Multi-modal analysis of vascular remodelling in human obliterative bronchiolitis after lung transplantation

Chronic lung allograft dysfunction (CLAD) is the leading cause of late morbidity and mortality after lung transplantation. Bronchiolitis obliterans syndrome (BOS), a major CLAD phenotype, is characterized by obliterative bronchiolitis (OB). While microvascular perturbations are described in BOS, their cellular and molecular characteristics remain unclear. Intact human CLAD lungs (n=3) underwent hierarchical phase-contrast tomography (HiP-CT) for three-dimensional vascular mapping. In OB-affected airways vs. controls (n=4 each), endothelial subtypes were characterized by multiplex immunofluorescence imaging, interactome changes quantified with CellChat and GeoMx spatial transcriptomics performed on CD31⁺ regions. HiP-CT revealed hypertrophic vessels infiltrating obliterated airways and forming web-like microvascular networks, absent in controls. Immunofluorescent imaging showed increased PLVAP⁺/VWA1⁺ systemic venous and PDPN⁺ lymphatic cells in OB lesions. Cell-cell interaction analysis showed that PLVAP⁺/VWA1⁺ systemic venous cells exhibited increased incoming and outgoing interaction strength, primarily directed toward other endothelial cells and fibroblasts, characterized by inflammatory and fibrotic signalling. Spatial transcriptomics demonstrated upregulation of inflammatory/matrix-remodelling genes and downregulation of homeostatic regulators in BOS endothelial cells. BOS lungs display altered vascular architecture and endothelial composition, with gene expression shifts consistent with pro-inflammatory and fibrogenic processes in CD31⁺ regions. These findings suggest a potential involvement of vascular remodelling in CLAD pathophysiology.