Spatial proteomic profiling of cardiac stromal populations and microenvironments in systemic sclerosis-associated primary myocardial involvement
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Objectives
To define cardiac stromal heterogeneity and spatial stromal-immune organization in systemic sclerosis-associated virus-negative myocarditis (SSc-VNM), a manifestation of primary SSc heart involvement.
Methods
Imaging mass cytometry, a single-cell multiplexed spatial proteomic technology, was applied to endomyocardial biopsies from ten patients with SSc-VNM, seven with VNM associated with other systemic autoimmune rheumatic diseases (a-VNM) and nine with isolated VNM (i-VNM). Single-cell phenotyping, cellular-neighborhood and spatial-interaction analyses, and correlations with histological fibrosis were performed at patient level.
Results
Among 82,673 segmented cells, we identified seven fibroblast phenotypes and one CD34 + stromal-cell population. The overall fibroblast/stromal compartment was expanded in SSc-VNM compared with i-VNM (adjusted p = 0.00973). The composition of fibroblasts/stromal cells differed profoundly in SSc-VNM compared with a-VNM and i-VNM, with numerical increases in CD34 + stromal cells and SOX9 + and PI16 high /POSTN high fibroblasts and de-enrichment of FAP high /POSTN high fibroblasts and myofibroblasts. Spatial analysis identified ten cellular neighborhoods and revealed distinct SSc-VNM-associated patterns, including altered stromal-immune interaction networks. These included enriched interactions of CD34 + stromal cells and PI16 high /POSTN high fibroblasts with CD163 high /HLA-DR + macrophages and CD68 + mast cells. PI16 high /POSTN high and PI16 - /POSTN high fibroblasts demonstrated the highest collagen type 1 expression among all stromal cell subsets across all VNM entities and PI16 high /POSTN high fibroblasts were numerically increased in biopsies with ≥10% fibrotic remodeling and were spatially associated with collagen-rich areas.
Conclusions
Our findings refine current concepts of human fibrotic myocardial remodeling and support therapeutic targeting of specific stromal players such as POSTN high fibroblast subpopulations.
Key messages
What is already known on this topic
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Primary SSc heart involvement contributes substantially to disease-related mortality, but its pathophysiology remains largely enigmatic, particularly lacking understanding of cellular interactions and spatial architecture.
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Single-cell studies have established fibroblast heterogeneity in both human cardiac disease and SSc skin and lung tissues.
What this study adds
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We provide a molecular characterization of cardiac stromal cell subpopulations using protein-based definitions and demonstrate dysregulation of several fibroblast/stromal subsets, with increases in CD34 + stromal cells and SOX9 + and PI16 high /POSTN high fibroblasts and de-enrichment of FAP high /POSTN high fibroblasts in SSc-VNM compared with a-VNM and i-VNM.
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We identify the SOX9 + fibroblast niche as a distinct microenvironment particularly enriched in immune cells, including monocyte and macrophage subsets.
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We demonstrate major differences in the spatial cellular interaction networks of cardiac fibroblasts/stromal cells in SSc-VNM compared with a-VNM and i-VNM.
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We identify PI16 high /POSTN high and PI16 - /POSTN high fibroblasts as the main collagen type 1 producing fibroblast subsets. Consistently, PI16 high /POSTN high fibroblasts were numerically increased in biopsies with ≥10% fibrotic remodeling and were associated with collagen-rich areas.
How this study might affect research, practice or policy
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Our study identifies defined cardiac stromal states and stromal-immune niches as potential candidates for future therapeutic intervention in primary SSc heart involvement.
Article activity feed
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Spatial proteomic profiling of cardiac stromal populations and microenvironments in systemic sclerosis-associated primary myocardial involvement #SingleCell 🧪🧬🖥️ https://www.biorxiv.org/content/10.64898/2026.10.17.758293v1
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Spatial proteomic profiling of cardiac stromal populations and microenvironments in systemic sclerosis-associated primary myocardial involvement https://www.biorxiv.org/content/10.64898/2026.10.17.758293v1
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Spatial proteomic profiling of cardiac stromal populations and microenvironments in systemic sclerosis-associated primary myocardial involvement https://www.biorxiv.org/content/10.64898/2026.10.17.758293v1
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