Single-cell multiomic analysis of mesenchymal cells reveals molecular signatures and regulators of lung allograft fibrosis.

Survival after lung transplantation is limited by chronic, progressive graft failure, termed chronic lung allograft dysfunction (CLAD). Graft-resident mesenchymal cells (MCs) drive CLAD pathogenesis and exhibit stable dysregulated signaling, yet the transcriptomic and epigenomic drivers underlying this fibrogenic transformation remain elusive. We used single-cell multiomic profiling to characterize gene expression and chromatin accessibility in MCs isolated from bronchoalveolar lavage fluid of lung transplant recipients with and without CLAD, collected early after transplantation or after disease onset. MCs obtained after CLAD onset demonstrated a distinct transcriptomic signature compared with non-CLAD controls, enabling classification of disease status at the single-cell level with greater than 98% accuracy using signature genes. Chromatin accessibility analyses identified enrichment of CCAAT-enhancer-binding protein family transcription factors, specifically CEBPD, in CLAD MCs. MCs early after transplantation showed minimal accessibility differences, suggesting that CEBPD-associated regulatory changes emerge over time. Integration analyses identified 8 MC states and a CLAD-specific shift toward a fibrotic state. CEBPD, SOX4, and FOXP2 were identified as putative regulators of this state with substantial overlap in predicted targets. Targeting CEBPD reversed fibrotic phenotypes of CLAD MCs (decreased ECM expression, contractility, proliferation, and migration). Together, these data provide insights into transcriptomic and epigenomic changes in posttransplant MCs, facilitating the nomination of biomarkers and therapeutic targets.