins affecting atheromatous plaque macrophages.

Härdtner C, Ehlert CA, Hilgendorf I
Purpose of review
Macrophages are key protagonists of atherosclerotic plaque development and hence represent targets of therapeutic intervention. Statins are the most potent widely used atheroprotective drugs. Therefore, whether and how statins influence atheromatous plaque macrophages has remained at the center of cardiovascular research for decades.
Recent findings
Because statins are capable of regulating macrophage functions in cell culture, largely independent of their cholesterol-lowering effect, it was assumed that these pleiotropic effects operate in vivo as well. Recent experimental data, in line with clinical observations, indicate, however, that statins do not interact with macrophages in atherosclerotic plaques, directly, and instead control their functions and assembly indirectly via changes to circulating lipid levels and endothelial activation.
Summary
Statin-mediated lipid lowering induces plaque regression which is characterized by a decline in plaque macrophage content. Understanding how statins provoke this protective phenotype may inspire conceptually new therapeutic approaches in cardiovascular medicine.

Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.

Curr Opin Lipidol: 31 Jul 2021; 32:258-264
Härdtner C, Ehlert CA, Hilgendorf I
Curr Opin Lipidol: 31 Jul 2021; 32:258-264 | PMID: 34054106
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Impact:
Abstract

Silencing Myeloid Netrin-1 Induces Inflammation Resolution and Plaque Regression.

Schlegel MP, Sharma M, Brown EJ, Newman AA, ... Fisher EA, Moore KJ
Rationale: Therapeutic efforts to decrease atherosclerotic cardiovascular disease risk have focused largely on reducing atherogenic lipoproteins, yet lipid lowering therapies alone are insufficient to fully regress plaque burden. We postulate that arterial repair requires resolution of a maladaptive immune response, and that targeting factors that hinder inflammation resolution will facilitate plaque regression. Objective: The guidance molecule netrin-1 is secreted by macrophages in atherosclerotic plaques, where it sustains inflammation by enhancing macrophage survival and blocking macrophage emigration. We tested whether silencing netrin-1 in advanced atherosclerosis could resolve arterial inflammation and regress plaques.
Methods and results:
To temporally silence netrin-1 in myeloid cells, we generated genetically modified mice in which Ntn1 could be selectively deleted in monocytes and macrophages using a tamoxifen-induced CX3CR1-driven cre-recombinase (Ntn1fl/flCx3cr1creERT2+) and littermate control mice (Ntn1fl/flCx3cr1WT). Mice were fed western diet in the setting of hepatic PCSK9 overexpression to render them atherosclerotic, and then treated with tamoxifen to initiate deletion of myeloid netrin-1 (MøΔNtn1) or not in controls (MøWT). Morphometric analyses performed 4 weeks later showed that myeloid Ntn1 silencing reduced plaque burden in the aorta (-50%) and plaque complexity in the aortic root. Monocyte-macrophage tracing experiments revealed lower monocyte recruitment, macrophage retention, and proliferation in MøΔNtn1 compared to MøWT plaques, indicating a restructuring of monocyte-macrophage dynamics in the artery wall upon netrin-1 silencing. Single cell RNA-sequencing of aortic immune cells prior to and after netrin-1 silencing revealed upregulation of gene pathways involved in macrophage phagocytosis and migration, including the Ccr7 chemokine receptor signaling pathway required for macrophage emigration from plaques and atherosclerosis regression. Additionally, plaques from MøΔNtn1 mice showed hallmarks of inflammation resolution, including higher levels of pro-resolving macrophages, interleukin-10, and efferocytosis, as compared to plaques from MøWT mice. Conclusions: Our data show that targeting netrin-1 in advanced atherosclerosis ameliorates atherosclerotic inflammation and promotes plaque regression.




Circ Res: 21 Jul 2021; epub ahead of print
Schlegel MP, Sharma M, Brown EJ, Newman AA, ... Fisher EA, Moore KJ
Circ Res: 21 Jul 2021; epub ahead of print | PMID: 34289717
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Abstract

Brain Hypoxia Is Associated With Neuroglial Injury in Humans Post-Cardiac Arrest.

Hoiland RL, Ainslie PN, Wellington CL, Cooper J, ... Griesdale D, Sekhon M
Rationale: Secondary brain hypoxia portends significant mortality in ischemic brain diseases, yet our understanding of hypoxic ischemic brain injury (HIBI) pathophysiology in humans remains rudimentary. Objective: To quantify the impact of secondary brain hypoxia on injury to the neurovascular unit in patients with HIBI.
Methods and results:
We conducted a prospective interventional study of invasive neuromonitoring in 18 post-cardiac arrest patients with HIBI. The partial pressures of brain tissue O2 (PbtO2) and intracranial pressure were directly measured via intra-parenchymal micro-catheters. To isolate the cerebrovascular bed, we conducted paired sampling of arterial and jugular venous bulb blood and calculated the trans-cerebral release of biomarkers of neurovascular injury and inflammation in the HIBI patients and 14 healthy volunteers for control comparisons. Ten HIBI patients exhibited secondary brain hypoxia (PbtO2<20mmHg), while eight exhibited brain normoxia (PbtO2≥20mmHg). In the patients with secondary brain hypoxia, we observed active cerebral release of glial fibrillary acidic protein (-161[ -3695 - -75] pg/mL; P=0.0078), neurofilament light chain (-231[-370 - -11] pg/mL; P=0.010), total tau (-32[-310 - -3] pg/mL; P=0.0039), neuron specific enolase (-14890[-148813 - -3311] pg/mL; P=0.0039), and ubiquitin carboxy-terminal hydrolase L1 (-14.7[-37.7 - -4.1] pg/mL; P=0.0059) indicating de novo neuroglial injury. This injury was unrelated to the systemic global ischemic burden or cerebral endothelial injury but rather was associated with cerebral release of interleukin-6 (-10.3[-43.0 - -4.2] pg/mL; P=0.0039). No cerebral release of the aforementioned biomarkers was observed in HIBI patients with brain normoxia or the healthy volunteers. Hyperosmolar therapy in the patients with secondary brain hypoxia reduced the partial pressure of jugular venous O2-to-PbtO2 gradient (39.6[34.1-51.1] vs. 32.0[24.5-39.2] mmHg; P=0.0078) and increased PbtO2 (17.0[9.1-19.7] vs. 20.2[11.9-22.7] mmHg; P=0.039) suggesting improved cerebrovascular-to-parenchymal O2 transport. Conclusions: Secondary brain hypoxia is associated with de novo neuroglial injury and cerebral release of interleukin-6. Mitigating cerebrovascular-to-parenchymal limitations to O2 transport is a promising therapeutic strategy for HIBI patients with secondary brain hypoxia.




Circ Res: 20 Jul 2021; epub ahead of print
Hoiland RL, Ainslie PN, Wellington CL, Cooper J, ... Griesdale D, Sekhon M
Circ Res: 20 Jul 2021; epub ahead of print | PMID: 34287000
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Abstract

Collecting duct-specific knockout of sphingosine-1-phosphate receptor 1 aggravates DOCA-salt hypertension in mice.

Hu G, Zhu Q, Wang W, Xie D, ... Ritter JK, Li N
Objective
We have previously reported that renal medullary sphingosine-1-phosphate (S1P) regulates sodium excretion via the S1P type-1 receptor (S1PR1). As S1PR1 is predominantly expressed in collecting ducts (CD), the present study tested the hypothesis that the CD-S1PR1 pathway plays a critical role in sodium excretion and contributes to salt-sensitive hypertension.
Methods
CD-specific S1PR1 knockout mice were generated by crossing aquaporin-2-Cre mice with S1PR1-floxed mice. Renal sodium excretion and arterial pressure were compared between wild type and KO mice in response to high-salt challenges and treatment of deoxycorticosterone acetate (DOCA) salt.
Results
Protein levels of renal medullary S1PR1 were increased by 100% after high-salt intake, whereas DOCA treatment with high-salt intake blocked the increase of S1PR1 levels. Urinary sodium excretions in knockout mice were decreased by 60% compared with wild type mice after acute intravenous sodium loading (0.84 ± 0.16 vs. 2.22 ± 0.62 μmole/min per g kwt). The pressure natriuresis was impaired in knockout mice compared with wild type mice (4.32 ± 1.04 vs. 8.73 ± 0.19 μmole/min per g kwt). The chronic high-salt intake-induced positive sodium balance was enhanced in knockout mice compared with wild type mice (5.27 ± 0.39 vs. 2.38 ± 1.04 mmol/100 g BW per 24 h). After 10-day DOCA-salt treatment, knockout mice developed more severe hypertension than wild type mice (SBP 142 ± 8 vs. 115 ± 4 mmHg).
Conclusion
The deletion of CD-S1PR1 reduced sodium excretion, promoted sodium retention, and accelerated DOCA-salt-induced salt-sensitive hypertension, suggesting that the CD-S1PR1 signaling is an important antihypertensive pathway by promoting sodium excretion and that impairment of renal medullary S1PR1 may represent a novel mechanism for salt-sensitive hypertension.

Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.

J Hypertens: 31 Jul 2021; 39:1559-1566
Hu G, Zhu Q, Wang W, Xie D, ... Ritter JK, Li N
J Hypertens: 31 Jul 2021; 39:1559-1566 | PMID: 33534341
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Abstract

Upregulation of miR-29a suppressed the migration and invasion of trophoblasts by directly targeting LOXL2 in preeclampsia.

Xu XH, Tang LC, Hao F, Jin LP
Objective
Preeclampsia is a pregnancy-specific disorder that is a major cause of maternal and foetal morbidity and mortality, with a prevalence of 6-8% of pregnancies. Although the downregulation of lysyl oxidase (LOX) and LOX-like protein 2 (LOXL2), which leads to reduced trophoblast cell migration and invasion through activation of the TGF-β1/Smad3/collagen pathway, is relevant to preeclampsia, the mechanisms regulating differences in the gene expression of LOX and LOXL2 in placentas are not yet understood. This study aimed to investigate the mechanisms regulating differences in the gene expression of LOX and LOXL2 in placentas.
Methods
The expression of miRNAs, LOX and LOXL2 in preeclamptic placentas and control placentas was analysed by qPCR. Localisation of miR29a and LOXL2 in preeclamptic placentas was performed by RNA-Fluorescence in-situ hybridization assay. The direct regulation of LOXL2 by miR-29a was assessed by dual-luciferase reporter assays in human extravillous trophoblast cells (HTR8/SVneo). Cell migration and invasion were evaluated by Transwell assays in HTR8/SVneo cells.
Results
miR-29a expression was upregulated in preeclamptic placentas and negatively correlated with LOXL2 mRNA expression levels. RNA-Fluorescence in-situ hybridization assay revealed a clear overlap between miR-29a and LOXL2 in the placentas of preeclampic women. LOXL2 was a direct target gene of miR-29a, as confirmed by a dual-luciferase reporter assay in HTR8/SVneo trophoblast cells. miR-29a suppressed HTR8/SVneo trophoblast cell migration and invasion. LOXL2 overexpression reversed the inhibitory effects of miR-29a on HTR8/SVneo trophoblast cell migration and invasion.
Conclusion
Our results suggest that the upregulation of miR-29a suppresses the migration and invasion of HTR8/SVneo trophoblast cells by directly targeting LOXL2 in preeclampsia.

Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.

J Hypertens: 31 Jul 2021; 39:1642-1651
Xu XH, Tang LC, Hao F, Jin LP
J Hypertens: 31 Jul 2021; 39:1642-1651 | PMID: 33657581
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Abstract

Klotho supplementation attenuates blood pressure and albuminuria in murine model of IgA nephropathy.

Takenaka T, Hasan A, Marumo T, Kobori H, ... Ishii N, Hayashi M
Background
Klotho interacts with various membrane proteins, such as transforming growth factor-β (TGFβ) and insulin-like growth factor (IGF) receptors. The renal expression of klotho is diminished in chronic kidney disease.
Method
In this study, we assessed the effects of klotho supplementation on a murine model of IgA nephropathy. Twenty-four-week-old hyper serum IgA (HIGA) mice were subcutaneously injected daily with recombinant human klotho protein (20 μg/kg per day) or the vehicle. After 2 months, the mice were killed using an anesthesia overdose and their kidneys were harvested for analysis.
Results
Supplementation of exogenous klotho protein reduced SBP, albuminuria, 8-epi-prostaglandin F2α excretion, glomerular filtration rate, renal angiotensin II concentration, and angiotensinogen expression in HIGA mice. Additionally, it enhanced renal expression of superoxide dismutase (SOD) and renal klotho itself. The findings using laser-manipulated microdissection demonstrated that klotho supplementation reduced the glomerular expression of TGFβ, fibronectin, and IGF, and increased the glomerular expression of connexin (Cx) 40.
Conclusion
These results indicate that klotho supplementation reduces blood pressure by suppressing the renin--angiotensin system in HIGA mice. Klotho inhibits IGF signaling to preserve glomerular Cx40 levels, ameliorating albuminuria in HIGA mice. Klotho protein supplementation attenuates mesangial expansion by inhibiting TGFβ signaling in HIGA mice.

Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.

J Hypertens: 31 Jul 2021; 39:1567-1576
Takenaka T, Hasan A, Marumo T, Kobori H, ... Ishii N, Hayashi M
J Hypertens: 31 Jul 2021; 39:1567-1576 | PMID: 33758157
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Abstract

Pyridostigmine ameliorates preeclamptic features in pregnant rats by inhibiting tumour necrosis factor-α synthetsis and antagonizing tumour necrosis factor-α-related effects.

Issotina Zibrila A, Wang Z, Ali MA, Osei JA, ... Kang Y, Liu J
Objective
Preeclampsia is a hypertensive disorder of pregnancy marked by an excessive inflammatory response. The anti-inflammatory effect of pyridostigmine (PYR) was previously reported; however, its role in hypertensive pregnancies remains unclear. We hypothesized that PYR could attenuate increased blood pressure and other pathological features in preeclampsia models.
Methods
The expression of tumour necrosis factor (TNF)-α was evaluated in normal and preeclampsia pregnant women. PYR (20 mg/kg) was administered daily to reduced uterine perfusion pressure (RUPP) and TNF-α (150 ng/day) infused rats from gestation day 14 to GD19. In a cell culture experiment, the effect of acetylcholine (ACh) on TNF-α-stimulated primary human umbilical endothelial cells (HUVEC) was assessed.
Results
Preeclampsia women had higher placental TNF-α expression than normal pregnant women. Mean arterial pressure (MAP) in the RUPP group was higher than in the Sham group. PYR inhibited serum and placental acetylcholinesterase activity in rats, and reduced MAP, placental oxidative stress, apoptosis and inflammation in the RUPP group but not in the Sham group. In addition, PYR significantly attenuated the TNF-α-induced increase in MAP, placental oxidative stress and apoptosis. Moreover, TNF-α decreased cell viability and increased the number of TUNEL-positive nuclei of HUVEC, which could largely be abolished by ACh treatment.
Conclusion
Collectively, PYR ameliorated hypertension and other preeclampsia-like symptoms in rat models of preeclampsia not only by inhibiting the synthesis of TNF-α but also by acting against TNF-α-induced detrimental effects directly, which is worthy of further investigation and may be used as a potential agent for preeclampsia management.

Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.

J Hypertens: 05 Jul 2021; epub ahead of print
Issotina Zibrila A, Wang Z, Ali MA, Osei JA, ... Kang Y, Liu J
J Hypertens: 05 Jul 2021; epub ahead of print | PMID: 34232157
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Impact:
Abstract

Activation of the intestinal tissue renin--angiotensin system by transient sodium loading in salt-sensitive rats.

Ryuzaki M, Miyashita K, Sato M, Inoue H, ... Kinouchi K, Itoh H
Background
The renal tissue renin--angiotensin system is known to be activated by salt loading in salt-sensitive rats; however, the response in other organs remains unclear.
Method
Spontaneously hypertensive rats were subjected to normal tap water or transient high-salt-concentration water from 6 to 14 weeks of age and were thereafter given normal tap water. From 18 to 20 weeks of age, rats given water with a high salt concentration were treated with an angiotensin II type 1 receptor blocker, valsartan.
Results
Sustained blood pressure elevation by transient salt loading coincided with a persistent decrease in the fecal sodium content and sustained excess of the circulating volume in spontaneously hypertensive rats. Administration of valsartan sustainably reduced the blood pressure and normalized the fecal sodium levels. Notably, transient salt loading persistently induced the intestinal tissue renin--angiotensin system and enhanced sodium transporter expression exclusively in the small intestine of salt-sensitive rats, suggesting the potential connection of intestinal sodium absorption to salt sensitivity.
Conclusion
These results reveal the previously unappreciated contribution of the intestinal tissue renin--angiotensin system to sodium homeostasis and blood pressure regulation in the pathophysiology of salt-sensitive hypertension.

Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.

J Hypertens: 18 Jul 2021; epub ahead of print
Ryuzaki M, Miyashita K, Sato M, Inoue H, ... Kinouchi K, Itoh H
J Hypertens: 18 Jul 2021; epub ahead of print | PMID: 34285148
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Impact:
Abstract

Gender-affirming hormone treatment causes changes in gender phenotype in a 12-lead electrocardiogram.

Saito N, Nagahara D, Ichihara K, Masumori N, Miura T, Takahashi S
Background
Men and women have specific patterns in an electrocardiogram (ECG) differentiated by J-point elevation and ST-segment angle. Although gender-affirming hormone treatment is one of the treatments for gender dysphoria, its influence on an ECG has not been clarified yet.
Objective
The purpose of this study was to investigate ECG changes induced by gender-affirming hormone treatment.
Methods
The study population consisted of 29 transgender males and 8 transgender females and 37 age- and sex-matched cisgender females and males. Male pattern was defined as J-point elevation > 0.1 mV and ST-segment angle > 20° in precordial leads.
Results
In the comparison between 29 transgender males and cisgender females, the prevalence of the male pattern (89.7% vs 6.9%; P < .001), prevalence of the early repolarization pattern (51.7% vs 17.2%; P = .01), J-point elevation (leads V1-V6), T-wave amplitudes (leads V1-V6), QRS amplitudes (leads II, III, V1-V6), and P-wave amplitudes (leads V1-V3) were significantly higher in transgender males. The prevalence of the male pattern was lower in transgender females than in cisgender males (25.0% vs 87.5%; P = .04). In the analysis of transgender males for whom ECGs were available before and after gender-affirming hormone treatment (n = 13), J-point elevation and T-wave amplitudes significantly increased after gender-affirming hormone treatment, leading to a higher prevalence of the male pattern (23.1% vs 92.3%; P < .001). The prevalence of the early repolarization pattern and QRS amplitudes also significantly increased after the treatment, but the augmentation of P-wave amplitudes was modest.
Conclusion
Gender-affirming hormone treatment for gender dysphoria is accompanied by a change in ECG phenotype toward affirming gender, in which change in androgen level may be involved.

Copyright © 2021 Heart Rhythm Society. Published by Elsevier Inc. All rights reserved.

Heart Rhythm: 29 Jun 2021; 18:1203-1209
Saito N, Nagahara D, Ichihara K, Masumori N, Miura T, Takahashi S
Heart Rhythm: 29 Jun 2021; 18:1203-1209 | PMID: 33706005
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Impact:
Abstract

Arrhythmia-induced cardiomyopathy: A potentially reversible cause of refractory cardiogenic shock requiring venoarterial extracorporeal membrane oxygenation.

Hékimian G, Paulo N, Waintraub X, Bréchot N, ... Gandjbakhch E, Luyt CE
Background
The most severe form of arrhythmia-induced cardiomyopathy in adults- refractory cardiogenic shock requiring mechanical circulatory support-has rarely been reported.
Objective
The purpose of this study was to describe the management of critically ill patients admitted for acute, nonischemic, or worsening of previously known cardiac dysfunction and recent-onset supraventricular arrhythmia who developed refractory cardiogenic shock requiring venoarterial extracorporeal membrane oxygenation (VA-ECMO).
Methods
This study is a retrospective analysis of prospectively collected data.
Results
Between 2004 and 2018, 35 patients received VA-ECMO for acute, nonischemic cardiogenic shock and recent supraventricular arrhythmia (77% atrial fibrillation [AF]). Cardiogenic shock was the first disease manifestation in 21 patients (60%). Characteristics at ECMO implantation [median (interquartile range)] were Sequential Organ Failure Assessment score 10 (7-13); inotrope score 29 (11-80); left ventricular ejection (LVEF) fraction 10% (10%-15%); and lactate level 8 (4-11) mmol/L. For 12 patients, amiodarone and/or electric cardioversion successfully reduced arrhythmia, improved LVEF, and enabled weaning off VA-ECMO; 11 had long-term survival without transplantation or long-term assist device. Eight patients experiencing arrhythmia-reduction failure underwent ablation procedures (7 atrioventricular node [AVN] with pacing, 1 atrial tachycardia) and were weaned off VA-ECMO; 7 survived. Of the remaining 15 patients without arrhythmia reduction or ablation, only the 6 bridged to heart transplantation or left ventricular (LV) assist device survived.
Conclusion
Arrhythmia-induced cardiomyopathy, mainly AF-related, is an underrecognized cause of refractory cardiogenic shock and should be considered in patients with nonischemic cardiogenic shock and recent-onset supraventricular arrhythmia. VA-ECMO support allowed safe arrhythmia reduction or rate control by AVN ablation while awaiting recovery, even among those with severe LV dilation.

Copyright © 2021 Heart Rhythm Society. Published by Elsevier Inc. All rights reserved.

Heart Rhythm: 29 Jun 2021; 18:1106-1112
Hékimian G, Paulo N, Waintraub X, Bréchot N, ... Gandjbakhch E, Luyt CE
Heart Rhythm: 29 Jun 2021; 18:1106-1112 | PMID: 33722763
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Impact:
Abstract

Effect of ivabradine on cardiac arrhythmias: Antiarrhythmic or proarrhythmic?

Marciszek M, Paterek A, Oknińska M, Zambrowska Z, Mackiewicz U, Mączewski M
Cardiac arrhythmias are a major source of mortality and morbidity. Unfortunately, their treatment remains suboptimal. Major classes of antiarrhythmic drugs pose a significant risk of proarrhythmia, and their side effects often outweigh their benefits. Therefore, implantable devices remain the only truly effective antiarrhythmic therapy, and new strategies of antiarrhythmic treatment are required. Ivabradine is a selective heart rate-reducing agent, an inhibitor of hyperpolarization-activated, cyclic nucleotide-gated (HCN) channels, currently approved for treatment of coronary artery disease and chronic heart failure. In this review, we focus on the clinical and basic science evidence for the antiarrhythmic and proarrhythmic effects of ivabradine. We attempt to dissect the mechanisms behind the effects of ivabradine and indicate the focus of future studies.

Copyright © 2021 Heart Rhythm Society. Published by Elsevier Inc. All rights reserved.

Heart Rhythm: 29 Jun 2021; 18:1230-1238
Marciszek M, Paterek A, Oknińska M, Zambrowska Z, Mackiewicz U, Mączewski M
Heart Rhythm: 29 Jun 2021; 18:1230-1238 | PMID: 33737235
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Abstract

Significance of manifest localized staining during ethanol infusion into the vein of Marshall.

Takagi T, Pambrun T, Nakashima T, Vlachos K, ... Jaïs P, Derval N
Background
Localized staining due to venule injury is attributable to ethanol infusion into the vein of Marshall (Et-VOM).
Objective
The purpose of this study was to investigate adverse outcomes of localized staining during Et-VOM in patients undergoing ablation for atrial fibrillation.
Methods
Two hundred four patients (age 64 ± 10 years; 153 male) were sorted based on the aspect of localized staining. Staining of atrial myocardium that spread uniformly along the VOM vascular tree following selective VOM venography was considered normal, in contrast to predominantly localized staining that spread concentrically from a focal point due to vascular injury. Outcomes between the 2 groups were compared.
Results
Localized staining was observed in 27% of patients. No patients developed clinically significant pericardial effusions during Et-VOM; however, 7 patients developed pericardial effusions on the first postprocedural day (3.6% in patients with vs 3.4% in patients without localized staining). No significant difference was found in achievement of acute mitral isthmus (MI) block (96% vs 98%) and size of the endocardial low-voltage area (8.5 ± 4.1 cm2 vs 9.3 ± 5.3 cm2) in patients with and without localized staining, respectively. Long-term follow-up was not impacted by localized staining. Freedom from recurrent atrial tachyarrhythmias (66% vs 76%) and durability of MI block (57% vs 54%) were not significantly different with and without localized staining. There were no cases of rehospitalization for pericarditis, chronic pericardial effusion, or heart failure.
Conclusion
In our study, localized staining was frequent but was not associated with clinically relevant impact or disadvantages.

Copyright © 2021 Heart Rhythm Society. Published by Elsevier Inc. All rights reserved.

Heart Rhythm: 29 Jun 2021; 18:1057-1063
Takagi T, Pambrun T, Nakashima T, Vlachos K, ... Jaïs P, Derval N
Heart Rhythm: 29 Jun 2021; 18:1057-1063 | PMID: 33741483
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Impact:
Abstract

Identification of a SCN5A Founder Mutation Causing Sudden Death, Brugada Syndrome and Conduction Blocks in Southern Italy.

Curcio A, Malovini A, Mazzanti A, Memmi M, ... Bellazzi R, Napolitano C
Background
The genetic architecture of Brugada syndrome (BrS) is emerging as an increasingly complex area of investigation. The identification of genetically homogeneous populations can provide mechanistic insights and improve genotype-phenotype correlation.
Objective
To characterize and define the clinical implications of a novel BrS founder mutation. Using a haplotype-based approach we investigated whether two SCN5A genetic variants could derive from founder events.
Methods
Single nucleotide polymorphisms were genotyped in 201 subjects, haplotypes reconstructed and mutational age estimated. Clinical phenotypes and historical records were collected.
Results
A SCN5A variant (c.3352C>T; p.Gln1118Ter) was identified in 3 probands with BrS originating from South Italy. The same mutation was identified in a proband from central Italy and in one U.S. resident subject with Italian ancestry. The five individuals carried a common core haplotype, whose frequency was extremely low in local non-carrier probands and in population controls (0%-6.06%). The clinical presentation included multi-generational dominant transmission of Brugada electrocardiographic pattern, high incidence of sudden cardiac death (SCD) and cardiac conduction defects (CCD). We reconstructed seven-generation pedigrees with common geographic origin. Variant\'s age estimates suggested that origin of the p.Gln1118Ter dates back 76 generations (95% Confidence Interval: 28-200). A second SCN5A variant (c.5350G>A; p.Glu1784Lys) identified in the region did not show similar founder signal.
Conclusions
p.Gln1118Ter is a novel BrS/CCD/SCD founder mutation. We illustrate how these findings provide insights on the inheritance patterns and phenotypes associated with SCN5A mutation.

Copyright © 2021. Published by Elsevier Inc.

Heart Rhythm: 06 Jul 2021; epub ahead of print
Curcio A, Malovini A, Mazzanti A, Memmi M, ... Bellazzi R, Napolitano C
Heart Rhythm: 06 Jul 2021; epub ahead of print | PMID: 34245912
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Impact:
Abstract

Lipolytic enzymes and free fatty acids at the endothelial interface.

Goldberg IJ, Cabodevilla AG, Samovski D, Cifarelli V, Basu D, Abumrad NA
Lipids released from circulating lipoproteins by intravascular action of lipoprotein lipase (LpL) reach parenchymal cells in tissues with a non-fenestrated endothelium by transfer through or around endothelial cells. The actions of LpL are controlled at multiple sites, its synthesis and release by myocytes and adipocytes, its transit and association with the endothelial cell luminal surface, and finally its activation and inhibition by a number of proteins and by its product non-esterified fatty acids. Multiple pathways mediate endothelial transit of lipids into muscle and adipose tissues. These include movement of fatty acids via the endothelial cell fatty acid transporter CD36 and movement of whole or partially LpL-hydrolyzed lipoproteins via other apical endothelial cell receptors such as SR-B1and Alk1. Lipids also likely change the barrier function of the endothelium and operation of the paracellular pathway around endothelial cells. This review summarizes in vitro and in vivo support for the key role of endothelial cells in delivery of lipids and highlights incompletely understood processes that are the focus of active investigation.

Copyright © 2021. Published by Elsevier B.V.

Atherosclerosis: 29 Jun 2021; 329:1-8
Goldberg IJ, Cabodevilla AG, Samovski D, Cifarelli V, Basu D, Abumrad NA
Atherosclerosis: 29 Jun 2021; 329:1-8 | PMID: 34130222
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Impact:
Abstract

Apolipoprotein A-I mimetic peptide inhibits atherosclerosis by increasing tetrahydrobiopterin via regulation of GTP-cyclohydrolase 1 and reducing uncoupled endothelial nitric oxide synthase activity.

Ning DS, Ma J, Peng YM, Li Y, ... Ou ZJ, Ou JS
Background:
and aims
The apolipoprotein A-I mimetic peptide D-4F, among its anti-atherosclerotic effects, improves vasodilation through mechanisms not fully elucidated yet.
Methods
Low-density lipoprotein (LDL) receptor null (LDLr-/-) mice were fed Western diet with or without D-4F. We then measured atherosclerotic lesion formation, endothelial nitric oxide synthase (eNOS) phosphorylation and its association with heat shock protein 90 (HSP90), nitric oxide (NO) and superoxide anion (O2•-) production, and tetrahydrobiopterin (BH4) and GTP-cyclohydrolase 1 (GCH-1) concentration in the aorta. Human umbilical vein endothelial cells (HUVECs) and aortas were treated with oxidized LDL (oxLDL) with or without D-4F; subsequently, BH4 and GCH-1 concentration, NO and O2•- production, eNOS association with HSP90, and endothelium-dependent vasodilation were measured.
Results
Unexpectedly, eNOS phosphorylation, eNOS-HSP90 association, and O2•- production were increased, whereas BH4 and GCH-1 concentration and NO production were reduced in atherosclerosis. D-4F significantly inhibited atherosclerosis, eNOS phosphorylation, eNOS-HSP90 association, and O2•- generation but increased NO production and BH4 and GCH-1 concentration. OxLDL reduced NO production and BH4 and GCH-1 concentration but enhanced O2•- generation and eNOS association with HSP90, and impaired endothelium-dependent vasodilation. D-4F inhibited the overall effects of oxLDL.
Conclusions
Hypercholesterolemia enhanced uncoupled eNOS activity by decreasing GCH-1 concentration, thereby reducing BH4 levels. D-4F reduced uncoupled eNOS activity by increasing BH4 levels through GCH-1 expression and decreasing eNOS phosphorylation and eNOS-HSP90 association. Our findings elucidate a novel mechanism by which hypercholesterolemia induces atherosclerosis and D-4F inhibits it, providing a potential therapeutic approach.

Copyright © 2021 Elsevier B.V. All rights reserved.

Atherosclerosis: 29 Jun 2021; 328:83-91
Ning DS, Ma J, Peng YM, Li Y, ... Ou ZJ, Ou JS
Atherosclerosis: 29 Jun 2021; 328:83-91 | PMID: 34118596
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Impact:
Abstract

Intracellular glutamine level determines vascular smooth muscle cell-derived thrombogenicity.

Koyama S, Yamashita A, Matsuura Y, Saito Y, ... Kitamura K, Asada Y
Background:
and aims
The everolimus-eluting stent (EES), one of the effective stents for in-stent restenosis (ISR), has a lower incidence of stent thrombosis; however, the underlying mechanism remains unknown. This study aimed to identify the effects of everolimus on vascular metabolism and thrombogenicity and examine their mechanistic link.
Methods
EESs and bare-metal stents were implanted in rabbit iliac arteries with smooth muscle cell (SMC)-rich neointima induced by endothelial denudation. Four weeks after stent implantation, the stented arteries were examined for histological analysis and metabolomics. Additionally, everolimus effects in coronary artery SMCs metabolism, tissue factor (TF) expression, and procoagulant activity were assessed in vitro.
Results
EES-implanted arteries showed decreased neointima formation, less SMCs infiltration, and reduced TF expression. Concomitantly, they were metabolically characterized by increased levels of metabolites in amino acids, such as glutamine. Similarly, everolimus increased intracellular glutamine levels, decreased TF expression, and reduced procoagulant activity in SMCs in vitro. On the contrary, exogenous glutamine administration also increased intracellular glutamine level, decreased TF expression, and reduced procoagulant activity despite enhanced mammalian target of rapamycin (mTOR) activity.
Conclusions
Intracellular glutamine level is likely to determine vascular SMC-related thrombogenicity regardless of mTOR pathway activity. Therefore, increased intracellular glutamine level might contribute partially to the beneficial effect of EES use on stent thrombosis.

Copyright © 2021 Elsevier B.V. All rights reserved.

Atherosclerosis: 29 Jun 2021; 328:62-73
Koyama S, Yamashita A, Matsuura Y, Saito Y, ... Kitamura K, Asada Y
Atherosclerosis: 29 Jun 2021; 328:62-73 | PMID: 34102425
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Impact:
Abstract

Myeloid-associated lipin-1 transcriptional co-regulatory activity is atheroprotective.

Blackburn CMR, Schilke RM, Vozenilek AE, Chandran S, ... Finck BN, Woolard MD
Background:
and aims
Atherosclerosis is the most prominent underlying cause of cardiovascular disease (CVD). It is initiated by cholesterol deposition in the arterial intima, which causes macrophage recruitment and proinflammatory responses that promote plaque growth, necrotic core formation, and plaque rupture. Lipin-1 is a phosphatidic acid phosphohydrolase for glycerolipid synthesis. We have shown that lipin-1 phosphatase activity promotes macrophage pro-inflammatory responses when stimulated with modified low-density lipoprotein (modLDL) and accelerates atherosclerosis. Lipin-1 also independently acts as a transcriptional co-regulator where it enhances the expression of genes involved in β-oxidation. In hepatocytes and adipocytes, lipin-1 augments the activity of transcription factors such as peroxisome proliferator-activated receptor (PPARs). PPARs control the expression of anti-inflammatory genes in macrophages and slow or reduce atherosclerotic progression. Therefore, we hypothesize myeloid-derived lipin-1 transcriptional co-regulatory activity reduces atherosclerosis.
Methods
We used myeloid-derived lipin-1 knockout (lipin-1mKO) and littermate control mice and AAV8-PCSK9 along with high-fat diet to elicit atherosclerosis.
Results
Lipin-1mKO mice had larger aortic root plaques than littermate control mice after 8 and 12 weeks of a high-fat diet. Lipin-1mKO mice also had increased serum proinflammatory cytokine concentrations, reduced apoptosis in plaques, and larger necrotic cores in the plaques compared to control mice.
Conclusions
Combined, the data suggest lipin-1 transcriptional co-regulatory activity in myeloid cells is atheroprotective.

Copyright © 2021 Elsevier B.V. All rights reserved.

Atherosclerosis: 30 Jun 2021; 330:76-84
Blackburn CMR, Schilke RM, Vozenilek AE, Chandran S, ... Finck BN, Woolard MD
Atherosclerosis: 30 Jun 2021; 330:76-84 | PMID: 34256308
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Impact:
Abstract

APOE gene variants in primary dyslipidemia.

Khalil YA, Rabès JP, Boileau C, Varret M
Apolipoprotein E (apoE) is a major apolipoprotein involved in lipoprotein metabolism. It is a polymorphic protein and different isoforms are associated with variations in lipid and lipoprotein levels and thus cardiovascular risk. The isoform apoE4 is associated with an increase in LDL-cholesterol levels and thus a higher cardiovascular risk compared to apoE3. Whereas, apoE2 is associated with a mild decrease in LDL-cholesterol levels. In the presence of other risk factors, apoE2 homozygotes could develop type III hyperlipoproteinemia (familial dysbetalipoproteinemia or FD), an atherogenic disorder characterized by an accumulation of remnants of triglyceride-rich lipoproteins. Several rare APOE gene variants were reported in different types of dyslipidemias including FD, familial combined hyperlipidemia (FCH), lipoprotein glomerulopathy and bona fide autosomal dominant hypercholesterolemia (ADH). ADH is characterized by elevated LDL-cholesterol levels leading to coronary heart disease, and due to molecular alterations in three main genes: LDLR, APOB and PCSK9. The identification of the APOE-p.Leu167del variant as the causative molecular element in two different ADH families, paved the way to considering APOE as a candidate gene for ADH. Due to non mendelian interacting factors, common genetic and environmental factors and perhaps epigenetics, clinical presentation of lipid disorders associated with APOE variants often strongly overlap. More studies are needed to determine the spectrum of APOE implication in each of the diseases, notably ADH, in order to improve clinical and genetic diagnosis, prognosis and patient management. The purpose of this review is to comment on these APOE variants and on the molecular and clinical overlaps between dyslipidemias.

Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.

Atherosclerosis: 29 Jun 2021; 328:11-22
Khalil YA, Rabès JP, Boileau C, Varret M
Atherosclerosis: 29 Jun 2021; 328:11-22 | PMID: 34058468
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Impact:
Abstract

Triglyceride-lowering LPL alleles combined with LDL-C-lowering alleles are associated with an additively improved lipoprotein profile.

Ibi D, Blauw LL, Noordam R, Dollé MET, ... Karpe F, van Dijk KW
Background:
and aims
Mendelian randomization studies have shown that triglyceride (TG)- lowering lipoprotein lipase (LPL) alleles and low-density lipoprotein-cholesterol (LDL-C)-lowering alleles have independent beneficial associations on cardiovascular disease (CVD) risk. We aimed to provide further insight into this observation by applying Mendelian randomization analyses of genetically-influenced TG and LDL-C levels on plasma metabolomic profiles.
Methods
We quantified over 100 lipoprotein metabolomic measures in the Netherlands Epidemiology of Obesity (NEO) study (N = 4838) and Oxford Biobank (OBB) (N = 6999) by nuclear magnetic resonance (NMR) spectroscopy. Weighted genetic scores for TG via five LPL alleles and LDL-C via 19 alleles were calculated and dichotomized by the median, resulting in four genotype combinations of high/low TG and high/low LDL-C. We performed linear regression analyses using a two × two design with the group with genetically-influenced high TG and LDL-C as a reference.
Results
Compared to the individual groups with genetically-influenced lower TG or lower LDL-C only, the group with combined genetically-influenced lower TG and LDL-C showed an overall independent and additive pattern of changes in metabolomic measures. Over 100 measures were different (p < 1.35 × 10-3) compared to the reference, with effect sizes and directionality being similar in NEO and OBB. Most notably, levels of all very-low density lipoprotein (VLDL) and LDL sub-particles were lower.
Conclusions
Our findings provide evidence that TG-lowering on top of LDL-C-lowering has additive beneficial effects on the lipoprotein profile compared to TG-lowering or LDL-C-lowering only, which is in accordance with reported additive genetic effects on CVD risk reduction.

Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.

Atherosclerosis: 29 Jun 2021; 328:144-152
Ibi D, Blauw LL, Noordam R, Dollé MET, ... Karpe F, van Dijk KW
Atherosclerosis: 29 Jun 2021; 328:144-152 | PMID: 34053745
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Impact:
Abstract

Inhibition of myeloid HDAC2 upregulates glutaredoxin 1 expression, improves protein thiol redox state and protects against high-calorie diet-induced monocyte dysfunction and atherosclerosis.

Wang L, Ahn YJ, Asmis R
Background:
and aims
The thiol transferase glutaredoxin 1 controls redox signaling and cellular functions by regulating the S-glutathionylation status of critical protein thiols. Here we tested the hypothesis that by derepressing the expression of glutaredoxin 1, inhibition of histone deacetylase 2 prevents nutrient stress-induced protein S-glutathionylation and monocyte dysfunction and protects against atherosclerosis.
Methods
Using both a pharmacological inhibitor and shRNA-mediated knockdown of histone deacetylase 2, we determine the role of this deacetylase on glutaredoxin 1 expression and nutrient stress-induced inactivation of mitogen-activated protein kinase phosphatase 1 activity and monocyte and macrophage dysfunction. To assess whether histone deacetylase 2 inhibition in myeloid cells protects against atherosclerosis, we fed eight-week-old female and male HDAC2-/-MyeloidLDLR-/- mice and age and sex-matched LysMcretg/wtLDLR-/- control mice a high-calorie diet for 12 weeks and assessed monocyte function and atherosclerotic lesion size.
Results
Myeloid histone deacetylase 2 deficiency in high-calorie diet-fed LDLR-/- mice reduced atherosclerosis in males by 39% without affecting plasma lipid and lipoprotein profiles or blood glucose levels but had no effect on atherogenesis in female mice. Macrophage content in plaques of male mice was reduced by 31%. Histone deacetylase 2-deficient blood monocytes from male mice showed increased acetylation on histone 3, and increased Grx1 expression, and was associated with increased MKP-1 activity and reduced recruitment of monocyte-derived macrophages, whereas in females, myeloid HDAC2 deficiency had no effect on Grx1 expression, did not prevent nutrient stress-induced loss of MKP-1 activity in monocytes and was not atheroprotective.
Conclusions
Specific histone deacetylase 2 inhibitors may represent a potential novel therapeutic strategy for the prevention and treatment of atherosclerosis, but any benefits may be sexually dimorphic.

Copyright © 2021 Elsevier B.V. All rights reserved.

Atherosclerosis: 29 Jun 2021; 328:23-32
Wang L, Ahn YJ, Asmis R
Atherosclerosis: 29 Jun 2021; 328:23-32 | PMID: 34077868
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Impact:
Abstract

Improving detection accuracy of perfusion defect in standard dose SPECT-myocardial perfusion imaging by deep-learning denoising.

Liu J, Yang Y, Wernick MN, Pretorius PH, Slomka PJ, King MA
Background
We previously developed a deep-learning (DL) network for image denoising in SPECT-myocardial perfusion imaging (MPI). Here we investigate whether this DL network can be utilized for improving detection of perfusion defects in standard-dose clinical acquisitions.
Methods
To quantify perfusion-defect detection accuracy, we conducted a receiver-operating characteristic (ROC) analysis on reconstructed images with and without processing by the DL network using a set of clinical SPECT-MPI data from 190 subjects. For perfusion-defect detection hybrid studies were used as ground truth, which were created from clinically normal studies with simulated realistic lesions inserted. We considered ordered-subset expectation-maximization (OSEM) reconstruction with corrections for attenuation, resolution, and scatter and with 3D Gaussian post-filtering. Total perfusion deficit (TPD) scores, computed by Quantitative Perfusion SPECT (QPS) software, were used to evaluate the reconstructed images.
Results
Compared to reconstruction with optimal Gaussian post-filtering (sigma = 1.2 voxels), further DL denoising increased the area under the ROC curve (AUC) from 0.80 to 0.88 (P-value < 10-4). For reconstruction with less Gaussian post-filtering (sigma = 0.8 voxels), thus better spatial resolution, DL denoising increased the AUC value from 0.78 to 0.86 (P-value < 10-4) and achieved better spatial resolution in reconstruction.
Conclusions
DL denoising can effectively improve the detection of abnormal defects in standard-dose SPECT-MPI images over conventional reconstruction.

© 2021. American Society of Nuclear Cardiology.

J Nucl Cardiol: 18 Jul 2021; epub ahead of print
Liu J, Yang Y, Wernick MN, Pretorius PH, Slomka PJ, King MA
J Nucl Cardiol: 18 Jul 2021; epub ahead of print | PMID: 34282538
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Impact:
Abstract

S62798, a potent TAFIa inhibitor, accelerates endogenous fibrinolysis in a murine model of pulmonary thromboembolism.

Sansilvestri-Morel P, Rupin A, Schaffner AP, Bertin F, ... Tupinon-Mathieu I, Delerive P
Enhancement of fibrinolysis constitutes a promising approach to treat thrombotic diseases. Venous thrombosis and thromboembolism risks are associated with increased plasma levels of TAFI (Thrombin Activatable Fibrinolysis Inhibitor) as well as its active form TAFIa. A new TAFIa inhibitor, namely S62798 has been identified. Its ability to enhance fibrinolysis was investigated both in vitro and in vivo in a mouse model of pulmonary thromboembolism, as well as its effect on bleeding. S62798 is a highly selective human, mouse and rat TAFIa inhibitor (IC50 = 11; 270; 178 nmol/L, respectively). It accelerates lysis of a human clot in vitro, evaluated by thromboelastometry (EC50 = 27 nmol/L). In a rat tail bleeding model, no effect of S62798 treatment was observed up to 20 mg/kg. Enhancement of endogenous fibrinolysis by S62798 was investigated in a mouse model of Tissue Factor-induced pulmonary thromboembolism. Intravenous administration of S62798 decreased pulmonary fibrin clots with a minimal effective dose of 0.03 mg/kg. Finally, effect of S62798 in combination with heparin was evaluated. When treatment of heparin was done in a curative setting, no effect was observed whereas a significantly decreased pulmonary fibrin deposition was observed in response to S62798 alone or in combination with heparin. This study demonstrates that S62798 is a potent TAFIa inhibitor with minimal risk of bleeding. In vivo, curative S62798 intravenous treatment, alone or associated with heparin, accelerated clot lysis by potentiating endogenous fibrinolysis and thus decreased pulmonary fibrin clots. S62798 is expected to be a therapeutic option for pulmonary embolism patients on top of anticoagulants.

Copyright © 2021 Elsevier Ltd. All rights reserved.

Thromb Res: 30 Jul 2021; 204:81-87
Sansilvestri-Morel P, Rupin A, Schaffner AP, Bertin F, ... Tupinon-Mathieu I, Delerive P
Thromb Res: 30 Jul 2021; 204:81-87 | PMID: 34153648
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Impact:
Abstract

Euglobulin clot lysis time reveals a high frequency of fibrinolytic activation in trauma.

Ilich A, Kumar V, Ferrara MJ, Henderson MW, ... Park MS, Key NS
Activation of the fibrinolytic system plays a central role in the host response to trauma. There is significant heterogeneity in the degree of fibrinolysis activation at baseline that is usually assessed by whole blood thromboelastography (TEG). Few studies have focused on plasma markers of fibrinolysis that could add novel insights into the frequency and mechanisms of fibrinolytic activation in trauma. Global fibrinolysis in plasma was assessed using a modified euglobulin clot lysis time (ECLT) assay in 171 major trauma patients and compared to commonly assessed analytes of fibrinolysis. The median ECLT in trauma patients was significantly shorter at 8.5 h (IQR, 1.3-19.5) compared to 19.9 h (9.8-22.6) in healthy controls (p < 0.0001). ECLT values ≤2.5th percentile of the reference range were present in 83 (48.5%) of trauma patients, suggesting increased fibrinolytic activation. Shortened ECLT values were associated with elevated plasmin-antiplasmin (PAP) complexes and free tissue plasminogen activator (tPA) levels in plasma. Sixteen (9.2%) individuals met the primary outcome for massive transfusion, here defined as the critical administration threshold (CAT) of 3 units of packed red cells in any 60-minute period within the first 24 h. In a univariate screen, plasma biomarkers associated with CAT included D-dimer (p < 0.001), PAP (p < 0.05), free tPA (p < 0.05) and ECLT (p < 0.05). We conclude that fibrinolytic activation, measured by ECLT, is present in a high proportion of trauma patients at presentation. The shortened ECLT is partially driven by high tPA levels and is associated with high levels of circulating PAP complexes. Further studies are needed to determine whether ECLT is an independent predictor of trauma outcomes.

Copyright © 2021 Elsevier Ltd. All rights reserved.

Thromb Res: 30 Jul 2021; 204:22-28
Ilich A, Kumar V, Ferrara MJ, Henderson MW, ... Park MS, Key NS
Thromb Res: 30 Jul 2021; 204:22-28 | PMID: 34111811
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Impact:
Abstract

Differential procoagulatory response of microvascular, arterial and venous endothelial cells upon inflammation in vitro.

Brandtner AK, Lehner GF, Pircher A, Feistritzer C, Joannidis M
Introduction
Inflammation induces a procoagulant phenotype of endothelial cells (EC) with the exposure of tissue factor (TF), a potent initiator of the extrinsic coagulation cascade. Although systemic inflammation affects the whole vascular system, thrombotic lesions occur particularly in microcirculation. This raises the question of whether TF-procoagulant activity (TF-PCA) differs between EC from arterial, venous, and microvascular beds.
Materials and methods
Functional coagulation tests, including TF-PCA, and inflammatory responses were investigated on arterial, venous and microvascular endothelial cells. Interleukin-6 (IL-6) and TF-levels were determined in cohort of 59 septic patients.
Results
We found that tumor necrosis factor alpha (TNFα), lipopolysaccharide, and interleukin-1β induce a solid, dose-dependent increase in TF-PCA, which is highest in microvascular EC. A positive correlation of interleukin-6 (IL-6) with TF levels was observed in a cohort of 59 septic patients. In contrast, TF-PCA was independent of IL-6 concentrations in vitro. Re-analysis of publicly available gene expression data revealed that among the top 50 genes annotated to coagulation, TF is one of three regulated genes common to the three investigated EC subtypes. The response to inflammatory stimuli in terms of exposure of leukocyte-endothelial- and platelet-endothelial adhesion molecules (E-selectin and PECAM-1), remodeling of adherens junctions, co-exposure of negatively charged surfaces nor breakdown of the glycocalyx was comparable between the EC subtypes and did not explain the higher TF-PCA on microvascular cells. We found that the ratio of TF and TFPI exposure on the endothelial membrane significantly differs between the EC subtypes.
Conclusions
These findings indicate that the ratio of TF to its inhibitor TFPI is a determinant of endothelial TF-PCA, which is most pronounced on microvascular endothelial cells and might explain why the microvascular system is particularly susceptible to inflammation-induced thrombosis.

Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.

Thromb Res: 01 Jul 2021; 205:70-80
Brandtner AK, Lehner GF, Pircher A, Feistritzer C, Joannidis M
Thromb Res: 01 Jul 2021; 205:70-80 | PMID: 34265605
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Impact:
Abstract

Relations of arterial stiffness and endothelial dysfunction with incident venous thromboembolism.

Ko D, Preis SR, Johnson AD, Vasan RS, ... Hamburg NM, Mitchell GF
Introduction
Association between arterial vascular dysfunction and risk of venous thromboembolism (VTE) is uncertain. We determined the associations between comprehensive measures of arterial vascular function and risk of incident VTE in a community-based cohort study with robust longitudinal follow-up.
Materials and methods
In the Framingham Heart Study Original, Offspring, Third Generation, and Omni cohorts, we measured carotid-femoral pulse wave velocity and central pulse pressure (n = 8261, age 51.5 ± 15.5 years, 54% women), flow-mediated dilation and hyperemic velocity (n = 6540, age 47.9 ± 14.1 years, 54% women), and peripheral arterial tonometry ratio (n = 4998, age 54.3 ± 16.0 years, 52% women). Deep venous thrombosis and pulmonary embolism were diagnosed with imaging studies and adjudicated by three Framingham Heart Study physicians.
Results and conclusions
The rate of incident VTE was 1.6-2.1 per 1000 person-years during mean follow-up of 8.5-11.2 years. In age- and sex-adjusted Cox proportional hazards regression models, carotid-femoral pulse wave velocity was associated with increased risk of VTE (HR 1.32, 95% CI 1.05-1.66, p = 0.02), however the association was no longer statistically significant after multivariable adjustment (HR 1.24, 95% CI 0.96-1.61, p = 0.10). None of the other vascular variables were associated with the risk of VTE in any of the models. In our comprehensive examination of arterial vascular function and risk of VTE, we did not observe any association between select arterial function measures and risk of VTE after multivariable adjustment.

Copyright © 2021 Elsevier Ltd. All rights reserved.

Thromb Res: 30 Jul 2021; 204:108-113
Ko D, Preis SR, Johnson AD, Vasan RS, ... Hamburg NM, Mitchell GF
Thromb Res: 30 Jul 2021; 204:108-113 | PMID: 34175748
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Impact:
Abstract

Diverse functional responses to high glucose by primary and permanent hybrid endothelial cells in vitro.

Uruski P, Mikuła-Pietrasik J, Drzewiecki M, Budkiewicz S, ... Tykarski A, Książek K
Various types of human endothelial cells, including human umbilical vein endothelial cells (HUVECs) and the established hybrid EAhy926 cells, are used in experimental research. Here, we compared the biological properties of HUVECs and EAhy926 cells under normal (5 mM) and high glucose (30 mM; HG) conditions. The results showed that HG induced cellular senescence and a stronger DNA damage response in HUVECs than in EAhy926 cells. The magnitude of oxidative stress elicited in HUVECs by HG was also greater than that elicited in their established counterparts. Both endothelial cell types promoted the progression of breast (MCF7), ovarian (OVCAR-3), and lung (A549) cancer cells; however, the effects elicited by HG-treated HUVECs on adhesion (MCF7, OVCAR-3), proliferation (OVCAR-3), and migration (OVCAR-3) were more pronounced. Finally, HG stimulated the production of a higher number of proangiogenic agents in HUVECs than in EAhy926 cells. Collectively, our study shows that the functional properties of primary and established endothelial cells exposed to HG differ substantially, which seems to result from the higher sensitivity of the former to this stressor. The interchangeability of both types of endothelial cells in biomedical research should be considered with great care to avoid losing some biological effects due to the choice of cells with higher stress tolerance.

Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.

J Mol Cell Cardiol: 29 Jun 2021; 156:1-6
Uruski P, Mikuła-Pietrasik J, Drzewiecki M, Budkiewicz S, ... Tykarski A, Książek K
J Mol Cell Cardiol: 29 Jun 2021; 156:1-6 | PMID: 33731316
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Impact:
Abstract

Application of genetic cell-lineage tracing technology to study cardiovascular diseases.

Sun X, Lyu L, Zhong X, Ni Z, Xu Q
Cardiovascular diseases are leading causes that threaten people\'s life. To investigate cells that are involved in disease development and tissue repair, various technologies have been introduced. Among these technologies, lineage tracing is a powerful tool to track the fate of cells in vivo, providing deep insights into cellular behavior and plasticity. In cardiac diseases, newly formed cardiomyocytes and endothelial cells are found from proliferation of local cells, while fibroblasts and macrophages are originated from diverse cell sources. Similarly, in response to vascular injury, various sources of cells including media smooth muscle cells, endothelium, resident progenitors and bone marrow cells are involved in lesion formation and/or vessel regeneration. In summary, current review summarizes the development of lineage tracing techniques and their utilizations in investigating roles of different cell types in cardiovascular diseases.

Copyright © 2021. Published by Elsevier Ltd.

J Mol Cell Cardiol: 29 Jun 2021; 156:57-68
Sun X, Lyu L, Zhong X, Ni Z, Xu Q
J Mol Cell Cardiol: 29 Jun 2021; 156:57-68 | PMID: 33745891
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Impact:
Abstract

New calcification model for intact murine aortic valves.

Kruithof BPT, van de Pol V, Los T, Lodder K, ... Goumans MJ, Ajmone Marsan N
Calcific aortic valve disease (CAVD) is a common progressive disease of the aortic valves, for which no medical treatment exists and surgery represents currently the only therapeutic solution. The development of novel pharmacological treatments for CAVD has been hampered by the lack of suitable test-systems, which require the preservation of the complex valve structure in a mechanically and biochemical controllable system. Therefore, we aimed at establishing a model which allows the study of calcification in intact mouse aortic valves by using the Miniature Tissue Culture System (MTCS), an ex vivo flow model for whole mouse hearts. Aortic valves of wild-type mice were cultured in the MTCS and exposed to osteogenic medium (OSM, containing ascorbic acid, β-glycerophosphate and dexamethasone) or inorganic phosphates (PI). Osteogenic calcification occurred in the aortic valve leaflets that were cultured ex vivo in the presence of PI, but not of OSM. In vitro cultured mouse and human valvular interstitial cells calcified in both OSM and PI conditions, revealing in vitro-ex vivo differences. Furthermore, endochondral differentiation occurred in the aortic root of ex vivo cultured mouse hearts near the hinge of the aortic valve in both PI and OSM conditions. Dexamethasone was found to induce endochondral differentiation in the aortic root, but to inhibit calcification and the expression of osteogenic markers in the aortic leaflet, partly explaining the absence of calcification in the aortic valve cultured with OSM. The osteogenic calcifications in the aortic leaflet and the endochondral differentiation in the aortic root resemble calcifications found in human CAVD. In conclusion, we have established an ex vivo calcification model for intact wild-type murine aortic valves in which the initiation and progression of aortic valve calcification can be studied. The in vitro-ex vivo differences found in our studies underline the importance of ex vivo models to facilitate pre-clinical translational studies.

Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.

J Mol Cell Cardiol: 29 Jun 2021; 156:95-104
Kruithof BPT, van de Pol V, Los T, Lodder K, ... Goumans MJ, Ajmone Marsan N
J Mol Cell Cardiol: 29 Jun 2021; 156:95-104 | PMID: 33744308
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Impact:
Abstract

YAP1/TEAD1 upregulate platelet-derived growth factor receptor beta to promote vascular smooth muscle cell proliferation and neointima formation.

Osman I, Dong K, Kang X, Yu L, ... Zhang W, Zhou J
We have previously demonstrated that the transcription co-factor yes-associated protein 1 (YAP1) promotes vascular smooth muscle cell (VSMC) de-differentiation. Yet, the role and underlying mechanisms of YAP1 in neointima formation in vivo remain unclear. The goal of this study was to investigate the role of VSMC-expressed YAP1 in vascular injury-induced VSMC proliferation and delineate the mechanisms underlying its action. Experiments employing gain- or loss-of-function of YAP1 demonstrated that YAP1 promotes human VSMC proliferation. Mechanistically, we identified platelet-derived growth factor receptor beta (PDGFRB) as a novel YAP1 target gene that confers the YAP1-dependent hyper-proliferative effects in VSMCs. Furthermore, we identified TEA domain transcription factor 1 (TEAD1) as a key transcription factor that mediates YAP1-dependent PDGFRβ expression. ChIP assays demonstrated that TEAD1 is enriched at a PDGFRB gene enhancer. Luciferase reporter assays further demonstrated that YAP1 and TEAD1 co-operatively activate the PDGFRB enhancer. Consistent with these observations, we found that YAP1 expression is upregulated after arterial injury and correlates with PDGFRβ expression and VSMC proliferation in vivo. Using a novel inducible SM-specific Yap1 knockout mouse model, we found that the specific deletion of Yap1 in adult VSMCs is sufficient to attenuate arterial injury-induced neointima formation, largely due to inhibited PDGFRβ expression and VSMC proliferation. Our study unravels a novel mechanism by which YAP1/TEAD1 promote VSMC proliferation via transcriptional induction of PDGFRβ, thereby enhancing PDGF-BB downstream signaling and promoting neointima formation.

Copyright © 2021 Elsevier Ltd. All rights reserved.

J Mol Cell Cardiol: 29 Jun 2021; 156:20-32
Osman I, Dong K, Kang X, Yu L, ... Zhang W, Zhou J
J Mol Cell Cardiol: 29 Jun 2021; 156:20-32 | PMID: 33753119
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Abstract

Impact of etiology on force and kinetics of left ventricular end-stage failing human myocardium.

Mashali MA, Saad NS, Canan BD, Elnakish MT, ... Mohler PJ, Janssen PML
Background
Heart failure (HF) is associated with highly significant morbidity, mortality, and health care costs. Despite the significant advances in therapies and prevention, HF remains associated with poor clinical outcomes. Understanding the contractile force and kinetic changes at the level of cardiac muscle during end-stage HF in consideration of underlying etiology would be beneficial in developing targeted therapies that can help improve cardiac performance.
Objective
Investigate the impact of the primary etiology of HF (ischemic or non-ischemic) on left ventricular (LV) human myocardium force and kinetics of contraction and relaxation under near-physiological conditions.
Methods and results
Contractile and kinetic parameters were assessed in LV intact trabeculae isolated from control non-failing (NF; n = 58) and end-stage failing ischemic (FI; n = 16) and non-ischemic (FNI; n = 38) human myocardium under baseline conditions, length-dependent activation, frequency-dependent activation, and response to the β-adrenergic stimulation. At baseline, there were no significant differences in contractile force between the three groups; however, kinetics were impaired in failing myocardium with significant slowing down of relaxation kinetics in FNI compared to NF myocardium. Length-dependent activation was preserved and virtually identical in all groups. Frequency-dependent activation was clearly seen in NF myocardium (positive force frequency relationship [FFR]), while significantly impaired in both FI and FNI myocardium (negative FFR). Likewise, β-adrenergic regulation of contraction was significantly impaired in both HF groups.
Conclusions
End-stage failing myocardium exhibited impaired kinetics under baseline conditions as well as with the three contractile regulatory mechanisms. The pattern of these kinetic impairments in relation to NF myocardium was mainly impacted by etiology with a marked slowing down of kinetics in FNI myocardium. These findings suggest that not only force development, but also kinetics should be considered as a therapeutic target for improving cardiac performance and thus treatment of HF.

Copyright © 2021 Elsevier Ltd. All rights reserved.

J Mol Cell Cardiol: 29 Jun 2021; 156:7-19
Mashali MA, Saad NS, Canan BD, Elnakish MT, ... Mohler PJ, Janssen PML
J Mol Cell Cardiol: 29 Jun 2021; 156:7-19 | PMID: 33766524
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Abstract

Creld1 regulates myocardial development and function.

Beckert V, Rassmann S, Kayvanjoo AH, Klausen C, ... Mass E, Wachten D
CRELD1 (Cysteine-Rich with EGF-Like Domains 1) is a risk gene for non-syndromic atrioventricular septal defects in human patients. In a mouse model, Creld1 has been shown to be essential for heart development, particularly in septum and valve formation. However, due to the embryonic lethality of global Creld1 knockout (KO) mice, its cell type-specific function during peri- and postnatal stages remains unknown. Here, we generated conditional Creld1 KO mice lacking Creld1 either in the endocardium (KOTie2) or the myocardium (KOMyHC). Using a combination of cardiac phenotyping, histology, immunohistochemistry, RNA-sequencing, and flow cytometry, we demonstrate that Creld1 function in the endocardium is dispensable for heart development. Lack of myocardial Creld1 causes extracellular matrix remodeling and trabeculation defects by modulation of the Notch1 signaling pathway. Hence, KOMyHC mice die early postnatally due to myocardial hypoplasia. Our results reveal that Creld1 not only controls the formation of septa and valves at an early stage during heart development, but also cardiac maturation and function at a later stage. These findings underline the central role of Creld1 in mammalian heart development and function.

Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.

J Mol Cell Cardiol: 29 Jun 2021; 156:45-56
Beckert V, Rassmann S, Kayvanjoo AH, Klausen C, ... Mass E, Wachten D
J Mol Cell Cardiol: 29 Jun 2021; 156:45-56 | PMID: 33773996
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Abstract

Role of myeloid-derived chemokine CCL5/RANTES at an early stage of atherosclerosis.

Jongstra-Bilen J, Tai K, Althagafi MG, Siu A, ... Hyduk SJ, Cybulsky MI
One of the hallmarks of atherosclerosis is ongoing accumulation of macrophages in the artery intima beginning at disease onset. Monocyte recruitment contributes to increasing macrophage abundance at early stages of atherosclerosis. Although the chemokine CCL5 (RANTES) has been studied in atherosclerosis, its role in the recruitment of monocytes to early lesions has not been elucidated. We show that expression of Ccl5 mRNA, as well as other ligands of the CCR5 receptor (Ccl3 and Ccl4), is induced in the aortic intima of Ldlr-/- mice 3 weeks after the initiation of cholesterol-rich diet (CRD)-induced hypercholesterolemia. En face immunostaining revealed that CCL5 protein expression is also upregulated at 3 weeks of CRD. Blockade of CCR5 significantly reduced monocyte recruitment to 3-week lesions, suggesting that chemokine signaling through CCR5 is critical. However, we observed that Ccl5-deficiency had no effect on early lesion formation and CCL5-blockade did not affect monocyte recruitment in Ldlr-/- mice. Immunostaining of the lesions in Ldlr-/- mice and reciprocal bone marrow transplantation (BMT) of Ccl5+/+ and Ccl5-/- mice revealed that CCL5 is expressed by both myeloid and endothelial cells. BMT experiments were carried out to determine if CCL5 produced by distinct cells has functions that may be concealed in Ccl5-/-Ldlr-/- mice. We found that hematopoietic cell-derived CCL5 regulates monocyte recruitment and the abundance of intimal macrophages in 3-week lesions of Ldlr-/- mice but plays a minor role in 6-week lesions. Our findings suggest that there is a short window in early lesion formation during which myeloid cell-derived CCL5 has a critical role in monocyte recruitment and macrophage abundance.

Copyright © 2021 Elsevier Ltd. All rights reserved.

J Mol Cell Cardiol: 29 Jun 2021; 156:69-78
Jongstra-Bilen J, Tai K, Althagafi MG, Siu A, ... Hyduk SJ, Cybulsky MI
J Mol Cell Cardiol: 29 Jun 2021; 156:69-78 | PMID: 33781821
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Abstract

Amino terminus of cardiac myosin binding protein-C regulates cardiac contractility.

Lynch TL, Kumar M, McNamara JW, Kuster DWD, ... Warshaw DM, Sadayappan S
Phosphorylation of cardiac myosin binding protein-C (cMyBP-C) regulates cardiac contraction through modulation of actomyosin interactions mediated by the protein\'s amino terminal (N\')-region (C0-C2 domains, 358 amino acids). On the other hand, dephosphorylation of cMyBP-C during myocardial injury results in cleavage of the 271 amino acid C0-C1f region and subsequent contractile dysfunction. Yet, our current understanding of amino terminus region of cMyBP-C in the context of regulating thin and thick filament interactions is limited. A novel cardiac-specific transgenic mouse model expressing cMyBP-C, but lacking its C0-C1f region (cMyBP-C∆C0-C1f), displayed dilated cardiomyopathy, underscoring the importance of the N\'-region in cMyBP-C. Further exploring the molecular basis for this cardiomyopathy, in vitro studies revealed increased interfilament lattice spacing and rate of tension redevelopment, as well as faster actin-filament sliding velocity within the C-zone of the transgenic sarcomere. Moreover, phosphorylation of the unablated phosphoregulatory sites was increased, likely contributing to normal sarcomere morphology and myoarchitecture. These results led us to hypothesize that restoration of the N\'-region of cMyBP-C would return actomyosin interaction to its steady state. Accordingly, we administered recombinant C0-C2 (rC0-C2) to permeabilized cardiomyocytes from transgenic, cMyBP-C null, and human heart failure biopsies, and we found that normal regulation of actomyosin interaction and contractility was restored. Overall, these data provide a unique picture of selective perturbations of the cardiac sarcomere that either lead to injury or adaptation to injury in the myocardium.

Copyright © 2021 Elsevier Ltd. All rights reserved.

J Mol Cell Cardiol: 29 Jun 2021; 156:33-44
Lynch TL, Kumar M, McNamara JW, Kuster DWD, ... Warshaw DM, Sadayappan S
J Mol Cell Cardiol: 29 Jun 2021; 156:33-44 | PMID: 33781820
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Abstract

LITAF acts as a novel regulator for pathological cardiac hypertrophy.

Xiang M, Yang F, Zhou Y, Li W, ... Wang PX, Chen M
Pathological hypertrophy generally progresses to heart failure. Exploring effective and promising therapeutic targets might lead to progress in preventing its detrimental outcomes. Our current knowledge about lipopolysaccharide-induced tumor necrosis factor-α factor (LITAF) is mainly limited to regulate inflammation. However, the role of LITAF in other settings that are not that relevant to inflammation, such as cardiac remodeling and heart failure, remains largely unknown. In the present study, we found that the expression of LITAF decreased in hypertrophic hearts and cardiomyocytes. Meanwhile, LITAF protected cultured neonatal rat cardiomyocytes against phenylephrine-induced hypertrophy. Moreover, using LITAF knockout mice, we demonstrated that LITAF deficiency exacerbated cardiac hypertrophy and fibrosis compared with wild-type mice. Mechanistically, LITAF directly binds to the N-terminal of ASK1, thus disrupting the dimerization of ASK1 and blocking ASK1 activation, ultimately inhibiting ASK1-JNK/p38 signaling over-activation and protecting against cardiac hypertrophy. Furthermore, AAV9-mediated LITAF overexpression attenuated cardiac hypertrophy in vivo. Conclusions: Our findings uncover the novel role of LITAF as a negative regulator of cardiac remodeling. Targeting the interaction between LITAF and ASK1 could be a promising therapeutic strategy for pathological cardiac remodeling.

Copyright © 2021 Elsevier Ltd. All rights reserved.

J Mol Cell Cardiol: 29 Jun 2021; 156:82-94
Xiang M, Yang F, Zhou Y, Li W, ... Wang PX, Chen M
J Mol Cell Cardiol: 29 Jun 2021; 156:82-94 | PMID: 33823186
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Abstract

Sarcoplasmic reticulum-mitochondria communication; implications for cardiac arrhythmia.

Hamilton S, Terentyeva R, Clements RT, Belevych AE, Terentyev D
Sudden cardiac death due to ventricular tachyarrhythmias remains the major cause of mortality in the world. Heart failure, diabetic cardiomyopathy, old age-related cardiac dysfunction and inherited disorders are associated with enhanced propensity to malignant cardiac arrhythmias. Both defective mitochondrial function and abnormal intracellular Ca2+ homeostasis have been established as the key contributing factors in the pathophysiology and arrhythmogenesis in these conditions. This article reviews current advances in understanding of bidirectional control of ryanodine receptor-mediated sarcoplasmic reticulum Ca2+ release and mitochondrial function, and how defects in crosstalk between these two organelles increase arrhythmic risk in cardiac disease.

Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.

J Mol Cell Cardiol: 29 Jun 2021; 156:105-113
Hamilton S, Terentyeva R, Clements RT, Belevych AE, Terentyev D
J Mol Cell Cardiol: 29 Jun 2021; 156:105-113 | PMID: 33857485
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Abstract

FNDC5/Irisin attenuates diabetic cardiomyopathy in a type 2 diabetes mouse model by activation of integrin αV/β5-AKT signaling and reduction of oxidative/nitrosative stress.

Lin C, Guo Y, Xia Y, Li C, ... Yan W, Tao L
Irisin, the cleaved form of the fibronectin type III domain containing 5 (FNDC5) protein, is involved in metabolism and inflammation. Recent findings indicated that irisin participated in cardiovascular physiology and pathology. In this study, we investigated the effects of FNDC5/irisin on diabetic cardiomyopathy (DCM) in type 2 diabetic db/db mice. Downregulation of myocardial FNDC5/irisin protein expression and plasma irisin levels was observed in db/db mice compared to db/+ controls. Moreover, echocardiography revealed that db/db mice exhibited normal cardiac systolic function and impaired diastolic function. Adverse structural remodeling, including cardiomyocyte apoptosis, myocardial fibrosis, and cardiac hypertrophy were observed in the hearts of db/db mice. Sixteen-week-old db/db mice were intramyocardially injected with adenovirus encoding FNDC5 or treated with recombinant human irisin via a peritoneal implant osmotic pump for 4 weeks. Both overexpression of myocardial FNDC5 and exogenous irisin administration attenuated diastolic dysfunction and cardiac structural remodeling in db/db mice. Results from in vitro studies revealed that FNDC5/irisin protein expression was decreased in high glucose (HG)/high fat (HF)-treated cardiomyocytes. Increased levels of inducible nitric oxide synthase (iNOS), NADPH oxidase 2 (NOX2), 3-nitrotyrosine (3-NT), reactive oxygen species (ROS), and peroxynitrite (ONOO-) in HG/HF-treated H9C2 cells provided evidence of oxidative/nitrosative stress, which was alleviated by treatment with FNDC5/irisin. Moreover, the mitochondria membrane potential (ΔΨm) was decreased and cytochrome C was released from mitochondria with increased levels of cleaved caspase-3 in HG/HF-treated H9C2 cells, indicating the presence of mitochondria-dependent apoptosis, which was partially reversed by FNDC5/irisin treatment. Mechanistic studies showed that activation of integrin αVβ5-AKT signaling and attenuation of oxidative/nitrosative stress were responsible for the cardioprotective effects of FNDC5/irisin. Therefore, FNDC5/irisin mediates cardioprotection in DCM by inhibiting myocardial apoptosis, myocardial fibrosis, and cardiac hypertrophy. These findings implicate that FNDC5/irisin as a potential therapeutic intervention for DCM, especially in type 2 diabetes mellitus (T2DM).

Copyright © 2021 Elsevier Ltd. All rights reserved.

J Mol Cell Cardiol: 02 Jul 2021; 160:27-41
Lin C, Guo Y, Xia Y, Li C, ... Yan W, Tao L
J Mol Cell Cardiol: 02 Jul 2021; 160:27-41 | PMID: 34224725
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Abstract

An organ-on-a-chip model for pre-clinical drug evaluation in progressive non-genetic cardiomyopathy.

Wang EY, Kuzmanov U, Smith JB, Dou W, ... Gramolini A, Radisic M
Angiotensin II (Ang II) presents a critical mediator in various pathological conditions such as non-genetic cardiomyopathy. Osmotic pump infusion in rodents is a commonly used approach to model cardiomyopathy associated with Ang II. However, profound differences in electrophysiology and pharmacokinetics between rodent and human cardiomyocytes may limit predictability of animal-based experiments. This study investigates the application of an Organ-on-a-chip (OOC) system in modeling Ang II-induced progressive cardiomyopathy. The disease model is constructed to recapitulate myocardial response to Ang II in a temporal manner. The long-term tissue cultivation and non-invasive functional readouts enable monitoring of both acute and chronic cardiac responses to Ang II stimulation. Along with mapping of cytokine secretion and proteomic profiles, this model presents an opportunity to quantitatively measure the dynamic pathological changes that could not be otherwise identified in animals. Further, we present this model as a testbed to evaluate compounds that target Ang II-induced cardiac remodeling. Through assessing the effects of losartan, relaxin, and saracatinib, the drug screening data implicated multifaceted cardioprotective effects of relaxin in restoring contractile function and reducing fibrotic remodeling. Overall, this study provides a controllable platform where cardiac activities can be explicitly observed and tested over the pathological process. The facile and high-content screening can facilitate the evaluation of potential drug candidates in the pre-clinical stage.

Copyright © 2021. Published by Elsevier Ltd.

J Mol Cell Cardiol: 29 Jun 2021; epub ahead of print
Wang EY, Kuzmanov U, Smith JB, Dou W, ... Gramolini A, Radisic M
J Mol Cell Cardiol: 29 Jun 2021; epub ahead of print | PMID: 34216608
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Abstract

Early adaptive chromatin remodeling events precede pathologic phenotypes and are reinforced in the failing heart.

Chapski DJ, Cabaj M, Morselli M, Mason RJ, ... Vondriska TM, Rosa-Garrido M
The temporal nature of chromatin structural changes underpinning pathologic transcription are poorly understood. We measured chromatin accessibility and DNA methylation to study the contribution of chromatin remodeling at different stages of cardiac hypertrophy and failure. ATAC-seq and reduced representation bisulfite sequencing were performed in cardiac myocytes after transverse aortic constriction (TAC) or depletion of the chromatin structural protein CTCF. Early compensation to pressure overload showed changes in chromatin accessibility and DNA methylation preferentially localized to intergenic and intronic regions. Most methylation and accessibility changes observed in enhancers and promoters at the late phase (3 weeks after TAC) were established at an earlier time point (3 days after TAC), before heart failure manifests. Enhancers were paired with genes based on chromatin conformation capture data: while enhancer accessibility generally correlated with changes in gene expression, this feature, nor DNA methylation, was alone sufficient to predict transcription of all enhancer interacting genes. Enrichment of transcription factors and active histone marks at these regions suggests that enhancer activity coordinates with other epigenetic factors to determine gene transcription. In support of this hypothesis, ChIP-qPCR demonstrated increased enhancer and promoter occupancy of GATA4 and NKX2-5 at Itga9 and Nppa, respectively, concomitant with increased transcription of these genes in the diseased heart. Lastly, we demonstrate that accessibility and DNA methylation are imperfect predictors of chromatin structure at the scale of A/B compartmentalization-rather, accessibility, DNA methylation, transcription factors and other histone marks work within these domains to determine gene expression. These studies establish that chromatin reorganization during early compensation after pathologic stimuli is maintained into the later decompensatory phases of heart failure. The findings reveal the rules for how local chromatin features govern gene expression in the context of global genomic structure and identify chromatin remodeling events for therapeutic targeting in disease.

Copyright © 2021. Published by Elsevier Ltd.

J Mol Cell Cardiol: 13 Jul 2021; epub ahead of print
Chapski DJ, Cabaj M, Morselli M, Mason RJ, ... Vondriska TM, Rosa-Garrido M
J Mol Cell Cardiol: 13 Jul 2021; epub ahead of print | PMID: 34273410
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Abstract

Non-invasive evaluation of retinal vascular remodeling and hypertrophy in humans: intricate effect of ageing, blood pressure and glycaemia.

Gallo A, Dietenbeck T, Giron A, Paques M, Kachenoura N, Girerd X
Background
Ageing, hypertension and diabetes have an intricate effect on microvascular structure. In the retina, the respective contribution of remodeling and hypertrophy in such process is still unclear. We aimed at disentangling age, blood pressure and glycaemia effects on retinal microcirculation using the non-invasive adaptive optics ophthalmoscopy (AOO).
Methods
We included 429 subjects, distributed into 4 groups according to normal (nBP) or high blood pressure (hBP) and/or normal (nGly) or high fasting glycaemia (hGly). The nBP/nGly group was stratified in age tertiles to isolate the effect of ageing. AOO was used to measure arteriolar wall thickness (WT, µm), arteriolar (aID, µm) and venular internal diameter (vID, µm) and calculate arteriolar wall-to-lumen ratio (WLR), wall cross-sectional area (WCSA, µm2). One-way ANOVA for parametric variables and Kruskal-Wallis test for non-parametric variables were used for comparison among groups. A multivariate regression analysis including age, gender, BP, hGly and antihypertensive treatment was performed to calculate independent predictors of retinal remodeling.
Results
WT was increased with ageing (tertile1: 22.5 ± 3.2, tertile2: 24.2 ± 3.5, tertile 3: 25.2 ± 3.8, p = 0.001) and BP (hBP: 25.2 ± 4.1 vs nBP: 23.9 ± 3.7, p = 0.003). aID decreased with BP (hBP: 90.2 ± 13.4 vs nBP: 93.6 ± 11.6, p = 0.013) and increased with glycaemia (hGly: 97.7 ± 12.5 vs nGly: 93.6 ± 11.6, p = 0.002). A multivariate analysis showed independent association of hBP with WLR; hGly with WCSA; ageing with WLR and WCSA.
Conclusions
AOO non-invasively identifies retinal structural changes in human confirming that microvascular remodeling is exclusively related to hypertension, whereas vascular growth is related to ageing and hyperglycaemia.



Clin Res Cardiol: 29 Jun 2021; 110:959-970
Gallo A, Dietenbeck T, Giron A, Paques M, Kachenoura N, Girerd X
Clin Res Cardiol: 29 Jun 2021; 110:959-970 | PMID: 32494923
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Abstract

Activation of Aryl Hydrocarbon Receptor by ITE Improves Cardiac Function in Mice After Myocardial Infarction.

Seong E, Lee JH, Lim S, Park EH, ... Chung WS, Chang K

Background:
The immune and inflammatory responses play a considerable role in left ventricular remodeling after myocardial infarction (MI). Binding of AhR (aryl hydrocarbon receptor) to its ligands modulates immune and inflammatory responses; however, the effects of AhR in the context of MI are unknown. Therefore, we evaluated the potential association between AhR and MI by treating mice with a nontoxic endogenous AhR ligand, ITE (2-[1\'H-indole-3\'-carbonyl]-thiazole-4-carboxylic acid methyl ester). We hypothesized that activation of AhR by ITE in MI mice would boost regulatory T-cell differentiation, modulate macrophage activity, and facilitate infarct healing. Methods and Results Acute MI was induced in C57BL/6 mice by ligation of the left anterior descending coronary artery. Then, the mice were randomized to daily intraperitoneal injection of ITE (200 µg/mouse, n=19) or vehicle (n=16) to examine the therapeutic effects of ITE during the postinfarct healing process. Echocardiographic and histopathological analyses revealed that ITE-treated mice exhibited significantly improved systolic function (P<0.001) and reduced infarct size compared with control mice (P<0.001). In addition, we found that ITE increased regulatory T cells in the mediastinal lymph node, spleen, and infarcted myocardium, and shifted the M1/M2 macrophage balance toward the M2 phenotype in vivo, which plays vital roles in the induction and resolution of inflammation after acute MI. In vitro, ITE expanded the Foxp3+ (forkhead box protein P3-positive) regulatory T cells and tolerogenic dendritic cell populations.
Conclusions:
Activation of AhR by a nontoxic endogenous ligand, ITE, improves cardiac function after MI. Post-MI mice treated with ITE have a significantly lower risk of developing advanced left ventricular systolic dysfunction than nontreated mice. Thus, the results imply that ITE has a potential as a stimulator of cardiac repair after MI to prevent heart failure.




J Am Heart Assoc: 05 Jul 2021; 10:e020502
Seong E, Lee JH, Lim S, Park EH, ... Chung WS, Chang K
J Am Heart Assoc: 05 Jul 2021; 10:e020502 | PMID: 34157850
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Abstract

Can polarization of macrophage metabolism enhance cardiac regeneration?

Lantz C, Becker A, Thorp EB
While largely appreciated for their antimicrobial and repair functions, macrophages have emerged as indispensable for the development, homeostasis, and regeneration of tissue, including regeneration of the neonatal heart. Upon activation, mammalian neonatal macrophages express and secrete factors that coordinate angiogenesis, resolution of inflammation, and ultimately cardiomyocyte proliferation. This is contrary to adult macrophages in the adult heart, which are incapable of inducing significant levels of cardiac regeneration. The underlying mechanisms by which pro-regenerative macrophages are activated and regulated remain vague. A timely hypothesis is that macrophage metabolism contributes to this proliferative and regenerative potential. This is because we now appreciate the significant contributions of metabolites to immune cell programming and function, beyond solely bioenergetics. After birth, the metabolic milieu of the neonate is subject to significant alterations in oxygenation and nutrient supply, which will affect how metabolic substrates are catabolized. In this context, we discuss potential roles for select macrophage metabolic pathways during cardiac regeneration.

Copyright © 2018. Published by Elsevier Ltd.

J Mol Cell Cardiol: 18 Jul 2021; epub ahead of print
Lantz C, Becker A, Thorp EB
J Mol Cell Cardiol: 18 Jul 2021; epub ahead of print | PMID: 34293342
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Abstract

Proteomic Signature of Dysfunctional Circulating Endothelial Colony-Forming Cells of Young Adults.

Tan CMJ, Lewandowski AJ, Williamson W, Huckstep OJ, ... Leeson P, Bertagnolli M

Background:
A subpopulation of endothelial progenitor cells called endothelial colony-forming cells (ECFCs) may offer a platform for cellular assessment in clinical studies because of their remarkable angiogenic and expansion potentials in vitro. Despite endothelial cell function being influenced by cardiovascular risk factors, no studies have yet provided a comprehensive proteomic profile to distinguish functional (ie, more angiogenic and expansive cells) versus dysfunctional circulating ECFCs of young adults. The aim of this study was to provide a detailed proteomic comparison between functional and dysfunctional ECFCs. Methods and Results Peripheral blood ECFCs were isolated from 11 subjects (45% men, aged 27±5 years) using Ficoll density gradient centrifugation. ECFCs expressed endothelial and progenitor surface markers and displayed cobblestone-patterned morphology with clonal and angiogenic capacities in vitro. ECFCs were deemed dysfunctional if <1 closed tube formed during the in vitro tube formation assay and proliferation rate was <20%. Hierarchical functional clustering revealed distinct ECFC proteomic signatures between functional and dysfunctional ECFCs with changes in cellular mechanisms involved in exocytosis, vesicle transport, extracellular matrix organization, cell metabolism, and apoptosis. Targeted antiangiogenic proteins in dysfunctional ECFCs included SPARC (secreted protein acidic and rich in cysteine), CD36 (cluster of differentiation 36), LUM (lumican), and PTX3 (pentraxin-related protein PYX3).
Conclusions:
Circulating ECFCs with impaired angiogenesis and expansion capacities have a distinct proteomic profile and significant phenotype changes compared with highly angiogenic endothelial cells. Impaired angiogenesis in dysfunctional ECFCs may underlie the link between endothelial dysfunction and cardiovascular disease risks in young adults.




J Am Heart Assoc: 18 Jul 2021:e021119; epub ahead of print
Tan CMJ, Lewandowski AJ, Williamson W, Huckstep OJ, ... Leeson P, Bertagnolli M
J Am Heart Assoc: 18 Jul 2021:e021119; epub ahead of print | PMID: 34275329
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Impact:
Abstract

Intranasal delivery of interleukin-4 attenuates chronic cognitive deficits via beneficial microglial responses in experimental traumatic brain injury.

Pu H, Ma C, Zhao Y, Wang Y, ... Bennett MV, Chen J
Traumatic brain injury (TBI) is commonly followed by long-term cognitive deficits that severely impact the quality of life in survivors. Recent studies suggest that microglial/macrophage (Mi/MΦ) polarization could have multidimensional impacts on post-TBI neurological outcomes. Here, we report that repetitive intranasal delivery of interleukin-4 (IL-4) nanoparticles for 4 weeks after controlled cortical impact improved hippocampus-dependent spatial and non-spatial cognitive functions in adult C57BL6 mice, as assessed by a battery of neurobehavioral tests for up to 5 weeks after TBI. IL-4-elicited enhancement of cognitive functions was associated with improvements in the integrity of the hippocampus at the functional (e.g., long-term potentiation) and structural levels (CA3 neuronal loss, diffusion tensor imaging of white matter tracts, etc.). Mechanistically, IL-4 increased the expression of PPARγ and arginase-1 within Mi/MΦ, thereby driving microglia toward a global inflammation-resolving phenotype. Notably, IL-4 failed to shift microglial phenotype after TBI in Mi/MΦ-specific PPARγ knockout (mKO) mice, indicating an obligatory role for PPARγ in IL-4-induced Mi/MΦ polarization. Accordingly, post-TBI treatment with IL-4 failed to improve hippocampal integrity or cognitive functions in PPARγ mKO mice. These results demonstrate that administration of exogenous IL-4 nanoparticles stimulates PPARγ-dependent beneficial Mi/MΦ responses, and improves hippocampal function after TBI.



J Cereb Blood Flow Metab: 13 Jul 2021:271678X211028680; epub ahead of print
Pu H, Ma C, Zhao Y, Wang Y, ... Bennett MV, Chen J
J Cereb Blood Flow Metab: 13 Jul 2021:271678X211028680; epub ahead of print | PMID: 34259069
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Impact:
Abstract

Modulation of Sirt1 and FoxO1 on Hypothalamic Leptin-Mediated Sympathetic Activation and Inflammation in Diet-Induced Obese Rats.

Liu X, Zheng H

Background:
Hypothalamic leptin-mediated signaling contributes to the exaggerated sympatho-excitation and increased blood pressure in obesity-associated hypertension. The aim of the study was to investigate the roles of energy-sensing enzyme sirtuin1 (Sirt1) and forkhead box protein O1 (FoxO1) on the hypothalamic leptin-mediated high sympathetic nerve activity and inflammation in obesity. Methods and Results Sprague Dawley rats were fed with high-fat diet (HFD) for 12 weeks. In vivo, the potential of Srit1 and FoxO1 in the sympathetic effects of leptin was investigated via siRNA injection to knockdown Sirt1 or FoxO1 gene in the arcuate nucleus (ARCN) of hypothalamus in rats. In vitro, the effects of Sirt1 or FoxO1 on leptin-mediated inflammation were observed in proopiomelanocortin (POMC) and microglial cells. Knockdown Sirt1 by siRNA significantly reduced the renal sympathetic nerve activity (RSNA) and blood pressure responses to leptin injection in the ARCN in the HFD rats. Conversely, knockdown FoxO1 significantly enhanced the RSNA and blood pressure responses to leptin injection in the HFD rats. Knockdown Sirt1 reduced the levels of pro-inflammatory cytokines interleukin 6 (IL-6), tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β), C1q/TNF-related protein-1 (CTRP1), and immune cell infiltration in the ARCN in the HFD rats. Knockdown FoxO1 significantly increased the level of IL-6 in the ARCN of HFD rats. In cultured hypothalamic POMC and microglial cells, knockdown Sirt1 significantly reduced leptin-induced IL-6 expression, affected the levels of AMP-activated protein kinase (AMPK) and serine/threonine-specific protein kinase (Akt). Knockdown FoxO1 significantly increased leptin-induced IL-6 in both POMC cells and microglial cells.
Conclusions:
These data suggest that both Sirt1 and FoxO1 are the key modulators of leptin signaling in the hypothalamus contributed to the over sympathetic activation and inflammation in obesity.




J Am Heart Assoc: 19 Jul 2021; 10:e020667
Liu X, Zheng H
J Am Heart Assoc: 19 Jul 2021; 10:e020667 | PMID: 34259031
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Impact:
Abstract

Endothelial Spns2 and ApoM Regulation of Vascular Tone and Hypertension Via Sphingosine-1-Phosphate.

Del Gaudio I, Rubinelli L, Sasset L, Wadsack C, Hla T, Di Lorenzo A

Background:
Most of the circulating sphingosine-1-phosphate (S1P) is bound to ApoM (apolipoprotein M) of high-density lipoprotein (HDL) and mediates many beneficial effects of HDL on the vasculature via G protein-coupled S1P receptors. HDL-bound S1P is decreased in atherosclerosis, myocardial infarction, and diabetes mellitus. In addition to being the target, the endothelium is a source of S1P, which is transported outside of the cells by Spinster-2, contributing to circulating S1P as well as to local signaling. Mice lacking endothelial S1P receptor 1 are hypertensive, suggesting a vasculoprotective role of S1P signaling. This study investigates the role of endothelial-derived S1P and ApoM-bound S1P in regulating vascular tone and blood pressure. Methods and Results ApoM knockout (ApoM KO) mice and mice lacking endothelial Spinster-2 (ECKO-Spns2) were infused with angiotensin II for 28 days. Blood pressure, measured by telemetry and tail-cuff, was significantly increased in both ECKO-Spns2 and ApoM KO versus control mice, at baseline and following angiotensin II. Notably, ECKO-Spns2 presented an impaired vasodilation to flow and blood pressure dipping, which is clinically associated with increased risk for cardiovascular events. In hypertension, both groups presented reduced flow-mediated vasodilation and some degree of impairment in endothelial NO production, which was more evident in ECKO-Spns2. Increased hypertension in ECKO-Spns2 and ApoM KO mice correlated with worsened cardiac hypertrophy versus controls.
Conclusions:
Our study identifies an important role for Spinster-2 and ApoM-HDL in blood pressure homeostasis via S1P-NO signaling and dissects the pathophysiological impact of endothelial-derived S1P and ApoM of HDL-bound S1P in hypertension and cardiac hypertrophy.




J Am Heart Assoc: 08 Jul 2021:e021261; epub ahead of print
Del Gaudio I, Rubinelli L, Sasset L, Wadsack C, Hla T, Di Lorenzo A
J Am Heart Assoc: 08 Jul 2021:e021261; epub ahead of print | PMID: 34240614
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