Sara Samari

Cardiovascular disease is the leading cause of death worldwide. Advanced insights into disease mechanisms and therapeutic strategies require a deeper understanding of the molecular processes involved in the healthy heart. Knowledge of the full repertoire of cardiac cells and their gene expression profiles is a fundamental first step in this endeavour. Here, using state-of-the-art analyses of large-scale single-cell and single-nucleus transcriptomes, we characterize six anatomical adult heart regions. Our results highlight the cellular heterogeneity of cardiomyocytes, pericytes and fibroblasts, and reveal distinct atrial and ventricular subsets of cells with diverse developmental origins and specialized properties. We define the complexity of the cardiac vasculature and its changes along the arterio-venous axis. In the immune compartment, we identify cardiac-resident macrophages with inflammatory and protective transcriptional signatures. Furthermore, analyses of cell-to-cell interactions highlight different networks of macrophages, fibroblasts and cardiomyocytes between atria and ventricles that are distinct from those of skeletal muscle. Our human cardiac cell atlas improves our understanding of the human heart and provides a valuable reference for future studies.

Pathogenic variants in genes that cause dilated cardiomyopathy (DCM) and arrhythmogenic cardiomyopathy (ACM) convey high risks for the development of heart failure through unknown mechanisms. Using single-nucleus RNA sequencing, we characterized the transcriptome of 880,000 nuclei from 18 control and 61 failing, nonischemic human hearts with pathogenic variants in DCM and ACM genes or idiopathic disease. We performed genotype-stratified analyses of the ventricular cell lineages and transcriptional states. The resultant DCM and ACM ventricular cell atlas demonstrated distinct right and left ventricular responses, highlighting genotype-associated pathways, intercellular interactions, and differential gene expression at single-cell resolution. Together, these data illuminate both shared and distinct cellular and molecular architectures of human heart failure and suggest candidate therapeutic targets.

Summary Cardiovascular disease is the leading cause of death worldwide. Advanced insights into disease mechanisms and strategies to improve therapeutic opportunities require deeper understanding of the molecular processes of the normal heart. Knowledge of the full repertoire of cardiac cells and their gene expression profiles is a fundamental first step in this endeavor. Here, using large-scale single cell and nuclei transcriptomic profiling together with state-of-the-art analytical techniques, we characterise the adult human heart cellular landscape covering six anatomical cardiac regions (left and right atria and ventricles, apex and interventricular septum). Our results highlight the cellular heterogeneity of cardiomyocytes, pericytes and fibroblasts, revealing distinct subsets in the atria and ventricles indicative of diverse developmental origins and specialized properties. Further we define the complexity of the cardiac vascular network which includes clusters of arterial, capillary, venous, lymphatic endothelial cells and an atrial-enriched population. By comparing cardiac cells to skeletal muscle and kidney, we identify cardiac tissue resident macrophage subsets with transcriptional signatures indicative of both inflammatory and reparative phenotypes. Further, inference of cell-cell interactions highlight a macrophage-fibroblast-cardiomyocyte network that differs between atria and ventricles, and compared to skeletal muscle. We expect this reference human cardiac cell atlas to advance mechanistic studies of heart homeostasis and disease.

The recent development of single cell gene expression technologies, and especially single cell transcriptomics, have revolutionized the way biologists and clinicians investigate organs and organisms, allowing an unprecedented level of resolution to the description of cell demographics in both healthy and diseased states. Single cell transcriptomics provide information on prevalence, heterogeneity, and gene co-expression at the individual cell level. This enables a cell-centric outlook to define intracellular gene regulatory networks and to bridge toward the definition of intercellular pathways otherwise masked in bulk analysis. The technologies have developed at a fast pace producing a multitude of different approaches, with several alternatives to choose from at any step, including single cell isolation and capturing, lysis, RNA reverse transcription and cDNA amplification, library preparation, sequencing, and computational analyses. Here, we provide guidelines for the experimental design of single cell RNA sequencing experiments, exploring the current options for the crucial steps. Furthermore, we provide a complete overview of the typical data analysis workflow, from handling the raw sequencing data to making biological inferences. Significantly, advancements in single cell transcriptomics have already contributed to outstanding exploratory and functional studies of cardiac development and disease models, as summarized in this review. In conclusion, we discuss achievable outcomes of single cell transcriptomics' applications in addressing unanswered questions and influencing future cardiac clinical applications.

Ischemic heart disease remains the foremost cause of death globally, with survivors at risk for subsequent heart failure. Paradoxically, cell therapies to offset cardiomyocyte loss after ischemic injury improve long-term cardiac function despite a lack of durable engraftment. An evolving consensus, inferred preponderantly from non-human models, is that transplanted cells benefit the heart via early paracrine signals. Here, we tested the impact of paracrine signals on human cardiomyocytes, using human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) as the target of mouse and human cardiac mesenchymal stromal cells (cMSC) with progenitor-like features. In co-culture and conditioned medium studies, cMSCs markedly inhibited human cardiomyocyte death. Little or no protection was conferred by mouse tail tip or human skin fibroblasts. Consistent with the results of transcriptomic profiling, functional analyses showed that the cMSC secretome suppressed apoptosis and preserved cardiac mitochondrial transmembrane potential. Protection was independent of exosomes under the conditions tested. In mice, injecting cMSC-conditioned media into the infarct border zone reduced apoptotic cardiomyocytes > 70% locally. Thus, hPSC-CMs provide an auspicious, relevant human platform to investigate extracellular signals for cardiac muscle survival, substantiating human cardioprotection by cMSCs, and suggesting the cMSC secretome or its components as potential cell-free therapeutic products.