Background The failing human heart is characterized by metabolic abnormalities, but these defects remains incompletely understood. In animal models of HF there is a switch from a predominance of fatty acid utilization to the more oxygen-sparing carbohydrate metabolism. Recent studies have reported decreases in myocardial lipid content, but inclusion of diabetics and nondiabetics obscures the distinction of adapations to metabolic derangements from adaptations to heart failure per se. Methods and Results We performed both unbiased and targeted myocardial lipid surveys using liquid chromatography-mass spectroscopy in non-diabetic, lean, predominantly non-ischemic advanced HF patients at the time of heart transplantation or LVAD implantation. We identified significantly decreased concentrations of the majority of myocardial lipid intermediates, including long-chain acylcarnitines, the primary subset of energetic lipid substrate for mitochondrial fatty acid oxidation. We report for the first time significantly reduced levels of intermediate and anaplerotic acyl-CoA species incorporated into Krebs cycle, while the myocardial concentration of acetyl-CoA was significantly increased in end-stage heart failure. In contrast, we observed an increased abundance of ketogenic β-hydroxybutyryl CoA, in association with increased myocardial utilization of β-hydroxybutyrate. We observed a significant increase in the expression of the gene encoding succinyl-CoA: 3oxoacid-CoA transferase (SCOT), the rate limiting enzyme for myocardial oxidation of βOHB and acetoacetate. Conclusions These findings indicate increased ketone utilization in the severely failing human heart independent of diabetes, support the role of ketone bodies as an alternative fuel and myocardial ketone oxidation as a key metabolic adaptation in the failing human heart.

Background: The human heart requires a complex ensemble of specialized cell types to perform its essential function. A greater knowledge of the intricate cellular milieu of the heart is critical to increase our understanding of cardiac homeostasis and pathology. As recent advances in low input RNA-sequencing have allowed definitions of cellular transcriptomes at single cell resolution at scale, here we have applied these approaches to assess the cellular and transcriptional diversity of the non-failing human heart. Methods: Microfluidic encapsulation and barcoding was used to perform single nuclear RNA sequencing with samples from seven human donors, selected for their absence of overt cardiac disease. Individual nuclear transcriptomes were then clustered based upon transcriptional profiles of highly variable genes. These clusters were used as the basis for between-chamber and between-sex differential gene expression analyses and intersection with genetic and pharmacologic data. Results: We sequenced the transcriptomes of 287,269 single cardiac nuclei, revealing a total of 9 major cell types and 20 subclusters of cell types within the human heart. Cellular subclasses include two distinct groups of resident macrophages, four endothelial subtypes, and two fibroblasts subsets. Comparisons of cellular transcriptomes by cardiac chamber or sex reveal diversity not only in cardiomyocyte transcriptional programs, but also in subtypes involved in extracellular matrix remodeling and vascularization. Using genetic association data, we identified strong enrichment for the role of cell subtypes in cardiac traits and diseases. Finally, intersection of our dataset with genes on cardiac clinical testing panels and the druggable genome reveals striking patterns of cellular specificity. Conclusions: Using large-scale single nuclei RNA sequencing, we have defined the transcriptional and cellular diversity in the normal human heart. Our identification of discrete cell subtypes and differentially expressed genes within the heart will ultimately facilitate the development of new therapeutics for cardiovascular diseases.

Fibrosis is observed in nearly every form of myocardial disease 1. Upon injury, cardiac fibroblasts (CF) in the heart begin to remodel the myocardium via extracellular matrix deposition, resulting in increased tissue stiffness and reduced compliance. Excessive cardiac fibrosis is an important factor in the progression of various forms of cardiac disease and heart failure 2. However, clinical interventions and therapies targeting fibrosis remain limited 3. In this study, we demonstrate the efficacy of redirected T-cell immunotherapy to specifically target pathologic cardiac fibrosis. We find that cardiac fibroblasts expressing a xenogeneic antigen can be effectively targeted and ablated by adoptive transfer of antigen-specific CD8 + T cells. Through expression analysis of cardiac fibroblast gene signatures from healthy versus diseased human hearts, we identified an endogenous CF target; fibroblast activation protein (FAP). Adoptive transfer of T cells expressing a chimeric antigen receptor (CAR) against FAP, results in a significant reduction in cardiac fibrosis and restoration of function after injury in mice. These results provide the proof-of-principle basis for a novel immunotherapeutic avenue for the treatment of cardiac disease.

Detyrosinated microtubules (MTs) provide mechanical resistance that can impede the motion of contracting cardiomyocytes. However, the functional effects of MT detyrosination in heart failure or in human hearts have not previously been studied. Here we utilize mass spectrometry and single-myocyte mechanical assays to characterize changes to the cardiomyocyte cytoskeleton and their functional consequences in human heart failure. Proteomic analysis of left ventricle tissue reveals a consistent upregulation and stabilization of intermediate filaments and MTs in failing human hearts. As revealed by super-resolution imaging, failing cardiomyocytes are characterized by a dense, heavily detyrosinated MT network, which is associated with increased myocyte stiffness and impaired contractility. Pharmacological suppression of detyrosinated MTs lowers the viscoelasticity of failing myocytes and restores 40–50% of lost contractile function; reduction of MT detyrosination using a genetic approach also softens cardiomyocytes and improves contractile kinetics. Together, these data demonstrate that a modified cytoskeletal network impedes contractile function in cardiomyocytes from failing human hearts and that targeting detyrosinated MTs could represent a new inotropic strategy for improving cardiac function.