Single Nucleus RNA-sequencing Reveals Altered Intercellular Communication and Dendritic Cell Activation in Nonobstructive Hypertrophic Cardiomyopathy

Abstract

End stage, nonobstructive hypertrophic cardiomyopathy (HCM) is an intractable condition with no disease-specific therapies. To gain insights into the pathogenesis of nonobstructive HCM, we performed single nucleus RNA-sequencing (snRNA-seq) on human HCM hearts explanted at the time of cardiac transplantation and organ donor hearts serving as controls. Differential gene expression analysis revealed 64 differentially expressed genes linked to specific cell types and molecular functions. Analysis of ligand-receptor pair gene expression to delineate potential intercellular communication revealed significant reductions in expressed ligand-receptor pairs likely affecting the extracellular matrix, growth factor binding, peptidase regulator activity, platelet-derived growth factor binding and protease binding in the HCM tissue. Changes in Integrin-β1 receptor expression were responsible for many observed changes related to extracellular matrix interactions, by increasing in dendritic, smooth muscle and pericyte cells while decreasing in endothelial and fibroblast cells, suggesting potential mechanisms for fibrosis and microvascular disease in HCM and a potential role for dendritic cells. In contrast, there was an increase in ligand-receptor pair expression associated with adenylate cyclase binding, calcium channel molecular functions, channel inhibitor activity, ion channel inhibitor activity, phosphatase activator activity, protein kinase activator activity and titin binding, suggesting important shifts in various signaling cascades in nonobstructive, end stage HCM.

Single Cell Transcriptomic Analysis in a Mouse Model of Barth Syndrome Reveals Cell-Specific Alterations in Gene Expression and Intercellular Communication

“…Given the sensitivity of this cell-cell communication analysis to differences in the product of a ligand and receptor's expression across many cell type pairs, the contrast in the number of identified dysregulated L-R pairs compared to the number of identified DEGs per individual cell type may be expected. The predicted loss of cell-cell communication has also been noted in other conditions such as hypertrophic cardiomyopathy [17,19,20].…”

“…Whole hearts from four WT and four Taz-KO mice were minced, cryopreserved, and then processed for snRNA-seq library generation and next generation sequencing as described previously [17][18][19][20] using commercially available reagents (10x Genomics). The Tafazzin-knockout mouse used in this study has been described previously [22,34].…”

“…The clustering resolution of 0.9 was chosen using the subsampling-based approach ChooseR [42]. The expression of known cell-specific gene markers was used to identify major cell types, as has been done previously [14,[16][17][18][19]. To further refine cell assignments, such as distinguishing specific myeloid or lymphoid cell populations, Seurat's FindSubCluster function was utilized [26][27][28][29].…”

“…Single-cell RNA-sequencing methods have facilitated the analysis of cell-specific gene expression, cellular diversity and intercellular communication in a wide variety of tissues and across a wide variety of species-including human and mouse hearts-for both normal and diseased conditions [12][13][14][15][16][17][18][19][20][21]. As a first step in identifying transcriptional networks and pathological pathways that mediate the effects of Tafazzin deficiency in the heart at the single-cell level, we have performed single-nuclei transcriptomics on heart tissue extracted from a Tafazzin knockout (Taz-KO) mouse model of Barth Syndrome [22].…”

Spatial Transcriptomic Analysis of Focal and Normal Areas of Myocyte Disarray in Human Hypertrophic Cardiomyopathy

“…Since sarcomere gene mutation-negative patients have similar phenotypes to sarcomere gene mutation-positive patients, it is likely that there are final common pathological pathways independent of sarcomere gene mutations that are involved, but these final common pathways are incompletely understood. Recent reports using single-nucleus RNA sequencing of human HCM tissue have identified potential alterations in cell-to-cell communication involving extracellular matrix proteins, integrin receptors and the activation of immune cells as potential contributors to the HCM phenotype [ 4 , 5 , 6 ]. These studies, however, did not determine how these alterations in single-cell transcription and intercellular communication are spatially organized in the context of known histopathological features of HCM.…”