Abstract
Analyses on the cellular level are indispensable to expand our understanding of complex tissues like the mammalian heart. Single-nucleus sequencing (snRNA-seq) allows for the exploration of cellular composition and cell features without major hurdles of single-cell sequencing. We used snRNA-seq to investigate for the first time an entire adult mammalian heart. Single-nucleus quantification and clustering led to an accurate representation of cell types, revealing 24 distinct clusters with endothelial cells (28.8%), fibroblasts (25.3%), and cardiomyocytes (22.8%) constituting the major cell populations. An additional RNA velocity analysis allowed us to study transcription kinetics and was utilized to visualize the transitions between mature and nascent cellular states of the cell types. We identified subgroups of cardiomyocytes with distinct marker profiles. For example, the expression of Hand2os1 distinguished immature cardiomyocytes from differentiated cardiomyocyte populations. Moreover, we found a cell population that comprises endothelial markers as well as markers clearly related to cardiomyocyte function. Our velocity data support the idea that this population is in a trans-differentiation process from an endothelial cell-like phenotype towards a cardiomyocyte-like phenotype. In summary, we present the first report of sequencing an entire adult mammalian heart, providing realistic cell-type distributions combined with RNA velocity kinetics hinting at interrelations.
Highlights
Single-cell sequencing allows for an in-depth characterization of complex tissues and their cell types [1]
The largest attributed to populations of endothelial cells (28.8%), fibroblasts (25.3%), and cardiomyocytes (22.8%)
Clusters can be attributed to populations of endothelial cells (28.8%), fibroblasts (25.3%), and containing ~2500, ~2200, and ~2000 nuclei, respectively
Summary
Single-cell sequencing allows for an in-depth characterization of complex tissues and their cell types [1]. (i) the difficulty of dissociating the adult mammalian heart tissue without damaging constituent cells and (ii) technical limitations regarding cell capture techniques leading to an underrepresentation of individual cell types (i.e., cardiomyocytes) due to their large cell size and irregular shape [2]. Ribonucleic acid sequencing (RNA-seq) and instead conducted single-nucleus RNA-seq (snRNA-seq), which has been shown to present similar transcriptomic results [3]. We present the first snRNA-seq analysis of an entire adult mammalian heart. A method was established to predict even future states of individual cells using single-cell or single-nucleus data. The RNA velocity analysis of our snRNA-seq data allowed us to study transcription kinetics and revealed details about the dynamics and interconnectedness of our identified cell clusters
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