Journal: Circ Res

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Abstract

COVID-19 and Cardiovascular Disease: From Bench to Bedside.

Chung MK, Zidar DA, Bristow MR, Cameron SJ, ... Barnard J, Loscalzo J
A pandemic of historic impact, coronavirus disease 2019 (COVID-19) has potential consequences on the cardiovascular health of millions of people who survive infection worldwide. Severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2), the etiologic agent of COVID-19, can infect the heart, vascular tissues, and circulating cells through ACE2 (angiotensin-converting enzyme 2), the host cell receptor for the viral spike protein. Acute cardiac injury is a common extrapulmonary manifestation of COVID-19 with potential chronic consequences. This update provides a review of the clinical manifestations of cardiovascular involvement, potential direct SARS-CoV-2 and indirect immune response mechanisms impacting the cardiovascular system, and implications for the management of patients after recovery from acute COVID-19 infection.



Circ Res: 15 Apr 2021; 128:1214-1236
Chung MK, Zidar DA, Bristow MR, Cameron SJ, ... Barnard J, Loscalzo J
Circ Res: 15 Apr 2021; 128:1214-1236 | PMID: 33856918
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Abstract

An Immuno-Cardiac Model for Macrophage-Mediated Inflammation in COVID-19 Hearts.

Yang L, Han Y, Jaffre F, Nilsson-Payant BE, ... Chen Z, Chen S
Rationale: While respiratory failure is a frequent and clinically significant outcome of COVID-19, cardiac complications are a common feature in hospitalized COVID-19 patients and are associated with worse patient outcomes. The cause of cardiac injury in COVID-19 patients is not yet known. Case reports of COVID-19 autopsy heart samples have demonstrated abnormal inflammatory infiltration of macrophages in heart tissues.Objective: Generate an immuno-cardiac co-culture platform to model macrophage-mediated hyper-inflammation in COVID-19 hearts and screen for drugs that can block the macrophage-mediated inflammation.
Methods and results:
We systematically compared autopsy samples from non-COVID-19 donors and COVID-19 patients using RNA-seq and immunohistochemistry. We observed strikingly increased expression levels of CCL2 as well as macrophage infiltration in heart tissues of COVID-19 patients. We generated an immuno-cardiac co-culture platform containing human pluripotent stem cell (hPSC)-derived cardiomyocytes (CMs) and macrophages. We found that macrophages induce increased reactive oxygen species (ROS) and apoptosis in CMs by secreting IL-6 and TNF-α after SARS-CoV-2 exposure. Using this immuno-cardiac co-culture platform, we performed a high content screen and identified ranolazine and tofacitinib as compounds that protect CMs from macrophage-induced cardiotoxicity. Conclusions: We established an immuno-host co-culture system to study macrophage-induced host cell damage following SARS-CoV-2 infection and identified FDA-approved drug candidates that alleviate the macrophage-mediated hyper-inflammation and cellular injury.




Circ Res: 14 Apr 2021; epub ahead of print
Yang L, Han Y, Jaffre F, Nilsson-Payant BE, ... Chen Z, Chen S
Circ Res: 14 Apr 2021; epub ahead of print | PMID: 33853355
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Abstract

Intrinsic Electrical Remodeling Underlies Atrioventricular Block in Athletes.

Mesirca P, Nakao S, Nissen SD, Forte G, ... Boyett MR, D\'Souza A
Rationale: Athletes present with atrioventricular node (AV node) dysfunction manifesting as AV block. This can necessitate electronic pacemaker implantation, known to be more frequent in athletes with a long training history. Objective: AV block in athletes is attributed to high vagal tone. Here we investigated the alternative hypothesis that electrical remodeling of the AV node is responsible.
Methods and results:
Radio-telemetry ECG data and AV node biopsies were collected in sedentary and trained Standardbred racehorses, a large-animal model of the athlete\'s heart. Trained horses presented with longer PR intervals (that persisted under complete autonomic block) versus sedentary horses, concomitant with reduced expression of key ion channels involved in AV node conduction: L-type Ca2+ channel subunit CaV1.2 and the hyperpolarization-activated cyclic nucleotide gated channel 4 (HCN4). AV node electrophysiology was explored further in mice; prolongation of the PR interval (in vivo and ex vivo), Wenckebach cycle length and AV node refractory period was observed in mice trained by swimming versus sedentary mice. Transcriptional profiling in laser-capture microdissected AV node revealed striking reduction in pacemaking ion channels in trained mice, translating into protein downregulation of CaV1.2 and HCN4. Correspondingly, patch clamp recordings in isolated AV node myocytes demonstrated a training-induced reduction in ICa,L and If density that likely contributed to the observed lower frequency of action potential firing in trained cohorts. microRNA (miR) profiling and in vitro studies revealed miR-211-5p and miR-432 as direct regulators of CaV1.2 and HCN4. In vivo miRs suppression or detraining restored training-induced PR prolongation and ion channel remodeling. Conclusions: Training-induced AV node dysfunction is underscored by likely miR-mediated transcriptional remodeling that translates into reduced current density of key ionic currents involved in impulse generation and conduction. We conclude that electrical remodeling is a key mechanism underlying AV block in athletes.




Circ Res: 13 Apr 2021; epub ahead of print
Mesirca P, Nakao S, Nissen SD, Forte G, ... Boyett MR, D'Souza A
Circ Res: 13 Apr 2021; epub ahead of print | PMID: 33849278
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Abstract

Over-Expression of Intestinal Alkaline Phosphatase Attenuates Atherosclerosis.

Ghosh SS, Wang J, Yannie PJ, Cooper RC, ... Korzun W, Ghosh S
Rationale: Intestinal Alkaline Phosphatase (IAP) is secreted by enterocytes and is present on the apical surface. It not only detoxifies bacterial endotoxin lipopolysaccharide (LPS) in the gut lumen and limits intestinal inflammation but also restricts translocation of LPS into systemic circulation. Diet-induced intestinal barrier dysfunction and subsequent development of metabolic endotoxemia seen in diabetes and heart disease is associated with reduced IAP levels. To examine the direct effects of increased IAP expression on barrier function and development of metabolic diseases, we developed intestine-specific IAP transgenic mice (IAPTg) over-expressing human chimeric IAP. Objective:The aim of this study was to evaluate the effects of intestine-specific IAP overexpression on Western-type diet (WD)-induced atherosclerosis in Ldlr-/- mice.
Methods and results:
IAPTg mice crossed into Ldlr-/- background (Ldlr-/-IAPTg) and Ldlr-/- littermates were fed WD for 16 weeks. Intestinal barrier dysfunction was assessed by monitoring plasma LPS levels and histological examination of colon. Over-expression of IAP attenuated WD-induced disruption of the colonic mucous layer, reducing intestinal barrier dysfunction and plasma LPS levels. Significant reduction in body, liver and adipose tissue weight was also seen in WD-fed Ldlr-/-IAPTg mice. Plasma and hepatic lipids were also significantly reduced in WD-fed Ldlr-/-IAPTg mice. Consistently, intestinal lipid absorption was attenuated in Ldlr-/-IAPTg mice with reduced expression of apical lipid transporters (CD36, FATP4 and NPC1L1) and intracellular lipid transport proteins (FABP1/2, SCP2). Attenuation of WD-induced atherosclerosis in Ldlr-/-IAPTg mice was demonstrated by significant reduction in arch and total aortic lesions as seen by enface analyses as well as significantly reduced atherosclerotic lesions in the ascending aorta of these mice.Conclusions: IAP overexpression improves intestinal barrier function by maintaining the integrity of the mucin layer in WD fed Ldlr-/-IAPTg mice and attenuates intestinal lipid absorption. Thus, by limiting translocation of gut-derived LPS and/or reducing plasma lipids, over-expression of IAP attenuates development of WD-induced atherosclerosis.




Circ Res: 08 Apr 2021; epub ahead of print
Ghosh SS, Wang J, Yannie PJ, Cooper RC, ... Korzun W, Ghosh S
Circ Res: 08 Apr 2021; epub ahead of print | PMID: 33834851
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Abstract

Randomized Trial of Etelcalcetide for Cardiac Hypertrophy in Hemodialysis.

Dörr K, Kammer M, Reindl-Schwaighofer R, Lorenz M, ... Erben RG, Oberbauer R
Rationale: Left ventricular hypertrophy (LVH) is highly prevalent in patients with chronic kidney disease and increases their risk of cardiac events and mortality. Fibroblast growth factor 23 (FGF23) and parathyroid hormone (PTH) levels, which are associated with the development of LVH, rise progressively with declining renal function. Objective: To determine whether FGF23 suppression by calcimimetic therapy may reduce LVH progression in comparison to FGF23 elevation under vitamin D analogs at equal PTH suppression under either therapy as well as tight volume control.
Methods and results:
We conducted a single-blinded trial with 1:1 block randomization to investigate the effect of the intravenous treatment with etelcalcetide (ETL) versus alfacalcidol (ALFA) on LVH progression in 62 maintenance hemodialysis patients with secondary hyperparathyroidism and LVH. In the intention-to-treat analysis of 59 patients, the mean difference in the change of left ventricular mass index (LVMI) determined by cardiac magnetic resonance imaging from baseline to 12 months of treatment was -6.9 g/m² (95% confidence interval [CI] -12.6 to -1.2, p=0.022) in the ETL compared to the ALFA group. The effect estimate was -8.2 g/m² (95% CI -14 to -2.4) in the per-protocol analysis on 52 patients. The trajectories of PTH, phosphate and Klotho were similar in both groups throughout follow-up. FGF23 levels, which showed a strong positive association with LVMI, were decreasing under ETL and increasing under ALFA at similar PTH suppression. Mild hypocalcemia was the most common adverse event under ETL. Blood pressure and the distribution of antihypertensive medications were similar between groups. Conclusions: In this trial we were able to show that FGF23 suppression by ETL inhibited the progression of LVH compared to ALFA in hemodialysis patients. A successful prevention of increasing hypertrophy may reduce the risk of sudden cardiac death in this population.




Circ Res: 06 Apr 2021; epub ahead of print
Dörr K, Kammer M, Reindl-Schwaighofer R, Lorenz M, ... Erben RG, Oberbauer R
Circ Res: 06 Apr 2021; epub ahead of print | PMID: 33825489
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Abstract

Prevention of Fibrosis and Pathological Cardiac Remodeling by Salinomycin.

Burke RM, Dirkx RA, Quijada P, Lighthouse JK, ... Ashton J, Small EM
Rationale: Cardiomyopathy is characterized by the deposition of extracellular matrix by activated resident cardiac fibroblasts, called myofibroblasts. There are currently no therapeutic approaches to blunt the development of pathological fibrosis and ventricle chamber stiffening that ultimately leads to heart failure. Objective: We undertook a high-throughput screen to identify small molecule inhibitors of myofibroblast activation that might limit the progression of heart failure. We evaluated the therapeutic efficacy of the polyether ionophore salinomycin in patient derived cardiac fibroblasts and pre-clinical mouse models of ischemic and non-ischemic heart failure.
Methods and results:
Here, we demonstrate that salinomycin displays potent anti-fibrotic activity in cardiac fibroblasts obtained from heart failure patients. In pre-clinical studies, salinomycin prevents cardiac fibrosis and functional decline in mouse models of ischemic and non-ischemic heart disease. Remarkably, interventional treatment with salinomycin attenuates pre-established pathological cardiac remodeling secondary to hypertension, and limits scar expansion when administered after a severe myocardial infarction. Mechanistically, salinomycin inhibits cardiac fibroblast activation by preventing p38/MAPK and Rho signaling. Salinomycin also promotes cardiomyocyte survival and improves coronary vessel density, suggesting that cardioprotection conferred by salinomycin occurs via the integration of multiple mechanisms in multiple relevant cardiac cell types. Conclusions: These data establish salinomycin as an anti-fibrotic agent that targets multiple cardioprotection pathways, thereby holding promise for the treatment of heart failure patients.




Circ Res: 06 Apr 2021; epub ahead of print
Burke RM, Dirkx RA, Quijada P, Lighthouse JK, ... Ashton J, Small EM
Circ Res: 06 Apr 2021; epub ahead of print | PMID: 33825488
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Abstract

Hypertension in Low- and Middle-Income Countries.

Schutte AE, Srinivasapura Venkateshmurthy N, Mohan S, Prabhakaran D
In recent decades low- and middle-income countries (LMICs) have been witnessing a significant shift toward raised blood pressure; yet in LMICs, only 1 in 3 are aware of their hypertension status, and ≈8% have their blood pressure controlled. This rising burden widens the inequality gap, contributes to massive economic hardships of patients and carers, and increases costs to the health system, facing challenges such as low physician-to-patient ratios and lack of access to medicines. Established risk factors include unhealthy diet (high salt and low fruit and vegetable intake), physical inactivity, tobacco and alcohol use, and obesity. Emerging risk factors include pollution (air, water, noise, and light), urbanization, and a loss of green space. Risk factors that require further in-depth research are low birth weight and social and commercial determinants of health. Global actions include the HEARTS technical package and the push for universal health care. Promising research efforts highlight that successful interventions are feasible in LMICs. These include creation of health-promoting environments by introducing salt-reduction policies and sugar and alcohol tax; implementing cost-effective screening and simplified treatment protocols to mitigate treatment inertia; pooled procurement of low-cost single-pill combination therapy to improve adherence; increasing access to telehealth and mHealth (mobile health); and training health care staff, including community health workers, to strengthen team-based care. As the blood pressure trajectory continues creeping upward in LMICs, contextual research on effective, safe, and cost-effective interventions is urgent. New emergent risk factors require novel solutions. Lowering blood pressure in LMICs requires urgent global political and scientific priority and action.



Circ Res: 01 Apr 2021; 128:808-826
Schutte AE, Srinivasapura Venkateshmurthy N, Mohan S, Prabhakaran D
Circ Res: 01 Apr 2021; 128:808-826 | PMID: 33793340
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Abstract

Artificial Intelligence in Hypertension: Seeing Through a Glass Darkly.

Padmanabhan S, Tran TQB, Dominiczak AF
Hypertension remains the largest modifiable cause of mortality worldwide despite the availability of effective medications and sustained research efforts over the past 100 years. Hypertension requires transformative solutions that can help reduce the global burden of the disease. Artificial intelligence and machine learning, which have made a substantial impact on our everyday lives over the last decade may be the route to this transformation. However, artificial intelligence in health care is still in its nascent stages and realizing its potential requires numerous challenges to be overcome. In this review, we provide a clinician-centric perspective on artificial intelligence and machine learning as applied to medicine and hypertension. We focus on the main roadblocks impeding implementation of this technology in clinical care and describe efforts driving potential solutions. At the juncture, there is a critical requirement for clinical and scientific expertise to work in tandem with algorithmic innovation followed by rigorous validation and scrutiny to realize the promise of artificial intelligence-enabled health care for hypertension and other chronic diseases.



Circ Res: 01 Apr 2021; 128:1100-1118
Padmanabhan S, Tran TQB, Dominiczak AF
Circ Res: 01 Apr 2021; 128:1100-1118 | PMID: 33793339
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Abstract

Role of the Peroxisome Proliferator Activated Receptors in Hypertension.

Fang S, Livergood MC, Nakagawa P, Wu J, Sigmund CD
Nuclear receptors represent a large family of ligand-activated transcription factors which sense the physiological environment and make long-term adaptations by mediating changes in gene expression. In this review, we will first discuss the fundamental mechanisms by which nuclear receptors mediate their transcriptional responses. We will focus on the PPAR (peroxisome proliferator-activated receptor) family of adopted orphan receptors paying special attention to PPARγ, the isoform with the most compelling evidence as an important regulator of arterial blood pressure. We will review genetic data showing that rare mutations in PPARγ cause severe hypertension and clinical trial data which show that PPARγ activators have beneficial effects on blood pressure. We will detail the tissue- and cell-specific molecular mechanisms by which PPARs in the brain, kidney, vasculature, and immune system modulate blood pressure and related phenotypes, such as endothelial function. Finally, we will discuss the role of placental PPARs in preeclampsia, a life threatening form of hypertension during pregnancy. We will close with a viewpoint on future research directions and implications for developing novel therapies.



Circ Res: 01 Apr 2021; 128:1021-1039
Fang S, Livergood MC, Nakagawa P, Wu J, Sigmund CD
Circ Res: 01 Apr 2021; 128:1021-1039 | PMID: 33793338
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Abstract

Hypertension and Prohypertensive Antineoplastic Therapies in Cancer Patients.

van Dorst DCH, Dobbin SJH, Neves KB, Herrmann J, ... Danser AHJ, Lang NN
The development of a wide range of novel antineoplastic therapies has improved the prognosis for patients with a wide range of malignancies, which has increased the number of cancer survivors substantially. Despite the oncological benefit, cancer survivors are exposed to short- and long-term adverse cardiovascular toxicities associated with anticancer therapies. Systemic hypertension, the most common comorbidity among cancer patients, is a major contributor to the increased risk for developing these adverse cardiovascular events. Cancer and hypertension have common risk factors, have overlapping pathophysiological mechanisms and hypertension may also be a risk factor for some tumor types. Many cancer therapies have prohypertensive effects. Although some of the mechanisms by which these antineoplastic agents lead to hypertension have been characterized, further preclinical and clinical studies are required to investigate the exact pathophysiology and the optimal management of hypertension associated with anticancer therapy. In this way, monitoring and management of hypertension before, during, and after cancer treatment can be improved to minimize cardiovascular risks. This is vital to optimize cardiovascular health in patients with cancer and survivors, and to ensure that advances in terms of cancer survivorship do not come at the expense of increased cardiovascular toxicities.



Circ Res: 01 Apr 2021; 128:1040-1061
van Dorst DCH, Dobbin SJH, Neves KB, Herrmann J, ... Danser AHJ, Lang NN
Circ Res: 01 Apr 2021; 128:1040-1061 | PMID: 33793337
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Abstract

Hypertension: Do Inflammation and Immunity Hold the Key to Solving this Epidemic?

Madhur MS, Elijovich F, Alexander MR, Pitzer A, ... Laffer CL, Kirabo A
Elevated cardiovascular risk including stroke, heart failure, and heart attack is present even after normalization of blood pressure in patients with hypertension. Underlying immune cell activation is a likely culprit. Although immune cells are important for protection against invading pathogens, their chronic overactivation may lead to tissue damage and high blood pressure. Triggers that may initiate immune activation include viral infections, autoimmunity, and lifestyle factors such as excess dietary salt. These conditions activate the immune system either directly or through their impact on the gut microbiome, which ultimately produces chronic inflammation and hypertension. T cells are central to the immune responses contributing to hypertension. They are activated in part by binding specific antigens that are presented in major histocompatibility complex molecules on professional antigen-presenting cells, and they generate repertoires of rearranged T-cell receptors. Activated T cells infiltrate tissues and produce cytokines including interleukin 17A, which promote renal and vascular dysfunction and end-organ damage leading to hypertension. In this comprehensive review, we highlight environmental, genetic, and microbial associated mechanisms contributing to both innate and adaptive immune cell activation leading to hypertension. Targeting the underlying chronic immune cell activation in hypertension has the potential to mitigate the excess cardiovascular risk associated with this common and deadly disease.



Circ Res: 01 Apr 2021; 128:908-933
Madhur MS, Elijovich F, Alexander MR, Pitzer A, ... Laffer CL, Kirabo A
Circ Res: 01 Apr 2021; 128:908-933 | PMID: 33793336
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Abstract

Oxidative Stress and Hypertension.

Griendling KK, Camargo LL, Rios FJ, Alves-Lopes R, Montezano AC, Touyz RM
A link between oxidative stress and hypertension has been firmly established in multiple animal models of hypertension but remains elusive in humans. While initial studies focused on inactivation of nitric oxide by superoxide, our understanding of relevant reactive oxygen species (superoxide, hydrogen peroxide, and peroxynitrite) and how they modify complex signaling pathways to promote hypertension has expanded significantly. In this review, we summarize recent advances in delineating the primary and secondary sources of reactive oxygen species (nicotinamide adenine dinucleotide phosphate oxidases, uncoupled endothelial nitric oxide synthase, endoplasmic reticulum, and mitochondria), the posttranslational oxidative modifications they induce on protein targets important for redox signaling, their interplay with endogenous antioxidant systems, and the role of inflammasome activation and endoplasmic reticular stress in the development of hypertension. We highlight how oxidative stress in different organ systems contributes to hypertension, describe new animal models that have clarified the importance of specific proteins, and discuss clinical studies that shed light on how these processes and pathways are altered in human hypertension. Finally, we focus on the promise of redox proteomics and systems biology to help us fully understand the relationship between ROS and hypertension and their potential for designing and evaluating novel antihypertensive therapies.



Circ Res: 01 Apr 2021; 128:993-1020
Griendling KK, Camargo LL, Rios FJ, Alves-Lopes R, Montezano AC, Touyz RM
Circ Res: 01 Apr 2021; 128:993-1020 | PMID: 33793335
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Abstract

Renin Cells, the Kidney, and Hypertension.

Sequeira-Lopez MLS, Gomez RA
Renin cells are essential for survival perfected throughout evolution to ensure normal development and defend the organism against a variety of homeostatic threats. During embryonic and early postnatal life, they are progenitors that participate in the morphogenesis of the renal arterial tree. In adult life, they are capable of regenerating injured glomeruli, control blood pressure, fluid-electrolyte balance, tissue perfusion, and in turn, the delivery of oxygen and nutrients to cells. Throughout life, renin cell descendants retain the plasticity or memory to regain the renin phenotype when homeostasis is threatened. To perform all of these functions and maintain well-being, renin cells must regulate their identity and fate. Here, we review the major mechanisms that control the differentiation and fate of renin cells, the chromatin events that control the memory of the renin phenotype, and the major pathways that determine their plasticity. We also examine how chronic stimulation of renin cells alters their fate leading to the development of a severe and concentric hypertrophy of the intrarenal arteries and arterioles. Lastly, we provide examples of additional changes in renin cell fate that contribute to equally severe kidney disorders.



Circ Res: 01 Apr 2021; 128:887-907
Sequeira-Lopez MLS, Gomez RA
Circ Res: 01 Apr 2021; 128:887-907 | PMID: 33793334
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Abstract

Vascular Stress Signaling in Hypertension.

Cicalese SM, da Silva JF, Priviero F, Webb RC, Eguchi S, Tostes RC
Cells respond to stress by activating a variety of defense signaling pathways, including cell survival and cell death pathways. Although cell survival signaling helps the cell to recover from acute insults, cell death or senescence pathways induced by chronic insults can lead to unresolved pathologies. Arterial hypertension results from chronic physiological maladaptation against various stressors represented by abnormal circulating or local neurohormonal factors, mechanical stress, intracellular accumulation of toxic molecules, and dysfunctional organelles. Hypertension and aging share common mechanisms that mediate or prolong chronic cell stress, such as endoplasmic reticulum stress and accumulation of protein aggregates, oxidative stress, metabolic mitochondrial stress, DNA damage, stress-induced senescence, and proinflammatory processes. This review discusses common adaptive signaling mechanisms against these stresses including unfolded protein responses, antioxidant response element signaling, autophagy, mitophagy, and mitochondrial fission/fusion, STING (signaling effector stimulator of interferon genes)-mediated responses, and activation of pattern recognition receptors. The main molecular mechanisms by which the vasculature copes with hypertensive and aging stressors are presented and recent advancements in stress-adaptive signaling mechanisms as well as potential therapeutic targets are discussed.



Circ Res: 01 Apr 2021; 128:969-992
Cicalese SM, da Silva JF, Priviero F, Webb RC, Eguchi S, Tostes RC
Circ Res: 01 Apr 2021; 128:969-992 | PMID: 33793333
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Abstract

The Gut Microbiome in Hypertension: Recent Advances and Future Perspectives.

Avery EG, Bartolomaeus H, Maifeld A, Marko L, ... Forslund SK, Müller DN
The pathogenesis of hypertension is known to involve a diverse range of contributing factors including genetic, environmental, hormonal, hemodynamic and inflammatory forces, to name a few. There is mounting evidence to suggest that the gut microbiome plays an important role in the development and pathogenesis of hypertension. The gastrointestinal tract, which houses the largest compartment of immune cells in the body, represents the intersection of the environment and the host. Accordingly, lifestyle factors shape and are modulated by the microbiome, modifying the risk for hypertensive disease. One well-studied example is the consumption of dietary fibers, which leads to the production of short-chain fatty acids and can contribute to the expansion of anti-inflammatory immune cells, consequently protecting against the progression of hypertension. Dietary interventions such as fasting have also been shown to impact hypertension via the microbiome. Studying the microbiome in hypertensive disease presents a variety of unique challenges to the use of traditional model systems. Integrating microbiome considerations into preclinical research is crucial, and novel strategies to account for reciprocal host-microbiome interactions, such as the wildling mouse model, may provide new opportunities for translation. The intricacies of the role of the microbiome in hypertensive disease is a matter of ongoing research, and there are several technical considerations which should be accounted for moving forward. In this review we provide insights into the host-microbiome interaction and summarize the evidence of its importance in the regulation of blood pressure. Additionally, we provide recommendations for ongoing and future research, such that important insights from the microbiome field at large can be readily integrated in the context of hypertension.



Circ Res: 01 Apr 2021; 128:934-950
Avery EG, Bartolomaeus H, Maifeld A, Marko L, ... Forslund SK, Müller DN
Circ Res: 01 Apr 2021; 128:934-950 | PMID: 33793332
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Abstract

Device Therapy of Hypertension.

Mahfoud F, Schlaich MP, Lobo MD
In the past decade, efforts to improve blood pressure control have looked beyond conventional approaches of lifestyle modification and drug therapy to embrace interventional therapies. Based upon animal and human studies clearly demonstrating a key role for the sympathetic nervous system in the etiology of hypertension, the newer technologies that have emerged are predominantly aimed at neuromodulation of peripheral nervous system targets. These include renal denervation, baroreflex activation therapy, endovascular baroreflex amplification therapy, carotid body ablation, and pacemaker-mediated programmable hypertension control. Of these, renal denervation is the most mature, and with a recent series of proof-of-concept trials demonstrating the safety and efficacy of radiofrequency and more recently ultrasound-based renal denervation, this technology is poised to become available as a viable treatment option for hypertension in the foreseeable future. With regard to baroreflex activation therapy, endovascular baroreflex amplification, carotid body ablation, and programmable hypertension control, these are developing technologies for which more human data are required. Importantly, central nervous system control of the circulation remains a poorly understood yet vital component of the hypertension pathway and mandates further investigation. Technology to improve blood pressure control through deep brain stimulation of key cardiovascular control territories is, therefore, of interest. Furthermore, alternative nonsympathomodulatory intervention targeting the hemodynamics of the circulation may also be worth exploring for patients in whom sympathetic drive is less relevant to hypertension perpetuation. Herein, we review the aforementioned technologies with an emphasis on the preclinical data that underpin their rationale and the human evidence that supports their use.



Circ Res: 01 Apr 2021; 128:1080-1099
Mahfoud F, Schlaich MP, Lobo MD
Circ Res: 01 Apr 2021; 128:1080-1099 | PMID: 33793330
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Abstract

Pathophysiology of Hypertension: The Mosaic Theory and Beyond.

Harrison DG, Coffman TM, Wilcox CS
Dr Irvine Page proposed the Mosaic Theory of Hypertension in the 1940s advocating that hypertension is the result of many factors that interact to raise blood pressure and cause end-organ damage. Over the years, Dr Page modified his paradigm, and new concepts regarding oxidative stress, inflammation, genetics, sodium homeostasis, and the microbiome have arisen that allow further refinements of the Mosaic Theory. A constant feature of this approach to understanding hypertension is that the various nodes are interdependent and that these almost certainly vary between experimental models and between individuals with hypertension. This review discusses these new concepts and provides an introduction to other reviews in this compendium of Circulation Research.



Circ Res: 01 Apr 2021; 128:847-863
Harrison DG, Coffman TM, Wilcox CS
Circ Res: 01 Apr 2021; 128:847-863 | PMID: 33793328
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Abstract

Obesity, Adipose Tissue and Vascular Dysfunction.

Koenen M, Hill MA, Cohen P, Sowers JR
Cardiovascular diseases are the leading cause of death worldwide. Overweight and obesity are strongly associated with comorbidities such as hypertension and insulin resistance, which collectively contribute to the development of cardiovascular diseases and resultant morbidity and mortality. Forty-two percent of adults in the United States are obese, and a total of 1.9 billion adults worldwide are overweight or obese. These alarming numbers, which continue to climb, represent a major health and economic burden. Adipose tissue is a highly dynamic organ that can be classified based on the cellular composition of different depots and their distinct anatomical localization. Massive expansion and remodeling of adipose tissue during obesity differentially affects specific adipose tissue depots and significantly contributes to vascular dysfunction and cardiovascular diseases. Visceral adipose tissue accumulation results in increased immune cell infiltration and secretion of vasoconstrictor mediators, whereas expansion of subcutaneous adipose tissue is less harmful. Therefore, fat distribution more than overall body weight is a key determinant of the risk for cardiovascular diseases. Thermogenic brown and beige adipose tissue, in contrast to white adipose tissue, is associated with beneficial effects on the vasculature. The relationship between the type of adipose tissue and its influence on vascular function becomes particularly evident in the context of the heterogenous phenotype of perivascular adipose tissue that is strongly location dependent. In this review, we address the abnormal remodeling of specific adipose tissue depots during obesity and how this critically contributes to the development of hypertension, endothelial dysfunction, and vascular stiffness. We also discuss the local and systemic roles of adipose tissue derived secreted factors and increased systemic inflammation during obesity and highlight their detrimental impact on cardiovascular health.



Circ Res: 01 Apr 2021; 128:951-968
Koenen M, Hill MA, Cohen P, Sowers JR
Circ Res: 01 Apr 2021; 128:951-968 | PMID: 33793327
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Abstract

Guideline-Driven Management of Hypertension: An Evidence-Based Update.

Carey RM, Wright JT, Taler SJ, Whelton PK
Several important findings bearing on the prevention, detection, and management of hypertension have been reported since publication of the 2017 American College of Cardiology/American Heart Association Blood Pressure Guideline. This review summarizes and places in context the results of relevant observational studies, randomized clinical trials, and meta-analyses published between January 2018 and March 2021. Topics covered include blood pressure measurement, patient evaluation for secondary hypertension, cardiovascular disease risk assessment and blood pressure threshold for drug therapy, lifestyle and pharmacological management, treatment target blood pressure goal, management of hypertension in older adults, diabetes, chronic kidney disease, resistant hypertension, and optimization of care using patient, provider, and health system approaches. Presenting new information in each of these areas has the potential to increase hypertension awareness, treatment, and control which remain essential for the prevention of cardiovascular disease and mortality in the future.



Circ Res: 01 Apr 2021; 128:827-846
Carey RM, Wright JT, Taler SJ, Whelton PK
Circ Res: 01 Apr 2021; 128:827-846 | PMID: 33793326
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Abstract

Arterial Stiffness and Cardiovascular Risk in Hypertension.

Boutouyrie P, Chowienczyk P, Humphrey JD, Mitchell GF
Arterial stiffness, a leading marker of risk in hypertension, can be measured at material or structural levels, with the latter combining effects of the geometry and composition of the wall, including intramural organization. Numerous studies have shown that structural stiffness predicts outcomes in models that adjust for conventional risk factors. Elastic arteries, nearer to the heart, are most sensitive to effects of blood pressure and age, major determinants of stiffness. Stiffness is usually considered as an index of vascular aging, wherein individuals excessively affected by risk factor exposure represent early vascular aging, whereas those resistant to risk factors represent supernormal vascular aging. Stiffness affects the function of the brain and kidneys by increasing pulsatile loads within their microvascular beds, and the heart by increasing left ventricular systolic load; excessive pressure pulsatility also decreases diastolic pressure, necessary for coronary perfusion. Stiffness promotes inward remodeling of small arteries, which increases resistance, blood pressure, and in turn, central artery stiffness, thus creating an insidious feedback loop. Chronic antihypertensive treatments can reduce stiffness beyond passive reductions due to decreased blood pressure. Preventive drugs, such as lipid-lowering drugs and antidiabetic drugs, have additional effects on stiffness, independent of pressure. Newer anti-inflammatory drugs also have blood pressure independent effects. Reduction of stiffness is expected to confer benefit beyond the lowering of pressure, although this hypothesis is not yet proven. We summarize different steps for making arterial stiffness measurement a keystone in hypertension management and cardiovascular prevention as a whole.



Circ Res: 01 Apr 2021; 128:864-886
Boutouyrie P, Chowienczyk P, Humphrey JD, Mitchell GF
Circ Res: 01 Apr 2021; 128:864-886 | PMID: 33793325
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Abstract

Cardiac Tissues From Stem Cells: New Routes to Maturation and Cardiac Regeneration.

Campostrini G, Windt LM, van Meer BJ, Bellin M, Mummery CL
The ability of human pluripotent stem cells to form all cells of the body has provided many opportunities to study disease and produce cells that can be used for therapy in regenerative medicine. Even though beating cardiomyocytes were among the first cell types to be differentiated from human pluripotent stem cell, cardiac applications have advanced more slowly than those, for example, for the brain, eye, and pancreas. This is, in part, because simple 2-dimensional human pluripotent stem cell cardiomyocyte cultures appear to need crucial functional cues normally present in the 3-dimensional heart structure. Recent tissue engineering approaches combined with new insights into the dialogue between noncardiomyocytes and cardiomyocytes have addressed and provided solutions to issues such as cardiomyocyte immaturity and inability to recapitulate adult heart values for features like contraction force, electrophysiology, or metabolism. Three-dimensional bioengineered heart tissues are thus poised to contribute significantly to disease modeling, drug discovery, and safety pharmacology, as well as provide new modalities for heart repair. Here, we review the current status of 3-dimensional engineered heart tissues.



Circ Res: 18 Mar 2021; 128:775-801
Campostrini G, Windt LM, van Meer BJ, Bellin M, Mummery CL
Circ Res: 18 Mar 2021; 128:775-801 | PMID: 33734815
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Abstract

MIR503HG Loss Promotes Endothelial-to-Mesenchymal Transition in Vascular Disease.

Monteiro JP, Rodor J, Caudrillier A, Scanlon JP, ... Brandes RP, Baker AH
Rationale: Endothelial-to-mesenchymal transition (EndMT) is a dynamic biological process involved in pathological vascular remodelling. However, the molecular mechanisms that govern this transition remain largely unknown, including the contribution of long non-coding RNAs (lncRNAs). Objective: To investigate the role of lncRNAs in EndMT and their relevance to vascular remodelling.
Methods and results:
To study EndMT in vitro, primary endothelial cells (EC) were treated with transforming growth factor-β2 and interleukin-1β. Single-cell and bulk RNA-sequencing were performed to investigate the transcriptional architecture of EndMT and identify regulated lncRNAs. The functional contribution of seven lncRNAs during EndMT was investigated based on a DsiRNA screening assay. The loss of lncRNA MIR503HG was identified as a common signature across multiple human EC types undergoing EndMT in vitro. MIR503HG depletion induced a spontaneous EndMT phenotype, while its overexpression repressed hallmark EndMT changes, regulating 29% of its transcriptome signature. Importantly, the phenotypic changes induced by MIR503HG were independent of miR-424 and miR-503, which overlap the lncRNA locus. The pathological relevance of MIR503HG down-regulation was confirmed in vivo using Sugen/Hypoxia (SuHx)-induced pulmonary hypertension (PH) in mouse, as well as in human clinical samples, in lung sections and blood outgrowth endothelial cells (BOECs) from pulmonary arterial hypertension (PAH) patients. Overexpression of human MIR503HG in SuHx mice led to reduced mesenchymal marker expression, suggesting MIR503HG therapeutic potential. We also revealed that MIR503HG interacts with the Polypyrimidine Tract Binding Protein 1 (PTB1) and regulates its protein level. PTBP1 regulation of EndMT markers suggests that the role of MIR503HG in EndMT might be mediated in part by PTBP1. Conclusions: This study reports a novel lncRNA transcriptional profile associated with EndMT and reveals the crucial role of the loss of MIR503HG in EndMT and its relevance to pulmonary hypertension.




Circ Res: 10 Mar 2021; epub ahead of print
Monteiro JP, Rodor J, Caudrillier A, Scanlon JP, ... Brandes RP, Baker AH
Circ Res: 10 Mar 2021; epub ahead of print | PMID: 33703914
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Abstract

BH4 Increases nNOS Activity and Preserves Left Ventricular Function in Diabetes.

Carnicer R, Duglan D, Ziberna K, Recalde A, ... Channon KM, Casadei B
Rationale
In diabetic patients, heart failure with predominant left ventricular (LV) diastolic dysfunction is a common complication for which there is no effective treatment. Oxidation of the NOS (nitric oxide synthase) cofactor tetrahydrobiopterin (BH4) and dysfunctional NOS activity have been implicated in the pathogenesis of the diabetic vascular and cardiomyopathic phenotype.
Objective
Using mice models and human myocardial samples, we evaluated whether and by which mechanism increasing myocardial BH4 availability prevented or reversed LV dysfunction induced by diabetes.
Methods and results
In contrast to the vascular endothelium, BH4 levels, superoxide production, and NOS activity (by liquid chromatography) did not differ in the LV myocardium of diabetic mice or in atrial tissue from diabetic patients. Nevertheless, the impairment in both cardiomyocyte relaxation and [Ca2+]i (intracellular calcium) decay and in vivo LV function (echocardiography and tissue Doppler) that developed in wild-type mice 12 weeks post-diabetes induction (streptozotocin, 42-45 mg/kg) was prevented in mGCH1-Tg (mice with elevated myocardial BH4 content secondary to trangenic overexpression of GTP-cyclohydrolase 1) and reversed in wild-type mice receiving oral BH4 supplementation from the 12th to the 18th week after diabetes induction. The protective effect of BH4 was abolished by CRISPR/Cas9-mediated knockout of nNOS (the neuronal NOS isoform) in mGCH1-Tg. In HEK (human embryonic kidney) cells, S-nitrosoglutathione led to a PKG (protein kinase G)-dependent increase in plasmalemmal density of the insulin-independent glucose transporter GLUT-1 (glucose transporter-1). In cardiomyocytes, mGCH1 overexpression induced a NO/sGC (soluble guanylate cyclase)/PKG-dependent increase in glucose uptake via GLUT-1, which was instrumental in preserving mitochondrial creatine kinase activity, oxygen consumption rate, LV energetics (by 31phosphorous magnetic resonance spectroscopy), and myocardial function.
Conclusions
We uncovered a novel mechanism whereby myocardial BH4 prevents and reverses LV diastolic and systolic dysfunction associated with diabetes via an nNOS-mediated increase in insulin-independent myocardial glucose uptake and utilization. These findings highlight the potential of GCH1/BH4-based therapeutics in human diabetic cardiomyopathy. Graphic Abstract: A graphic abstract is available for this article.



Circ Res: 04 Mar 2021; 128:585-601
Carnicer R, Duglan D, Ziberna K, Recalde A, ... Channon KM, Casadei B
Circ Res: 04 Mar 2021; 128:585-601 | PMID: 33494625
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Abstract

Generation of Vascular Smooth Muscle Cells From Induced Pluripotent Stem Cells: Methods, Applications, and Considerations.

Shen M, Quertermous T, Fischbein MP, Wu JC
The developmental origin of vascular smooth muscle cells (VSMCs) has been increasingly recognized as a major determinant for regional susceptibility or resistance to vascular diseases. As a human material-based complement to animal models and human primary cultures, patient induced pluripotent stem cell iPSC-derived VSMCs have been leveraged to conduct basic research and develop therapeutic applications in vascular diseases. However, iPSC-VSMCs (induced pluripotent stem cell VSMCs) derived by most existing induction protocols are heterogeneous in developmental origins. In this review, we summarize signaling networks that govern in vivo cell fate decisions and in vitro derivation of distinct VSMC progenitors, as well as key regulators that terminally specify lineage-specific VSMCs. We then highlight the significance of leveraging patient-derived iPSC-VSMCs for vascular disease modeling, drug discovery, and vascular tissue engineering and discuss several obstacles that need to be circumvented to fully unleash the potential of induced pluripotent stem cells for precision vascular medicine.



Circ Res: 04 Mar 2021; 128:670-686
Shen M, Quertermous T, Fischbein MP, Wu JC
Circ Res: 04 Mar 2021; 128:670-686 | PMID: 33818124
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Abstract

Kynurenine Relaxes Arteries of Normotensive Women and Those with Preeclampsia.

Worton SA, Pritchard HA, Greenwood SL, Alakrawi M, ... Greenstein AS, Myers JE
Rationale: Activation of the kynurenine pathway of tryptophan catabolism by infection and inflammation contributes to the development of systemic hypotension. Commercially-available kynurenine has direct vasorelaxant effects on arteries from several species and reduces systemic blood pressure when administered to normotensive or hypertensive rats. Objective: To determine whether kynurenine promotes relaxation of human resistance arteries from normotensive and hypertensive pregnant women and to identify the vascular mechanism of its effects.
Methods and results:
In isolated omental and myometrial resistance arteries from normotensive pregnant women, kynurenine (1 mmol/L) significantly reduced U46619-induced constriction (omentum N=14, p=2.4x10-3; myometrium N=21-25, p=2.6x10-4) and relaxed pre-constricted arteries (N=53, p=1.0x10-11; N=20, p=8.8x10-3). Vasorelaxation persisted following endothelium removal (N=7, p=1.6x10-4), but was completely prevented by inhibition of large-conductance Ca2+-activated (BKCa) channels with iberiotoxin (N=9, p=5.7x10-4) or paxilline (N=10, p=2.1x10-17). Accordingly, in isolated vascular smooth muscle cells from omental arteries, kynurenine increased the BKCa current (n=5-8, p=0.022) and the amplitude of spontaneous transient outward currents (STOCs; n=6, p=0.031), but did not affect STOC frequency. Kynurenine also increased Ca2+ spark frequency of pressurised omental arteries (n=8, p=0.031). Vasorelaxant effects of kynurenine persisted following inhibition of ryanodine receptors (N=7, p=0.48), but were moderately reduced by inhibition of adenylate cyclase (N=9, p=0.024). In arteries from women with preeclampsia, kynurenine similarly attenuated vasoconstriction (N=15, p=1.3x10-5) and induced BKCa-mediated vasodilation (N=16, p=2.0x10-4). Vasorelaxation in response to kynurenine and a specific BKCa activator, NS11021, was absent in fetal-derived placental resistance arteries in normal pregnancy and preeclampsia. Conclusions: Kynurenine dilates systemic arteries from multiple territories via BKCa activation. Notably, the vasorelaxatory capacity of kynurenine is preserved in preeclampsia, suggesting this approach may have translational potential for the treatment of hypertension in pregnancy. The data warrants further investigation of the potential to exploit this endogenous vasorelaxant as a new treatment for hypertensive pathologies.




Circ Res: 02 Mar 2021; epub ahead of print
Worton SA, Pritchard HA, Greenwood SL, Alakrawi M, ... Greenstein AS, Myers JE
Circ Res: 02 Mar 2021; epub ahead of print | PMID: 33656370
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Abstract

Innervation and Neuronal Control of the Mammalian Sinoatrial Node: A Comprehensive Atlas.

Hanna P, Dacey MJ, Brennan J, Moss A, ... Ardell JL, Shivkumar K
Rationale: Cardiac function is under exquisite intrinsic cardiac neural control. Neuroablative techniques to modulate control of cardiac function are currently being studied in patients, albeit with variable and sometimes deleterious results. Objective: Recognizing the major gaps in our understanding of cardiac neural control, we sought to evaluate neural regulation of impulse initiation in the sinoatrial node as an initial discovery step.
Methods and results:
We report an in-depth, multi-scale structural and functional characterization of the innervation of the sinoatrial node (SAN) by the right atrial ganglionated plexus (RAGP) in porcine and human hearts. Combining intersectional strategies including tissue clearing, immunohistochemical and ultrastructural techniques, we have delineated a comprehensive neuroanatomic atlas of the RAGP-SAN complex. The RAGP shows significant phenotypic diversity of neurons while maintaining predominant cholinergic innervation. Cellular and tissue-level electrophysiologic mapping and ablation studies demonstrate interconnected ganglia with synaptic convergence within the RAGP to modulate SAN automaticity, atrioventricular (AV) conduction and left ventricular (LV) contractility. Using this approach, we comprehensively demonstrate that intrinsic cardiac neurons influence the pacemaking site in the heart. Conclusions: This report provides an experimental demonstration of a discrete neuronal population controlling a specific geographic region of the heart (SAN) that can serve as a framework for further exploration of other parts of the intrinsic cardiac nervous system (ICNS) in mammalian hearts and for developing targeted therapies.




Circ Res: 24 Feb 2021; epub ahead of print
Hanna P, Dacey MJ, Brennan J, Moss A, ... Ardell JL, Shivkumar K
Circ Res: 24 Feb 2021; epub ahead of print | PMID: 33629877
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Abstract

CARMN Loss Regulates Smooth Muscle Cells and Accelerates Atherosclerosis in Mice.

Vacante F, Rodor J, Lalwani MK, Mahmoud AD, ... Sluimer JC, Baker AH
Rationale: In the microenvironment of atherosclerotic lesions, vascular smooth muscle cells (vSMCs) switch to a dedifferentiated state but the underlying molecular mechanisms driving this switch are not fully understood. Long noncoding RNAs (lncRNAs) are dysregulated during vascular pathology, but relatively little is known about their involvement in controlling vSMCs function. CARMN is a lncRNA located immediately upstream of the microRNAs (miRNAs) miR-143 and miR-145, both involved in vSMCs function. Objective: We investigated the role of the lncRNA CARMN, independent from miR-143 and miR-145, as potential a regulator of vSMC phenotypes in vitro and the consequences of its loss during the development of atherosclerosis in vivo. We hypothesized that loss of CARMN is a primary event controlling the functional switch towards pro-atherogenic vSMC phenotype and accelerates the development of the plaques in vivo.
Methods and results:
Expression of CARMN lncRNA was silenced using GapmeRs in human coronary arterial smooth muscle cells (hCASMCs), revealing that GapmeR-mediated loss of CARMN negatively affects miR-143 and miR-145 miRNA expression. RNA sequencing of CARMN-depleted hCASMCs revealed large transcriptomic changes, associated with vSMC proliferation, migration, inflammation, lipid metabolism and dedifferentiation. The use of miR-143 and miR-145 mimics revealed that CARMN regulates hCASMC proliferation in a miRNA-independent manner. In human and mouse, CARMN and associated miRNAs were downregulated in advanced versus early atherosclerotic lesions. Using a CRISPR-Cas9 knock-out approach, we explored the implications of CARMN depletion during atherosclerosis in vivo. Consistent with in vitro results, the knock-out of CARMN impaired the expression of miR-143 and miR-145 under homeostatic conditions. Importantly, when atherosclerosis was induced in these mice, CARMN knock-out increased the volume, size, pro-inflammatory LGALS3-expressing cells content and altered plaque composition, yielding an advanced phenotype. Conclusions: We identified the early loss of CARMN lncRNA as critical event which primes vSMCs towards a pro-atherogenic phenotype in vitro and accelerates the development of atherosclerosis in vivo.




Circ Res: 23 Feb 2021; epub ahead of print
Vacante F, Rodor J, Lalwani MK, Mahmoud AD, ... Sluimer JC, Baker AH
Circ Res: 23 Feb 2021; epub ahead of print | PMID: 33622045
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Abstract

JNK2, a Newly-Identified SERCA2 Enhancer, Augments an Arrhythmic [Ca] Leak-Load Relationship.

Yan J, Bare DJ, DeSantiago J, Zhao W, ... Chen SRW, Ai X
Rationale
We recently discovered pivotal contributions of stress kinase JNK2 (c-Jun N-terminal kinase isoform 2) in increased risk of atrial fibrillation through enhanced diastolic sarcoplasmic reticulum (SR) calcium (Ca2+) leak via RyR2 (ryanodine receptor isoform 2). However, the role of JNK2 in the function of the SERCA2 (SR Ca2+-ATPase), essential in maintaining SR Ca2+ content cycling during each heartbeat, is completely unknown.
Objective
To test the hypothesis that JNK2 increases SERCA2 activity SR Ca2+ content and exacerbates an arrhythmic SR Ca2+ content leak-load relationship.
Methods and results
We used confocal Ca2+ imaging in myocytes and HEK-RyR2 (ryanodine receptor isoform 2-expressing human embryonic kidney 293 cells) cells, biochemistry, dual Ca2+/voltage optical mapping in intact hearts from alcohol-exposed or aged mice (where JNK2 is activated). We found that JNK2, but not JNK1 (c-Jun N-terminal kinase isoform 1), increased SERCA2 uptake and consequently elevated SR Ca2+ content load. JNK2 also associates with and phosphorylates SERCA2 proteins. JNK2 causally enhances SERCA2-ATPase activity via increased maximal rate, without altering Ca2+ affinity. Unlike the CaMKII (Ca2+/calmodulin-dependent kinase II)-dependent JNK2 action in SR Ca2+ leak, JNK2-driven SERCA2 function was CaMKII independent (not prevented by CaMKII inhibition). With CaMKII blocked, the JNK2-driven SR Ca2+ loading alone did not significantly raise leak. However, with JNK2-CaMKII-driven SR Ca2+ leak present, the JNK2-enhanced SR Ca2+ uptake limited leak-induced reduction in SR Ca2+, normalizing Ca2+ transient amplitude, but at a higher arrhythmogenic SR Ca2+ leak. JNK2-specific inhibition completely normalized SR Ca2+ handling, attenuated arrhythmic Ca2+ activities, and alleviated atrial fibrillation susceptibility in aged and alcohol-exposed myocytes and intact hearts.
Conclusions
We have identified a novel JNK2-induced activation of SERCA2. The dual action of JNK2 in CaMKII-dependent arrhythmic SR Ca2+ leak and a CaMKII-independent uptake exacerbates atrial arrhythmogenicity, while helping to maintain normal levels of Ca2+ transients and heart function. JNK2 modulation may be a novel therapeutic target for atrial fibrillation prevention and treatment.



Circ Res: 18 Feb 2021; 128:455-470
Yan J, Bare DJ, DeSantiago J, Zhao W, ... Chen SRW, Ai X
Circ Res: 18 Feb 2021; 128:455-470 | PMID: 33334123
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Abstract

Klotho Deficiency Causes Heart Aging via Impairing the Nrf2-GR Pathway.

Chen K, Wang S, Sun QW, Zhang B, Ullah M, Sun Z
Rationale
Cardiac aging is an important contributing factor for heart failure, which affects a large population but remains poorly understood.
Objective
The purpose of this study is to investigate whether Klotho plays a role in cardiac aging.
Methods and results
Heart function declined in old mice (24 months), as evidenced by decreases in fractional shortening, ejection fraction, and cardiac output. Heart size and weight, cardiomyocyte size, and cardiac fibrosis were increased in old mice, indicating that aging causes cardiac hypertrophy and remodeling. Circulating Klotho levels were dramatically decreased in old mice, which prompted us to investigate whether the Klotho decline may cause heart aging. We found that Klotho gene mutation (KL-/-) largely decreased serum klotho levels and impaired heart function. Interestingly, supplement of exogenous secreted Klotho prevented heart failure, hypertrophy, and remodeling in both old mice and KL (-/-) mice. Secreted Klotho treatment inhibited excessive cardiac oxidative stress, senescence and apoptosis in old mice and KL (-/-) mice. Serum phosphate levels in KL (-/-) mice were kept in the normal range, suggesting that Klotho deficiency-induced heart aging is independent of phosphate metabolism. Mechanistically, Klotho deficiency suppressed GR (glutathione reductase) expression and activity in the heart via inhibition of transcription factor Nrf2 (nuclear factor-erythroid 2 p45-related factor 2). Furthermore, cardiac-specific overexpression of GR prevented excessive oxidative stress, apoptosis, and heart failure in both old and KL (-/-) mice.
Conclusions
Klotho deficiency causes cardiac aging via impairing the Nrf2-GR pathway. Supplement of exogenous secreted Klotho represents a promising therapeutic strategy for aging-associated cardiomyopathy and heart failure.



Circ Res: 18 Feb 2021; 128:492-507
Chen K, Wang S, Sun QW, Zhang B, Ullah M, Sun Z
Circ Res: 18 Feb 2021; 128:492-507 | PMID: 33334122
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Abstract

Different DOACs Control Inflammation in Cardiac Ischemia-Reperfusion Differently.

Gadi I, Fatima S, Elwakiel A, Nazir S, ... Isermann B, Shahzad K
Rationale
While thrombin is the key protease in thrombus formation, other coagulation proteases, such as fXa (factor Xa) or aPC (activated protein C), independently modulate intracellular signaling via partially distinct receptors.
Objectives
To study the differential effects of fXa or fIIa (factor IIa) inhibition on gene expression and inflammation in myocardial ischemia-reperfusion injury.
Methods and results
Mice were treated with a direct fIIa inhibitor (fIIai) or direct fXa inhibitor (fXai) at doses that induced comparable anticoagulant effects ex vivo and in vivo (tail-bleeding assay and FeCl3-induced thrombosis). Myocardial ischemia-reperfusion injury was induced via left anterior descending ligation. We determined infarct size and in vivo aPC generation, analyzed gene expression by RNA sequencing, and performed immunoblotting and ELISA. The signaling-only 3K3A-aPC variant and inhibitory antibodies that blocked all or only the anticoagulant function of aPC were used to determine the role of aPC. Doses of fIIai and fXai that induced comparable anticoagulant effects resulted in a comparable reduction in infarct size. However, unbiased gene expression analyses revealed marked differences, including pathways related to sterile inflammation and inflammasome regulation. fXai but not fIIai inhibited sterile inflammation by reducing the expression of proinflammatory cytokines (IL [interleukin]-1β, IL-6, and TNFα [tumor necrosis factor alpha]), as well as NF-κB (nuclear factor kappa B) and inflammasome activation. This anti-inflammatory effect was associated with reduced myocardial fibrosis 28 days post-myocardial ischemia-reperfusion injury. Mechanistically, in vivo aPC generation was higher with fXai than with fIIai. Inhibition of the anticoagulant and signaling properties of aPC abolished the anti-inflammatory effect associated with fXai, while inhibiting only the anticoagulant function of aPC had no effect. Combining 3K3A-aPC with fIIai reduced the inflammatory response, mimicking the fXai-associated effect.
Conclusions
We showed that specific inhibition of coagulation via direct oral anticoagulants had differential effects on gene expression and inflammation, despite comparable anticoagulant effects and infarct sizes. Targeting individual coagulation proteases induces specific cellular responses unrelated to their anticoagulant effect.



Circ Res: 18 Feb 2021; 128:513-529
Gadi I, Fatima S, Elwakiel A, Nazir S, ... Isermann B, Shahzad K
Circ Res: 18 Feb 2021; 128:513-529 | PMID: 33353373
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Abstract

SIRT6 Protects Smooth Muscle Cells From Senescence and Reduces Atherosclerosis.

Grootaert MOJ, Finigan A, Figg NL, Uryga AK, Bennett MR
Rationale
Vascular smooth muscle cell (VSMC) senescence promotes atherosclerosis and features of plaque instability, in part, through lipid-mediated oxidative DNA damage and telomere dysfunction. SIRT6 (Sirtuin 6) is a nuclear deacetylase involved in DNA damage response signaling, inflammation, and metabolism; however, its role in regulating VSMC senescence and atherosclerosis is unclear.
Objective
We examined SIRT6 expression in human VSMCs, the role, regulation, and downstream pathways activated by SIRT6, and how VSMC SIRT6 regulates atherogenesis.
Methods and results
SIRT6 protein, but not mRNA, expression was markedly reduced in VSMCs in human and mouse atherosclerotic plaques, and in human VSMCs derived from plaques or undergoing replicative or palmitate-induced senescence versus healthy aortic VSMCs. The ubiquitin ligase CHIP (C terminus of HSC70-interacting protein) promoted SIRT6 stability, but CHIP expression was reduced in human and mouse plaque VSMCs and by palmitate in a p38- and c-Jun N-terminal kinase-dependent manner. SIRT6 bound to telomeres, while SIRT6 inhibition using shRNA or a deacetylase-inactive mutant (SIRT6H133Y) shortened human VSMC lifespan and induced senescence, associated with telomeric H3K9 (histone H3 lysine 9) hyperacetylation and 53BP1 (p53 binding protein 1) binding, indicative of telomere damage. In contrast, SIRT6 overexpression preserved telomere integrity, delayed cellular senescence, and reduced inflammatory cytokine expression and changes in VSMC metabolism associated with senescence. SIRT6, but not SIRT6H133Y, promoted proliferation and lifespan of mouse VSMCs, and prevented senescence-associated metabolic changes. ApoE-/- (apolipoprotein E) mice were generated that overexpress SIRT6 or SIRT6H133Y in VSMCs only. SM22α-hSIRT6/ApoE-/- mice had reduced atherosclerosis, markers of senescence and inflammation compared with littermate controls, while plaques of SM22α-hSIRT6H133Y/ApoE-/- mice showed increased features of plaque instability.
Conclusions
SIRT6 protein expression is reduced in human and mouse plaque VSMCs and is positively regulated by CHIP. SIRT6 regulates telomere maintenance and VSMC lifespan and inhibits atherogenesis, all dependent on its deacetylase activity. Our data show that endogenous SIRT6 deacetylase is an important and unrecognized inhibitor of VSMC senescence and atherosclerosis.



Circ Res: 18 Feb 2021; 128:474-491
Grootaert MOJ, Finigan A, Figg NL, Uryga AK, Bennett MR
Circ Res: 18 Feb 2021; 128:474-491 | PMID: 33353368
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Abstract

Single-Cell Analysis of Blood-Brain Barrier Response to Pericyte Loss.

Mäe MA, He L, Nordling S, Vazquez-Liebanas E, ... Keller A, Betsholtz C
Rationale
Pericytes are capillary mural cells playing a role in stabilizing newly formed blood vessels during development and tissue repair. Loss of pericytes has been described in several brain disorders, and genetically induced pericyte deficiency in the brain leads to increased macromolecular leakage across the blood-brain barrier (BBB). However, the molecular details of the endothelial response to pericyte deficiency remain elusive.
Objective
To map the transcriptional changes in brain endothelial cells resulting from lack of pericyte contact at single-cell level and to correlate them with regional heterogeneities in BBB function and vascular phenotype.
Methods and results
We reveal transcriptional, morphological, and functional consequences of pericyte absence for brain endothelial cells using a combination of methodologies, including single-cell RNA sequencing, tracer analyses, and immunofluorescent detection of protein expression in pericyte-deficient adult Pdgfbret/ret mice. We find that endothelial cells without pericyte contact retain a general BBB-specific gene expression profile, however, they acquire a venous-shifted molecular pattern and become transformed regarding the expression of numerous growth factors and regulatory proteins. Adult Pdgfbret/ret brains display ongoing angiogenic sprouting without concomitant cell proliferation providing unique insights into the endothelial tip cell transcriptome. We also reveal heterogeneous modes of pericyte-deficient BBB impairment, where hotspot leakage sites display arteriolar-shifted identity and pinpoint putative BBB regulators. By testing the causal involvement of some of these using reverse genetics, we uncover a reinforcing role for angiopoietin 2 at the BBB.
Conclusions
By elucidating the complexity of endothelial response to pericyte deficiency at cellular resolution, our study provides insight into the importance of brain pericytes for endothelial arterio-venous zonation, angiogenic quiescence, and a limited set of BBB functions. The BBB-reinforcing role of ANGPT2 (angiopoietin 2) is paradoxical given its wider role as TIE2 (TEK receptor tyrosine kinase) receptor antagonist and may suggest a unique and context-dependent function of ANGPT2 in the brain.



Circ Res: 18 Feb 2021; 128:e46-e62
Mäe MA, He L, Nordling S, Vazquez-Liebanas E, ... Keller A, Betsholtz C
Circ Res: 18 Feb 2021; 128:e46-e62 | PMID: 33375813
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Abstract

Ca-CaM Dependent Inactivation of RyR2 Underlies Ca Alternans in Intact Heart.

Wei J, Yao J, Belke D, Guo W, ... Echebarria B, Chen SRW
Rationale
Ca2+ alternans plays an essential role in cardiac alternans that can lead to ventricular fibrillation, but the mechanism underlying Ca2+ alternans remains undefined. Increasing evidence suggests that Ca2+ alternans results from alternations in the inactivation of cardiac RyR2 (ryanodine receptor 2). However, what inactivates RyR2 and how RyR2 inactivation leads to Ca2+ alternans are unknown.
Objective
To determine the role of CaM (calmodulin) on Ca2+ alternans in intact working mouse hearts.
Methods and results
We used an in vivo local gene delivery approach to alter CaM function by directly injecting adenoviruses expressing CaM-wild type, a loss-of-function CaM mutation, CaM (1-4), and a gain-of-function mutation, CaM-M37Q, into the anterior wall of the left ventricle of RyR2 wild type or mutant mouse hearts. We monitored Ca2+ transients in ventricular myocytes near the adenovirus-injection sites in Langendorff-perfused intact working hearts using confocal Ca2+ imaging. We found that CaM-wild type and CaM-M37Q promoted Ca2+ alternans and prolonged Ca2+ transient recovery in intact RyR2 wild type and mutant hearts, whereas CaM (1-4) exerted opposite effects. Altered CaM function also affected the recovery from inactivation of the L-type Ca2+ current but had no significant impact on sarcoplasmic reticulum Ca2+ content. Furthermore, we developed a novel numerical myocyte model of Ca2+ alternans that incorporates Ca2+-CaM-dependent regulation of RyR2 and the L-type Ca2+ channel. Remarkably, the new model recapitulates the impact on Ca2+ alternans of altered CaM and RyR2 functions under 9 different experimental conditions. Our simulations reveal that diastolic cytosolic Ca2+ elevation as a result of rapid pacing triggers Ca2+-CaM dependent inactivation of RyR2. The resultant RyR2 inactivation diminishes sarcoplasmic reticulum Ca2+ release, which, in turn, reduces diastolic cytosolic Ca2+, leading to alternations in diastolic cytosolic Ca2+, RyR2 inactivation, and sarcoplasmic reticulum Ca2+ release (ie, Ca2+ alternans).
Conclusions
Our results demonstrate that inactivation of RyR2 by Ca2+-CaM is a major determinant of Ca2+ alternans, making Ca2+-CaM dependent regulation of RyR2 an important therapeutic target for cardiac alternans.



Circ Res: 18 Feb 2021; 128:e63-e83
Wei J, Yao J, Belke D, Guo W, ... Echebarria B, Chen SRW
Circ Res: 18 Feb 2021; 128:e63-e83 | PMID: 33375811
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Abstract

Radiation Impacts Early Atherosclerosis by Suppressing Intimal LDL Accumulation.

Ikeda J, Scipione CA, Hyduk SJ, Althagafi MG, ... MacParland SA, Cybulsky MI
Rationale
Bone marrow transplantation (BMT) is used frequently to study the role of hematopoietic cells in atherosclerosis, but aortic arch lesions are smaller in mice after BMT.
Objective
To identify the earliest stage of atherosclerosis inhibited by BMT and elucidate potential mechanisms.
Methods and results
Ldlr-/- mice underwent total body γ-irradiation, bone marrow reconstitution, and 6-week recovery. Atherosclerosis was studied in the ascending aortic arch and compared with mice without BMT. In BMT mice, neutral lipid and myeloid cell topography were lower in lesions after feeding a cholesterol-rich diet for 3, 6, and 12 weeks. Lesion coalescence and height were suppressed dramatically in mice post-BMT, whereas lateral growth was inhibited minimally. Targeted radiation to the upper thorax alone reproduced the BMT phenotype. Classical monocyte recruitment, intimal myeloid cell proliferation, and apoptosis did not account for the post-BMT phenotype. Neutral lipid accumulation was reduced in 5-day lesions, thus we developed quantitative assays for LDL (low-density lipoprotein) accumulation and paracellular leakage using DiI-labeled human LDL and rhodamine B-labeled 70 kD dextran. LDL accumulation was dramatically higher in the intima of Ldlr-/- relative to Ldlr+/+ mice, and was inhibited by injection of HDL mimics, suggesting a regulated process. LDL, but not dextran, accumulation was lower in mice post-BMT both at baseline and in 5-day lesions. Since the transcript abundance of molecules implicated in LDL transcytosis was not significantly different in the post-BMT intima, transcriptomics from whole aortic arch intima, and at single-cell resolution, was performed to give insights into pathways modulated by BMT.
Conclusions
Radiation exposure inhibits LDL entry into the aortic intima at baseline and the earliest stages of atherosclerosis. Single-cell transcriptomic analysis suggests that LDL uptake by endothelial cells is diverted to lysosomal degradation and reverse cholesterol transport pathways. This reduces intimal accumulation of lipid and impacts lesion initiation and growth.



Circ Res: 18 Feb 2021; 128:530-543
Ikeda J, Scipione CA, Hyduk SJ, Althagafi MG, ... MacParland SA, Cybulsky MI
Circ Res: 18 Feb 2021; 128:530-543 | PMID: 33397122
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Abstract

KPT-330 Prevents Aortic Valve Calcification via a Novel C/EBPβ Signaling Pathway.

Dutta P, Kodigepalli MK, LaHaye S, Thompson JW, ... Hinton RB, Lincoln J
Rationale: Calcific Aortic Valve Disease (CAVD) affects more than 5.2 million people in the US. The only effective treatment is surgery and this comes with complications and no guarantee of long-term success. Objective: Outcomes from pharmacological initiatives remain unsubstantiated and therefore the aim of this study is to determine if repurposing a selective XPO1 inhibitor drug (KPT-330) is beneficial in the treatment of CAVD.
Methods and results:
We show that KPT-330 prevents, attenuates and mitigates calcific nodule formation in heart valve interstitial cells (VICs) in vitro, and prevents CAVD in Klotho-/- mice. Using RNA-sequencing and Mass Spectrometry we show that KPT-330\'s beneficial effect is mediated by inhibiting nuclear export of the transcription factor C/EBPβ in VICs, leading to repression of canonical Wnt signaling, in part through activation of the Wnt antagonist Axin1, and a subsequent decrease in pro-osteogenic markers and cell viability. Conclusions: Our findings have met a critical need to discover alterative, pharmacological-based therapies in the treatment of CAVD.




Circ Res: 18 Feb 2021; epub ahead of print
Dutta P, Kodigepalli MK, LaHaye S, Thompson JW, ... Hinton RB, Lincoln J
Circ Res: 18 Feb 2021; epub ahead of print | PMID: 33601919
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Abstract

Machine Learning in Arrhythmia and Electrophysiology.

Trayanova NA, Popescu DM, Shade JK
Machine learning (ML), a branch of artificial intelligence, where machines learn from big data, is at the crest of a technological wave of change sweeping society. Cardiovascular medicine is at the forefront of many ML applications, and there is a significant effort to bring them into mainstream clinical practice. In the field of cardiac electrophysiology, ML applications have also seen a rapid growth and popularity, particularly the use of ML in the automatic interpretation of ECGs, which has been extensively covered in the literature. Much lesser known are the other aspects of ML application in cardiac electrophysiology and arrhythmias, such as those in basic science research on arrhythmia mechanisms, both experimental and computational; in the development of better techniques for mapping of cardiac electrical function; and in translational research related to arrhythmia management. In the current review, we examine comprehensively such ML applications as they match the scope of this journal. The current review is organized in 3 parts. The first provides an overview of general ML principles and methodologies that will afford readers of the necessary information on the subject, serving as the foundation for inviting further ML applications in arrhythmia research. The basic information we provide can serve as a guide on how one might design and conduct an ML study. The second part is a review of arrhythmia and electrophysiology studies in which ML has been utilized, highlighting the broad potential of ML approaches. For each subject, we outline comprehensively the general topics, while reviewing some of the research advances utilizing ML under the subject. Finally, we discuss the main challenges and the perspectives for ML-driven cardiac electrophysiology and arrhythmia research.



Circ Res: 18 Feb 2021; 128:544-566
Trayanova NA, Popescu DM, Shade JK
Circ Res: 18 Feb 2021; 128:544-566 | PMID: 33600229
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Abstract

Restructuring the Gut Microbiota by Intermittent Fasting Lowers Blood Pressure.

Shi H, Zhang B, Abo-Hamzy T, Nelson JW, ... Bryan RM, Durgan DJ
Raionale:: In recent years, it has been demonstrated that a pathological change in the gut microbiota, termed gut dysbiosis, can be an underlying factor for the development of hypertension. Prevention of this dysbiosis can attenuate or abolish hypertension. Translational mechanisms to prevent gut dysbiosis as well as understanding of the mechanisms linking gut dysbiosis to hypertension are lacking. Objective: We first examined the efficacy of intermittent fasting (IF) in altering the gut microbiota and lowering blood pressure (BP). Next, we utilized a multi-omics approach to examine microbial influenced metabolites that may serve as the link between the gut microbiota and host BP regulation.
Methods and results:
We demonstrate that IF significantly altered the makeup of the gut microbiota, cecal and plasma metabolome, and prevented the development of hypertension in the spontaneously hypertensive stroke-prone rat (SHRSP). The beneficial effects of IF were shown to be due to alterations of the gut microbiota through germ-free (GF) transplantation studies. GF rats receiving microbiota from IF SHRSP had significantly lower BP as compared to GF rats receiving microbiota from ad libitum fed SHRSPs. Through whole genome shotgun sequence analysis of the microbiota and untargeted metabolomics of cecal content and plasma we identified bile acid (BA) metabolism as a potential mediator in BP regulation. Finally, we show supplementation with cholic acid, or activation of the G protein-coupled BA receptor (TGR5), significantly reduced BP of the SHRSP. Conclusions: These studies demonstrate the BP lowering effects of IF involves manipulation of the gut microbiota and metabolome, and implicates disrupted BA signaling as novel mechanisms by which gut dysbiosis contributes to hypertension.




Circ Res: 17 Feb 2021; epub ahead of print
Shi H, Zhang B, Abo-Hamzy T, Nelson JW, ... Bryan RM, Durgan DJ
Circ Res: 17 Feb 2021; epub ahead of print | PMID: 33596669
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Abstract

Loss of CASK Accelerates Heart Failure Development.

Mustroph J, Sag CM, Bähr F, Schmidtmann AL, ... Maier LS, Wagner S
Rationale: Increased myocardial activity of Ca/calmodulin-dependent kinase II (CaMKII) leads to heart failure (HF) and arrhythmias. In Drosophila neurons, interaction of CaMKII with Ca/CaM-dependent serine protein kinase (CASK) has been shown to inhibit CaMKII activity, but the consequences of this regulation for HF and ventricular arrhythmias are unknown. Objective: We hypothesize that CASK associates with CaMKII in human and mouse hearts thereby limiting CaMKII activity, and that altering CASK expression in mice changes CaMKII activity accordingly, with functional consequences for contractile function and arrhythmias.
Methods and results:
Immunoprecipitation revealed that CASK associates with CaMKII in human hearts. CASK expression is unaltered in HF but increased in patients with aortic stenosis. In mice, cardiomyocyte-specific knockout of CASK (CASK-KO) increased CaMKII auto-phosphorylation at the stimulatory T287 site, but reduced phosphorylation at the inhibitory T305/306 site. CASK-KO mice showed increased CaMKII-dependent sarcoplasmic reticulum (SR) Ca leak, reduced SR Ca-content, increased susceptibility to ventricular arrhythmias, greater loss of ejection fraction, and increased mortality after transverse aortic constriction. Intriguingly, stimulation of the cardiac glucagon-like peptide 1-receptor with exenatide increased CASK expression resulting in increased inhibitory CaMKII T305 phosphorylation, reduced CaMKII activity, and reduced SR Ca leak in WT but not CASK KO. Conclusions: CASK associates with CaMKII in the human heart. CASK-KO in mice increases CaMKII activity, leading to contractile dysfunction and arrhythmias. Increasing CASK expression reduces CaMKII activity, improves Ca handling and contractile function.




Circ Res: 16 Feb 2021; epub ahead of print
Mustroph J, Sag CM, Bähr F, Schmidtmann AL, ... Maier LS, Wagner S
Circ Res: 16 Feb 2021; epub ahead of print | PMID: 33593074
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Abstract

miR-33 Silencing Reprograms the Immune Cell Landscape in Atherosclerotic Plaques.

Afonso MS, Sharma M, Schlegel MP, van Solingen C, ... Fisher EA, Moore KJ
Rationale: MicroRNA-33 post-transcriptionally represses genes involved in lipid metabolism and energy homeostasis. Targeted inhibition of miR-33 increases plasma HDL cholesterol and promotes atherosclerosis regression, in part, by enhancing reverse cholesterol transport and dampening plaque inflammation. However, how miR-33 reshapes the immune microenvironment of plaques remains poorly understood. Objective: To define how miR-33 inhibition alters the dynamic balance and transcriptional landscape of immune cells in atherosclerotic plaques.
Methods and results:
We used single cell RNA-sequencing of aortic CD45+ cells, combined with immunohistologic, morphometric and flow cytometric analyses to define the changes in plaque immune cell composition, gene expression and function following miR-33 inhibition. We report that anti-miR-33 treatment of Ldlr-/- mice with advanced atherosclerosis reduced plaque burden and altered the plaque immune cell landscape by shifting the balance of pro- and anti-atherosclerotic macrophage and T cell subsets. By quantifying the kinetic processes that determine plaque macrophage burden, we found that anti-miR-33 reduced levels of circulating monocytes and splenic myeloid progenitors, decreased macrophage proliferation and retention, and promoted macrophage attrition by apoptosis and efferocytotic clearance. scRNA-sequencing of aortic arch plaques showed that anti-miR-33 reduced the frequency of MHCIIhi \"inflammatory\" and Trem2hi \"metabolic\" macrophages, but not tissue resident macrophages. Furthermore, anti-miR-33 led to derepression of distinct miR-33 target genes in the different macrophage subsets: in resident and Trem2hi macrophages, anti-miR-33 relieved repression of miR-33 target genes involved in lipid metabolism (e.g., Abca1, Ncoa1, Ncoa2, Crot), whereas in MHCIIhi macrophages, anti-miR-33 upregulated target genes involved in chromatin remodeling and transcriptional regulation. Anti-miR-33 also reduced the accumulation of aortic CD8+ T cells and CD4+ Th1 cells, and increased levels of FoxP3+ regulatory T cells in plaques, consistent with an immune-dampening effect on plaque inflammation. Conclusions: Our results provide insight into the immune mechanisms and cellular players that execute anti-miR-33\'s atheroprotective actions in the plaque.




Circ Res: 16 Feb 2021; epub ahead of print
Afonso MS, Sharma M, Schlegel MP, van Solingen C, ... Fisher EA, Moore KJ
Circ Res: 16 Feb 2021; epub ahead of print | PMID: 33593073
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Abstract

Mechanisms of Congenital Heart Disease Caused by NAA15 Haploinsufficiency.

Ward TL, Tai W, Morton SU, Impens F, ... Seidman CE, Seidman JG
Rationale: NAA15 is a component of the N-terminal (Nt) acetyltransferase complex, NatA. The mechanism by which NAA15 haploinsufficiency causes congenital heart disease (CHD) remains unknown. To better understand molecular processes by which NAA15 haploinsufficiency perturbs cardiac development, we introduced NAA15 variants into human induced pluripotent stem cells (iPSCs) and assessed the consequences of these mutations on RNA and protein expression. Objective: We aim to understand the role of NAA15 haploinsufficiency in cardiac development by investigating proteomic effects on NatA complex activity, and identifying proteins dependent upon a full amount of NAA15.
Methods and results:
We introduced heterozygous LoF, compound heterozygous and missense residues (R276W) in iPS cells using CRISPR/Cas9. Haploinsufficient NAA15 iPS cells differentiate into cardiomyocytes, unlike NAA15-null iPS cells, presumably due to altered composition of NatA. Mass spectrometry (MS) analyses reveal ~80% of identified iPS cell NatA targeted proteins displayed partial or complete Nt-acetylation. Between null and haploinsufficient NAA15 cells Nt-acetylation levels of 32 and 9 NatA-specific targeted proteins were reduced, respectively. Similar acetylation loss in few proteins occurred in NAA15 R276W iPSCs. In addition, steady-state protein levels of 562 proteins were altered in both null and haploinsufficient NAA15 cells; eighteen were ribosomal-associated proteins. At least four proteins were encoded by genes known to cause autosomal dominant CHD. Conclusions: These studies define a set of human proteins that requires a full NAA15 complement for normal synthesis and development. A 50% reduction in the amount of NAA15 alters levels of at least 562 proteins and Nt-acetylation of only 9 proteins. One or more modulated proteins are likely responsible for NAA15-haploinsufficiency mediated CHD. Additionally, genetically engineered iPS cells provide a platform for evaluating the consequences of amino acid sequence variants of unknown significance on NAA15 function.




Circ Res: 08 Feb 2021; epub ahead of print
Ward TL, Tai W, Morton SU, Impens F, ... Seidman CE, Seidman JG
Circ Res: 08 Feb 2021; epub ahead of print | PMID: 33557580
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Abstract

LncRNA ZNF593-AS Alleviates Contractile Dysfunction in Dilated Cardiomyopathy.

Fan J, Li H, Xie R, Zhang X, ... Chen C, Wang DW
Rationale: Previously, we identified the human cardiac long non-coding RNAs (lncRNAs) profile in dilated cardiomyopathy (DCM) patients, among which ZNF593-AS, also named as RP11-96L14.7 and ENST00000448923.2, showed good conservation among species. Objective: We aim to elucidate the mechanism underlying lncRNA in DCM and DCM that lead to heart failure, which might provide new insights into the mechanisms of DCM and possible treatment strategies in the future.
Methods and results:
lncRNA expression was measured by real-time PCR and in situ hybridization assays. Coding potential was verified by bioinformatic and biologic assays. Recombinant adeno-associated virus with cardiac specific promoter was used to deliver lncRNA in vivo, while cardiac structure and functions were assessed by echocardiography and catheter. Sarcomere shortening, calcium imaging, gene expression profiling, and pull-down assays were performed to investigate the underlying mechanisms. ZNF593-AS, which mainly localized in the cytoplasm of cardiomyocytes, was robustly decreased in the failing heart of DCM patients, as well as in phenylephrine-treated human cardiomyocytes. Overexpression of mmu-ZNF593-AS significantly improved transverse aortic constriction (TAC)-induced cardiac dysfunction in mice. Moreover, ZNF593-AS overexpression restored the aberrant Ca2+ handling and contractility of cardiomyocytes from TAC-treated mice. Further, we found that ZNF593-AS acted as a guide RNA scaffold and recruited HNRNPC to ryanodine receptor type 2 (RYR2) mRNA, which in turn facilitated RYR2 mRNA stability, contributed to the improvement of cardiac Ca2+ handling and contractile function in DCM. Conclusions: Our findings suggested that lncRNA-based therapeutics may protect against DCM.




Circ Res: 07 Feb 2021; epub ahead of print
Fan J, Li H, Xie R, Zhang X, ... Chen C, Wang DW
Circ Res: 07 Feb 2021; epub ahead of print | PMID: 33550812
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Abstract

Structural and Functional Characterization of a Na1.5-Mitochondrial Couplon.

Pérez-Hernández M, Leo-Macias A, Keegan S, Jouni M, ... Rothenberg E, Delmar M
Rationale
The cardiac sodium channel NaV1.5 has a fundamental role in excitability and conduction. Previous studies have shown that sodium channels cluster together in specific cellular subdomains. Their association with intracellular organelles in defined regions of the myocytes, and the functional consequences of that association, remain to be defined.
Objective
To characterize a subcellular domain formed by sodium channel clusters in the crest region of the myocytes and the subjacent subsarcolemmal mitochondria.
Methods and results
Through a combination of imaging approaches including super-resolution microscopy and electron microscopy we identified, in adult cardiac myocytes, a NaV1.5 subpopulation in close proximity to subjacent subsarcolemmal mitochondria; we further found that subjacent subsarcolemmal mitochondria preferentially host the mitochondrial NCLX (Na+/Ca2+ exchanger). This anatomic proximity led us to investigate functional changes in mitochondria resulting from sodium channel activity. Upon TTX (tetrodotoxin) exposure, mitochondria near NaV1.5 channels accumulated more Ca2+ and showed increased reactive oxygen species production when compared with interfibrillar mitochondria. Finally, crosstalk between NaV1.5 channels and mitochondria was analyzed at a transcriptional level. We found that SCN5A (encoding NaV1.5) and SLC8B1 (which encode NaV1.5 and NCLX, respectively) are negatively correlated both in a human transcriptome data set (Genotype-Tissue Expression) and in human-induced pluripotent stem cell-derived cardiac myocytes deficient in SCN5A.
Conclusions
We describe an anatomic hub (a couplon) formed by sodium channel clusters and subjacent subsarcolemmal mitochondria. Preferential localization of NCLX to this domain allows for functional coupling where the extrusion of Ca2+ from the mitochondria is powered, at least in part, by the entry of sodium through NaV1.5 channels. These results provide a novel entry-point into a mechanistic understanding of the intersection between electrical and structural functions of the heart.



Circ Res: 04 Feb 2021; 128:419-432
Pérez-Hernández M, Leo-Macias A, Keegan S, Jouni M, ... Rothenberg E, Delmar M
Circ Res: 04 Feb 2021; 128:419-432 | PMID: 33342222
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Abstract

Cell-Extracellular Matrix Interactions Play Multiple Essential Roles in Aortic Arch Development.

Warkala M, Chen D, Ramirez A, Jubran A, ... Zhao H, Astrof S
Rationale
Defects in the morphogenesis of the fourth pharyngeal arch arteries (PAAs) give rise to lethal birth defects. Understanding genes and mechanisms regulating PAA formation will provide important insights into the etiology and treatments for congenital heart disease.
Objective
Cell-ECM (extracellular matrix) interactions play essential roles in the morphogenesis of PAAs and their derivatives, the aortic arch artery and its major branches; however, their specific functions are not well-understood. Previously, we demonstrated that integrin α5β1 and Fn1 (fibronectin) expressed in the Isl1 lineages regulate PAA formation. The objective of the current studies was to investigate cellular mechanisms by which integrin α5β1 and Fn1 regulate aortic arch artery morphogenesis.
Methods and results
Using temporal lineage tracing, whole-mount confocal imaging, and quantitative analysis of the second heart field (SHF) and endothelial cell (EC) dynamics, we show that the majority of PAA EC progenitors arise by E7.5 in the SHF and contribute to pharyngeal arch endothelium between E7.5 and E9.5. Consequently, SHF-derived ECs in the pharyngeal arches form a plexus of small blood vessels, which remodels into the PAAs by 35 somites. The remodeling of the vascular plexus is orchestrated by signals dependent on the pharyngeal ECM microenvironment, extrinsic to the endothelium. Conditional ablation of integrin α5β1 or Fn1 in the Isl1 lineages showed that signaling by the ECM regulates aortic arch artery morphogenesis at multiple steps: (1) accumulation of SHF-derived ECs in the pharyngeal arches, (2) remodeling of the EC plexus in the fourth arches into the PAAs, and (3) differentiation of neural crest-derived cells adjacent to the PAA endothelium into vascular smooth muscle cells.
Conclusions
PAA formation is a multistep process entailing dynamic contribution of SHF-derived ECs to pharyngeal arches, the remodeling of endothelial plexus into the PAAs, and the remodeling of the PAAs into the aortic arch artery and its major branches. Cell-ECM interactions regulated by integrin α5β1 and Fn1 play essential roles at each of these developmental stages.



Circ Res: 04 Feb 2021; 128:e27-e44
Warkala M, Chen D, Ramirez A, Jubran A, ... Zhao H, Astrof S
Circ Res: 04 Feb 2021; 128:e27-e44 | PMID: 33249995
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Abstract

Lipid Profiles and Heart Failure Risk: Results From Two Prospective Studies.

Wittenbecher C, Eichelmann F, Toledo E, Guasch-Ferré M, ... Martínez-González MÁ, Hu FB
Rationale
Altered lipid metabolism has been implicated in heart failure (HF) development, but no prospective studies have examined comprehensive lipidomics data and subsequent risk of HF.
Objective
We aimed to link single lipid metabolites and lipidomics networks to the risk of developing HF.
Methods and results
Discovery analyses were based on 216 targeted lipids in a case-control study (331 incident HF cases and 507 controls, matched by age, sex, and study center), nested within the PREDIMED (Prevención con Dieta Mediterránea) study. Associations of single lipids were examined in conditional logistic regression models. Furthermore, lipidomics networks were linked to HF risk in a multistep workflow, including machine learning-based identification of the HF-related network clusters, and regression-based discovery of the HF-related lipid patterns within these clusters. If available, significant findings were externally validated in a subsample of the EPIC-Potsdam cohort (2414 at-risk participants, including 87 incident HF cases). After confounder-adjustments, 2 lipids were significantly associated with HF risk in both cohorts: CER (ceramide) 16:0 (relative risk [RR] per SD in PREDIMED, 1.28 [95% CI, 1.13-1.47]) and phosphatidylcholine 32_0 (RR per SD in PREDIMED, 1.23 [95% CI, 1.08-1.41]). Additionally, lipid patterns in several network clusters were associated with HF risk in PREDIMED. Adjusted for standard risk factors, an internally cross-validated score based on the significant HF-related lipids that were identified in the network analysis in PREDIMED was associated with a higher HF risk (20 lipids, RR per SD, 2.33 [95% CI, 1.93%-2.81%). Moreover, a lipid score restricted to the externally available lipids was significantly associated with HF incidence in both cohorts (6 lipids, RRs per SD, 1.30 [95% CI, 1.14-1.47] in PREDIMED, and 1.46 [95% CI, 1.17-1.82] in EPIC-Potsdam).
Conclusions
Our study identified and validated 2 lipid metabolites and several lipidomics patterns as potential novel biomarkers of HF risk. Lipid profiling may capture preclinical molecular alterations that predispose for incident HF. Registration: URL: https://www.isrctn.com/ISRCTN35739639; Unique identifier: ISRCTN35739639.



Circ Res: 04 Feb 2021; 128:309-320
Wittenbecher C, Eichelmann F, Toledo E, Guasch-Ferré M, ... Martínez-González MÁ, Hu FB
Circ Res: 04 Feb 2021; 128:309-320 | PMID: 33272114
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Abstract

Endothelial S1P Signaling Counteracts Infarct Expansion in Ischemic Stroke.

Nitzsche A, Poittevin M, Benarab A, Bonnin P, ... Hla T, Camerer E
Rationale
Cerebrovascular function is critical for brain health, and endogenous vascular protective pathways may provide therapeutic targets for neurological disorders. S1P (Sphingosine 1-phosphate) signaling coordinates vascular functions in other organs, and S1P1 (S1P receptor-1) modulators including fingolimod show promise for the treatment of ischemic and hemorrhagic stroke. However, S1P1 also coordinates lymphocyte trafficking, and lymphocytes are currently viewed as the principal therapeutic target for S1P1 modulation in stroke.
Objective
To address roles and mechanisms of engagement of endothelial cell S1P1 in the naive and ischemic brain and its potential as a target for cerebrovascular therapy.
Methods and results
Using spatial modulation of S1P provision and signaling, we demonstrate a critical vascular protective role for endothelial S1P1 in the mouse brain. With an S1P1 signaling reporter, we reveal that abluminal polarization shields S1P1 from circulating endogenous and synthetic ligands after maturation of the blood-neural barrier, restricting homeostatic signaling to a subset of arteriolar endothelial cells. S1P1 signaling sustains hallmark endothelial functions in the naive brain and expands during ischemia by engagement of cell-autonomous S1P provision. Disrupting this pathway by endothelial cell-selective deficiency in S1P production, export, or the S1P1 receptor substantially exacerbates brain injury in permanent and transient models of ischemic stroke. By contrast, profound lymphopenia induced by loss of lymphocyte S1P1 provides modest protection only in the context of reperfusion. In the ischemic brain, endothelial cell S1P1 supports blood-brain barrier function, microvascular patency, and the rerouting of blood to hypoperfused brain tissue through collateral anastomoses. Boosting these functions by supplemental pharmacological engagement of the endothelial receptor pool with a blood-brain barrier penetrating S1P1-selective agonist can further reduce cortical infarct expansion in a therapeutically relevant time frame and independent of reperfusion.
Conclusions
This study provides genetic evidence to support a pivotal role for the endothelium in maintaining perfusion and microvascular patency in the ischemic penumbra that is coordinated by S1P signaling and can be harnessed for neuroprotection with blood-brain barrier-penetrating S1P1 agonists.



Circ Res: 04 Feb 2021; 128:363-382
Nitzsche A, Poittevin M, Benarab A, Bonnin P, ... Hla T, Camerer E
Circ Res: 04 Feb 2021; 128:363-382 | PMID: 33301355
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Abstract

RYR2 Channel Inhibition Is the Principal Mechanism of Flecainide Action in CPVT.

Kryshtal DO, Blackwell DJ, Egly CL, Smith AN, ... Laver DR, Knollmann BC
Rationale
The class Ic antiarrhythmic drug flecainide prevents ventricular tachyarrhythmia in patients with catecholaminergic polymorphic ventricular tachycardia (CPVT), a disease caused by hyperactive RyR2 (cardiac ryanodine receptor) mediated calcium (Ca) release. Although flecainide inhibits single RyR2 channels in vitro, reports have claimed that RyR2 inhibition by flecainide is not relevant for its mechanism of antiarrhythmic action and concluded that sodium channel block alone is responsible for flecainide\'s efficacy in CPVT.
Objective
To determine whether RyR2 block independently contributes to flecainide\'s efficacy for suppressing spontaneous sarcoplasmic reticulum Ca release and for preventing ventricular tachycardia in vivo.
Methods and results
We synthesized N-methylated flecainide analogues (QX-flecainide and N-methyl flecainide) and showed that N-methylation reduces flecainide\'s inhibitory potency on RyR2 channels incorporated into artificial lipid bilayers. N-methylation did not alter flecainide\'s inhibitory activity on human cardiac sodium channels expressed in HEK293T cells. Antiarrhythmic efficacy was tested utilizing a Casq2 (cardiac calsequestrin) knockout (Casq2-/-) CPVT mouse model. In membrane-permeabilized Casq2-/- cardiomyocytes-lacking intact sarcolemma and devoid of sodium channel contribution-flecainide, but not its analogues, suppressed RyR2-mediated Ca release at clinically relevant concentrations. In voltage-clamped, intact Casq2-/- cardiomyocytes pretreated with tetrodotoxin to inhibit sodium channels and isolate the effect of flecainide on RyR2, flecainide significantly reduced the frequency of spontaneous sarcoplasmic reticulum Ca release, while QX-flecainide and N-methyl flecainide did not. In vivo, flecainide effectively suppressed catecholamine-induced ventricular tachyarrhythmias in Casq2-/- mice, whereas N-methyl flecainide had no significant effect on arrhythmia burden, despite comparable sodium channel block.
Conclusions
Flecainide remains an effective inhibitor of RyR2-mediated arrhythmogenic Ca release even when cardiac sodium channels are blocked. In mice with CPVT, sodium channel block alone did not prevent ventricular tachycardia. Hence, RyR2 channel inhibition likely constitutes the principal mechanism of antiarrhythmic action of flecainide in CPVT.



Circ Res: 04 Feb 2021; 128:321-331
Kryshtal DO, Blackwell DJ, Egly CL, Smith AN, ... Laver DR, Knollmann BC
Circ Res: 04 Feb 2021; 128:321-331 | PMID: 33297863
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Abstract

PPARγ-p53-Mediated Vasculoregenerative Program to Reverse Pulmonary Hypertension.

Hennigs JK, Cao A, Li CG, Shi M, ... Snyder MP, Rabinovitch M
Rationale
In pulmonary arterial hypertension (PAH), endothelial dysfunction and obliterative vascular disease are associated with DNA damage and impaired signaling of BMPR2 (bone morphogenetic protein type 2 receptor) via two downstream transcription factors, PPARγ (peroxisome proliferator-activated receptor gamma), and p53.
Objective
We investigated the vasculoprotective and regenerative potential of a newly identified PPARγ-p53 transcription factor complex in the pulmonary endothelium.
Methods and results
In this study, we identified a pharmacologically inducible vasculoprotective mechanism in pulmonary arterial and lung MV (microvascular) endothelial cells in response to DNA damage and oxidant stress regulated in part by a BMPR2 dependent transcription factor complex between PPARγ and p53. Chromatin immunoprecipitation sequencing and RNA-sequencing established an inducible PPARγ-p53 mediated regenerative program regulating 19 genes involved in lung endothelial cell survival, angiogenesis and DNA repair including, EPHA2 (ephrin type-A receptor 2), FHL2 (four and a half LIM domains protein 2), JAG1 (jagged 1), SULF2 (extracellular sulfatase Sulf-2), and TIGAR (TP53-inducible glycolysis and apoptosis regulator). Expression of these genes was partially impaired when the PPARγ-p53 complex was pharmacologically disrupted or when BMPR2 was reduced in pulmonary artery endothelial cells (PAECs) subjected to oxidative stress. In endothelial cell-specific Bmpr2-knockout mice unable to stabilize p53 in endothelial cells under oxidative stress, Nutlin-3 rescued endothelial p53 and PPARγ-p53 complex formation and induced target genes, such as APLN (apelin) and JAG1, to regenerate pulmonary microvessels and reverse pulmonary hypertension. In PAECs from BMPR2 mutant PAH patients, pharmacological induction of p53 and PPARγ-p53 genes repaired damaged DNA utilizing genes from the nucleotide excision repair pathway without provoking PAEC apoptosis.
Conclusions
We identified a novel therapeutic strategy that activates a vasculoprotective gene regulation program in PAECs downstream of dysfunctional BMPR2 to rehabilitate PAH PAECs, regenerate pulmonary microvessels, and reverse disease. Our studies pave the way for p53-based vasculoregenerative therapies for PAH by extending the therapeutic focus to PAEC dysfunction and to DNA damage associated with PAH progression.



Circ Res: 04 Feb 2021; 128:401-418
Hennigs JK, Cao A, Li CG, Shi M, ... Snyder MP, Rabinovitch M
Circ Res: 04 Feb 2021; 128:401-418 | PMID: 33322916
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Abstract

KLF5 Is Induced by FOXO1 and Causes Oxidative Stress and Diabetic Cardiomyopathy.

Kyriazis ID, Hoffman M, Gaignebet L, Lucchese AM, ... Kararigas G, Drosatos K
Rationale
Diabetic cardiomyopathy (DbCM) is a major complication in type-1 diabetes, accompanied by altered cardiac energetics, impaired mitochondrial function, and oxidative stress. Previous studies indicate that type-1 diabetes is associated with increased cardiac expression of KLF5 (Krüppel-like factor-5) and PPARα (peroxisome proliferator-activated receptor) that regulate cardiac lipid metabolism.
Objective
In this study, we investigated the involvement of KLF5 in DbCM and its transcriptional regulation.
Methods and results
KLF5 mRNA levels were assessed in isolated cardiomyocytes from cardiovascular patients with diabetes and were higher compared with nondiabetic individuals. Analyses in human cells and diabetic mice with cardiomyocyte-specific FOXO1 (Forkhead box protein O1) deletion showed that FOXO1 bound directly on the KLF5 promoter and increased KLF5 expression. Diabetic mice with cardiomyocyte-specific FOXO1 deletion had lower cardiac KLF5 expression and were protected from DbCM. Genetic, pharmacological gain and loss of KLF5 function approaches and AAV (adeno-associated virus)-mediated Klf5 delivery in mice showed that KLF5 induces DbCM. Accordingly, the protective effect of cardiomyocyte FOXO1 ablation in DbCM was abolished when KLF5 expression was rescued. Similarly, constitutive cardiomyocyte-specific KLF5 overexpression caused cardiac dysfunction. KLF5 caused oxidative stress via direct binding on NADPH oxidase (NOX)4 promoter and induction of NOX4 (NADPH oxidase 4) expression. This was accompanied by accumulation of cardiac ceramides. Pharmacological or genetic KLF5 inhibition alleviated superoxide formation, prevented ceramide accumulation, and improved cardiac function in diabetic mice.
Conclusions
Diabetes-mediated activation of cardiomyocyte FOXO1 increases KLF5 expression, which stimulates NOX4 expression, ceramide accumulation, and causes DbCM.



Circ Res: 04 Feb 2021; 128:335-357
Kyriazis ID, Hoffman M, Gaignebet L, Lucchese AM, ... Kararigas G, Drosatos K
Circ Res: 04 Feb 2021; 128:335-357 | PMID: 33539225
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Abstract

Endothelial Cell Receptors in Tissue Lipid Uptake and Metabolism.

Abumrad NA, Cabodevilla AG, Samovski D, Pietka T, Basu D, Goldberg IJ
Lipid uptake and metabolism are central to the function of organs such as heart, skeletal muscle, and adipose tissue. Although most heart energy derives from fatty acids (FAs), excess lipid accumulation can cause cardiomyopathy. Similarly, high delivery of cholesterol can initiate coronary artery atherosclerosis. Hearts and arteries-unlike liver and adrenals-have nonfenestrated capillaries and lipid accumulation in both health and disease requires lipid movement from the circulation across the endothelial barrier. This review summarizes recent in vitro and in vivo findings on the importance of endothelial cell receptors and uptake pathways in regulating FAs and cholesterol uptake in normal physiology and cardiovascular disease. We highlight clinical and experimental data on the roles of ECs in lipid supply to tissues, heart, and arterial wall in particular, and how this affects organ metabolism and function. Models of FA uptake into ECs suggest that receptor-mediated uptake predominates at low FA concentrations, such as during fasting, whereas FA uptake during lipolysis of chylomicrons may involve paracellular movement. Similarly, in the setting of an intact arterial endothelial layer, recent and historic data support a role for receptor-mediated processes in the movement of lipoproteins into the subarterial space. We conclude with thoughts on the need to better understand endothelial lipid transfer for fuller comprehension of the pathophysiology of hyperlipidemia, and lipotoxic diseases such as some forms of cardiomyopathy and atherosclerosis.



Circ Res: 04 Feb 2021; 128:433-450
Abumrad NA, Cabodevilla AG, Samovski D, Pietka T, Basu D, Goldberg IJ
Circ Res: 04 Feb 2021; 128:433-450 | PMID: 33539224
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Abstract

MCU Overexpression Rescues Inotropy and Reverses Heart Failure by Reducing SR Ca Leak.

Liu T, Yang N, Sidor A, O\'Rourke B
Rationale: In heart failure (HF), impaired sarcoplasmic reticulum (SR) Ca2+ release and cytosolic Na+ overload depress mitochondrial Ca2+ (mCa2+) signaling, resulting in a diminished ability to maintain matrix NAD(P)H redox potential, leading to increased oxidative stress when workload increases. Enhancing mCa2+ can reverse this defect but could potentially increase the likelihood of mitochondrial Ca2+ overload.Objective: To determine if moderate mitochondrial Ca2+ uniporter (MCU) overexpression has beneficial or detrimental effects on the development of HF and incident arrythmias in a guinea pig model (ACi) of HF and sudden cardiac death.
Methods and results:
In vivo viral gene transfer was used to increase MCU levels by ~30% in ACi hearts. Left ventricular myocytes from hearts with MCU overexpression (ACi+MCU) displayed enhanced mCa2+ uptake, decreased oxidative stress, and increased β-adrenergic- and frequency-dependent augmentation of Ca2+ transients and contractions, compared to myocytes from ACi hearts. MCU overexpression decreased SR Ca2+ leak in the ACi group and mitigated the elevated ryanodine receptor disulfide crosslinks in HF. β-adrenergic responses were blunted in isolated perfused ACi hearts and these deficiencies were normalized in ACI+MCU hearts. To examine the in vivo effects of MCU overexpression, ACi hearts were transduced with the MCU virus 2 3w after aortic constriction, at the onset of cardiac decompensation. Two weeks later, cardiac function worsened in the untreated ACi group (fractional shortening: 39{plus minus}1% at 2w and 32{plus minus}1% at 4w), whereas MCU overexpression significantly improved cardiac function (36{plus minus}1% at 2w and 42{plus minus}2% at 4w). MCU overexpression in the decompensating ACi heart also attenuated pulmonary edema and interstitial fibrosis and prevented triggered arrhythmias. Conclusions: Moderate MCU overexpression in failing hearts enhances contractility and responses to β-adrenergic stimulation in isolated myocytes and perfused hearts by inhibiting mitochondrial oxidative stress-induced SR Ca2+ leak. MCU overexpression also reversed HF and inhibited ectopic ventricular arrhythmias.




Circ Res: 31 Jan 2021; epub ahead of print
Liu T, Yang N, Sidor A, O'Rourke B
Circ Res: 31 Jan 2021; epub ahead of print | PMID: 33522833
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Impact:
Abstract

β2M Signals Monocytes Through Non-Canonical TGFβ Receptor Signal Transduction.

Hilt ZT, Maurya P, Tesoro L, Pariser DN, ... McGrath K, Morrell C
Rationale: Circulating monocytes can have pro-inflammatory or pro-reparative phenotypes. The endogenous signaling molecules and pathways that regulate monocyte polarization in vivo are poorly understood. We have shown that platelet derived beta-2 microglobulin (β2M) and transforming growth factor beta (TGFβ) have opposing effects on monocytes by inducing inflammatory and reparative phenotypes respectively, but each bind and signal through the same receptor. We now define the signaling pathways involved. Objective: To determine the molecular mechanisms and signal transduction pathways by which β2M and TGFβ regulate monocyte responses both in vitro and in vivo.
Methods and results:
Wild-type (WT) and platelet specific β2M knockout (Plt-β2M-/-) mice were treated intravenously with either β2M or TGFβ to increase plasma concentrations to those in cardiovascular diseases. Elevated plasma β2M increased pro-inflammatory monocytes, while increased plasma TGFβ increased pro-reparative monocytes. TGFβ receptor (TGFβR) inhibition blunted monocyte responses to both β2M and TGFβ in vivo. Using imaging flow cytometry, we found that β2M decreased monocyte SMAD2/3 nuclear localization, while TGFβ promoted SMAD nuclear translocation, but decreased non-canonical/inflammatory (JNK and NFκB nuclear localization). This was confirmed in vitro using both imaging flow cytometry and immunoblots. β2M, but not TGFβ, promoted ubiquitination of SMAD3 and SMAD4, that inhibited their nuclear trafficking. Inhibition of ubiquitin ligase activity blocked non-canonical SMAD-independent monocyte signaling and skewed monocytes towards a pro-reparative monocyte response. Conclusions: Our findings indicate that elevated plasma β2M and TGFβ dichotomously polarize monocytes. Furthermore, these immune molecules share a common receptor, but induce SMAD-dependent canonical signaling (TGFβ) versus non-canonical SMAD-independent signaling (β2M) in a ubiquitin ligase dependent manner. This work has broad implications as β2M is increased in several inflammatory conditions, while TGFβ is increased in fibrotic diseases.




Circ Res: 28 Jan 2021; epub ahead of print
Hilt ZT, Maurya P, Tesoro L, Pariser DN, ... McGrath K, Morrell C
Circ Res: 28 Jan 2021; epub ahead of print | PMID: 33508948
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Abstract

Chromatin Accessibility of Human Mitral Valves and Functional Assessment of MVP Risk Loci.

Kyryachenko S, Georges A, Yu M, Berrandou T, ... Voigt N, Bouatia-Naji N
Rationale: Mitral valve prolapse (MVP) is a common valvopathy that leads to mitral insufficiency, heart failure and sudden death. Functional genomic studies in mitral valves are needed to better characterize MVP associated variants and target genes.Objective: To establish the chromatin accessibility profiles and assess functionality of variants and narrow down target genes at MVP loci.
Methods and results:
We mapped the open chromatin regions in nuclei from 11 human pathogenic and 7 non-pathogenic mitral valves by an assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-Seq). Open chromatin peaks were globally similar between pathogenic and non-pathogenic valves. Compared to the heart tissue and cardiac fibroblasts, we found that MV-specific ATAC-Seq peaks are enriched near genes involved in extracellular matrix organization, chondrocyte differentiation, and connective tissue development. One of the most enriched motif in MV-specific open chromatin peaks was for the nuclear factor of activated T cells (NFATC) family of transcription factors, involved in valve endocardial and interstitial cells formation. We also found that MVP-associated variants were significantly enriched (p<0.05) in mitral valve open chromatin peaks. Integration of the ATAC-Seq data with risk loci, extensive functional annotation, and gene reporter assay suggest plausible causal variants for rs2641440 at the SMG6/SRR locus and rs6723013 at the IGFBP2/IGFBP5/TNS1 locus. CRISPR-Cas9 deletion of the sequence including rs6723013 in human fibroblasts correlated with increased expression only for TNS1. 4C-Seq experiments provided evidence for several target genes, including SRR, HIC1, and DPH1 at the SMG6/SRR locus and further supported TNS1 as the most likely target gene on Chr2.Conclusions: Here we describe unprecedented genome-wide open chromatin profiles from human pathogenic and non-pathogenic MVs and report specific gene regulation profiles, compared to the heart. We also report in vitro functional evidence for potential causal variants and target genes at MVP risk loci involving established and new biological mechanisms.




Circ Res: 27 Jan 2021; epub ahead of print
Kyryachenko S, Georges A, Yu M, Berrandou T, ... Voigt N, Bouatia-Naji N
Circ Res: 27 Jan 2021; epub ahead of print | PMID: 33508947
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Abstract

Influenza Causes MLKL-Driven Cardiac Proteome Remodeling During Convalescence.

Lin YH, Platt M, Gilley RP, Brown D, ... Yu Y, Gonzalez-Juarbe N
Rationale: Patients with and without cardiovascular diseases have been shown to be at risk of influenza-mediated cardiac complications. Recent clinical reports support the notion of a direct link between laboratory-confirmed influenza virus infections and adverse cardiac events. Objective: Define the molecular mechanisms underlying influenza virus-induced cardiac pathogenesis after resolution of pulmonary infection and the role of necroptosis in this process.
Methods and results:
Hearts from wild-type and necroptosis deficient (MLKL-KO) mice were dissected twelve days after initial Influenza A virus (IAV) infection when viral titers were undetectable in the lungs. Immunofluorescence microscopy and plaque assays showed presence of viable IAV particles in the myocardium without generation of interferon responses. Global proteome and phosphoproteome analyses using high resolution accurate mass based LC-MS/MS and label-free quantitation showed that the global proteome as well as the phosphoproteome profiles were significantly altered in IAV-infected mouse hearts in a strain independent manner. Necroptosis deficient mice had increased survival and reduced weight loss post-IAV infection, as well as increased antioxidant and mitochondrial function, indicating partial protection to IAV infection. These findings were confirmed in vitro by pre-treatment of human and rat myocytes with antioxidants or necroptosis inhibitors, which blunted oxidative stress and mitochondrial damage after IAV infection. Conclusions: This study provides the first evidence that the cardiac proteome and phosphoproteome are significantly altered post pulmonary influenza infection. Moreover, viral particles can persist in the heart after lung clearance, altering mitochondrial function and promoting cell death without active replication and interferon responses. Finally, our findings show inhibition of necroptosis or prevention of mitochondrial damage as possible therapeutic interventions to reduce cardiac damage during influenza infections.




Circ Res: 26 Jan 2021; epub ahead of print
Lin YH, Platt M, Gilley RP, Brown D, ... Yu Y, Gonzalez-Juarbe N
Circ Res: 26 Jan 2021; epub ahead of print | PMID: 33501852
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Abstract

Loss of Endogenously Cycling Adult Cardiomyocytes Worsens Myocardial Function.

Bradley LA, Young A, Li H, Billcheck HO, Wolf MJ
Rationale
Endogenously cycling adult cardiomyocytes increase after myocardial infarction (MI) but remain scarce and are generally thought not to contribute to myocardial function. However, this broadly held assumption has not been tested, mainly because of the lack of transgenic reporters that restrict Cre expression to adult cardiomyocytes that reenter the cell cycle.
Objective
We created and validated a new transgenic mouse, αMHC (alpha myosin heavy chain)-MerDreMer-Ki67p-RoxedCre (denoted αDKRC [cardiomyocyte-specific αMHC-MerDreMer-Ki67p-RoxedCre]) that restricts Cre expression to cycling adult cardiomyocytes and uniquely integrates spatial and temporal adult cardiomyocyte cycling events based on the DNA specificities of orthologous Dre and Cre recombinases. We then created αDKRC::DTA mice that expressed an inducible diphtheria toxin in adult cycling cardiomyocytes and examined the effects of ablating these endogenously cycling cardiomyocytes on myocardial function after ischemic-reperfusion (I/R) MI.
Methods and results
A tandem αDKRC transgene was designed, validated in cultured cells, and used to make transgenic mice. The αDKRC transgene integrated between MYH6 and MYH7 and did not disrupt expression of the surrounding genes. Compared with controls, αDKRC::RLTG (Rox-Lox-tdTomato-eGFP) mice treated with Tamoxifen expressed tdTomato+ in cardiomyocytes with rare Bromodeoxyuridine+, eGFP+ cardiomyocytes, consistent with reentry of the cell cycle. We then pretreated αDKRC::RLTG mice with Tamoxifen to activate the reporter before sham or reperfusion (I/R) MI surgeries. Compared with Sham surgery, the I/R MI group had increased single and paired eGFP+ (enhanced green fluorescent protein)+ cardiomyocytes predominantly in the border zones (5.8±0.5 versus 3.3±0.3 cardiomyocytes per 10-micron section, N=8-9 mice per group, n=16-24 sections per mouse), indicative of cycled cardiomyocytes. The single to paired eGFP+ cardiomyocyte ratio was ≈9 to 1 (5.2±0.4 single versus 0.6±0.2 paired cardiomyocytes) in the I/R MI group after MI, suggesting that cycling cardiomyocytes were more likely to undergo polyploidy than replication. The ablation of endogenously cycling adult cardiomyocytes in αDKRC::DTA (diphtheria) mice caused progressive worsening left ventricular chamber size and function after I/R MI, compared with controls.
Conclusions
Although scarce, endogenously cycling adult cardiomyocytes contribute to myocardial function after injury, suggesting that these cells may be physiologically relevant.



Circ Res: 21 Jan 2021; 128:155-168
Bradley LA, Young A, Li H, Billcheck HO, Wolf MJ
Circ Res: 21 Jan 2021; 128:155-168 | PMID: 33146578
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Abstract

Clonal Hematopoiesis-Driver DNMT3A Mutations Alter Immune Cells in Heart Failure.

Abplanalp WT, Cremer S, John D, Hoffmann J, ... Zeiher AM, Dimmeler S
Rationale
Clonal hematopoiesis driven by mutations of DNMT3A (DNA methyltransferase 3a) is associated with increased incidence of cardiovascular disease and poor prognosis of patients with chronic heart failure (HF) and aortic stenosis. Although experimental studies suggest that DNMT3A clonal hematopoiesis-driver mutations may enhance inflammation, specific signatures of inflammatory cells in humans are missing.
Objective
To define subsets of immune cells mediating inflammation in humans using single-cell RNA sequencing.
Methods and results
Transcriptomic profiles of peripheral blood mononuclear cells were analyzed in n=6 patients with HF harboring DNMT3A clonal hematopoiesis-driver mutations and n=4 patients with HF and no DNMT3A mutations by single-cell RNA sequencing. Monocytes of patients with HF carrying DNMT3A mutations demonstrated a significantly increased expression of inflammatory genes compared with monocytes derived from patients with HF without DNMT3A mutations. Among the specific upregulated genes were the prototypic inflammatory IL (interleukin) IL1B (interleukin 1B), IL6, IL8, the inflammasome NLRP3, and the macrophage inflammatory proteins CCL3 and CCL4 as well as resistin, which augments monocyte-endothelial adhesion. Silencing of DNMT3A in monocytes induced a paracrine proinflammatory activation and increased adhesion to endothelial cells. Furthermore, the classical monocyte subset of DNMT3A mutation carriers showed increased expression of T-cell stimulating immunoglobulin superfamily members CD300LB, CD83, SIGLEC12, as well as the CD2 ligand and cell adhesion molecule CD58, all of which may be involved in monocyte-T-cell interactions. DNMT3A mutation carriers were further characterized by increased expression of the T-cell alpha receptor constant chain and changes in T helper cell 1, T helper cell 2, T helper cell 17, CD8+ effector, CD4+ memory, and regulatory T-cell-specific signatures.
Conclusions
This study demonstrates that circulating monocytes and T cells of patients with HF harboring clonal hematopoiesis-driver mutations in DNMT3A exhibit a highly inflamed transcriptome, which may contribute to the aggravation of chronic HF.



Circ Res: 21 Jan 2021; 128:216-228
Abplanalp WT, Cremer S, John D, Hoffmann J, ... Zeiher AM, Dimmeler S
Circ Res: 21 Jan 2021; 128:216-228 | PMID: 33155517
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Impact:
Abstract

Eosinophils Protect Mice From Angiotensin-II Perfusion-Induced Abdominal Aortic Aneurysm.

Liu CL, Liu X, Zhang Y, Liu J, ... Zhang J, Shi GP
Rationale
Blood eosinophil count and ECP (eosinophil cationic protein) associate with human cardiovascular diseases. Yet, whether eosinophils play a role in cardiovascular disease remains untested. The current study detected eosinophil accumulation in human and murine abdominal aortic aneurysm (AAA) lesions, suggesting eosinophil participation in this aortic disease.
Objective
To test whether and how eosinophils affect AAA growth.
Methods and results
Population-based randomized clinically controlled screening trials revealed higher blood eosinophil count in 579 male patients with AAA than in 5063 non-AAA control (0.236±0.182 versus 0.211±0.154, 109/L, P<0.001). Univariate (odds ratio, 1.381, P<0.001) and multivariate (odds ratio, 1.237, P=0.031) logistic regression analyses indicated that increased blood eosinophil count in patients with AAA served as an independent risk factor of human AAA. Immunostaining and immunoblot analyses detected eosinophil accumulation and eosinophil cationic protein expression in human and murine AAA lesions. Results showed that eosinophil deficiency exacerbated AAA growth with increased lesion inflammatory cell contents, matrix-degrading protease activity, angiogenesis, cell proliferation and apoptosis, and smooth muscle cell loss using angiotensin-II perfusion-induced AAA in Apoe-/- and eosinophil-deficient Apoe-/-ΔdblGATA mice. Eosinophil deficiency increased lesion chemokine expression, muted lesion expression of IL (interleukin) 4 and eosinophil-associated-ribonuclease-1 (mEar1 [mouse EOS-associated-ribonuclease-1], human ECP homolog), and slanted M1 macrophage polarization. In cultured macrophages and monocytes, eosinophil-derived IL4 and mEar1 polarized M2 macrophages, suppressed CD11b+Ly6Chi monocytes, and increased CD11b+Ly6Clo monocytes. mEar1 treatment or adoptive transfer of eosinophil from wild-type and Il13-/- mice, but not eosinophil from Il4-/- mice, blocked AAA growth in Apoe-/-ΔdblGATA mice. Immunofluorescent staining and immunoblot analyses demonstrated a role for eosinophil IL4 and mEar1 in blocking NF-κB (nuclear factor-κB) activation in macrophages, smooth muscle cells, and endothelial cells.
Conclusions
Eosinophils play a protective role in AAA by releasing IL4 and cationic proteins such as mEar1 to regulate macrophage and monocyte polarization and to block NF-κB activation in aortic inflammatory and vascular cells.



Circ Res: 21 Jan 2021; 128:188-202
Liu CL, Liu X, Zhang Y, Liu J, ... Zhang J, Shi GP
Circ Res: 21 Jan 2021; 128:188-202 | PMID: 33153394
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Impact:
Abstract

Machine Learned Cellular Phenotypes in Cardiomyopathy Predict Sudden Death.

Rogers AJ, Selvalingam A, Alhusseini MI, Krummen DE, ... Zaharia M, Narayan SM
Rationale
Susceptibility to VT/VF (ventricular tachycardia/fibrillation) is difficult to predict in patients with ischemic cardiomyopathy either by clinical tools or by attempting to translate cellular mechanisms to the bedside.
Objective
To develop computational phenotypes of patients with ischemic cardiomyopathy, by training then interpreting machine learning of ventricular monophasic action potentials (MAPs) to reveal phenotypes that predict long-term outcomes.
Methods and results
We recorded 5706 ventricular MAPs in 42 patients with coronary artery disease and left ventricular ejection fraction ≤40% during steady-state pacing. Patients were randomly allocated to independent training and testing cohorts in a 70:30 ratio, repeated K=10-fold. Support vector machines and convolutional neural networks were trained to 2 end points: (1) sustained VT/VF or (2) mortality at 3 years. Support vector machines provided superior classification. For patient-level predictions, we computed personalized MAP scores as the proportion of MAP beats predicting each end point. Patient-level predictions in independent test cohorts yielded c-statistics of 0.90 for sustained VT/VF (95% CI, 0.76-1.00) and 0.91 for mortality (95% CI, 0.83-1.00) and were the most significant multivariate predictors. Interpreting trained support vector machine revealed MAP morphologies that, using in silico modeling, revealed higher L-type calcium current or sodium-calcium exchanger as predominant phenotypes for VT/VF.
Conclusions
Machine learning of action potential recordings in patients revealed novel phenotypes for long-term outcomes in ischemic cardiomyopathy. Such computational phenotypes provide an approach which may reveal cellular mechanisms for clinical outcomes and could be applied to other conditions.



Circ Res: 21 Jan 2021; 128:172-184
Rogers AJ, Selvalingam A, Alhusseini MI, Krummen DE, ... Zaharia M, Narayan SM
Circ Res: 21 Jan 2021; 128:172-184 | PMID: 33167779
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Impact:
Abstract

Targeting Mitochondria-Inflammation Circuit by β-Hydroxybutyrate Mitigates HFpEF.

Deng Y, Xie M, Li Q, Xu X, ... Tian R, Li T
Rationale
Over 50% of patients with heart failure have preserved ejection fraction (HFpEF), rather than reduced ejection fraction. Complexity of its pathophysiology and the lack of animal models hamper the development of effective therapy for HFpEF.
Objective
This study was designed to investigate the metabolic mechanisms of HFpEF and test therapeutic interventions using a novel animal model.
Methods and results
By combining the age, long-term high-fat diet, and desoxycorticosterone pivalate challenge in a mouse model, we were able to recapture the myriad features of HFpEF. In these mice, mitochondrial hyperacetylation exacerbated while increasing ketone body availability rescued the phenotypes. The HFpEF mice exhibited overproduction of IL (interleukin)-1β/IL-18 and tissue fibrosis due to increased assembly of NLPR3 inflammasome on hyperacetylated mitochondria. Increasing β-hydroxybutyrate level attenuated NLPR3 inflammasome formation and antagonized proinflammatory cytokine-triggered mitochondrial dysfunction and fibrosis. Moreover, β-hydroxybutyrate downregulated the acetyl-CoA pool and mitochondrial acetylation, partially via activation of CS (citrate synthase) and inhibition of fatty acid uptake.
Conclusions
Therefore, we identify the interplay of mitochondrial hyperacetylation and inflammation as a key driver in HFpEF pathogenesis, which can be ameliorated by promoting β-hydroxybutyrate abundance.



Circ Res: 21 Jan 2021; 128:232-245
Deng Y, Xie M, Li Q, Xu X, ... Tian R, Li T
Circ Res: 21 Jan 2021; 128:232-245 | PMID: 33176578
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Impact:
Abstract

Intracellular β-Adrenergic Receptors and Organic Cation Transporter 3 Mediate Phospholamban Phosphorylation to Enhance Cardiac Contractility.

Wang Y, Shi Q, Li M, Zhao M, ... Bers DM, Xiang YK
Rationale
β1ARs (β1-adrenoceptors) exist at intracellular membranes and OCT3 (organic cation transporter 3) mediates norepinephrine entry into cardiomyocytes. However, the functional role of intracellular β1AR in cardiac contractility remains to be elucidated.
Objective
Test localization and function of intracellular β1AR on cardiac contractility.
Methods and results
Membrane fractionation, super-resolution imaging, proximity ligation, coimmunoprecipitation, and single-molecule pull-down demonstrated a pool of β1ARs in mouse hearts that were associated with sarco/endoplasmic reticulum Ca2+-ATPase at the sarcoplasmic reticulum (SR). Local PKA (protein kinase A) activation was measured using a PKA biosensor targeted at either the plasma membrane (PM) or SR. Compared with wild-type, myocytes lacking OCT3 (OCT3-KO [OCT3 knockout]) responded identically to the membrane-permeant βAR agonist isoproterenol in PKA activation at both PM and SR. The same was true at the PM for membrane-impermeant norepinephrine, but the SR response to norepinephrine was suppressed in OCT3-KO myocytes. This differential effect was recapitulated in phosphorylation of the SR-pump regulator phospholamban. Similarly, OCT3-KO selectively suppressed calcium transients and contraction responses to norepinephrine but not isoproterenol. Furthermore, sotalol, a membrane-impermeant βAR-blocker, suppressed isoproterenol-induced PKA activation at the PM but permitted PKA activation at the SR, phospholamban phosphorylation, and contractility. Moreover, pretreatment with sotalol in OCT3-KO myocytes prevented norepinephrine-induced PKA activation at both PM and the SR and contractility.
Conclusions
Functional β1ARs exists at the SR and is critical for PKA-mediated phosphorylation of phospholamban and cardiac contractility upon catecholamine stimulation. Activation of these intracellular β1ARs requires catecholamine transport via OCT3.



Circ Res: 21 Jan 2021; 128:246-261
Wang Y, Shi Q, Li M, Zhao M, ... Bers DM, Xiang YK
Circ Res: 21 Jan 2021; 128:246-261 | PMID: 33183171
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Impact:
Abstract

Heterodimerization With 5-HTR Is Indispensable for βAR-Mediated Cardioprotection.

Song Y, Xu C, Liu J, Li Y, ... Woo AY, Xiao RP
Rationale
The β2-adrenoceptor (β2-AR), a prototypical GPCR (G protein-coupled receptor), couples to both Gs and Gi proteins. Stimulation of the β2-AR is beneficial to humans and animals with heart failure presumably because it activates the downstream Gi-PI3K-Akt cell survival pathway. Cardiac β2-AR signaling can be regulated by crosstalk or heterodimerization with other GPCRs, but the physiological and pathophysiological significance of this type of regulation has not been sufficiently demonstrated.
Objective
Here, we aim to investigate the potential cardioprotective effect of β2-adrenergic stimulation with a subtype-selective agonist, (R,R\')-4-methoxy-1-naphthylfenoterol (MNF), and to decipher the underlying mechanism with a particular emphasis on the role of heterodimerization of β2-ARs with another GPCR, 5-hydroxytryptamine receptors 2B (5-HT2BRs).
Methods and results
Using pharmacological, genetic and biophysical protein-protein interaction approaches, we studied the cardioprotective effect of the β2-agonist, MNF, and explored the underlying mechanism in both in vivo in mice and cultured rodent cardiomyocytes insulted with doxorubicin, hydrogen peroxide (H2O2) or ischemia/reperfusion. In doxorubicin (Dox)-treated mice, MNF reduced mortality and body weight loss, while improving cardiac function and cardiomyocyte viability. MNF also alleviated myocardial ischemia/reperfusion injury. In cultured rodent cardiomyocytes, MNF inhibited DNA damage and cell death caused by Dox, H2O2 or hypoxia/reoxygenation. Mechanistically, we found that MNF or another β2-agonist zinterol markedly promoted heterodimerization of β2-ARs with 5-HT2BRs. Upregulation of the heterodimerized 5-HT2BRs and β2-ARs enhanced β2-AR-stimulated Gi-Akt signaling and cardioprotection while knockdown or pharmacological inhibition of the 5-HT2BR attenuated β2-AR-stimulated Gi signaling and cardioprotection.
Conclusions
These data demonstrate that the β2-AR-stimulated cardioprotective Gi signaling depends on the heterodimerization of β2-ARs and 5-HT2BRs.



Circ Res: 21 Jan 2021; 128:262-277
Song Y, Xu C, Liu J, Li Y, ... Woo AY, Xiao RP
Circ Res: 21 Jan 2021; 128:262-277 | PMID: 33208036
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Impact:
Abstract

Beat-by-Beat Cardiomyocyte T-Tubule Deformation Drives Tubular Content Exchange.

Rog-Zielinska EA, Scardigli M, Peyronnet R, Zgierski-Johnston CM, ... Sacconi L, Kohl P
Rationale
The sarcolemma of cardiomyocytes contains many proteins that are essential for electromechanical function in general, and excitation-contraction coupling in particular. The distribution of these proteins is nonuniform between the bulk sarcolemmal surface and membrane invaginations known as transverse tubules (TT). TT form an intricate network of fluid-filled conduits that support electromechanical synchronicity within cardiomyocytes. Although continuous with the extracellular space, the narrow lumen and the tortuous structure of TT can form domains of restricted diffusion. As a result of unequal ion fluxes across cell surface and TT membranes, limited diffusion may generate ion gradients within TT, especially deep within the TT network and at high pacing rates.
Objective
We postulate that there may be an advective component to TT content exchange, wherein cyclic deformation of TT during diastolic stretch and systolic shortening serves to mix TT luminal content and assists equilibration with bulk extracellular fluid.
Methods and results
Using electron tomography, we explore the 3-dimensional nanostructure of TT in rabbit ventricular myocytes, preserved at different stages of the dynamic cycle of cell contraction and relaxation. We show that cellular deformation affects TT shape in a sarcomere length-dependent manner and on a beat-by-beat time-scale. Using fluorescence recovery after photobleaching microscopy, we show that apparent speed of diffusion is affected by the mechanical state of cardiomyocytes, and that cyclic contractile activity of cardiomyocytes accelerates TT diffusion dynamics.
Conclusions
Our data confirm the existence of an advective component to TT content exchange. This points toward a novel mechanism of cardiac autoregulation, whereby the previously implied increased propensity for TT luminal concentration imbalances at high electrical stimulation rates would be countered by elevated advection-assisted diffusion at high mechanical beating rates. The relevance of this mechanism in health and during pathological remodeling (eg, cardiac hypertrophy or failure) forms an exciting target for further research.



Circ Res: 21 Jan 2021; 128:203-215
Rog-Zielinska EA, Scardigli M, Peyronnet R, Zgierski-Johnston CM, ... Sacconi L, Kohl P
Circ Res: 21 Jan 2021; 128:203-215 | PMID: 33228470
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Impact:
Abstract

The Molecular Basis of Predicting Atherosclerotic Cardiovascular Disease Risk.

Nayor M, Brown KJ, Vasan RS
Atherosclerotic cardiovascular disease (ASCVD) proceeds through a series of stages: initiation, progression (or regression), and complications. By integrating known biology regarding molecular signatures of each stage with recent advances in high-dimensional molecular data acquisition platforms (to assay the genome, epigenome, transcriptome, proteome, metabolome, and gut microbiome), snapshots of each phase of atherosclerotic cardiovascular disease development can be captured. In this review, we will summarize emerging approaches for assessment of atherosclerotic cardiovascular disease risk in humans using peripheral blood molecular signatures and molecular imaging approaches. We will then discuss the potential (and challenges) for these snapshots to be integrated into a personalized movie providing dynamic readouts of an individual\'s atherosclerotic cardiovascular disease risk status throughout the life course.



Circ Res: 21 Jan 2021; 128:287-303
Nayor M, Brown KJ, Vasan RS
Circ Res: 21 Jan 2021; 128:287-303 | PMID: 33476202
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Impact:

This program is still in alpha version.