Cross-cohort single-nucleus transcriptomic identification of a MEOX1+FAP+ACTA2-low extracellular matrix-remodeling fibroblast state in dilated cardiomyopathy

Background Cardiac fibrosis is a major determinant of adverse remodeling in dilated cardiomyopathy (DCM), but the human disease-associated fibroblast state is incompletely resolved. We tested whether a non-classical MEOX1+FAP + ACTA2-low extracellular matrix (ECM)-remodeling state could be reproduced across independent human cardiac single-nucleus RNA-sequencing cohorts. Methods Public GSE183852 data (25 non-failing donor and 13 DCM hearts) were used for discovery, and SCP1303 (16 normal and 11 DCM hearts) for external validation. Fibroblasts were independently reclustered. All inferential analyses used donors as the statistical unit. State abundance was assessed with Wilcoxon tests, 10,000-iteration donor bootstrap confidence intervals, leave-one-donor-out analyses, and heteroscedasticity-robust covariate models. Raw fibroblast counts were aggregated by donor for pseudobulk differential expression with age-tertile and sex adjustment. ECM activity was compared using pseudobulk and cell-derived donor summaries. Results Discovery cluster two showed a MEOX1+FAP + ACTA2-low profile distinct from an ACTA2-high state and was enriched in DCM. Its mean abundance difference was 0.165 of fibroblasts (bootstrap 95% confidence interval 0.035–0.349), with a positive effect after omission of every donor. Age-tertile and sex adjustment preserved the effect magnitude, although the robust confidence interval marginally crossed zero. In SCP1303, the corresponding activated-fibroblast state was enriched in DCM (mean difference 0.192, bootstrap 95% confidence interval 0.144–0.256) and remained associated after sex adjustment. Donor-pseudobulk models identified higher DDAH1, MEOX1 and FAP expression in DCM after false-discovery-rate correction. Two ECM summaries were strongly correlated (Spearman ρ = 0.891), although group significance was method dependent. Conclusion A reproducible MEOX1+FAP+, relatively ACTA2-low fibroblast state is associated with ECM remodeling in the analyzed human DCM cohorts. DDAH1 is a prioritized state-associated candidate, not an established causal regulator. Spatial and perturbation studies are required for mechanistic validation.