Location and aetiology are determinants of fibroblast activation and heterogeneity in the failing human heart

BACKGROUND: Cardiac fibrosis is a key feature of pathological cardiac remodelling that significantly impacts heart function through contributing to stiffness, diastolic dysfunction, and arrhythmias, ultimately leading to heart failure (HF). Despite extensive research into fibrosis-related matrix alterations, therapeutic advancements are limited, in part owing to the different nature (reparative vs interstitial) and tissue distribution of fibrosis involved. To identify unique features of fibrosis phenotypes, we investigated fibroblast (FB) heterogeneity and spatial distribution in left ventricular myocardium in HF patients with ischemic (ICM) and dilated cardiomyopathy (DCM). Infarct scar was also analysed. METHODS: We performed single-nucleus RNA sequencing of 20 human left ventricular tissue samples: from non-failing, NF (N = 4), DCM (N = 6) and ICM (N = 5) hearts, and from the ICM scar region (N = 5). The data was subjected to bioinformatic analysis, included clustering, differential expression, ligand-receptor inference, and pseudotime trajectory mapping to delineate FB transitions and regional fibrosis signatures. To identify localisations of FB states and cellular neighbourhoods, data was integrated with publicly available spatial transcriptomics datasets. RESULTS: We identified distinct FB subpopulations across failing and non-failing hearts. Resident FB states showed preferential perivascular and interstitial distribution in NF and exhibited significant depletion in HF, giving rise to different disease states. We identified shared and unique activation ligands driving the onset of FB transitions as well as transcriptional differences between scar and interstitial fibrosis, and between ICM and DCM interstitial fibrosis. Trajectory analysis revealed distinct differentiation pathways for FB depending on its originating resident FB, with specific transcription factors guiding each transition. CONCLUSIONS: These findings provide a comprehensive framework for understanding fibroblast dynamics, highlighting the heterogeneity and spatial complexity of fibrosis in human end-stage HF, and offering potential therapeutic targets to mitigate fibrosis while preserving scar integrity.