Heart Failure with preserved ejection fraction (HFpEF) is a syndrome with high mortality and an increasing incidence, which is caused by various etiologies and molecular mechanisms. Due to the complex nature of the disease, therapies lowering mortality rates remain unavailable. Hence, there is an urgent need for a better understanding of the cellular disease mechanisms to elucidate therapeutic targets in HFpEF. However, the distinct roles of myocardial cell populations and their contribution to disease progression remain largely elusive.
Here, we performed single-nucleus RNA sequencing from endomyocardial biopsies obtained from six patients with HFpEF without an overt underlying disease-driving pathology and used a publicly available dataset from healthy donors as controls. The patients showed typical risk factors for HFpEF, including hypertension (83%) and diabetes mellitus (33%). All major myocardial cell populations were annotated, including cardiomyocytes (CM), endothelial cells (EC), fibroblasts (FB), and macrophages (MΦ).
Differential gene expression was analyzed using the MAST hurdle model incorporating a random effect to reduce the risk of pseudoreplication bias. Gene expression patterns were analyzed using scoring approaches, enrichment analyses and interaction analyses using CellChat DB.
The strongest transcriptomic alterations were observed in CM. The differentially expressed genes (DEG) in CM suggest a major metabolic shift characterised by the downregulation of genes involved in mitochondrial function and aerobic respiration. This was accompanied by an upregulation in Rho/GTPase pathway genes, which might associate with increased cellular stiffness. Validating this finding, ROCK1, a canonical downstream-effector of Rho/GTPase signaling was upregulated as indicated by immunofluorescence stainings.
Surprisingly, ECs from HFpEF myocardium showed signs of increased angiogenesis but also augmented anti-angiogenic Semaphorin 3 signaling, potentially reflecting compensatory mechanisms involved in the regulation of vascularization in HFpEF.
In MΦ, we observed an overall pro-inflammatory phenotype. Whereas there was no shift in the ratio of tissue resident to bone marrow-derived MΦ, we observed a robust upregulation of multiple MHC class II (MHC-II) genes, their upstream regulators (i.e. CIITA) and processing components, while MHC class I molecules was unaltered. The expression of MHC-II genes correlated with the expression of IFNγ response genes with MHC-IIhigh MΦ exhibited a particularly pro-inflammatory phenotype.
This study presents the first snRNA-Seq analysis of human HFpEF myocardium, uncovering cell type–specific transcriptomic signatures linked to metabolic dysfunction, vascular remodeling, and inflammation. The data confirm key hallmarks of HFpEF, including metabolic and inflammatory alterations, while revealing novel mechanisms such as enhanced interferon-γ signaling in immune cells and opposing angiogenic pathways in endothelial cells. These findings provide new insight into the cellular mechanisms driving HFpEF and highlight potential therapeutic targets.
