<div><h4>Association of genetic risk and outcomes in patients with atrial fibrillation: interactions with early rhythm control in the EAST-AFNET4 trial.</h4><i>Kany S, Al-Taie C, Roselli C, Pirruccello JP, ... Ellinor PT, Kirchhof P</i><br /><b>Aims</b><br />The randomized Early Treatment of Atrial Fibrillation for Stroke Prevention Trial found that early rhythm control reduces cardiovascular events in patients with recently diagnosed atrial fibrillation (AF) compared with usual care. How genetic predisposition to AF and stroke interacts with early rhythm-control therapy is not known.<br /><b>Methods and results</b><br />Array genotyping and imputation for common genetic variants were performed. Polygenic risk scores (PRS) were calculated for AF (PRS-AF) and ischaemic stroke risk (PRS-stroke). The effects of PRS-AF and PRS-stroke on the primary outcome (composite of cardiovascular death, stroke, and hospitalization for acute coronary syndrome or worsening heart failure), its components, and recurrent AF were determined.A total of 1567 of the 2789 trial patients were analysed [793 randomized to early rhythm control; 774 to usual care, median age 71 years (65-75), 704 (44%) women]. Baseline characteristics were similar between randomized groups. Early rhythm control reduced the primary outcome compared with usual care [HR 0.67, 95% CI: (0.53, 0.84), P < 0.001]. The randomized intervention, early rhythm control, did not interact with PRS-AF (interaction P = 0.806) or PRS-stroke (interaction P = 0.765). PRS-AF was associated with recurrent AF [HR 1.08 (01.0, 1.16), P = 0.047]. PRS-stroke showed an association with the primary outcome [HR 1.13 (1.0, 1.27), P = 0.048], driven by more heart failure events [HR 1.23 (1.05-1.43), P = 0.010] without differences in stroke [HR 1.0 (0.75, 1.34), P = 0.973] in this well-anticoagulated cohort. In a replication analysis, PRS-stroke was associated with incident AF [HR 1.16 (1.14, 1.67), P < 0.001] and with incident heart failure in the UK Biobank [HR 1.08 (1.06, 1.10), P < 0.001]. The association with heart failure was weakened when excluding AF patients [HR 1.03 (1.01, 1.05), P = 0.001].<br /><b>Conclusions</b><br />Early rhythm control is effective across the spectrum of genetic AF and stroke risk. The association between genetic stroke risk and heart failure calls for research to understand the interactions between polygenic risk and treatment.<br /><b>Registration</b><br />ISRCTN04708680, NCT01288352, EudraCT2010-021258-20, www.easttrial.org.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 02 Jun 2023; epub ahead of print</small></div>

<div><h4>Modulation of lncRNA links endothelial glycocalyx to vascular dysfunction of tyrosine kinase inhibitor.</h4><i>Nukala SB, Jousma J, Yan G, Han Z, ... Lee WH, Ong SG</i><br /><b>Aims</b><br />Novel cancer therapies leading to increased survivorship of cancer patients have been negated by a concomitant rise in cancer therapies-related cardiovascular toxicities. Sunitinib, a first line multi receptor tyrosine kinase inhibitor (TKI), has been reported to cause vascular dysfunction although the initiating mechanisms contributing to this side effect remain unknown. Long non-coding RNAs (lncRNAs) are emerging regulators of biological processes in endothelial cells (ECs); however, their roles in cancer therapies-related vascular toxicities remain underexplored.<br /><b>Methods and results</b><br />We performed lncRNA expression profiling to identify potential lncRNAs that are dysregulated in human induced pluripotent stem cell-derived ECs (iPSC-ECs) treated with sunitinib. We show that the lncRNA hyaluronan synthase 2 antisense 1 (HAS2-AS1) is significantly diminished in sunitinib-treated iPSC-ECs. Sunitinib was found to downregulate HAS2-AS1 by an epigenetic mechanism involving hypermethylation. Depletion of HAS2-AS1 recapitulated sunitinib-induced detrimental effects on iPSC-ECs, whereas CRISPR-mediated activation of HAS2-AS1 reversed sunitinib-induced dysfunction. We confirmed that HAS2-AS1 stabilizes the expression of its sense gene HAS2 via an RNA/mRNA heteroduplex formation. Knockdown of HAS2-AS1 led to reduced synthesis of hyaluronic acid (HA) and upregulation of ADAMTS5, an enzyme involved in extracellular matrix degradation, resulting in disruption of the endothelial glycocalyx which is critical for ECs. In vivo, sunitinib-treated mice showed reduced coronary flow reserve, accompanied by a reduction in Has2os and degradation of the endothelial glycocalyx. Finally, we identified that treatment with high molecular-weight HA can prevent the deleterious effects of sunitinib both in vitro and in vivo by preserving the endothelial glycocalyx.<br /><b>Conclusions</b><br />Our findings highlight the importance of lncRNA-mediated regulation of the endothelial glycocalyx as an important determinant of sunitinib-induced vascular toxicity and reveal potential novel therapeutic avenues to attenuate sunitinib-induced vascular dysfunction.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 02 Jun 2023; epub ahead of print</small></div>

<div><h4>Cardiovascular complications in chronic kidney disease - A review from the European Renal and Cardiovascular Medicine Working Group (EURECA-m) of the European Renal Association (ERA).</h4><i>Zoccali C, Mallamaci F, Adamczak M, de Oliveira RB, ... Vanholder R, Wiecek A</i><br /><AbstractText>Chronic kidney disease (CKD) is classified into 5 stages with kidney failure being the most severe stage (stage G5). CKD conveys a high risk for coronary artery disease, heart failure, arrhythmias, and sudden cardiac death. Cardiovascular complications are the most common causes of death in patients with kidney failure (stage G5) who are maintained on regular dialysis treatment. Because of the high death rate attributable to cardiovascular (CV) disease, most patients with progressive CKD die before reaching kidney failure. Classical risk factors implicated in CV disease are involved in the early stages of CKD. In intermediate and late stages, non-traditional risk factors, including iso-osmotic and non-osmotic sodium retention, volume expansion, anaemia, inflammation, malnutrition, sympathetic overactivity, mineral bone disorders, accumulation of a class of endogenous compounds called \"uremic toxins\" and a variety of hormonal disorders are the main factors that accelerate the progression of CV disease in these patients. Arterial disease in CKD patients is characterized by an almost unique propensity to calcification and vascular stiffness. Left ventricular hypertrophy, a major risk factor for heart failure occurs early in CKD and reaches a prevalence of 70%-80% in patients with kidney failure. Recent clinical trials have shown the potential benefits of hypoxia-inducible factor prolyl hydroxylase inhibitors, especially as an oral agent in CKD patients. Likewise, the value of proactively administered intravenous iron for safely treating anaemia in dialysis patients has been shown. Sodium/glucose cotransporter- 2 (SGLT2) inhibitors are now fully emerged as a class of drugs that substantially reduces the risk for CV complications in patients who are already being treated with adequate doses of inhibitors of the renin-angiotensin system. Concerted efforts are being made by major scientific societies to advance basic and clinical research on CV disease in patients with CKD, a research area that remains insufficiently explored.</AbstractText><br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 30 May 2023; epub ahead of print</small></div>

<div><h4>NLRP3 inflammasome and interleukin-1 contributions to COVID-19-associated coagulopathy and immunothrombosis.</h4><i>Potere N, Garrad E, Kanthi Y, Di Nisio M, ... De Caterina R, Abbate A</i><br /><AbstractText>Immunothrombosis - immune-mediated activation of coagulation - is protective against pathogens, but excessive immunothrombosis can result in pathological thrombosis and multiorgan damage, as in severe Coronavirus Disease 2019 (COVID-19). The NACHT-, LRR- and pyrin domain-containing protein 3 (NLRP3) inflammasome produces major proinflammatory cytokines of the interleukin (IL)-1 family, IL-1β and IL-18, and induces pyroptotic cell death. Activation of the NLRP3 inflammasome pathway also promotes immunothrombotic programs including release of neutrophil extracellular traps and tissue factor by leukocytes, and prothrombotic responses by platelets and the vascular endothelium. NLRP3 inflammasome activation occurs in patients with COVID-19 pneumonia. In preclinical models, NLRP3 inflammasome pathway blockade restrains COVID-19-like hyperinflammation and pathology. Anakinra, recombinant human IL-1 receptor antagonist, showed safety and efficacy, and is approved for the treatment of hypoxemic COVID-19 patients with early signs of hyperinflammation. The non-selective NLRP3 inhibitor colchicine reduced hospitalization and death in a subgroup of COVID-19 outpatients, but is not approved for the treatment of COVID-19. Additional COVID-19 trials testing NLRP3 inflammasome pathway blockers are inconclusive or ongoing. We herein outline the contribution of immunothrombosis to COVID-19-associated coagulopathy, and review preclinical and clinical evidence suggesting an engagement of the NLRP3 inflammasome pathway in the immunothrombotic pathogenesis of COVID-19. We also summarize current efforts to target the NLRP3 inflammasome pathway in COVID-19, and discuss challenges, unmet gaps and the therapeutic potential that inflammasome-targeted strategies may provide for inflammation-driven thrombotic disorders including COVID-19.</AbstractText><br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 30 May 2023; epub ahead of print</small></div>

<div><h4>Macrophage angiotensin-converting enzyme (ACE) reduces atherosclerosis by increasing PPARα and fundamentally changing lipid metabolism.</h4><i>Cao D, Khan Z, Li X, Saito S, ... Okwan-Duodu D, Bernstein KE</i><br /><b>Aims</b><br />The metabolic failure of macrophages to adequately process lipid is central to the etiology of atherosclerosis. Here, we examine the role of macrophage angiotensin converting enzyme (ACE) in a mouse model of PCSK9 induced atherosclerosis.<br /><b>Methods and results</b><br />Atherosclerosis in mice was induced with AAV-PCSK9 and a high fat diet. Animals with increased macrophage ACE (ACE 10/10 mice) have a marked reduction in atherosclerosis vs. WT mice. Macrophages from both the aorta and peritoneum of ACE 10/10 express increased PPARα and have a profoundly altered phenotype to process lipids characterized by higher levels of the surface scavenger receptor CD36, increased uptake of lipid, increased capacity to transport long chain fatty acids into mitochondria, higher oxidative metabolism and lipid β-oxidation as determined using 13C isotope tracing, increased cell ATP, increased capacity for efferocytosis, increased concentrations of the lipid transporters ABCA1 and ABCG1, and increased cholesterol efflux. These effects are mostly independent of angiotensin II. Human THP-1 cells, when modified to express more ACE, increase expression of PPARα, increase cell ATP and acetyl-CoA, and increase cell efferocytosis.<br /><br /><b>Conclusion:</b><br/>and translational perspective</b><br />Increased macrophage ACE expression enhances macrophage lipid metabolism, cholesterol efflux, efferocytosis, and it reduces atherosclerosis. This has implications for the treatment of cardiovascular disease with angiotensin II receptor antagonists (ARBs) vs. ACE inhibitors.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 24 May 2023; epub ahead of print</small></div>

<div><h4>Inhibition of the extracellular enzyme ADAMTS4 prevents cardiac fibrosis and dysfunction.</h4><i>Vistnes M, Erusappan PM, Sasi A, Nordén ES, ... Sjaastad I, Christensen G</i><br /><b>Aims</b><br />Heart failure is a condition with high mortality rates, and there is a lack of therapies that directly target maladaptive changes in the extracellular matrix (ECM), such as fibrosis. We investigated whether the ECM enzyme known as A disintegrin and metalloprotease with thrombospondin motif (ADAMTS) 4 might serve as a therapeutic target in treatment of heart failure and cardiac fibrosis.<br /><b>Methods and results</b><br />The effects of pharmacological ADAMTS4 inhibition on cardiac function and fibrosis were examined in rats exposed to cardiac pressure overload. Disease mechanisms affected by the treatment were identified based on changes in the myocardial transcriptome. Following aortic banding (AB), rats receiving an ADAMTS inhibitor, with high inhibitory capacity for ADAMTS4, showed substantially better cardiac function than vehicle-treated rats, including ∼30 % reduction in E/e\' and left atrial diameter, indicating an improvement in diastolic function. ADAMTS inhibition also resulted in a marked reduction in myocardial collagen content and a downregulation of transforming growth factor (TGF) β target genes. The mechanism for the beneficial effects of ADAMTS inhibition was further studied in cultured human cardiac fibroblasts producing mature ECM. ADAMTS4 caused a 50% increase in the TGF-β levels in the medium. Simultaneously, ADAMTS4 elicited a not previously known cleavage of TGF-β-binding proteins, i.e. latent binding protein of TGF-β (LTBP1) and extra domain A (EDA)-fibronectin. These effects were abolished by the ADAMTS inhibitor. In failing human hearts, we observed a marked increase in ADAMTS4 expression and cleavage activity.<br /><b>Conclusion</b><br />Inhibition of ADAMTS4 improves cardiac function and reduces collagen accumulation in rats with cardiac pressure overload, possibly through a not previously known cleavage of molecules that control TGF-β availability. Targeting ADAMTS4 may serve as a novel strategy in heart failure treatment, in particular in heart failure with fibrosis and diastolic dysfunction.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 22 May 2023; epub ahead of print</small></div>

<div><h4>Myeloid CD40 deficiency reduces atherosclerosis by impairing macrophages\' transition into a pro-inflammatory state.</h4><i>Bosmans LA, van Tiel CM, Aarts SABM, Willemsen L, ... Shami A, Lutgens E</i><br /><b>Aims</b><br />CD40 and its ligand, CD40L, play a critical role in driving atherosclerotic plaque development. Disrupted CD40-signalling reduces experimental atherosclerosis and induces a favourable stable plaque phenotype. We recently showed that small molecule-based inhibition of CD40-tumour necrosis factor receptor associated factor-6 interactions attenuates atherosclerosis in hyperlipidaemic mice via macrophage-driven mechanisms. The present study aims to detail the function of myeloid CD40 in atherosclerosis using myeloid-specific CD40-deficient mice.<br /><b>Method and results</b><br />Cd40flox/flox and LysM-cre Cd40flox/flox mice on an Apoe-/- background were generated (CD40wt and CD40mac-/-, respectively). Atherosclerotic lesion size, as well as plaque macrophage content, was reduced in CD40mac-/- compared to CD40wt mice, and their plaques displayed a reduction in necrotic core size. Transcriptomics analysis of the CD40mac-/- atherosclerotic aorta revealed downregulated pathways of immune pathways and inflammatory responses. Loss of CD40 in macrophages changed the representation of aortic macrophage subsets. Mass cytometry analysis revealed a higher content of a subset of alternative or resident-like CD206+CD209b- macrophages in the atherosclerotic aorta of CD40mac-/- compared to CD40wt mice. RNA-sequencing of bone marrow-derived macrophages of CD40mac-/- mice demonstrated upregulation of genes associated with alternatively activated macrophages (including Folr2, Thbs1, Sdc1, and Tns1).<br /><b>Conclusions</b><br />We here show that absence of CD40 signalling in myeloid cells reduces atherosclerosis and limits systemic inflammation by preventing a shift in macrophage polarization towards pro-inflammatory states. Our study confirms the merit of macrophage-targeted inhibition of CD40 as a valuable therapeutic strategy to combat atherosclerosis.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 22 May 2023; 119:1146-1160</small></div>

<div><h4>Transforming growth factor-β2 is associated with atherosclerotic plaque stability and lower risk for cardiovascular events.</h4><i>Edsfeldt A, Singh P, Matthes F, Tengryd C, ... Sun J, Gonçalves I</i><br /><b>Aims</b><br />Transforming growth factor-beta (TGF-β) exists in three isoforms TGF-β1, -β2 and -β3. TGF-β1 has been suggested to be important for maintaining plaque stability, yet the role of TGF-β2 and -β3 in atherosclerosis remains to be investigated.<br /><b>Objective</b><br />This study explores the association of these three isoforms of TGF-β with plaque stability in the human atherosclerotic disease.<br /><b>Methods and results</b><br />TGF-β1, -β2 and -β3 proteins were quantified in 223 human carotid plaques by immunoassays. Indications for the endarterectomy were: symptomatic carotid plaque with stenosis >70% or without symptoms and >80% stenosis. Plaque mRNA levels were assessed by RNA sequencing. Plaque components and extracellular matrix were measured histologically and biochemically. Matrix metalloproteinases were measured with ELISA. Monocyte chemoattractant protein-1 (MCP-1) was measured with immunoassays. The effect of TGF-β2 on inflammation and protease activity was investigated in vitro using THP-1 and RAW264.7 macrophages. Patients were followed longitudinally for cardiovascular events.TGF-β2 was the most abundant isoform and was increased at both protein and mRNA levels in asymptomatic plaques. TGF-β2 was the main determinant separating asymptomatic plaques in an Orthogonal Projections to Latent Structures Discriminant Analysis. TGF-β2 correlated positively to features of plaque stability and inversely to markers of plaque vulnerability. TGF-β2 was the only isoform inversely correlated to the matrix-degrading matrix metalloproteinase-9 and inflammation in the plaque tissue. In vitro, TGF-β2 pre-treatment reduced MCP-1 gene and protein levels as well as matrix metalloproteinase-9 gene levels and activity. Patients with plaques with high TGF-β2 levels had a lower risk to suffer from future cardiovascular events.<br /><b>Conclusions</b><br />TGF-β2 is the most abundant TGF-β isoform in human plaques and may maintain plaque stability by decreasing inflammation and matrix degradation.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 18 May 2023; epub ahead of print</small></div>

<div><h4>PRMT5-mediated arginine methylation stabilizes KLF4 to accelerate neointimal formation.</h4><i>Liu H, Dong X, Jia K, Yuan B, ... Sun J, Pan LL</i><br /><b>Aims</b><br />Accumulating evidence supports an indispensable role of protein arginine methyltransferase 5 (PRMT5) in the pathological progression of several human cancers. As an important enzyme regulating protein methylation, how PRMT5 participates in vascular remodeling remains unknown. To investigate the role and underlying mechanism of PRMT5 in neointimal formation and to evaluate its potential as an effective therapeutic target for the condition.<br /><b>Methods and results</b><br />Aberrant PRMT5 overexpression was positively correlated with clinical carotid arterial stenosis. Vascular smooth muscle cells (SMC)-specific PRMT5 knockout inhibited intimal hyperplasia with enhanced expression of contractile markers in mice. Conversely, PRMT5 overexpression inhibited SMC contractile markers and promoted intimal hyperplasia. Furthermore, we showed that PRMT5 promoted SMC phenotypic switching through stabilizing Kruppel-like factor 4 (KLF4). Mechanistically, PRMT5-mediated KLF4 methylation inhibited ubiquitin-dependent proteolysis of KLF4, leading to a disruption of myocardin (MYOCD) -serum response factor (SRF) interaction and MYOCD-SRF-mediated the transcription of SMC contractile markers.<br /><b>Conclusions</b><br />Our data demonstrated that PRMT5 critically mediated vascular remodeling by promoting KLF4-mediated SMC phenotypic conversion and consequently the progression of intimal hyperplasia. Therefore, PRMT5 may represent a potential therapeutic target for intimal hyperplasia-associated vascular diseases.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 18 May 2023; epub ahead of print</small></div>

<div><h4>Takotsubo syndrome: getting closer to its causes.</h4><i>Akhtar MM, Cammann VL, Templin C, Ghadri JR, Lüscher TF</i><br /><AbstractText>Takotsubo syndrome (TTS) accounts for between 1 and 4% of cases presenting clinically as an acute coronary syndrome. It typically presents as a transient cardiac phenotype of left ventricular dysfunction with spontaneous recovery. More dramatic presentations may include cardiogenic shock or cardiac arrest. Despite progress in the understanding of the condition since its first description in 1990, considerable questions remain into understanding underlying pathomechanisms. In this review article, we describe the current published data on potential underlying mechanisms associated with the onset of TTS including sympathetic nervous system over-stimulation, structural and functional alterations in the central nervous system, catecholamine secretion, alterations in the balance and distribution of adrenergic receptors, the additive impact of hormones including oestrogen, epicardial coronary or microvascular spasm, endothelial dysfunction, and genetics as potentially contributing to the cascade of events leading to the onset. These pathomechanisms provide suggestions for novel potential therapeutic strategies in patients with TTS including the role of cognitive behavioural therapy, beta-blockers, and endothelin-A antagonists. The underlying mechanism of TTS remains elusive. In reality, physical or emotional stressors likely trigger through the amygdala and hippocampus a central neurohumoral activation with the local and systemic secretion of excess catecholamine and other neurohormones, which exert its effect on the myocardium through a metabolic switch, altered cellular signalling, and endothelial dysfunction. These complex pathways exert a regional activation in the myocardium through the altered distribution of adrenoceptors and density of autonomic innervation as a protective mechanism from myocardial apoptosis. More research is needed to understand how these different complex mechanisms interact with each other to bring on the TTS phenotype.</AbstractText><br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 15 May 2023; epub ahead of print</small></div>

<div><h4>Human antigen R regulates autophagic flux by stabilizing autophagy-associated mRNA in calcific aortic valve disease.</h4><i>Fang J, Qian Y, Chen J, Xu D, ... Wang J, Liu X</i><br /><b>Aims</b><br />The incidence of calcific aortic valve disease (CAVD) has risen over the last decade and is expected to continue rising; however, pharmacological approaches have proven ineffective. In this study, we evaluated the role and underlying mechanisms of human antigen R (HuR)-mediated post-transcriptional regulation in CAVD.<br /><b>Methods and results</b><br />We found that HuR was significantly upregulated in human calcified aortic valves and primary aortic valvular interstitial cells (VICs) following osteogenic stimulation. Subsequent functional studies revealed that HuR silencing ameliorated calcification both in vitro and in vivo. For the first time, we demonstrated that HuR directly interacted with the transcript of phosphatidylinositol-5-phosphate 4-kinase, type II, alpha (PIP4K2A), which mediates phosphatidylinositol signaling, facilitates autophagy, and act as an mRNA stabilizer. HuR positively modulated PIP4K2A expression at the post-transcriptional level, and consequently influenced the AKT/mTOR/ATG13 pathway to regulate autophagy and CAVD progression.<br /><b>Conclusion</b><br />Our study provides new insights into the post-transcriptional regulatory role of HuR in modulating autophagy-positive factors to regulate the pathogenesis of CAVD. Our findings highlight the potential of HuR as an innovative therapeutic target in CAVD treatment.<br /><b>Translational perspective</b><br />As the incidence of CAVD has been rising over the past decade with only invasive, expensive, and risky interventions available, it is imperative to explore novel approaches to halt aortic valve calcification. Currently, regulation of CAVD pathogenesis is not fully understood. HuR, an RNA-binding protein, and autophagy are thought to play key roles in CAVD; however, the precise underlying mechanisms have not been elucidated. Therefore, we believe that the findings of this study are relevant to the field because they provide direct evidence of a critical role for HuR, hVIC phenotypic transition, and autophagy in CAVD and suggest a novel target for hindering CAVD progression.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 15 May 2023; epub ahead of print</small></div>

<div><h4>Monocyte Heterogeneity in Cardiovascular Disease.</h4><i>Ruder AV, Wetzels SMW, Temmerman L, Biessen EAL, Goossens P</i><br /><AbstractText>Monocytes circulate the vasculature at steady state and are recruited to sites of inflammation where they differentiate into macrophages (MФ) to replenish tissue-resident MФ populations and engage in the development of cardiovascular disease (CVD). Monocytes display considerable heterogeneity, currently reflected by a nomenclature based on their expression of cluster of differentiation (CD) 14 and CD16, distinguishing CD14++CD16- classical (cMo), CD14++CD16+ intermediate (intMo) and CD14 + CD16++ non-classical (ncMo) monocytes. Several reports point to shifted subset distributions in the context of CVD, with significant association of intMo numbers with atherosclerosis, myocardial infarction, and heart failure. However, clear indications of their causal involvement as well as their predictive value for CVD are lacking. As recent high-parameter cytometry and single-cell RNA sequencing (scRNA-Seq) studies suggest an even higher degree of heterogeneity, better understanding of the functionalities of these subsets is pivotal. Considering their high heterogeneity, surprisingly little is known about functional differences between MФ originating from monocytes belonging to different subsets, and implications thereof for CVD pathogenesis. This paper provides an overview of recent findings on monocyte heterogeneity in the context of homeostasis and disease as well as functional differences between the subsets and their potential to differentiate into MФ, focusing on their role in vessels and the heart. The emerging paradigm of monocyte heterogeneity transcending the current tripartite subset division argues for an updated nomenclature and functional studies to substantiate marker-based subdivision and to clarify subset-specific implications for CVD.</AbstractText><br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 10 May 2023; epub ahead of print</small></div>

<div><h4>Nedd4-2 upregulation is associated with ACE2 ubiquitination in hypertension.</h4><i>Mohammed M, Ogunlade B, Elgazzaz M, Berdasco C, ... Lazartigues E, Filipeanu CM</i><br /><b>Background</b><br />ACE2 is a critical component of the compensatory renin-angiotensin system that is downregulated during the development of hypertension, possibly via ubiquitination. However, little is known about the mechanisms involved in ACE2 ubiquitination in neurogenic hypertension. This study aimed at identifying ACE2 ubiquitination partners, establish causal relationships, clinical relevance and test a gene therapy strategy to mitigate ACE2 ubiquitination in neurogenic hypertension.<br /><b>Methods and results</b><br />Bioinformatics and proteomics were combined to identify E3 ubiquitin ligases associated with ACE2 ubiquitination in chronically hypertensive mice. In vitro gain/loss of function experiments assessed ACE2 expression and activity to validate the interaction between ACE2 and the identified E3 ligase. Mutation experiments were further used to generate a ubiquitination-resistant ACE2 mutant (ACE2-5R). Optogenetics, blood pressure telemetry, pharmacological blockade of GABAA receptors in mice expressing ACE2-5R in the bed nucleus of the stria terminalis (BNST) and capillary Western were used to assess the role of ACE2 ubiquitination in neurogenic hypertension.Ubiquitination was first validated as leading to ACE2 downregulation and Nedd4-2 identified as a E3 ligase up-regulated in hypertension and promoting ACE2 ubiquitination. Mutation of lysine residues in the C-terminal of ACE2 was associated with increased activity and resistance to Ang-II-mediated degradation. Mice transfected with ACE2-5R in the BNST exhibited enhanced GABAergic input to the paraventricular nucleus (PVN) and reduction of hypertension. ACE2-5R expression was associated with reduced Nedd4-2 levels in the BNST.<br /><b>Conclusions</b><br />Our data identify Nedd4-2 as the first E3 ubiquitin ligase involved in ACE2 ubiquitination in Ang-II-mediated hypertension. We demonstrate the pivotal role of ACE2 on GABAergic neurons in the maintenance of a tonic inhibitory tone to the PVN, and the regulation of pre-sympathetic activity. These findings provide a new working model where Nedd4-2 could contribute to ACE2 ubiquitination leading to the development of neurogenic hypertension and highlight potential novel therapeutic strategies.<br /><b>Translational perspective</b><br />While ACE2 conversion of Ang-II to Ang-(1-7) is supposed to limit the overactivity of the renin-angiotensin system (RAS), the enzyme is downregulated during the development of hypertension. As antihypertensive RAS blockers on the market only provide limited control of BP among hypertensive patients, understanding the mechanisms responsible for this blunted compensation provides new possible targets for the treatment of hypertension. In this study we show that Nedd4-2 upregulation is associated with ACE2 ubiquitination while prevention of this post-translational modification prevents the development of hypertension. Accordingly, targeting of ACE2 ubiquitination provides a new treatment strategy to reduce hypertension.<br /><br />Published by Oxford University Press on behalf of the European Society of Cardiology 2023.<br /><br /><small>Cardiovasc Res: 10 May 2023; epub ahead of print</small></div>

<div><h4>Endothelial mechanobiology in atherosclerosis.</h4><i>Wang X, Shen Y, Shang M, Liu X, Munn LL</i><br /><AbstractText>Cardiovascular disease (CVD) is a serious health challenge, causing more deaths world-wide than cancer. The vascular endothelium, which forms the inner lining of blood vessels, plays a central role in maintaining vascular integrity and homeostasis and is in direct contact with the blood flow. Research over the past century has shown that mechanical perturbations of the vascular wall contribute to formation and progression of atherosclerosis. While the straight part of the artery is exposed to sustained laminar flow and physiological high shear stress, flow near branch points or in curved vessels can exhibit \"disturbed\" flow. Clinical studies as well as carefully controlled in vitro analyses have confirmed that these regions of disturbed flow, which can include low shear stress, recirculation, oscillation, or lateral flow, are preferential sites of atherosclerotic lesion formation. Because of their critical role in blood flow homeostasis, vascular endothelial cells (ECs) have mechanosensory mechanisms that allow them to react rapidly to changes in mechanical forces, and to execute context-specific adaptive responses to modulate EC functions. This review summarizes the current understanding of endothelial mechanobiology, which can guide the identification of new therapeutic targets to slow or reverse the progression of atherosclerosis.</AbstractText><br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 10 May 2023; epub ahead of print</small></div>

<div><h4>An interferon gamma response signature links myocardial aging and immunosenescence.</h4><i>Ashour D, Rebs S, Arampatzi P, Saliba AE, ... Streckfuß-Bömeke K, Ramos GC</i><br /><b>Aims</b><br />Aging entails profound immunological transformations that can impact myocardial homeostasis and predispose to heart failure. However, preclinical research in the immune-cardiology field is mostly conducted in young healthy animals, which potentially weakens its translational relevance. Herein, we sought to investigate how the aging T-cell compartment associates with changes in myocardial cell biology in aged mice.<br /><b>Methods and results</b><br />We phenotyped the antigen-experienced effector/memory T cells purified from heart-draining lymph nodes of 2-, 6-, 12-, and 18-month-old C57BL/6J mice using single-cell RNA/T cell receptor (TCR) sequencing (sc-seq). Simultaneously, we profiled all non-cardiomyocyte cell subsets purified from 2- and 18-month-old hearts and integrated our data with publicly available cardiomyocyte sc-seq datasets. Some of these findings were confirmed at the protein level by flow cytometry. With aging, the heart-draining lymph node and myocardial T cells underwent clonal expansion and exhibited an up-regulated pro-inflammatory transcription signature, marked by an increased interferon-γ (IFN-γ) production. In parallel, all major myocardial cell populations showed increased IFN-γ responsive signature with aging. In the aged cardiomyocytes, a stronger IFN-γ response signature was paralleled by the dampening of expression levels of transcripts related to most metabolic pathways, especially oxidative phosphorylation. Likewise, induced pluripotent stem cells-derived cardiomyocytes (iPSC-CM) exposed to chronic, low grade IFN-γ treatment showed a similar inhibition of metabolic activity.<br /><b>Conclusions</b><br />By investigating the paired age-related alterations in the T cells found in the heart and its draining lymph nodes, we provide evidence for increased myocardial IFN-γ signaling with age, which is associated with inflammatory and metabolic shifts typically seen in heart failure.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 04 May 2023; epub ahead of print</small></div>

<div><h4>Targeting DNA damage response in cardiovascular diseases: from pathophysiology to therapeutic implications.</h4><i>Wu L, Sowers JR, Zhang Y, Ren J</i><br /><AbstractText>Cardiovascular diseases (CVDs) arise from a complex interplay among genomic, proteomic, and metabolomic abnormalities. Emerging evidence has recently consolidated the presence of robust DNA damage in a variety of cardiovascular disorders. DNA damage triggers a series of cellular responses termed DNA damage response (DDR) including detection of DNA lesions, cell cycle arrest, DNA repair, cellular senescence, and apoptosis, in all organ systems including hearts and vasculature. Although transient DDR in response to temporary DNA damage can be beneficial for cardiovascular function, persistent activation of DDR promotes the onset and development of CVDs. Moreover, therapeutic interventions that target DNA damage and DDR have the potential to attenuate cardiovascular dysfunction and improve disease outcome. In this review, we will discuss molecular mechanisms of DNA damage and repair in the onset and development of CVDs, and explore how DDR in specific cardiac cell types contributes to CVDs. Moreover, we will highlight the latest advances regarding the potential therapeutic strategies targeting DNA damage signalling in CVDs.</AbstractText><br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 02 May 2023; 119:691-709</small></div>

<div><h4>Cellular interplay between cardiomyocytes and non-myocytes in diabetic cardiomyopathy.</h4><i>Phang RJ, Ritchie RH, Hausenloy DJ, Lees JG, Lim SY</i><br /><AbstractText>Patients with Type 2 diabetes mellitus (T2DM) frequently exhibit a distinctive cardiac phenotype known as diabetic cardiomyopathy. Cardiac complications associated with T2DM include cardiac inflammation, hypertrophy, fibrosis, and diastolic dysfunction in the early stages of the disease, which can progress to systolic dysfunction and heart failure. Effective therapeutic options for diabetic cardiomyopathy are limited and often have conflicting results. The lack of effective treatments for diabetic cardiomyopathy is due in part, to our poor understanding of the disease development and progression, as well as a lack of robust and valid preclinical human models that can accurately recapitulate the pathophysiology of the human heart. In addition to cardiomyocytes, the heart contains a heterogeneous population of non-myocytes including fibroblasts, vascular cells, autonomic neurons, and immune cells. These cardiac non-myocytes play important roles in cardiac homeostasis and disease, yet the effect of hyperglycaemia and hyperlipidaemia on these cell types is often overlooked in preclinical models of diabetic cardiomyopathy. The advent of human-induced pluripotent stem cells provides a new paradigm in which to model diabetic cardiomyopathy as they can be differentiated into all cell types in the human heart. This review will discuss the roles of cardiac non-myocytes and their dynamic intercellular interactions in the pathogenesis of diabetic cardiomyopathy. We will also discuss the use of sodium-glucose cotransporter 2 inhibitors as a therapy for diabetic cardiomyopathy and their known impacts on non-myocytes. These developments will no doubt facilitate the discovery of novel treatment targets for preventing the onset and progression of diabetic cardiomyopathy.</AbstractText><br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 02 May 2023; 119:668-690</small></div>

<div><h4>Lipoprotein(a) and calcific aortic valve disease initiation and progression: a systematic review and meta-analysis.</h4><i>Pantelidis P, Oikonomou E, Lampsas S, Zakynthinos GE, ... Tousoulis D, Vavouranakis M</i><br /><AbstractText>Although evidence indicates the association of lipoprotein(a) [Lp(a)] with atherosclerosis, the link with calcific aortic valve disease (CAVD) is unclear. This systematic review and meta-analysis explores the connection between Lp(a) and aortic valve calcification (AVC) and stenosis (AVS). We included all relevant studies, indexed in eight databases, up to February 2023. A total of 44 studies (163,139 subjects) were included, with 16 of them being further meta-analysed. Despite considerable heterogeneity, most studies support the relationship between Lp(a) and CAVD, especially in younger populations, with evidence of early aortic valve micro-calcification in elevated-Lp(a) populations. The quantitative synthesis showed higher Lp(a) levels, by 22.63 nmol/L (95% CI: 9.98-35.27), for patients with AVS, while meta-regressing the data revealed smaller Lp(a) differences for older populations with a higher proportion of females. The meta-analysis of eight studies providing genetic data, revealed that the minor alleles of both rs10455872 and rs3798220 LPA gene loci were associated with higher risk for AVS (pooled odds ratio 1.42; 95% CI: 1.34-1.50 and 1.27; 95% CI: 1.09-1.48, respectively). Importantly, high-Lp(a) individuals displayed not only faster AVS progression, by a mean difference of 0.09 m/s/year (95% CI: 0.09-0.09), but also a higher risk of serious adverse outcomes, including death (pooled hazard ratio 1.39; 95% CI: 1.01-1.90). These summary findings highlight the effect of Lp(a) on CAVD initiation, progression and outcomes, and support the early onset of Lp(a)-related subclinical lesions before clinical evidence.</AbstractText><br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 20 Apr 2023; epub ahead of print</small></div>

<div><h4>Arterial wall inflammation assessed by 18F-FDG-PET/CT is higher in individuals with type 1 diabetes and associated with circulating inflammatory proteins.</h4><i>Janssen AWM, van Heck JIP, Stienstra R, Aarntzen EHJG, ... Riksen NP, Tack CJ</i><br /><b>Aims</b><br />Here we investigate whether chronic hyperglycaemia in T1D is associated with a pro-inflammatory immune signature and with arterial wall inflammation, driving the development of atherosclerosis.<br /><b>Methods and results</b><br />We recruited patients with T1D (n = 41), and healthy age-, sex- and BMI-matched controls (n = 20). Arterial wall inflammation and hematopoietic activity was measured with 2\'-deoxy-2\'-(18F)-fluoro-D-glucose (18F-FDG) positron emission tomography/computed tomography (PET/CT). In addition, flow cytometry of circulating leukocytes was performed as well as targeted proteomics to measure circulating inflammatory markers. 18F-FDG uptake in the wall of the abdominal aorta, carotid arteries and iliac arteries was higher in T1D compared to the healthy controls. Also, 18F-FDG uptake in the bone marrow and spleen was higher in T1D patients. CCR2 and CD36 expression on circulating monocytes was higher in T1D patients, as well as several circulating inflammatory proteins. In addition, several circulating inflammatory markers (OPG, TGF-alpha, CX3CL1 and CSF-1) displayed a positive correlation with FDG uptake. Within T1D, no differences were found between people with a high and low HbA1c.<br /><b>Conclusion</b><br />Our findings strengthen the concept that chronic hyperglycaemia in T1D induces inflammatory changes that fuel arterial wall inflammation leading to atherosclerosis. The degree of hyperglycaemia appears to play a minor role in driving this inflammatory response in patients with T1D.<br /><b>Translational perspective</b><br />Arterial wall inflammation is associated with increased levels of several circulating inflammatory markers suggesting that these proteins are directly involved in driving this process yet may also serve as future biomarkers to identify patients with T1D at risk for the development of CVD. And could potentially be future treatment targets in reducing the risk of CVD in people with T1D.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 20 Apr 2023; epub ahead of print</small></div>

<div><h4>Lem2 is essential for cardiac development by maintaining nuclear integrity.</h4><i>Ross JA, Arcos-Villacis N, Battey E, Boogerd C, ... Shah AM, Stroud MJ</i><br /><b>Aims</b><br />Nuclear envelope integrity is essential for compartmentalisation of nucleus and cytoplasm. Importantly, mutations in genes encoding nuclear envelope and associated proteins are the second-highest cause of familial dilated cardiomyopathy. One such nuclear envelope protein that causes cardiomyopathy in humans and affects mouse heart development is Lem2. However, its role in heart remains poorly understood.<br /><b>Methods and results</b><br />We generated mice in which Lem2 was specifically ablated either in embryonic cardiomyocytes (Lem2 cKO) or adult cardiomyocytes (Lem2 iCKO) and carried out detailed physiological, tissue and cellular analyses. High resolution episcopic microscopy was used for 3D reconstructions and detailed morphological analyses. RNA-sequencing and immunofluorescence identified altered pathways and cellular phenotypes, and cardiomyocytes were isolated to interrogate nuclear integrity in more detail. In addition, echocardiography provided physiological assessment of Lem2 iCKO adult mice.We found that Lem2 was essential for cardiac development, and hearts from Lem2 cKO mice were morphologically and transcriptionally underdeveloped. Lem2 cKO hearts displayed high levels of DNA damage, nuclear rupture, and apoptosis. Crucially, we found that these defects were driven by muscle contraction as they were ameliorated by inhibiting myosin contraction and L-type calcium channels. Conversely, reducing Lem2 levels to ∼45% in adult cardiomyocytes did not lead to overt cardiac dysfunction up to 18 months of age.<br /><b>Conclusions</b><br />Our data suggest that Lem2 is critical for integrity at the nascent nuclear envelope in fetal hearts, and protects the nucleus from the mechanical forces of muscle contraction. In contrast, the adult heart is not detectably affected by partial Lem2 depletion, perhaps owing to a more established nuclear envelope and increased adaptation to mechanical stress. Taken together, these data provide insights into mechanisms underlying cardiomyopathy in patients with mutations in Lem2 and cardio-laminopathies in general.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 17 Apr 2023; epub ahead of print</small></div>

<div><h4>The circadian protein BMAL1 supports endothelial cell cycle during angiogenesis.</h4><i>Astone M, Oberkersch RE, Tosi G, Biscontin A, Santoro MM</i><br /><b>Aims</b><br />The circadian clock is an internal biological timer that coordinates physiology and gene expression with the 24-hour solar day. Circadian clock perturbations have been associated to vascular dysfunctions in mammals and a function of the circadian clock in angiogenesis has been suggested. However, the functional role of the circadian clock in endothelial cells (EC) and in the regulation of angiogenesis is widely unexplored.<br /><b>Methods and results</b><br />Here, we used both in vivo and in vitro approaches to demonstrate that EC possess an endogenous molecular clock and show robust circadian oscillations of core clock genes. By impairing the EC-specific function of the circadian clock transcriptional activator BMAL1 in vivo, we detect angiogenesis defects in mouse neonatal vascular tissues, as well as in adult tumor angiogenic settings. We then investigate the function of circadian clock machinery in cultured EC and show evidence that BMAL and CLOCK knock-down impair EC cell cycle progression. By using an RNA- and ChIP-seq genome-wide approaches we identified that BMAL1 binds the promoters of CCNA1 and CDK1 genes and controls their expression in EC.<br /><b>Conclusions</b><br />Our findings show that EC display a robust circadian clock and that BMAL1 regulates EC physiology in both developmental and pathological contexts. Genetic alteration of BMAL1 can affect angiogenesis in in vivo and in vitro settings.<br /><b>Translational perspective</b><br />These findings support the need to explore the manipulation of the circadian clock in vascular diseases. Further investigation of the behavior of BMAL1 and its target genes in the tumor endothelium can aim to discover novel therapeutic interventions to interfere with the endothelial circadian clock in the tumor context.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 13 Apr 2023; epub ahead of print</small></div>

<div><h4>The association between kidney function, cognitive function, and structural brain abnormalities in community-dwelling individuals aged 50 + is mediated by age and biomarkers of cardiovascular disease.</h4><i>Nowak N, De Looze C, O\'Halloran A, Kenny RA, Sexton DJ</i><br /><b>Aim</b><br />Cognitive impairment has been associated with kidney function and chronic kidney disease. Whether this association is due to accelerated cardiovascular disease or an independent specific kidney function effect related to toxins is unclear. We investigated the impact of an array of clinical factors, inflammatory biomarkers, and cardiovascular biomarkers on the association between kidney function, cognitive function, and structural brain abnormalities.<br /><b>Methods and results</b><br />We used data from the first and third waves of the TILDA Study, a population-representative prospective cohort of Irish adults aged 50 years and over, based on stratified random sampling (n = 3,774). The MRI sub-study included participants who consented to MRI brain imaging in addition to the health assessment. Multivariable linear and mixed-effect longitudinal regression models were fitted separately for each kidney marker/eGFR-equation after adjusting for baseline age and demographics, clinical vascular risk factors, and biomarkers. Unadjusted analyses showed an association between low eGFR, cognitive dysfunction, and cognitive decline (p < 0.001 for all kidney markers). Kidney function markers were also associated with white matter disease (OR = 3.32 (95% CI: 1.11, 9.98)), total grey matter volume (β=-0.17, 95% CI -0.27 to -0.07), and regional grey matter volumes within areas particularly susceptible to hypoxia (p < 0.001 for all). All the associations decreased after adjusting for age and were also diminished after adjusting for CVD biomarkers. Age and CVD-biomarker score were significant mediators of the adjusted associations between eGFR and cognitive status. These results remained consistent for cross-sectional and longitudinal outcomes and specific cognitive domains.<br /><b>Conclusions</b><br />Decreased kidney function was associated with cerebrovascular disease. The association appeared to be mediated predominantly by age and the combination of CVD markers (namely NT-proBNP and GDF15), supporting the idea that shared biological pathways underline both diseases. Further mechanistic studies of the specific molecular mechanisms that lead to both kidney and cognitive decline are warranted.<br /><b>Translational perspective</b><br />In our prospective cohort study, age, and concentrations of biomarkers previously related to CVD were found to be mediators of the association between mild to moderately decreased kidney function and neurocognitive abnormalities. In particular, a score composed of combined levels of GDF15 and NT-proBNP, was associated with cystatin C; and was mediating the adjusted association between kidney function and cognitive decline. Inflammation biomarker score showed borderline mediating effect. These results suggest potential causal vascular pathways that underline both development of early kidney function decline and cognitive decline. Additionally, CVD biomarkers could be evaluated in discriminating individuals at particular risk of dementia in the general population, who may benefit from intensive CVD risk factor modification.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 13 Apr 2023; epub ahead of print</small></div>

<div><h4>CHARGE syndrome-associated CHD7 acts at ISL1-regulated enhancers to modulate second heart field gene expression.</h4><i>Stathopoulou A, Wang P, Thellier C, Kelly RG, Zheng D, Scambler PJ</i><br /><b>Aims</b><br />Haploinsufficiency of the chromo-domain protein CHD7 underlies most cases of CHARGE syndrome, a multisystem birth defect including congenital heart malformation. Context specific roles for CHD7 in various stem, progenitor and differentiated cell lineages have been reported. Previously we showed severe defects when Chd7 is absent from cardiopharyngeal mesoderm (CPM). Here we investigate altered gene expression in the CPM and identify specific CHD7-bound target genes with known roles in the morphogenesis of affected structures.<br /><b>Methods and results</b><br />We generated conditional KO of Chd7 in CPM and analysed cardiac progenitor cells using transcriptomic and epigenomic analyses, in vivo expression analysis, and bioinformatic comparisons with existing datasets. We show CHD7 is required for correct expression of several genes established as major players in cardiac development, especially within the second heart field (SHF). We identified CHD7 binding sites in cardiac progenitor cells and found strong association with histone marks suggestive of dynamically regulated enhancers during the mesodermal to cardiac progenitor transition of mESC differentiation. Moreover, CHD7 shares a subset of its target sites with ISL1, a pioneer transcription factor in the cardiogenic gene regulatory network, including one enhancer modulating Fgf10 expression in SHF progenitor cells versus differentiating cardiomyocytes.<br /><b>Conclusion</b><br />We show that CHD7 interacts with ISL1, binds ISL1-regulated cardiac enhancers and modulates gene expression across the mesodermal heart fields during cardiac morphogenesis.<br /><b>Translational perspective</b><br />CHD7 is a chromatin remodeller haploinsufficient in CHARGE syndrome and implicated in autism spectrum disorder and various cancers. Heart defects in the syndrome are recapitulated by murine loss-of-function in two linages, neural crest and cardiopharyngeal mesoderm (CPM). CHD7 regulates vital cardiogenic genes via binding predominantly to enhancers distant from the target gene at sites often shared with the pioneer transcription factor ISL1. CHD7 bound enhancer elements show highly dynamic switching of histone modifications during the mesodermal to cardiac progenitor cell transition. Thus, manipulation of CHD7 activity may assist in directed differentiation of distinct cardiovascular progenitors for use in regenerative/repair therapeutics.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 13 Apr 2023; epub ahead of print</small></div>

<div><h4>Corticotropin-releasing hormone neurons in the central nucleus of amygdala are required for chronic stress-induced hypertension.</h4><i>Sheng ZF, Zhang H, Phaup JG, Zheng P, ... Pan HL, Li DP</i><br /><b>Aims</b><br />Chronic stress is a well-known risk factor for the development of hypertension. However, the underlying mechanisms remain unclear. Corticotropin-releasing hormone (CRH) neurons in the central nucleus of the amygdala (CeA) are involved in the autonomic responses to chronic stress. Here, we determined the role of CeA-CRH neurons in chronic stress-induced hypertension.<br /><b>Methods and results</b><br />Borderline hypertensive rats (BHRs) and Wistar-Kyoto (WKY) rats were subjected to chronic unpredictable stress (CUS). Firing activity and M-currents of CeA-CRH neurons were assessed, and a CRH-Cre-directed chemogenetic approach was used to suppress CeA-CRH neurons. CUS induced a sustained elevation of arterial blood pressure (ABP) and heart rate (HR) in BHRs, while in WKY rats, CUS-induced increases in ABP and HR quickly returned to baseline levels after CUS ended. CeA-CRH neurons displayed significantly higher firing activities in CUS-treated BHRs than unstressed BHRs. Selectively suppressing CeA-CRH neurons by chemogenetic approach attenuated CUS-induced hypertension and decreased elevated sympathetic outflow in CUS-treated BHRs. Also, CUS significantly decreased protein and mRNA levels of Kv7.2 and Kv7.3 channels in the CeA of BHRs. M-currents in CeA-CRH neurons were significantly decreased in CUS-treated BHRs compared with unstressed BHRs. Blocking Kv7 channel with its blocker XE-991 increased the excitability of CeA-CRH neurons in unstressed BHRs but not in CUS-treated BHRs. Microinjection of XE-991 into the CeA increased sympathetic outflow and ABP in unstressed BHRs but not in CUS-treated BHRs.<br /><b>Conclusions</b><br />CeA-CRH neurons are required for chronic stress-induced sustained hypertension. The hyperactivity of CeA-CRH neurons may be due to impaired Kv7 channel activity, which represents a new mechanism involved in chronic stress-induced hypertension.<br /><b>Translational perspective</b><br />We found that hyperactivity of CRH neurons in the CeA, likely due to diminished Kv7 channel activity, play a major role in the development of chronic stress-induced hypertension. Our study suggests that CRH neurons in the brain may be targeted for treating chronic stress-induced hypertension. Thus, increasing Kv7 channel activity or overexpressing Kv7 channels in the CeA may reduce stress-induced hypertension. Further studies are needed to delineate how chronic stress diminishes Kv7 channel activity in the brain.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 12 Apr 2023; epub ahead of print</small></div>

<div><h4>Microtubule plus-end tracking proteins: novel modulators of cardiac sodium channels and arrhythmogenesis.</h4><i>Marchal GA, Galjart N, Portero V, Remme CA</i><br /><AbstractText>The cardiac sodium channel NaV1.5 is an essential modulator of cardiac excitability, with decreased NaV1.5 levels at the plasma membrane and consequent reduction in sodium current (INa) leading to potentially lethal cardiac arrhythmias. NaV1.5 is distributed in a specific pattern at the plasma membrane of cardiomyocytes, with localization at the crests, grooves, and T-tubules of the lateral membrane, and particularly high levels at the intercalated disc region. NaV1.5 forms a large macromolecular complex with and is regulated by interacting proteins, some of which are specifically localised at either the lateral membrane or intercalated disc. One of the NaV1.5 trafficking routes is via microtubules (MTs), which are regulated by MT plus-end tracking proteins (+TIPs). In our search for mechanisms involved in targeted delivery of NaV1.5, we here provide an overview of previously demonstrated interactions between NaV1.5 interacting proteins and +TIPs, which potentially (in)directly impact on NaV1.5 trafficking. Strikingly, +TIPs interact extensively with several intercalated disc- and lateral membrane-specific NaV1.5 interacting proteins. Recent work indicates that this interplay of +TIPs and NaV1.5 interacting proteins mediates the targeted delivery of NaV1.5 at specific cardiomyocyte subcellular domains, while also being potentially relevant for the trafficking of other ion channels. These observations are especially relevant for diseases associated with loss of NaV1.5 specifically at the lateral membrane (such as Duchenne muscular dystrophy), or at the intercalated disc (for example, arrhythmogenic cardiomyopathy), and open up potential avenues for development of new anti-arrhythmic therapies.</AbstractText><br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 11 Apr 2023; epub ahead of print</small></div>

<div><h4>FOXK1 regulates Wnt signaling to promote cardiogenesis.</h4><i>Sierra-Pagan JE, Dsouza N, Das S, Larson TA, ... Gong W, Garry DJ</i><br /><b>Aims</b><br />Congenital heart disease (CHD) is the most common genetic birth defect, which has considerable morbidity and mortality. We focused on deciphering key regulators that govern cardiac progenitors and cardiogenesis. FOXK1 is a forkhead/winged helix transcription factor known to regulate cell cycle kinetics and is restricted to mesodermal progenitors, somites and the heart. In the present study, we define an essential role for FOXK1 during cardiovascular development.<br /><b>Methods & results</b><br />We used the mouse embryoid body system to differentiate control and Foxk1 KO ESCs into mesodermal, cardiac progenitor cells and mature cardiac cells. Using flow cytometry, immunohistochemistry, cardiac beating, transcriptional and ChIP qPCR assays, bulk RNAseq and ATACseq analyses, FOXK1 was observed to be an important regulator of cardiogenesis. Flow cytometry analyses revealed perturbed cardiogenesis in Foxk1 KO EBs. Bulk RNAseq analysis at two developmental stages showed a significant reduction of the cardiac molecular program in Foxk1 KO EBs compared to the control EBs. ATACseq analysis during EB differentiation demonstrated that the chromatin landscape nearby known important regulators of cardiogenesis was significantly relaxed in control EBs compared to Foxk1 KO EBs. Furthermore, we demonstrated that in the absence of FOXK1, cardiac differentiation was markedly impaired by assaying for cTnT expression and cardiac contractility. We demonstrate that FOXK1 is an important regulator of cardiogenesis by repressing the Wnt/β-catenin signaling pathway and thereby promoting differentiation.<br /><b>Conclusions</b><br />These results identify FOXK1 as an essential transcriptional and epigenetic regulator of cardiovascular development. Mechanistically, FOXK1 represses Wnt signaling to promote the development of cardiac progenitor cells.<br /><b>Translational perspective</b><br />Congenital heart disease is the most common birth defect. Deciphering the networks that govern cardiomyocyte specification, proliferation and differentiation will provide insights regarding therapeutic interventions for cardiovascular disease. The winged helix/forkhead family of transcription factors have been shown to have critical roles in epigenetics, organogenesis, cellular proliferation and differentiation. FOXK1 is an important transcription factor that regulates cardiovascular development through the Wnt signaling pathway. This FOXK1-Wnt pathway defines a network that may be therapeutically targeted to promote cardiogenesis.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 10 Apr 2023; epub ahead of print</small></div>

<div><h4>Inducible apelin receptor knockdown reduces differentiation efficiency and contractility of hESC-derived cardiomyocytes.</h4><i>Macrae RGC, Colzani MT, Williams TL, Bayraktar S, ... Sinha S, Davenport AP</i><br /><b>Aims</b><br />The apelin receptor, a G protein-coupled receptor, has emerged as a key regulator of cardiovascular development, physiology, and disease. However, there is a lack of suitable human in vitro models to investigate the apelinergic system in cardiovascular cell types. For the first time we have used human embryonic stem cell-derived cardiomyocytes (hESC-CMs) and a novel inducible knockdown system to examine the role of the apelin receptor in both cardiomyocyte development and to determine the consequences of loss of apelin receptor function as a model of disease.<br /><b>Methods and results</b><br />Expression of the apelin receptor and its ligands in hESCs and hESC-CMs was determined. hESCs carrying a tetracycline-inducible short hairpin RNA targeting the apelin receptor were generated using the sOPTiKD system. Phenotypic assays characterized the consequences of either apelin receptor knockdown before hESC-CM differentiation (early knockdown) or in 3D engineered heart tissues as a disease model (late knockdown). hESC-CMs expressed the apelin signalling system at a similar level to the adult heart. Early apelin receptor knockdown decreased cardiomyocyte differentiation efficiency and prolonged voltage sensing, associated with asynchronous contraction. Late apelin receptor knockdown had detrimental consequences on 3D engineered heart tissue contractile properties, decreasing contractility and increasing stiffness.<br /><b>Conclusions</b><br />We have successfully knocked down the apelin receptor, using an inducible system, to demonstrate a key role in hESC-CM differentiation. Knockdown in 3D engineered heart tissues recapitulated the phenotype of apelin receptor down-regulation in a failing heart, providing a potential platform for modelling heart failure and testing novel therapeutic strategies.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 31 Mar 2023; 119:587-598</small></div>

<div><h4>No sex-related differences in infarct size, no-reflow, and protection by ischaemic pre-conditioning in Göttingen minipigs.</h4><i>Kleinbongard P, Lieder H, Skyschally A, Heusch G</i><br /><b>Aims</b><br />Female sex has been proposed to be cardioprotective per se. Studies with myocardial ischaemia/reperfusion and infarct size as endpoint have demonstrated cardioprotection in female, castrated male, and male pigs. These studies are difficult to compare, given the different pig strains, models, durations of ischaemia, and methods of infarct size quantification. The few studies using both female and male pigs reported no differences in infarct size and cardioprotection. We, therefore, prospectively compared infarct size in Göttingen minipigs undergoing ischaemia/reperfusion (I/R) without and with ischaemic pre-conditioning (IPC) between female, castrated male, and male pigs.<br /><b>Methods and results</b><br />In a prospective, randomized approach, 28 Göttingen open-chest, anaesthetized minipigs underwent 60 min ischaemia by distal left anterior descending artery (LAD) occlusion and 180 min reperfusion without and with IPC by three cycles of 5 min LAD occlusion/10 min reperfusion. Infarct size with I/R was not different between female, castrated male, and male pigs (45 ± 8 vs. 45 ± 13 vs. 41 ± 9% area at risk), as was the reduction in infarct size with IPC (25 ± 11 vs. 30 ± 8 vs. 19 ± 10% area at risk). In addition, the area of no-reflow was not different between female, castrated male, and male pigs with I/R (57 ± 13 vs. 35 ± 7 vs. 47 ± 26% infarct size) or IPC (4 ± 10 vs.12 ± 20 vs. 0 ± 0% infarct size). Phosphorylation of signal transducer and activator of transcription 3 was increased at 10 min reperfusion by IPC but not by I/R to the same extent in female, castrated male, and male pigs (198 ± 30 vs. 230 ± 165 vs. 179 ± 107% of baseline).<br /><b>Conclusion</b><br />Our data do not support the notion of sex- or castration-related differences in infarct size, coronary microvascular injury, and cardioprotection by IPC.<br /><b>Translational perspective</b><br />The translation of successful preclinical studies on cardioprotection to the benefit of patients with reperfused myocardial infarction has been difficult. The difficulties have been attributed to confounders such as co-morbidities and co-medications which patients typically have but animals don´t, but also to age and sex. Notably, female sex has been considered as protective per se. We have now, using our established and clinically relevant pig model of reperfused acute myocardial infarction and ischaemic preconditioning as the most robust cardioprotective intervention looked for sex-related differences of infarct size, no-reflow and cardioprotection by ischaemic preconditioning in a prospectively powered approach but found none such difference.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 31 Mar 2023; 119:561-570</small></div>

<div><h4>An antiplatelet response gene expression signature is associated with bleeding.</h4><i>Friede KA, Myers RA, Gales J, Zhbannikov I, ... Ginsburg GS, Voora D</i><br /><b>Aims</b><br />Gene expression biosignatures may hold promise to individualize antiplatelet therapy in conjunction with current guidelines and risk scores. The Aspirin Response Signature (ARS) score is comprised of a weighted sum of correlated, pro-thrombotic gene transcripts measured in whole blood. In prior work where volunteers were exposed to aspirin 325 mg daily, higher ARS score was associated with lower platelet function; separately, in a clinical cohort of patients, higher ARS scores were associated with increased risk of adverse cardiovascular events. To better understand this apparent paradox, we measured ARS gene expression and score in volunteers to determine aspirin dose-response and ticagrelor relationships with ARS score and separately in patients to assess whether ARS is associated with incident bleeding.<br /><b>Methods and results</b><br />Blood samples were collected from volunteers (N = 188) who were exposed to 4 weeks of daily aspirin 81 mg, daily aspirin 325 mg, and/or twice-daily ticagrelor 90 mg. ARS scores were calculated from whole blood RNA qPCR, and platelet function and protein expression were assessed in platelet-rich plasma. In mixed linear regression models, aspirin 81 mg exposure was not associated with changes in ARS gene expression or score. Aspirin 325 mg exposure resulted in a 6.0% increase in ARS gene expression (P = 7.5 × 10-9 vs. baseline, P = 2.1 × 10-4 vs. aspirin 81 mg) and an increase in expression of platelet proteins corresponding to ARS genes. Ticagrelor exposure resulted in a 30.7% increase in ARS gene expression (P < 1 × 10-10 vs. baseline and each aspirin dose) and ARS score (P = 7.0 × 10-7 vs. baseline, P = 3.6 × 10-6 and 5.59 × 10-4 vs. aspirin 81 and 325 mg, respectively). Increases in ARS gene expression or score were associated with the magnitude of platelet inhibition across agents. To assess the association between ARS scores and incident bleeding, ARS scores were calculated in patients undergoing cardiac catheterization (N = 1421), of whom 25.4% experienced bleeding events over a median 6.2 years of follow-up. In a Cox model adjusting for demographics and baseline antithrombotic medication use, patients with ARS scores above the median had a higher risk of incident bleeding [hazard ratio 1.26 (95% CI 1.01-1.56), P = 0.038].<br /><b>Conclusions</b><br />The ARS is an Antiplatelet Response Signature that increases in response to greater platelet inhibition due to antiplatelet therapy and may represent a homeostatic mechanism to prevent bleeding. ARS scores could inform future strategies to prevent bleeding while maintaining antiplatelet therapy\'s benefit of ischaemic cardiovascular event protection.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 31 Mar 2023; 119:551-560</small></div>

<div><h4>Frequent premature atrial contractions as a signalling marker of atrial cardiomyopathy, incident atrial fibrillation, and stroke.</h4><i>Farinha JM, Gupta D, Lip GYH</i><br /><AbstractText>Premature atrial contractions are a common cardiac phenomenon. Although previously considered a benign electrocardiographic finding, they have now been associated with a higher risk of incident atrial fibrillation (AF) and other adverse outcomes such as stroke and all-cause mortality. Since premature atrial contractions can be associated with these adverse clinical outcomes independently of AF occurrence, different explanations have being proposed. The concept of atrial cardiomyopathy, where AF would be an epiphenomenon outside the causal pathway between premature atrial contractions and stroke has received traction recently. This concept suggests that structural, functional, and biochemical changes in the atria lead to arrhythmia occurrence and thromboembolic events. Some consensus about diagnosis and treatment of this condition have been published, but this is based on scarce evidence, highlighting the need for a clear definition of excessive premature atrial contractions and for prospective studies regarding antiarrhythmic therapies, anticoagulation or molecular targets in this group of patients.</AbstractText><br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 31 Mar 2023; 119:429-439</small></div>

<div><h4>Four decades of experience of prosthetic valve endocarditis reflect a high variety of diverse pathogens.</h4><i>Oberbach A, Schlichting N, Hagl C, Lehmann S, ... Saha S, Bagaev E</i><br /><AbstractText>Prosthetic valve endocarditis (PVE) remains a serious condition with a high mortality rate. Precise identification of the PVE-associated pathogen/s and their virulence is essential for successful therapy and patient survival. The commonly described PVE-associated pathogens are staphylococci, streptococci, and enterococci, with Staphylococcus aureus being the most frequently diagnosed species. Furthermore, multi-drug resistance pathogens are increasing in prevalence and continue to pose new challenges mandating a personalized approach. Blood cultures in combination with echocardiography are the most common methods to diagnose PVE, often being the only indication, it exists. In many cases, the diagnostic strategy recommended in the clinical guidelines does not identify the precise microbial agent, and frequently, false-negative blood cultures are reported. Despite the fact that blood culture findings are not always a good indicator of the actual PVE agent in the valve tissue, only a minority of re-operated prostheses are subjected to microbiological diagnostic evaluation. In this review, we focus on the diversity and the complete spectrum of PVE-associated bacterial, fungal, and viral pathogens in blood and prosthetic heart valve, their possible virulence potential, and their challenges in making a microbial diagnosis. We are curious to understand if the unacceptable high mortality of PVE is associated with the high number of negative microbial findings in connection with a possible PVE. Herein, we discuss the possibilities and limits of the diagnostic methods conventionally used and make recommendations for enhanced pathogen identification. We also show possible virulence factors of the most common PVE-associated pathogens and their clinical effects. Based on blood culture, molecular biological diagnostics, and specific valve examination, better derivations for the antibiotic therapy as well as possible preventive intervention can be established in the future.</AbstractText><br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 31 Mar 2023; 119:410-428</small></div>

<div><h4>Metaboloepigenetics in cancer, immunity, and cardiovascular disease.</h4><i>Keating ST, El-Osta A</i><br /><AbstractText>The influence of cellular metabolism on epigenetic pathways is well documented but misunderstood. Scientists have long known of the metabolic impact on epigenetic determinants. More often than not, that title role for DNA methylation was portrayed by the metabolite S-adenosylmethionine. Technically speaking, there are many other metabolites that drive epigenetic processes that instruct seemingly distant-yet highly connect pathways-and none more so than our understanding of the cancer epigenome. Recent studies have shown that available energy links the extracellular environment to influence cellular responses. This focused review examines the recent interest in epigenomics and casts cancer, metabolism, and immunity in unfamiliar roles-cooperating. There are not only language lessons from cancer research, we have come round to appreciate that reaching into areas previously thought of as too distinct are also object lessons in understanding health and disease. The Warburg effect is one such signature of how glycolysis influences metabolic shift during oncogenesis. That shift in metabolism-now recognized as central to proliferation in cancer biology-influences core enzymes that not only control gene expression but are also central to replication, condensation, and the repair of nucleic acid. These nuclear processes rely on metabolism, and with glucose at centre stage, the role of respiration and oxidative metabolism is now synonymous with the mitochondria as the powerhouses of metaboloepigenetics. The emerging evidence for metaboloepigenetics in trained innate immunity has revealed recognizable signalling pathways with antecedent extracellular stimulation. With due consideration to immunometabolism, we discuss the striking signalling similarities influencing these core pathways. The immunometabolic-epigenetic axis in cardiovascular disease has deeply etched connections with inflammation, and we examine the chromatin template as a carrier of epigenetic indices that determine the expression of genes influencing atherosclerosis and vascular complications of diabetes.</AbstractText><br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 31 Mar 2023; 119:357-370</small></div>

<div><h4>VEGF-B hypertrophy predisposes to transition from diastolic to systolic heart failure in hypertensive rats.</h4><i>Samuelsson AM, Bartolomaeus TUP, Anandakumar H, Thowsen I, ... Muller DN, Wiig H</i><br /><b>Aims</b><br />Cardiac energy metabolism is centrally involved in heart failure (HF), although the direction of the metabolic alterations is complex and likely dependent on the particular stage of HF progression. Vascular endothelial growth factor B (VEGF-B) has been shown to modulate metabolic processes and to induce physiological cardiac hypertrophy; thus, it could be cardioprotective in the failing myocardium. This study investigates the role of VEGF-B in cardiac proteomic and metabolic adaptation in HF during aldosterone and high-salt hypertensive challenges.<br /><b>Methods and results</b><br />Male rats overexpressing the cardiac-specific VEGF-B transgene (VEGF-B TG) were treated for 3 or 6 weeks with deoxycorticosterone-acetate combined with a high-salt (HS) diet (DOCA + HS) to induce hypertension and cardiac damage. Extensive longitudinal echocardiographic studies of HF progression were conducted, starting at baseline. Sham-treated rats served as controls. To evaluate the metabolic alterations associated with HF, cardiac proteomics by mass spectrometry was performed. Hypertrophic non-treated VEGF-B TG hearts demonstrated high oxygen and adenosine triphosphate (ATP) demand with early onset of diastolic dysfunction. Administration of DOCA + HS to VEGF-B TG rats for 6 weeks amplified the progression from cardiac hypertrophy to HF, with a drastic drop in heart ATP concentration. Dobutamine stress echocardiographic analyses uncovered a significantly impaired systolic reserve. Mechanistically, the hallmark of the failing TG heart was an abnormal energy metabolism with decreased mitochondrial ATP, preceding the attenuated cardiac performance and leading to systolic HF.<br /><b>Conclusions</b><br />This study shows that the VEGF-B TG accelerates metabolic maladaptation which precedes structural cardiomyopathy in experimental hypertension and ultimately leads to systolic HF.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 23 Mar 2023; epub ahead of print</small></div>

<div><h4>The translational landscape of human vascular smooth muscle cells identifies novel short ORF-encoded peptide regulators for phenotype alteration.</h4><i>Li K, Li B, Zhang D, Du T, ... Yang JH, Huang ZP</i><br /><b>Aims</b><br />The plasticity of vascular smooth muscle cells (VSMCs) enables them to alter phenotypes under various physiological and pathological stimuli. The alteration of VSMC phenotype is a key step in vascular diseases, including atherosclerosis. Although the transcriptome shift during VSMC phenotype alteration has been intensively investigated, uncovering multiple key regulatory signaling pathways, the translatome dynamics in this cellular process remain largely unknown. Here, we explored the genome-wide regulation at the translational level of human VSMCs during phenotype alteration.<br /><b>Methods and results</b><br />We generated nucleotide-resolution translatome and transcriptome data from human VSMCs undergoing phenotype alteration. Deep sequencing of ribosome-protected fragments (Ribo-seq) revealed alterations in protein synthesis independent of changes in mRNA levels. Increased translational efficiency of many translational machinery components, including ribosomal proteins, eukaryotic translation elongation factors and initiation factors, were observed during the phenotype alteration of VSMCs. In addition, hundreds of candidates for short ORF-encoded polypeptides (SEPs), a class of peptides containing 200 aa or less, were identified in a combined analysis of translatome and transcriptome data with a high positive rate in validating their coding capability. Three evolutionarily conserved SEPs were further detected endogenously by customized antibodies and suggested to participate in the pathogenesis of atherosclerosis by analyzing the transcriptome and single cell RNA-seq data from patient atherosclerotic artery samples. Gain- and loss-of-function studies in human VSMCs and genetically engineered mice showed that these SEPs modulate the alteration of VSMC phenotype through different signaling pathways, including the mitogen-activated protein kinase (MAPK) pathway and p53 pathway.<br /><b>Conclusion</b><br />Our study indicates that an increase in the capacity of translation, which is attributable to an increased quantity of translational machinery components, mainly controls alterations of VSMC phenotype at the level of translational regulation. In addition, SEPs could function as important regulators in the phenotype alteration of human VSMCs.<br /><b>Translational perspective</b><br />Alterations of VSMC status are tightly associated with vascular diseases, including atherosclerosis. By incorporating Ribo-seq and RNA-seq of human VSMCs, we revealed that the increased translational capacity dominated the translational regulation of gene expression during alterations of VSMC phenotype. This finding of regulation in the translational level could provide new strategies of treating vascular diseases in the future. In addition, novel short ORF-encoded polypeptide (SEP) regulators for phenotype alteration were identified in the incorporated analyses. With the uncovered mechanisms, these SEPs may represent a new type of potential therapeutic target for clinical intervention of vascular diseases.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 21 Mar 2023; epub ahead of print</small></div>

<div><h4>Post-transcriptional control of hemostatic genes: mechanisms and emerging therapeutic concepts in thrombo-inflammatory disorders.</h4><i>Danckwardt S, Trégouët DA, Castoldi E</i><br /><AbstractText>The hemostatic system is pivotal to maintaining vascular integrity. Multiple components involved in blood coagulation have central functions in inflammation and immunity. A derailed hemostasis is common in prevalent pathologies such as sepsis, cardiovascular disorders and, lately, COVID-19. Physiological mechanisms limit the deleterious consequences of a hyperactivated hemostatic system through adaptive changes in gene expression. While this is mainly regulated at the level of transcription, co- and posttranscriptional mechanisms are increasingly perceived as central hubs governing multiple facets of the hemostatic system. This layer of regulation modulates the biogenesis of hemostatic components, for example in situations of increased turnover and demand. However, they can also be \'hijacked\' in disease processes, thereby perpetuating and even causally entertaining associated pathologies. This review summarizes examples and emerging concepts that illustrate the importance of posttranscriptional mechanisms in hemostatic control and crosstalk with the immune system. It also discusses how such regulatory principles can be used to usher in new therapeutic concepts to combat global medical threats such as sepsis or cardiovascular disorders.</AbstractText><br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 21 Mar 2023; epub ahead of print</small></div>

<div><h4>Periodontitis exacerbates atherosclerosis through Fusobacterium nucleatum-promoted hepatic glycolysis and lipogenesis.</h4><i>Zhou LJ, Lin WZ, Meng XQ, Zhu H, ... Zhu YQ, Duan SZ</i><br /><b>Aims</b><br />Positive associations between periodontitis (PD) and atherosclerosis have been established, but the causality and mechanisms are not clear. We aimed to explore the causal roles of PD in atherosclerosis and dissect the underlying mechanisms.<br /><b>Methods and results</b><br />A mouse model of PD was established by ligation of molars in combination with application of subgingival plaques collected from PD patients, and then combined with atherosclerosis model induced by treating atheroprone mice with a high-cholesterol diet (HCD). PD significantly aggravated atherosclerosis in HCD-fed atheroprone mice, including increased en face plaque areas in whole aortas and lesion size at aortic roots. PD also increased circulating levels of triglycerides and cholesterol, hepatic levels of cholesterol, and hepatic expression of rate-limiting enzymes for lipogenesis. Using 16S rRNA gene sequencing, F. nucleatum was identified as the most enriched PD-associated pathobiont that present in both oral cavity and livers. Co-culture experiments demonstrated that F. nucleatum directly stimulated lipid biosynthesis in primary mouse hepatocytes. Moreover, oral inoculation of F. nucleatum markedly elevated plasma levels of triglycerides and cholesterol and promoted atherogenesis in HCD-fed ApoE-/- mice. Results of RNA-seq and Seahorse assay indicated that F. nucleatum activated glycolysis, inhibition of which by 2-deoxyglucose in turn suppressed F. nucleatum-induced lipogenesis in hepatocytes. Finally, interrogation of the molecular mechanisms revealed that F. nucleatum induced glycolysis and lipogenesis by activating PI3K/Akt/mTOR signaling pathway in hepatocytes.<br /><b>Conclusions</b><br />PD exacerbates atherosclerosis and impairs lipid metabolism in mice, which may be mediated by F. nucleatum-promoted glycolysis and lipogenesis through PI3K/Akt/mTOR signaling in hepatocytes. Treatment of PD and specific targeting of F. nucleatum are promising strategies to improve therapeutic effectiveness of hyperlipidemia and atherosclerosis.<br /><b>Translational perspective</b><br />This study has reinforced the causal relationship between PD and atherosclerosis, and identified F. nucleatum-mediated hepatic glycolysis and lipogenesis as a new mechanism underlying the causal relationship. These findings support that intervention of PD or F. nucleatum may improve lipid homeostasis and contribute to alleviation of atherosclerosis and improvement of cardiovascular health.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 21 Mar 2023; epub ahead of print</small></div>

<div><h4>Aldosterone and cardiovascular diseases.</h4><i>Parksook WW, Williams GH</i><br /><AbstractText>Aldosterone\'s role in the kidney and its pathophysiologic actions in hypertension are well known. However, its role or that of its receptor [minieralocorticoid receptor (MR)] in other cardiovascular (CV) disease are less well described. To identify their potential roles in six CV conditions (heart failure, myocardial infarction, atrial fibrillation, stroke, atherosclerosis, and thrombosis), we assessed these associations in the following four areas: (i) mechanistic studies in rodents and humans; (ii) pre-clinical studies of MR antagonists; (iii) clinical trials of MR antagonists; and (iv) genetics. The data were acquired from an online search of the National Library of Medicine using the PubMed search engine from January 2011 through June 2021. There were 3702 publications identified with 200 publications meeting our inclusion and exclusion criteria. Data strongly supported an association between heart failure and dysregulated aldosterone/MR. This association is not surprising given aldosterone/MR\'s prominent role in regulating sodium/volume homeostasis. Atrial fibrillation and myocardial infarction are also associated with dysregulated aldosterone/MR, but less strongly. For the most part, the data were insufficient to determine whether there was a relationship between atherosclerosis, stroke, or thrombosis and aldosterone/MR dysregulation. This review clearly documented an expanding role for aldosterone/MR\'s dysregulation in CV diseases beyond hypertension. How expansive it might be is limited by the currently available data. It is anticipated that with an increased focus on aldosterone/MR\'s potential roles in these diseases, additional clinical and pre-clinical data will clarify these relationships, thereby, opening approaches to use modulators of aldosterone/MR\'s action to more precisely treat these CV conditions.</AbstractText><br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:28-44</small></div>

<div><h4>Age-related enhanced degeneration of bioprosthetic valves due to leaflet calcification, tissue crosslinking, and structural changes.</h4><i>Xue Y, Kossar AP, Abramov A, Frasca A, ... Levy RJ, Ferrari G</i><br /><b>Aims</b><br />Bioprosthetic heart valves (BHVs), made from glutaraldehyde-fixed heterograft materials, are subject to more rapid structural valve degeneration (SVD) in paediatric and young adult patients. Differences in blood biochemistries and propensity for disease accelerate SVD in these patients, which results in multiple re-operations with compounding risks. The goal of this study is to investigate the mechanisms of BHV biomaterial degeneration and present models for studying SVD in young patients and juvenile animal models.<br /><b>Methods and results</b><br />We studied SVD in clinical BHV explants from paediatric and young adult patients, juvenile sheep implantation model, rat subcutaneous implants, and an ex vivo serum incubation model. BHV biomaterials were analysed for calcification, collagen microstructure (alignment and crimp), and crosslinking density. Serum markers of calcification and tissue crosslinking were compared between young and adult subjects. We demonstrated that immature subjects were more susceptible to calcification, microstructural changes, and advanced glycation end products formation. In vivo and ex vivo studies comparing immature and mature subjects mirrored SVD in clinical observations. The interaction between host serum and BHV biomaterials leads to significant structural and biochemical changes which impact their functions.<br /><b>Conclusions</b><br />There is an increased risk for accelerated SVD in younger subjects, both experimental animals and patients. Increased calcification, altered collagen microstructure with loss of alignment and increased crimp periods, and increased crosslinking are three main characteristics in BHV explants from young subjects leading to SVD. Together, our studies establish a basis for assessing the increased susceptibility of BHV biomaterials to accelerated SVD in young patients.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:302-315</small></div>

<div><h4>Deciphering endothelial heterogeneity in health and disease at single-cell resolution: progress and perspectives.</h4><i>Becker LM, Chen SH, Rodor J, de Rooij LPMH, Baker AH, Carmeliet P</i><br /><AbstractText>Endothelial cells (ECs) constitute the inner lining of vascular beds in mammals and are crucial for homeostatic regulation of blood vessel physiology, but also play a key role in pathogenesis of many diseases, thereby representing realistic therapeutic targets. However, it has become evident that ECs are heterogeneous, encompassing several subtypes with distinct functions, which makes EC targeting and modulation in diseases challenging. The rise of the new single-cell era has led to an emergence of studies aimed at interrogating transcriptome diversity along the vascular tree, and has revolutionized our understanding of EC heterogeneity from both a physiological and pathophysiological context. Here, we discuss recent landmark studies aimed at teasing apart the heterogeneous nature of ECs. We cover driving (epi)genetic, transcriptomic, and metabolic forces underlying EC heterogeneity in health and disease, as well as current strategies used to combat disease-enriched EC phenotypes, and propose strategies to transcend largely descriptive heterogeneity towards prioritization and functional validation of therapeutically targetable drivers of EC diversity. Lastly, we provide an overview of the most recent advances and hurdles in single EC OMICs.</AbstractText><br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:6-27</small></div>

<div><h4>Histologic and proteomic remodeling of the pulmonary veins and arteries in a porcine model of chronic pulmonary venous hypertension.</h4><i>Fayyaz AU, Sabbah MS, Dasari S, Griffiths LG, ... Edwards WD, Redfield MM</i><br /><b>Aims</b><br />In heart failure (HF), pulmonary venous hypertension (PVH) produces pulmonary hypertension (PH) with remodeling of pulmonary veins (PV) and arteries (PA). In a porcine PVH model, we performed proteomic-based bioinformatics to investigate unique pathophysiologic mechanisms mediating PA and PV remodeling.<br /><b>Methods and results</b><br />Large PV were banded (PVH, n = 10) or not (Sham, n = 9) in piglets. At sacrifice, PV and PA were perfusion labelled for vessel-specific histology and proteomics. The PA and PV were separately sampled with laser-capture micro-dissection for mass spectrometry. Pulmonary vascular resistance [Wood Units; 8.6 (95% confidence interval: 6.3, 12.3) vs. 2.0 (1.7, 2.3)] and PA [19.9 (standard error of mean, 1.1) vs. 10.3 (1.1)] and PV [14.2 (1.2) vs. 7.6 (1.1)] wall thickness/external diameter (%) were increased in PVH (P < 0.05 for all). Similar numbers of proteins were identified in PA (2093) and PV (2085) with 94% overlap, but biological processes differed. There were more differentially expressed proteins (287 vs. 161), altered canonical pathways (17 vs. 3), and predicted upstream regulators (PUSR; 22 vs. 6) in PV than PA. In PA and PV, bioinformatics indicated activation of the integrated stress response and mammalian target of rapamycin signalling with dysregulated growth. In PV, there was also activation of Rho/Rho-kinase signalling with decreased actin cytoskeletal signalling and altered tight and adherens junctions, ephrin B, and caveolae-mediated endocytosis signalling; all indicating disrupted endothelial barrier function. Indeed, protein biomarkers and the top PUSR in PV (transforming growth factor-beta) suggested endothelial to mesenchymal transition in PV. Findings were similar in human autopsy specimens.<br /><b>Conclusion</b><br />These findings provide new therapeutic targets to oppose pulmonary vascular remodeling in HF-related PH.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:268-282</small></div>

<div><h4>Arterial myeloperoxidase in the detection and treatment of vulnerable atherosclerotic plaque: a new dawn for an old light.</h4><i>Nadel J, Jabbour A, Stocker R</i><br /><AbstractText>Intracellular myeloperoxidase (MPO) plays a specific role in the innate immune response; however, upon release into the extracellular space in the setting of inflammation, drives oxidative tissue injury. Extracellular MPO has recently been shown to be abundant in unstable atheroma and causally linked to plaque destabilization, erosion, and rupture, identifying it as a potential target for the surveillance and treatment of vulnerable atherosclerosis. Through the compartmentalization of MPO\'s protective and deleterious effects, extracellular MPO can be selectively detected using non-invasive molecular imaging and targeted by burgeoning pharmacotherapies. Given its causal relationship to plaque destabilization coupled with an ability to preserve its beneficial properties, MPO is potentially a superior translational inflammatory target compared with other immunomodulatory therapies and imaging biomarkers utilized to date. This review explores the role of MPO in plaque destabilization and provides insights into how it can be harnessed in the management of patients with vulnerable atherosclerotic plaque.</AbstractText><br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:112-120</small></div>

<div><h4>Single-cell transcriptomes in the heart: when every epigenome counts.</h4><i>Gromova T, Gehred ND, Vondriska TM</i><br /><AbstractText>The response of an organ to stimuli emerges from the actions of individual cells. Recent cardiac single-cell RNA-sequencing studies of development, injury, and reprogramming have uncovered heterogeneous populations even among previously well-defined cell types, raising questions about what level of experimental resolution corresponds to disease-relevant, tissue-level phenotypes. In this review, we explore the biological meaning behind this cellular heterogeneity by undertaking an exhaustive analysis of single-cell transcriptomics in the heart (including a comprehensive, annotated compendium of studies published to date) and evaluating new models for the cardiac function that have emerged from these studies (including discussion and schematics that depict new hypotheses in the field). We evaluate the evidence to support the biological actions of newly identified cell populations and debate questions related to the role of cell-to-cell variability in development and disease. Finally, we present emerging epigenomic approaches that, when combined with single-cell RNA-sequencing, can resolve basic mechanisms of gene regulation and variability in cell phenotype.</AbstractText><br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:64-78</small></div>

<div><h4>Methods for the identification and characterization of extracellular vesicles in cardiovascular studies: from exosomes to microvesicles.</h4><i>Davidson SM, Boulanger CM, Aikawa E, Badimon L, ... Witwer K, Sluijter JPG</i><br /><AbstractText>Extracellular vesicles (EVs) are nanosized vesicles with a lipid bilayer that are released from cells of the cardiovascular system, and are considered important mediators of intercellular and extracellular communications. Two types of EVs of particular interest are exosomes and microvesicles, which have been identified in all tissue and body fluids and carry a variety of molecules including RNAs, proteins, and lipids. EVs have potential for use in the diagnosis and prognosis of cardiovascular diseases and as new therapeutic agents, particularly in the setting of myocardial infarction and heart failure. Despite their promise, technical challenges related to their small size make it challenging to accurately identify and characterize them, and to study EV-mediated processes. Here, we aim to provide the reader with an overview of the techniques and technologies available for the separation and characterization of EVs from different sources. Methods for determining the protein, RNA, and lipid content of EVs are discussed. The aim of this document is to provide guidance on critical methodological issues and highlight key points for consideration for the investigation of EVs in cardiovascular studies.</AbstractText><br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:45-63</small></div>

<div><h4>Rapid neutrophil mobilization by VCAM-1+ endothelial cell-derived extracellular vesicles.</h4><i>Akbar N, Braithwaite AT, Corr EM, Koelwyn GJ, ... Anthony DC, Choudhury RP</i><br /><b>Aims</b><br />Acute myocardial infarction rapidly increases blood neutrophils (<2 h). Release from bone marrow, in response to chemokine elevation, has been considered their source, but chemokine levels peak up to 24 h after injury, and after neutrophil elevation. This suggests that additional non-chemokine-dependent processes may be involved. Endothelial cell (EC) activation promotes the rapid (<30 min) release of extracellular vesicles (EVs), which have emerged as an important means of cell-cell signalling and are thus a potential mechanism for communicating with remote tissues.<br /><b>Methods and results</b><br />Here, we show that injury to the myocardium rapidly mobilizes neutrophils from the spleen to peripheral blood and induces their transcriptional activation prior to arrival at the injured tissue. Time course analysis of plasma-EV composition revealed a rapid and selective increase in EVs bearing VCAM-1. These EVs, which were also enriched for miRNA-126, accumulated preferentially in the spleen where they induced local inflammatory gene and chemokine protein expression, and mobilized splenic-neutrophils to peripheral blood. Using CRISPR/Cas9 genome editing, we generated VCAM-1-deficient EC-EVs and showed that its deletion removed the ability of EC-EVs to provoke the mobilization of neutrophils. Furthermore, inhibition of miRNA-126 in vivo reduced myocardial infarction size in a mouse model.<br /><b>Conclusions</b><br />Our findings show a novel EV-dependent mechanism for the rapid mobilization of neutrophils to peripheral blood from a splenic reserve and establish a proof of concept for functional manipulation of EV-communications through genetic alteration of parent cells.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:236-251</small></div>

<div><h4>Ryanodine receptor 2 (RYR2) dysfunction activates the unfolded protein response and perturbs cardiomyocyte maturation.</h4><i>Guo Y, Cao Y, Jardin BD, Zhang X, ... Dong E, Pu WT</i><br /><b>Aims</b><br />Calcium-handling capacity is a major gauge of cardiomyocyte maturity. Ryanodine receptor 2 (RYR2) is the pre-dominant calcium channel that releases calcium from the sarcoplasmic reticulum/endoplasmic reticulum (SR/ER) to activate cardiomyocyte contraction. Although RYR2 was previously implied as a key regulator of cardiomyocyte maturation, the mechanisms remain unclear. The aim of this study is to solve this problem.<br /><b>Methods and results</b><br />We performed Cas9/AAV9-mediated somatic mutagenesis to knockout RYR2 specifically in cardiomyocytes in mice. We conducted a genetic mosaic analysis to dissect the cell-autonomous function of RYR2 during cardiomyocyte maturation. We found that RYR2 depletion triggered ultrastructural and transcriptomic defects relevant to cardiomyocyte maturation. These phenotypes were associated with the drastic activation of ER stress pathways. The ER stress alleviator tauroursodeoxycholic acid partially rescued the defects in RYR2-depleted cardiomyocytes. Overexpression of ATF4, a key ER stress transcription factor, recapitulated defects in RYR2-depleted cells. Integrative analysis of RNA-Seq and bioChIP-Seq data revealed that protein biosynthesis-related genes are the major direct downstream targets of ATF4.<br /><b>Conclusion</b><br />RYR2-regulated ER homeostasis is essential for cardiomyocyte maturation. Severe ER stress perturbs cardiomyocyte maturation primarily through ATF4 activation. The major downstream effector genes of ATF4 are related to protein biosynthesis.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:221-235</small></div>

<div><h4>Renin, aldosterone, the aldosterone-to-renin ratio, and incident hypertension among normotensive subjects from the general population.</h4><i>Arnold N, Hermanns IM, Schulz A, Hahad O, ... Münzel T, Wild PS</i><br /><b>Aims</b><br />To investigate the predictive ability of direct plasma renin and aldosterone concentrations as well as their ratio [aldosterone-to-renin (ARR)] for incident hypertension in the general population.<br /><b>Methods and results</b><br />Concentration of renin and aldosterone were measured by a chemiluminescence immunoassay using the fully automated LIAISON® platform (DiaSorin) among 5362 participants of the population-based Gutenberg Health Study, who were normotensive and had no clinically overt cardiovascular disease at baseline. During a follow-up period of 5 years, 18.6% (n = 996) developed a new-onset hypertension. Comparing extreme quartiles of biomarker distribution, the relative risk (RR) for incident arterial hypertension was found to be 1.58 [95% confidence interval (CI) 1.25-2.00; P = 0.00015; Q1 vs. Q4ref] for renin; 1.29 (95% CI 1.05-1.59, P = 0.018; Q4 vs. Q1ref) for aldosterone and 1.70 (95% CI 1.33-2.12; P < 0.0001; Q4 vs. Q1ref) for ARR after multivariable adjustment in men. In females, only high ARR was independently predictive for incident hypertension over 5 years [RR 1.29 (95% CI 1.04-1.62); P = 0.024]. Even in the subgroup of individuals having biomarker concentrations within the reference range, high ARR was predictive for new-onset hypertension in men [RR 1.44 (95% CI 1.13-1.83); P = 0.003]. Finally, synergistic effects of co-prevalent obesity and ARR on incident hypertension were also demonstrated, resulting in markedly higher risk estimates as seen for biomarker alone [RR of 2.70 (95% CI 2.05-3.6) for Q4 of ARR and having body mass index ≥ 30 kg/m2 vs. low ARR (Q1ref) and normal weight; P < 0.0001].<br /><b>Conclusion</b><br />Among normotensives from the general population ARR possesses a stronger predictive value for incident hypertension than renin or aldosterone alone. The prediction of arterial hypertension by ARR was even stronger in obese subjects.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:294-301</small></div>

<div><h4>Properties and fate of human mesenchymal stem cells upon miRNA let-7f-promoted recruitment to atherosclerotic plaques.</h4><i>Egea V, Megens RTA, Santovito D, Wantha S, ... Weber C, Ries C</i><br /><b>Aims</b><br />Atherosclerosis is a chronic inflammatory disease of the arteries leading to the formation of atheromatous plaques. Human mesenchymal stem cells (hMSCs) are recruited from the circulation into plaques where in response to their environment they adopt a phenotype with immunomodulatory properties. However, the mechanisms underlying hMSC function in these processes are unclear. Recently, we described that miRNA let-7f controls hMSC invasion guided by inflammatory cytokines and chemokines. Here, we investigated the role of let-7f in hMSC tropism to human atheromas and the effects of the plaque microenvironment on cell fate and release of soluble factors.<br /><b>Methods and results</b><br />Incubation of hMSCs with LL-37, an antimicrobial peptide abundantly found in plaques, increased biosynthesis of let-7f and N-formyl peptide receptor 2 (FPR2), enabling chemotactic invasion of the cells towards LL-37, as determined by qRT-PCR, flow cytometry, and cell invasion assay analysis. In an Apoe-/- mouse model of atherosclerosis, circulating hMSCs preferentially adhered to athero-prone endothelium. This property was facilitated by elevated levels of let-7f in the hMSCs, as assayed by ex vivo artery perfusion and two-photon laser scanning microscopy. Exposure of hMSCs to homogenized human atheromatous plaque material considerably induced the production of various cytokines, chemokines, matrix metalloproteinases, and tissue inhibitors of metalloproteinases, as studied by PCR array and western blot analysis. Moreover, exposure to human plaque extracts elicited differentiation of hMSCs into cells of the myogenic lineage, suggesting a potentially plaque-stabilizing effect.<br /><b>Conclusions</b><br />Our findings indicate that let-7f promotes hMSC tropism towards atheromas through the LL-37/FPR2 axis and demonstrate that hMSCs upon contact with human plaque environment develop a potentially athero-protective signature impacting the pathophysiology of atherosclerosis.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:155-166</small></div>

<div><h4>CRISPR and cardiovascular diseases.</h4><i>Musunuru K</i><br /><AbstractText>CRISPR technologies have progressed by leaps and bounds over the past decade, not only having a transformative effect on biomedical research but also yielding new therapies that are poised to enter the clinic. In this review, I give an overview of (i) the various CRISPR DNA-editing technologies, including standard nuclease gene editing, base editing, prime editing, and epigenome editing, (ii) their impact on cardiovascular basic science research, including animal models, human pluripotent stem cell models, and functional screens, and (iii) emerging therapeutic applications for patients with cardiovascular diseases, focusing on the examples of hypercholesterolaemia, transthyretin amyloidosis, and Duchenne muscular dystrophy.</AbstractText><br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:79-93</small></div>

<div><h4>Activating P2Y1 receptors improves function in arteries with repressed autophagy.</h4><i>Cho JM, Park SK, Kwon OS, Taylor La Salle D, ... Trinity JD, Symons JD</i><br /><b>Aim</b><br />The importance of endothelial cell (EC) autophagy to vascular homeostasis in the context of health and disease is evolving. Earlier, we reported that intact EC autophagy is requisite to maintain shear-stress-induced nitric oxide (NO) generation via glycolysis-dependent purinergic signalling to endothelial NO synthase (eNOS). Here, we illustrate the translational and functional significance of these findings.<br /><b>Methods and results</b><br />First, we assessed translational relevance using older male humans and mice that exhibit blunted EC autophagy and impaired arterial function vs. adult controls. Active hyperaemia evoked by rhythmic handgrip exercise-elevated radial artery shear-rate similarly from baseline in adult and older subjects for 60 min. Compared with baseline, indexes of autophagy initiation, p-eNOSS1177 activation, and NO generation, occurred in radial artery ECs obtained from adult but not older volunteers. Regarding mice, indexes of autophagy and p-eNOSS1177 activation were robust in ECs from adult but not older animals that completed 60-min treadmill-running. Furthermore, 20 dyne • cm2 laminar shear stress × 45-min increased autophagic flux, glycolysis, ATP production, and p-eNOSS1177 in primary arterial ECs obtained from adult but not older mice. Concerning functional relevance, we next questioned whether the inability to initiate EC autophagy, glycolysis, and p-eNOSS1177in vitro precipitates arterial dysfunction ex vivo. Compromised intraluminal flow-mediated vasodilation displayed by arteries from older vs. adult mice was recapitulated in vessels from adult mice by (i) NO synthase inhibition; (ii) acute autophagy impairment using 3-methyladenine (3-MA); (iii) EC Atg3 depletion (iecAtg3KO mice); (iv) purinergic 2Y1-receptor (P2Y1-R) blockade; and (v) germline depletion of P2Y1-Rs. Importantly, P2Y1-R activation using 2-methylthio-ADP (2-Me-ADP) improved vasodilatory capacity in arteries from (i) adult mice treated with 3-MA; (ii) adult iecAtg3KO mice; and (iii) older animals with repressed EC autophagy.<br /><b>Conclusions</b><br />Arterial dysfunction concurrent with pharmacological, genetic, and age-associated EC autophagy compromise is improved by activating P2Y1-Rs.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf the of European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:252-267</small></div>

<div><h4>Maturation of hiPSC-derived cardiomyocytes promotes adult alternative splicing of SCN5A and reveals changes in sodium current associated with cardiac arrhythmia.</h4><i>Campostrini G, Kosmidis G, Ward-van Oostwaard D, Davis RP, ... Mummery CL, Bellin M</i><br /><b>Aims</b><br />Human-induced pluripotent stem cell-cardiomyocytes (hiPSC-CMs) are widely used to study arrhythmia-associated mutations in ion channels. Among these, the cardiac sodium channel SCN5A undergoes foetal-to-adult isoform switching around birth. Conventional hiPSC-CM cultures, which are phenotypically foetal, have thus far been unable to capture mutations in adult gene isoforms. Here, we investigated whether tri-cellular cross-talk in a three-dimensional (3D) cardiac microtissue (MT) promoted post-natal SCN5A maturation in hiPSC-CMs.<br /><b>Methods and results</b><br />We derived patient hiPSC-CMs carrying compound mutations in the adult SCN5A exon 6B and exon 4. Electrophysiological properties of patient hiPSC-CMs in monolayer were not altered by the exon 6B mutation compared with isogenic controls since it is not expressed; further, CRISPR/Cas9-mediated excision of the foetal exon 6A did not promote adult SCN5A expression. However, when hiPSC-CMs were matured in 3D cardiac MTs, SCN5A underwent isoform switch and the functional consequences of the mutation located in exon 6B were revealed. Up-regulation of the splicing factor muscleblind-like protein 1 (MBNL1) drove SCN5A post-natal maturation in microtissues since its overexpression in hiPSC-CMs was sufficient to promote exon 6B inclusion, whilst knocking-out MBNL1 failed to foster isoform switch.<br /><b>Conclusions</b><br />Our study shows that (i) the tri-cellular cardiac microtissues promote post-natal SCN5A isoform switch in hiPSC-CMs, (ii) adult splicing of SCN5A is driven by MBNL1 in these tissues, and (iii) this model can be used for examining post-natal cardiac arrhythmias due to mutations in the exon 6B.<br /><b>Translational perspective</b><br />The cardiac sodium channel is essential for conducting the electrical impulse in the heart. Postnatal alternative splicing regulation causes mutual exclusive inclusion of fetal or adult exons of the corresponding gene, SCN5A. Typically, immature hiPSCCMs fall short in studying the effect of mutations located in the adult exon. We describe here that an innovative tri-cellular three-dimensional cardiac microtissue culture promotes hiPSC-CMs maturation through upregulation of MBNL1, thus revealing the effect of a pathogenic genetic variant located in the SCN5A adult exon. These results help advancing the use of hiPSC-CMs in studying adult heart disease and for developing personalized medicine applications.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:167-182</small></div>

<div><h4>Intravascular imaging assessment of pharmacotherapies targeting atherosclerosis: advantages and limitations in predicting their prognostic implications.</h4><i>Tufaro V, Serruys PW, Räber L, Bennett MR, ... Baumbach A, Bourantas CV</i><br /><AbstractText>Intravascular imaging has been often used over the recent years to examine the efficacy of emerging therapies targeting plaque evolution. Serial intravascular ultrasound, optical coherence tomography, or near-infrared spectroscopy-intravascular ultrasound studies have allowed us to evaluate the effects of different therapies on plaque burden and morphology, providing unique mechanistic insights about the mode of action of these treatments. Plaque burden reduction, a decrease in necrotic core component or macrophage accumulation-which has been associated with inflammation-and an increase in fibrous cap thickness over fibroatheromas have been used as surrogate endpoints to assess the value of several drugs in inhibiting plaque evolution and improving clinical outcomes. However, some reports have demonstrated weak associations between the effects of novel treatments on coronary atheroma and composition and their prognostic implications. This review examines the value of invasive imaging in assessing pharmacotherapies targeting atherosclerosis. It summarizes the findings of serial intravascular imaging studies assessing the effects of different drugs on atheroma burden and morphology and compares them with the results of large-scale trials evaluating their impact on clinical outcome. Furthermore, it highlights the limited efficacy of established intravascular imaging surrogate endpoints in predicting the prognostic value of these pharmacotherapies and introduces alternative imaging endpoints based on multimodality/hybrid intravascular imaging that may enable more accurate assessment of the athero-protective and prognostic effects of emerging therapies.</AbstractText><br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:121-135</small></div>

<div><h4>RNF207 exacerbates pathological cardiac hypertrophy via post-translational modification of TAB1.</h4><i>Yuan L, Bu S, Du M, Wang Y, ... Yang L, Huang K</i><br /><b>Aims</b><br />The heart undergoes pathological remodelling, featured by the hypertrophic growth of cardiomyocytes and increased cardiac fibrosis, under biomechanical stress such as haemodynamic overload. Ring Finger Protein 207 (RNF207) is an E3 ubiquitin ligase that is predominantly expressed in the heart, but its function remains elusive. In this study, we aimed to explore the role of RNF207 in the development of pathological cardiac hypertrophy and dysfunction.<br /><b>Methods and results</b><br />Transverse aortic constriction (TAC) surgery was performed on mice to induce cardiac hypertrophy. Cardiac function and remodelling were evaluated by echocardiography, histological assessment, and molecular analyses. Our data indicated that RNF207 overexpression (OE) exacerbated cardiac hypertrophy, fibrosis, and systolic dysfunction. In contrast, TAC-induced cardiac remodelling was profoundly blunted in RNF207 knockdown (KD) hearts. In line with the in vivo findings, RNF207 OE augmented, whereas RNF207 KD alleviated, phenylephrine-induced cardiomyocyte hypertrophy in vitro. Mechanistically, we demonstrated that RNF207 elicited detrimental effects by promoting K63-linked ubiquitination of TAK1-binding protein 1 (TAB1), which triggered the autophosphorylation of transforming growth factor-β activated kinase 1 (TAK1) and the activation of downstream p38 and c-Jun N-terminal kinase (JNK)1/2 signalling pathways. In the TAB1-KD cardiomyocytes, RNF207-OE-induced cell hypertrophy was significantly attenuated, indicating that RNF207-induced hypertrophy is, at least in part, TAB1-dependent.<br /><b>Conclusions</b><br />This study demonstrates that RNF207 exacerbates pressure overload-induced cardiac hypertrophy and dysfunction via post-translational modification of TAB1.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:183-194</small></div>

<div><h4>Eosinophils protect pressure overload- and β-adrenoreceptor agonist-induced cardiac hypertrophy.</h4><i>Yang C, Li J, Deng Z, Luo S, ... Guo J, Shi GP</i><br /><b>Aims</b><br />Blood eosinophil (EOS) counts and EOS cationic protein (ECP) levels associate positively with major cardiovascular disease (CVD) risk factors and prevalence. This study investigates the role of EOS in cardiac hypertrophy.<br /><b>Methods and results</b><br />A retrospective cross-section study of 644 consecutive inpatients with hypertension examined the association between blood EOS counts and cardiac hypertrophy. Pressure overload- and β-adrenoreceptor agonist isoproterenol-induced cardiac hypertrophy was produced in EOS-deficient ΔdblGATA mice. This study revealed positive correlations between blood EOS counts and left ventricular (LV) mass and mass index in humans. ΔdblGATA mice showed exacerbated cardiac hypertrophy and dysfunction, with increased LV wall thickness, reduced LV internal diameter, and increased myocardial cell size, death, and fibrosis. Repopulation of EOS from wild-type (WT) mice, but not those from IL4-deficient mice ameliorated cardiac hypertrophy and cardiac dysfunctions. In ΔdblGATA and WT mice, administration of ECP mEar1 improved cardiac hypertrophy and function. Mechanistic studies demonstrated that EOS expression of IL4, IL13, and mEar1 was essential to control mouse cardiomyocyte hypertrophy and death and cardiac fibroblast TGF-β signalling and fibrotic protein synthesis. The use of human cardiac cells yielded the same results. Human ECP, EOS-derived neurotoxin, human EOS, or murine recombinant mEar1 reduced human cardiomyocyte death and hypertrophy and human cardiac fibroblast TGF-β signalling.<br /><b>Conclusion</b><br />Although blood EOS counts correlated positively with LV mass or LV mass index in humans, this study established a cardioprotective role for EOS IL4 and cationic proteins in cardiac hypertrophy and tested a therapeutic possibility of ECPs in this human CVD.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:195-212</small></div>

<div><h4>The sympathetic nervous system exacerbates carotid body sensitivity in hypertension.</h4><i>Felippe ISA, Zera T, da Silva MP, Moraes DJA, McBryde F, Paton JFR</i><br /><b>Aims</b><br />The carotid bodies (CBs) of spontaneously hypertensive (SH) rats exhibit hypertonicity and hyperreflexia contributing to heightened peripheral sympathetic outflow. We hypothesized that CB hyperexcitability is driven by its own sympathetic innervation.<br /><b>Methods and results</b><br />To test this, the chemoreflex was activated (NaCN 50-100 µL, 0.4 µg/µL) in SH and Wistar rats in situ before and after: (i) electrical stimulation (ES; 30 Hz, 2 ms, 10 V) of the superior cervical ganglion (SCG), which innervates the CB; (ii) unilateral resection of the SCG (SCGx); (iii) CB injections of an α1-adrenergic receptor agonist (phenylephrine, 50 µL, 1 mmol/L), and (iv) α1-adrenergic receptor antagonist prazosin (40 µL, 1 mmol/L) or tamsulosin (50 µL, 1 mmol/L). ES of the SCG enhanced CB-evoked sympathoexcitation by 40-50% (P < 0.05) with no difference between rat strains. Unilateral SCGx attenuated the CB-evoked sympathoexcitation in SH (62%; P < 0.01) but was without effect in Wistar rats; it also abolished the ongoing firing of chemoreceptive petrosal neurones of SH rats, which became hyperpolarized. In Wistar rats, CB injections of phenylephrine enhanced CB-evoked sympathoexcitation (33%; P < 0.05), which was prevented by prazosin (26%; P < 0.05) in SH rats. Tamsulosin alone reproduced the effects of prazosin in SH rats and prevented the sensitizing effect of the SCG following ES. Within the CB, α1A- and α1B-adrenoreceptors were co-localized on both glomus cells and blood vessels. In conscious SH rats instrumented for recording blood pressure (BP), the CB-evoked pressor response was attenuated after SCGx, and systolic BP fell by 16 ± 4.85 mmHg.<br /><b>Conclusions</b><br />The sympathetic innervation of the CB is tonically activated and sensitizes the CB of SH but not Wistar rats. Furthermore, sensitization of CB-evoked reflex sympathoexcitation appears to be mediated by α1-adrenoceptors located either on the vasculature and/or glomus cells. The SCG is novel target for controlling CB pathophysiology in hypertension.<br /><br />© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 17 Mar 2023; 119:316-331</small></div>

<div><h4>Clinical and prognostic associations of autoantibodies recognizing adrenergic/muscarinic receptors in patients with heart failure.</h4><i>Markousis-Mavrogenis G, Minich WB, Al-Mubarak AA, Anker SD, ... Schomburg L, Bomer N</i><br /><b>Aims</b><br />The importance of autoantibodies (AABs) against adrenergic/muscarinic receptors in heart failure (HF) is not well-understood. We investigated the prevalence and clinical/prognostic associations of four AABs recognizing the M2-muscarinic receptor or the β1-, β2-, or β3-adrenergic receptor in a large and well-characterized cohort of patients with HF.<br /><b>Methods and results</b><br />Serum samples from 2256 patients with HF from the BIOSTAT-CHF cohort and 299 healthy controls were analyzed using newly established chemiluminescence immunoassays. The primary outcome was a composite of all-cause mortality and HF-rehospitalization at 2-year follow-up, and each outcome was also separately investigated. Collectively, 382 (16.9%) patients and 37 (12.4%) controls were seropositive for ≥1 AAB (p=0.045). Seropositivity occurred more frequently only for anti-M2 AABs (p=0.025). Amongst patients with HF, seropositivity was associated with the presence of comorbidities (renal disease, chronic obstructive pulmonary disease, stroke, atrial fibrillation), and with medication use. Only anti-β1 AAB seropositivity was associated with the primary outcome [hazard ratio (95% confidence interval): 1.37 (1.04-1.81), p=0.024] and HF-rehospitalization [1.57 (1.13-2.19), p=0.010] in univariable analyses, but remained associated only with HF-rehospitalization after multivariable adjustment for the BIOSTAT-CHF risk model [1.47 (1.05-2.07), p=0.030]. Principal component analyses showed considerable overlap in B-lymphocyte activity between seropositive and seronegative patients, based on 31 circulating biomarkers related to B-lymphocyte function.<br /><b>Conclusions</b><br />AAB seropositivity was not strongly associated with adverse outcomes in HF and was mostly related to the presence of comorbidities and medication use. Only anti-β1 AABs were independently associated with HF-rehospitalization. The exact clinical value of AABs remains to be elucidated.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 08 Mar 2023; epub ahead of print</small></div>

<div><h4>Cardiomyocyte-specific PCSK9 deficiency compromises mitochondrial bioenergetics and heart function.</h4><i>Laudette M, Lindbom M, Arif M, Cinato M, ... Levin MC, Borén J</i><br /><b>Aims</b><br />PCSK9, which is expressed mainly in the liver and at low levels in the heart, regulates cholesterol levels by directing low-density lipoprotein receptors to degradation. Studies to determine the role of PCSK9 in the heart are complicated by the close link between cardiac function and systemic lipid metabolism. Here, we sought to elucidate the function of PCSK9 specifically in the heart by generating and analysing mice with cardiomyocyte-specific Pcsk9 deficiency (CM-Pcsk9-/- mice) and by silencing Pcsk9 acutely in a cell culture model of adult cardiomyocyte-like cells.<br /><b>Methods and results</b><br />Mice with cardiomyocyte-specific deletion of Pcsk9 had reduced contractile capacity, impaired cardiac function and left ventricular dilatation at 28 weeks of age and died prematurely. Transcriptomic analyses revealed alterations of signalling pathways linked to cardiomyopathy and energy metabolism in hearts from CM-Pcsk9-/- mice versus wildtype littermates. In agreement, levels of genes and proteins involved in mitochondrial metabolism were reduced in CM-Pcsk9-/- hearts. By using a Seahorse flux analyser, we showed that mitochondrial but not glycolytic function was impaired in cardiomyocytes from CM-Pcsk9-/- mice. We further showed that assembly and activity of electron transport chain (ETC) complexes were altered in isolated mitochondria from CM-Pcsk9-/- mice. Circulating lipid levels were unchanged in CM-Pcsk9-/- mice, but the lipid composition of mitochondrial membranes was altered. In addition, cardiomyocytes from CM-Pcsk9-/- mice had an increased number of mitochondria-ER contacts and alterations in the morphology of cristae, the physical location of the ETC complexes. We also showed that acute Pcsk9 silencing in adult cardiomyocyte-like cells reduced the activity of ETC complexes and impaired mitochondrial metabolism.<br /><b>Conclusion</b><br />PCSK9, despite its low expression in cardiomyocytes, contributes to cardiac metabolic function, and PCSK9 deficiency in cardiomyocytes is linked to cardiomyopathy, impaired heart function, and compromised energy production.<br /><b>Translational perspective</b><br />PCSK9 is mainly present in the circulation where it regulates plasma cholesterol levels. Here we show that PCSK9 mediates intracellular functions that differ from its extracellular functions. We further show that intracellular PCSK9 in cardiomyocytes, despite low expression levels, is important for maintaining physiological cardiac metabolism and function.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 06 Mar 2023; epub ahead of print</small></div>

<div><h4>VEGF-D plasma levels and VEGFD genetic variants are independently associated with outcomes in patients with cardiovascular disease.</h4><i>Davidsson P, Eketjäll S, Eriksson N, Walentinsson A, ... White H, Wallentin L</i><br /><b>Aims</b><br />The vascular endothelial growth factor (VEGF) family is involved in pathophysiological mechanisms underlying cardio vascular (CV) diseases. The aim of this study was to investigate the associations between circulating VEGF ligands and/or soluble receptors and CV outcome in patients with acute coronary syndrome (ACS) and chronic coronary syndrome (CCS).<br /><b>Methods and results</b><br />Levels of VEGF biomarkers, including bFGF, Flt-1, KDR (VEGFR2), PlGF, Tie-2, VEGF-A, VEGF-C, and VEGF-D, were measured in the PLATO ACS cohort (n = 2,091, discovery cohort). Subsequently, VEGF-D was also measured in the STABILITY CCS cohort (n = 4,015, confirmation cohort) to verify associations with CV outcomes. Associations between plasma VEGF-D and outcomes were analyzed by multiple Cox regression models with hazard ratios (HR [95% CI]) comparing the upper versus the lower quartile of VEGF-D. GWAS of VEGF-D in PLATO identified SNPs that were used as genetic instruments in Mendelian randomization (MR) meta-analyses versus clinical endpoints. GWAS and MR was performed in patients with ACS from PLATO (n = 10,013) and FRISC-II (n = 2,952), and with CCS from the STABILITY trial (n = 10,786). VEGF-D, KDR, Flt-1, and PlGF showed significant association with CV outcomes. VEGF-D was most strongly associated with CV death (p = 3.73e-05, HR 1.892 [1.419, 2.522]). Genome wide significant associations with VEGF-D levels were identified at the VEGFD locus on chromosome Xp22. MR analyses of the combined top ranked SNPs (GWAS p-values; rs192812042, p = 5.82e-20; rs234500, p = 1.97e-14) demonstrated a significant effect on CV mortality (p = 0.0257, HR 1.81 [1.07, 3.04] per increase of one unit in log VEGF-D).<br /><b>Conclusion</b><br />This is the first large-scale cohort study to demonstrate that both VEGF-D plasma levels and VEGFD genetic variants are independently associated with CV outcomes in patients with ACS and CCS. Measurements of VEGF-D levels and/or VEGFD genetic variants may provide incremental prognostic information in patients with ACS and CCS.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.<br /><br /><small>Cardiovasc Res: 03 Mar 2023; epub ahead of print</small></div>

<div><h4>Phospholamban pentamerization increases sensitivity and dynamic range of cardiac relaxation.</h4><i>Funk F, Kronenbitter A, Hackert K, Oebbeke M, ... Koch D, Schmitt JP</i><br /><b>Aims</b><br />A key event in the regulation of cardiac contraction and relaxation is the phosphorylation of phospholamban (PLN) that relieves the inhibition of the sarco/endoplasmic Ca2+-ATPase SERCA2a. PLN exists in an equilibrium between monomers and pentamers. While only monomers can inhibit SERCA2a by direct interaction, the functional role of pentamers is still unclear. This study investigates the functional consequences of PLN pentamerization.<br /><b>Methods and results</b><br />We generated transgenic mouse models expressing either a PLN mutant that cannot form pentamers (TgAFA-PLN) or wildtype PLN (TgPLN) in a PLN-deficient background. TgAFA-PLN hearts demonstrated 3-fold stronger phosphorylation of monomeric PLN, accelerated Ca2+ cycling of cardiomyocytes and enhanced contraction and relaxation of sarcomeres and whole hearts in vivo. All of these effects were observed under baseline conditions and abrogated upon inhibition of protein kinase A (PKA). Mechanistically, far western kinase assays revealed that PLN pentamers are phosphorylated by PKA directly and independent of any subunit exchange for free monomers. In vitro-phosphorylation of synthetic PLN demonstrated that pentamers even provide a preferred PKA substrate and compete with monomers for the kinase, thereby reducing monomer phosphorylation and maximizing SERCA2a inhibition. However, β-adrenergic stimulation induced strong PLN monomer phosphorylation in TgPLN hearts and sharp acceleration of cardiomyocyte Ca2+ cycling and hemodynamic values that now were indistinguishable from TgAFA-PLN and PLN-KO hearts. The pathophysiological relevance of PLN pentamerization was evaluated using transverse aortic constriction (TAC) to induce left ventricular pressure overload. Compared to TgPLN, TgAFA-PLN mice demonstrated reduced survival after TAC, impaired cardiac hemodynamics, failure to respond to adrenergic stimulation, higher heart weight, and increased myocardial fibrosis.<br /><b>Conclusions</b><br />The findings show that PLN pentamerization greatly impacts on SERCA2a activity as it mediates the full range of PLN effects from maximum inhibition to full release of SERCA2a. function. This regulation is important for myocardial adaptation to sustained pressure overload.<br /><b>Translational perspective</b><br />Pentamerization of PLN adds to the regulation of cardiac contractile function and facilitates myocardial transition to an energy saving mode during resting phases. Thus, PLN pentamers would protect cardiomyocytes from energetic deficits, and they improve stress adaptation of the heart as shown for sustained pressure overload in this study. Strategies that target PLN pentamerization promise therapeutic potential in the treatment of myocardial maladaptation to stress as well as cardiac pathologies associated with altered monomer-to-pentamer ratios, e.g., cardiomyopathies due to PLN mutations, certain types of heart failure, and aged hearts.<br /><br />© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.<br /><br /><small>Cardiovasc Res: 03 Mar 2023; epub ahead of print</small></div>