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Single-Nucleus Sequencing of an Entire Mammalian Heart: Cell Type Composition and Velocity

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Single-Nucleus Sequencing of an Entire Mammalian Heart: Cell Type Composition and Velocity

Markus Wolfien et al. Cells.

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.

Keywords: RNA velocity; cardiomyocytes; cluster analysis; seurat; snRNA-seq.

Conflict of interest statement

The authors declare no conflict of interest. The funders were not involved in study design, data collection and interpretation, and manuscript preparation.

Figures

Figure 1
Figure 1
Single-nucleus transcriptome characteristics of pooled whole Fzt:DU mice hearts (n = 4). UMAP clustering of snRNA-seq data (8635 nuclei) reveals 24 distinct clusters for the indicated cell types. The arrows represent RNA velocity kinetics visualizing the direction and acceleration between mature and nascent mRNA. The percentages represent the nuclei ratio.
Figure 2
Figure 2
Dot-plot representation of the gene expression marker genes for the identified cell types. The size of dots represents the relative gene expression in percent for each cluster, e.g., a value of 100 means that each cell within this cell type expressed this gene. The color indicates the average expression level for the indicated gene per cell type. The color of the clusters is taken from Figure 1. A dot plot for the most significant gene per cluster as well as an extended visualization of the top 10 markers per cluster can be obtained at our FairdomHub/iRhythmics instance.

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