Basic Research in Cardiology最新文献

The ambiguous results of multiple CD34⁺ cell-based therapeutic trials for patients with heart disease have halted the large-scale application of stem/progenitor cell treatment. This study aimed to delineate the biological functions of heterogenous CD34⁺ cell populations and investigate the net effect of CD34⁺ cell intervention on cardiac remodeling. We confirmed, by combining single-cell RNA sequencing on human and mouse ischemic hearts and an inducible Cd34 lineage-tracing mouse model, that Cd34⁺ cells mainly contributed to the commitment of mesenchymal cells, endothelial cells (ECs), and monocytes/macrophages during heart remodeling with distinct pathological functions. The Cd34⁺-lineage-activated mesenchymal cells were responsible for cardiac fibrosis, while CD34⁺Sca-1^high was an active precursor and intercellular player that facilitated Cd34⁺-lineage angiogenic EC-induced postinjury vessel development. We found through bone marrow transplantation that bone marrow-derived CD34⁺ cells only accounted for inflammatory response. We confirmed using a Cd34-CreER^T2; R26-DTA mouse model that the depletion of Cd34⁺ cells could alleviate the severity of ventricular fibrosis after ischemia/reperfusion (I/R) injury with improved cardiac function. This study provided a transcriptional and cellular landscape of CD34⁺ cells in normal and ischemic hearts and illustrated that the heterogeneous population of Cd34⁺ cell-derived cells served as crucial contributors to cardiac remodeling and function after the I/R injury, with their capacity to generate diverse cellular lineages.

The number of "omics" approaches is continuously growing. Among others, epigenetics has appeared as an attractive area of investigation by the cardiovascular research community, notably considering its association with disease development. Complex diseases such as cardiovascular diseases have to be tackled using methods integrating different omics levels, so called "multi-omics" approaches. These approaches combine and co-analyze different levels of disease regulation. In this review, we present and discuss the role of epigenetic mechanisms in regulating gene expression and provide an integrated view of how these mechanisms are interlinked and regulate the development of cardiac disease, with a particular attention to heart failure. We focus on DNA, histone, and RNA modifications, and discuss the current methods and tools used for data integration and analysis. Enhancing the knowledge of these regulatory mechanisms may lead to novel therapeutic approaches and biomarkers for precision healthcare and improved clinical outcomes.

Calcium transfer into the mitochondrial matrix during sarcoplasmic reticulum (SR) Ca²⁺ release is essential to boost energy production in ventricular cardiomyocytes (VCMs) and match increased metabolic demand. Mitochondria from female hearts exhibit lower mito-[Ca²⁺] and produce less reactive oxygen species (ROS) compared to males, without change in respiration capacity. We hypothesized that in female VCMs, more efficient electron transport chain (ETC) organization into supercomplexes offsets the deficit in mito-Ca²⁺ accumulation, thereby reducing ROS production and stress-induced intracellular Ca²⁺ mishandling. Experiments using mitochondria-targeted biosensors confirmed lower mito-ROS and mito-[Ca²⁺] in female rat VCMs challenged with β-adrenergic agonist isoproterenol compared to males. Biochemical studies revealed decreased mitochondria Ca²⁺ uniporter expression and increased supercomplex assembly in rat and human female ventricular tissues vs male. Importantly, western blot analysis showed higher expression levels of COX7RP, an estrogen-dependent supercomplex assembly factor in female heart tissues vs males. Furthermore, COX7RP was decreased in hearts from aged and ovariectomized female rats. COX7RP overexpression in male VCMs increased mitochondrial supercomplexes, reduced mito-ROS and spontaneous SR Ca²⁺ release in response to ISO. Conversely, shRNA-mediated knockdown of COX7RP in female VCMs reduced supercomplexes and increased mito-ROS, promoting intracellular Ca²⁺ mishandling. Compared to males, mitochondria in female VCMs exhibit higher ETC subunit incorporation into supercomplexes, supporting more efficient electron transport. Such organization coupled to lower levels of mito-[Ca²⁺] limits mito-ROS under stress conditions and lowers propensity to pro-arrhythmic spontaneous SR Ca²⁺ release. We conclude that sexual dimorphism in mito-Ca²⁺ handling and ETC organization may contribute to cardioprotection in healthy premenopausal females.

Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are increasingly used for personalised medicine and preclinical cardiotoxicity testing. Reports on hiPSC-CM commonly describe heterogenous functional readouts and underdeveloped or immature phenotypical properties. Cost-effective, fully defined monolayer culture is approaching mainstream adoption; however, the optimal age at which to utilise hiPSC-CM is unknown. In this study, we identify, track and model the dynamic developmental behaviour of key ionic currents and Ca²⁺-handling properties in hiPSC-CM over long-term culture (30-80 days). hiPSC-CMs > 50 days post differentiation show significantly larger I_Ca,L density along with an increased I_Ca,L-triggered Ca²⁺-transient. I_Na and I_K1 densities significantly increase in late-stage cells, contributing to increased upstroke velocity and reduced action potential duration, respectively. Importantly, our in silico model of hiPSC-CM electrophysiological age dependence confirmed I_K1 as the key ionic determinant of action potential shortening in older cells. We have made this model available through an open source software interface that easily allows users to simulate hiPSC-CM electrophysiology and Ca²⁺-handling and select the appropriate age range for their parameter of interest. This tool, together with the insights from our comprehensive experimental characterisation, could be useful in future optimisation of the culture-to-characterisation pipeline in the field of hiPSC-CM research.

The coronary circulation has an innate ability to maintain constant blood flow over a wide range of perfusion pressures. However, the mechanisms responsible for coronary autoregulation remain a fundamental and highly contested question. This study interrogated the local metabolic hypothesis of autoregulation by testing the hypothesis that hypoxemia-induced exaggeration of the metabolic error signal improves the autoregulatory response. Experiments were performed on open-chest anesthetized swine during stepwise changes in coronary perfusion pressure (CPP) from 140 to 40 mmHg under normoxic (n = 15) and hypoxemic (n = 8) conditions, in the absence and presence of dobutamine-induced increases in myocardial oxygen consumption (MVO₂) (n = 5-7). Hypoxemia (PaO₂ < 40 mmHg) decreased coronary venous PO₂ (CvPO₂) ~ 30% (P < 0.001) and increased coronary blood flow ~ 100% (P < 0.001), sufficient to maintain myocardial oxygen delivery (P = 0.14) over a wide range of CPPs. Autoregulatory responsiveness during hypoxemia-induced reductions in CvPO₂ were associated with increases of autoregulatory gain (Gc; P = 0.033) but not slope (P = 0.585) over a CPP range of 120 to 60 mmHg. Preservation of autoregulatory Gc (P = 0.069) and slope (P = 0.264) was observed during dobutamine administration ± hypoxemia. Reductions in coronary resistance in response to decreases in CPP predominantly occurred below CvPO₂ values of ~ 25 mmHg, irrespective of underlying vasomotor reserve. These findings support the presence of an autoregulatory threshold under which oxygen-sensing pathway(s) act to preserve sufficient myocardial oxygen delivery as CPP is reduced during increases in MVO₂ and/or reductions in arterial oxygen content.

Coronary microvascular dysfunction (CMD) is associated with cardiac dysfunction and predictive of cardiac mortality in obesity, especially in females. Clinical data further support that CMD associates with development of heart failure with preserved ejection fraction and that mineralocorticoid receptor (MR) antagonism may be more efficacious in obese female, versus male, HFpEF patients. Accordingly, we examined the impact of smooth muscle cell (SMC)-specific MR deletion on obesity-associated coronary and cardiac diastolic dysfunction in female mice. Obesity was induced in female mice via western diet (WD) feeding alongside littermates fed standard diet. Global MR blockade with spironolactone prevented coronary and cardiac dysfunction in obese females and specific deletion of SMC-MR was sufficient to prevent obesity-associated coronary and cardiac diastolic dysfunction. Cardiac gene expression profiling suggested reduced cardiac inflammation in WD-fed mice with SMC-MR deletion independent of blood pressure, aortic stiffening, and cardiac hypertrophy. Further mechanistic studies utilizing single-cell RNA sequencing of non-cardiomyocyte cell populations revealed novel impacts of SMC-MR deletion on the cardiac cellulome in obese mice. Specifically, WD feeding induced inflammatory gene signatures in non-myocyte populations including B/T cells, macrophages, and endothelium as well as increased coronary VCAM-1 protein expression, independent of cardiac fibrosis, that was prevented by SMC-MR deletion. Further, SMC-MR deletion induced a basal reduction in cardiac mast cells and prevented WD-induced cardiac pro-inflammatory chemokine expression and leukocyte recruitment. These data reveal a central role for SMC-MR signaling in obesity-associated coronary and cardiac dysfunction, thus supporting the emerging paradigm of a vascular origin of cardiac dysfunction in obesity.

The prospective use of human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) for cardiac regenerative medicine strongly depends on the electro-mechanical properties of these cells, especially regarding the Ca²⁺-dependent excitation-contraction (EC) coupling mechanism. Currently, the immature structural and functional features of hiPSC-CM limit the progression towards clinical applications. Here, we show that a specific microarchitecture is essential for functional maturation of hiPSC-CM. Structural remodelling towards a cuboid cell shape and induction of BIN1, a facilitator of membrane invaginations, lead to transverse (t)-tubule-like structures. This transformation brings two Ca²⁺ channels critical for EC coupling in close proximity, the L-type Ca²⁺ channel at the sarcolemma and the ryanodine receptor at the sarcoplasmic reticulum. Consequently, the Ca²⁺-dependent functional interaction of these channels becomes more efficient, leading to improved spatio-temporal synchronisation of Ca²⁺ transients and higher EC coupling gain. Thus, functional maturation of hiPSC-cardiomyocytes by optimised cell microarchitecture needs to be considered for future cardiac regenerative approaches.

Precision-based molecular phenotyping of heart failure must overcome limited access to cardiac tissue. Although epigenetic alterations have been found to underlie pathological cardiac gene dysregulation, the clinical utility of myocardial epigenomics remains narrow owing to limited clinical access to tissue. Therefore, the current study determined whether patient plasma confers indirect phenotypic, transcriptional, and/or epigenetic alterations to ex vivo cardiomyocytes to mirror the failing human myocardium. Neonatal rat ventricular myocytes (NRVMs) and single-origin human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) and were treated with blood plasma samples from patients with dilated cardiomyopathy (DCM) and donor subjects lacking history of cardiovascular disease. Following plasma treatments, NRVMs and hiPSC-CMs underwent significant hypertrophy relative to non-failing controls, as determined via automated high-content screening. Array-based DNA methylation analysis of plasma-treated hiPSC-CMs and cardiac biopsies uncovered robust, and conserved, alterations in cardiac DNA methylation, from which 100 sites were validated using an independent cohort. Among the CpG sites identified, hypo-methylation of the ATG promoter was identified as a diagnostic marker of HF, wherein cg03800765 methylation (AUC = 0.986, P < 0.0001) was found to out-perform circulating NT-proBNP levels in differentiating heart failure. Taken together, these findings support a novel approach of indirect epigenetic testing in human HF.

A modern-day physician is faced with a vast abundance of clinical and scientific data, by far surpassing the capabilities of the human mind. Until the last decade, advances in data availability have not been accompanied by analytical approaches. The advent of machine learning (ML) algorithms might improve the interpretation of complex data and should help to translate the near endless amount of data into clinical decision-making. ML has become part of our everyday practice and might even further change modern-day medicine. It is important to acknowledge the role of ML in prognosis prediction of cardiovascular disease. The present review aims on preparing the modern physician and researcher for the challenges that ML might bring, explaining basic concepts but also caveats that might arise when using these methods. Further, a brief overview of current established classical and emerging concepts of ML disease prediction in the fields of omics, imaging and basic science is presented.

Whereas cardiomyocytes (CMs) in the fetal heart divide, postnatal CMs fail to undergo karyokinesis and/or cytokinesis and therefore become polyploid or binucleated, a key process in terminal CM differentiation. This switch from a diploid proliferative CM to a terminally differentiated polyploid CM remains an enigma and seems an obstacle for heart regeneration. Here, we set out to identify the transcriptional landscape of CMs around birth using single cell RNA sequencing (scRNA-seq) to predict transcription factors (TFs) involved in CM proliferation and terminal differentiation. To this end, we established an approach combining fluorescence activated cell sorting (FACS) with scRNA-seq of fixed CMs from developing (E16.5, P1, and P5) mouse hearts, and generated high-resolution single-cell transcriptomic maps of in vivo diploid and tetraploid CMs, increasing the CM resolution. We identified TF-networks regulating the G2/M phases of developing CMs around birth. ZEB1 (Zinc Finger E-Box Binding Homeobox 1), a hereto unknown TF in CM cell cycling, was found to regulate the highest number of cell cycle genes in cycling CMs at E16.5 but was downregulated around birth. CM ZEB1-knockdown reduced proliferation of E16.5 CMs, while ZEB1 overexpression at P0 after birth resulted in CM endoreplication. These data thus provide a ploidy stratified transcriptomic map of developing CMs and bring new insight to CM proliferation and endoreplication identifying ZEB1 as a key player in these processes.