IntroductionSarcoplasmic reticulum Ca2+ATPase 2a (SERCA2a) plays a critical role in regulating cardiac contraction. Changes in the levels of SERCA2a and its main regulator, phospholamban (PLN), have ...been implicated in heart failure (HF). Previous studies in small and heterogenous cohorts of HF patients have reported decreased gene expression levels of SERCA2a and PLN but findings at the protein level have been less consistent. Thus, we aimed to investigate SERCA2a and PLN protein levels in a large cohort of patients with HF.MethodsWe examined the transcript and protein levels of SERCA2a and PLN in cardiac explants obtained from 114 HF patients with dilated cardiomyopathy (DCM), 65 HF patients with ischaemic heart disease (IHD) and 57 non-failing controls. Transcript levels were assessed via quantitative polymerase chain reaction. Protein levels were measured by untargeted multiplexed quantitative proteomics using tandem mass tags (TMT). For validation, we developed a targeted proteomics assay using parallel reaction monitoring (PRM) on LV tissues from 10 patients carrying the single arginine deletion in PLN (R14del) mutation and compared to 13 non-failing controls.ResultsTranscript levels of SERCA2a were reduced in DCM patients compared to controls (FC=0.81, P=0.005), as were PLN levels (FC=0.78, P<0.001). No significant changes were observed in IHD patients. However, TMT proteomics revealed no change in SERCA2a protein levels in both groups of human HF. Similarly, PLN levels were unaltered between groups. Even in carriers of the R14del PLN mutation, SERCA2a protein levels were unaltered. The targeted PRM approach was highly specific and able to discern PLN peptides that are affected by the R14del PLN mutation. Total PLN levels were decreased in R14del patients compared to controls (FC=0.72, P=0.002). When the targeted PRM method was applied to our larger cohort of patients with end-stage HF, it confirmed the absence of significant differences in SERCA2a and PLN protein levels between patients with DCM, IHD and controls.ConclusionIn this largest proteomics analysis of cardiac tissues from patients with end-stage HF to date, we observed no changes in SERCA2a and PLN protein abundance compared to control hearts. Our findings challenge the existing paradigm that a reduction of SERCA2a is a hallmark of human HF. While similar protein abundance does not rule out altered SERCA2a activity, our findings provide a possible explanation for the failure of SERCA2a gene therapy in clinical trials.Conflict of InterestNone
Gene expression in human tissue has primarily been studied on the transcriptional level, largely neglecting translational regulation. Here, we analyze the translatomes of 80 human hearts to identify ...new translation events and quantify the effect of translational regulation. We show extensive translational control of cardiac gene expression, which is orchestrated in a process-specific manner. Translation downstream of predicted disease-causing protein-truncating variants appears to be frequent, suggesting inefficient translation termination. We identify hundreds of previously undetected microproteins, expressed from lncRNAs and circRNAs, for which we validate the protein products in vivo. The translation of microproteins is not restricted to the heart and prominent in the translatomes of human kidney and liver. We associate these microproteins with diverse cellular processes and compartments and find that many locate to the mitochondria. Importantly, dozens of microproteins are translated from lncRNAs with well-characterized noncoding functions, indicating previously unrecognized biology.
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•Ribosome profiling reveals the principles of translational control in human tissue•Ribosomes translate mRNAs downstream of protein-truncating variants•Functionally characterized lncRNAs and circRNAs produce microproteins in vivo•Microproteins can be implicated in mitochondrial and other cellular processes
Translational profiling in a primary human tissue reveals frequent translation downstream of predicted disease-causing variants as well as translation of hundreds of microproteins from long noncoding RNAs and circular RNAs.
New technologies make their way into education and one of the most prominent of them is the immersive Virtual Reality (IVR). But the scientific data about its efficiency in the educational process is ...controversial. In the present study three randomized groups of students, who did not have biological and medical classes amongst their courses, studied human heart anatomy using three different learning methods – a paper (text and images); a 3D interactive human heart model presented on a computer display; an IVR human heart model. Prior and after learning session students performed the test of human heart anatomy with 28 open questions. The IVR group showed the increase of correct answers within the group and compared with other groups. Also, the subjects with lesser baseline knowledge in IVR group showed the greater increase of correct answers following IVR session. The structure and the way how the learning material should be presented in IVR and how it affects the learning outcomes are discussed.
Hypoplastic left heart syndrome (HLHS) is a complex congenital heart disease characterized by abnormalities in the left ventricle, associated valves, and ascending aorta. Studies have shown intrinsic ...myocardial defects but do not sufficiently explain developmental defects in the endocardial-derived cardiac valve, septum, and vasculature. Here, we identify a developmentally impaired endocardial population in HLHS through single-cell RNA profiling of hiPSC-derived endocardium and human fetal heart tissue with an underdeveloped left ventricle. Intrinsic endocardial defects contribute to abnormal endothelial-to-mesenchymal transition, NOTCH signaling, and extracellular matrix organization, key factors in valve formation. Endocardial abnormalities cause reduced cardiomyocyte proliferation and maturation by disrupting fibronectin-integrin signaling, consistent with recently described de novo HLHS mutations associated with abnormal endocardial gene and fibronectin regulation. Together, these results reveal a critical role for endocardium in HLHS etiology and provide a rationale for considering endocardial function in regenerative strategies.
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•Single-cell RNA-seq identifies an abnormal endocardial population in HLHS•HLHS endocardial cells show impaired endothelial to mesenchymal transition•Endocardial defects in HLHS contribute to suppressed myocardial growth and maturation•Reduction of FN1 underlies endocardial and myocardial dysfunctions in HLHS
Gu and colleagues provide direct evidence that a developmentally impaired endocardium underlies the ventricular and valvular hypoplasia in hypoplastic left heart syndrome (HLHS) and relate recently discovered de novo mutations to the pathogenesis of this condition.
The cardiomyocyte (CM) subtypes in the mammalian heart derive from distinct lineages known as the first heart field (FHF), the anterior second heart field (aSHF), and the posterior second heart field ...(pSHF) lineages that are specified during gastrulation. We modeled human heart field development from human pluripotent stem cells (hPSCs) by using single-cell RNA-sequencing to delineate lineage specification and progression. Analyses of hPSC-derived and mouse mesoderm transcriptomes enabled the identification of distinct human FHF, aSHF, and pSHF mesoderm subpopulations. Through staged manipulation of signaling pathways identified from transcriptomics, we generated myocyte populations that display molecular characteristics of key CM subtypes. The developmental trajectory of the human cardiac lineages recapitulated that of the mouse, demonstrating conserved cardiovascular programs. These findings establish a comprehensive landscape of human embryonic cardiogenesis that provides access to a broad spectrum of cardiomyocytes for modeling congenital heart diseases and chamber-specific cardiomyopathies as well as for developing new therapies to treat them.
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•ScRNA-seq profiling of human heart field development spanning various stages•Comparisons of hPSC-derived, human fetal-, and mouse fetal-cardiac populations•Identification of pathways involved in the development of heart field lineages•Generation of functionally and transcriptionally distinct cardiomyocyte (CM) subtypes
We describe hPSC-derived models of FHF, aSHF, and pSHF development spanning the mesoderm, progenitor, and cardiomyocyte stages. Comparison between the hPSC-derived and the corresponding mouse populations revealed conserved developmental trajectories across the species. Access to cardiomyocyte subtypes derived from different heart fields enables modeling chamber-specific diseases and developing new therapies.
A major limiting factor with heart allotransplantation remains the availability of organs from deceased donors. Porcine heart xenotransplantation could serve as an alternative source of organs for ...patients with terminal heart failure. A first‐in‐human porcine xenotransplantation that occurred in January 2022 at the University of Maryland Medical Center provided an opportunity to examine several ethical issues to guide selection criteria for future xenotransplantation clinical trials. In this article, the authors, who are clinicians at UMMC, discuss the appropriate balancing of risks and benefits and the significance, if any, of clinical equipoise. The authors also review the alleged role of the psychosocial evaluation in identifying patients at an elevated risk of posttransplant noncompliance, and they consider how the evaluation's implementation might enhance inequities among diverse populations. The authors argue that, based on the principle of reciprocity, psychosocial criteria should be used, not to exclude patients, but instead to identify patients who need additional support. Finally, the authors discuss the requirements for and the proper assessment of informed and voluntary consent from patients being considered for xenotransplantation.
In this paper a highly parallel method is developed for simulating the elastodynamics of a four-chamber human heart with patient-specific geometry. The heterogeneous hyperelastic model is discretized ...by a finite element method in space and a fully implicit adaptive method in time, and the resulting nonlinear algebraic systems are solved by a scalable domain decomposition algorithm. The deformations of the cardiac muscles are quite complex due to the realistic geometry, the heterogeneous hyperelasticity of the cardiac tissue, and the myocardial fibers with active stresses. Moreover, the deformations in different chambers and at different phases of the cardiac cycle are very different. To simulate all the muscle movements including the atrial diastole, the atrial systole, the isovolumic contraction, the ventricular ejection, the isovolumic relaxation, and the ventricular filling, the temporal-spatial mesh needs to be sufficiently fine, but not too fine so that the overall computing time is manageable, we introduce a baseline mesh in space and a two-level time stepping strategy including a uniform baseline time step size to obtain the desired time accuracy and an adaptive time stepping method within a baseline time step to guarantee the convergence of the nonlinear solver. Through numerical experiments, we investigate the performance of the proposed method with respect to the material coefficients, the fiber orientations, as well as the mesh sizes and the time step sizes. For an unstructured tetrahedral mesh with more than 200 million degree of freedoms, the method scales well for up to 16,384 processor cores for all steps of an entire cardiac cycle.
•Introduce a highly parallel method for simulating elastodynamics of a human heart.•A 4-chamber patient-specific heart is modeled by heterogeneous material with fibers.•A fully unstructured finite element method is used for the complex geometry.•A nonlinear domain decomposition algorithm is developed for the algebraic systems.•A 2-level adaptive time integration scheme is introduced for accuracy and robustness.
Global bottom-up mass spectrometry (MS)-based proteomics is widely used for protein identification and quantification to achieve a comprehensive understanding of the composition, structure, and ...function of the proteome. However, traditional sample preparation methods are time-consuming, typically including overnight tryptic digestion, extensive sample cleanup to remove MS-incompatible surfactants, and offline sample fractionation to reduce proteome complexity prior to online liquid chromatography–tandem mass spectrometry (LC-MS/MS) analysis. Thus, there is a need for a fast, robust, and reproducible method for protein identification and quantification from complex proteomes. Herein, we developed an ultrafast bottom-up proteomics method enabled by Azo, a photocleavable, MS-compatible surfactant that effectively solubilizes proteins and promotes rapid tryptic digestion, combined with the Bruker timsTOF Pro, which enables deeper proteome coverage through trapped ion mobility spectrometry (TIMS) and parallel accumulation–serial fragmentation (PASEF) of peptides. We applied this method to analyze the complex human cardiac proteome and identified nearly 4000 protein groups from as little as 1 mg of human heart tissue in a single one-dimensional LC-TIMS-MS/MS run with high reproducibility. Overall, we anticipate this ultrafast, robust, and reproducible bottom-up method empowered by both Azo and the timsTOF Pro will be generally applicable and greatly accelerate the throughput of large-scale quantitative proteomic studies. Raw data are available via the MassIVE repository with identifier MSV000087476.
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Ventricular tachycardia is the leading cause of sudden arrhythmic death in the U.S. Recently, the moderate IK1 channel activator, zacopride, was shown to suppress triggered ...ventricular tachycardia in rats. Nonetheless, concerns were raised about the possibility of pro-arrhythmic activity after IK1 channel stimulation based on the promising anti-arrhythmic strategy of IK1 blockade in other animal models. Therefore, the goal of the current study was to investigate the ex-vivo effects of zacopride on triggered arrhythmia and contractility in ventricular human myocardium in order to validate data that was solely obtained from animal models. Application of 100nmol/L isoproterenol and 0.5mmol/L caffeine led to triggered arrhythmia in isolated cardiac muscles from non-failing and end-stage failing hearts. However, the occurrence of arrhythmia in muscles of non-failing hearts was markedly higher than those of end-stage failing hearts. Interestingly, zacopride eliminated the ex-vivo triggered arrhythmia in these muscles of non-failing and failing hearts in a concentration-dependent manner, with an effective IC50 in the range of 28–40μmol/L. Conversely, in the absence of isoproterenol/caffeine, zacopride led to a negative inotropic effect in a concentration-dependent manner. Reduced cardiac contraction was clearly observed at high zacopride concentration of 200μmol/L, along with the occurrence of contractile alternans in muscles of non-failing and failing hearts. Zacopride shows promising antiarrhythmic effects against triggered arrhythmia in human ventricular myocardium. However, in the absence of Ca2+ overload/arrhythmia, zacopride, albeit at high concentrations, decreases the force of contraction and increases the likelihood of occurrence of contractile alternans, which may predispose the heart to contractile dysfunction and/or arrhythmia. Overall, our results represent a key step in translating this drug from the benchtop to the bedside in the research area.
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