Spatial Transcriptomics Reveals Region-Specific Remodeling in Vein Grafts After Peripheral Arterial Bypass

Objective Vein graft failure due to intimal hyperplasia and maladaptive remodeling remains a major limitation of peripheral bypass surgery. Although vascular remodeling is recognized as a multilayered process, layer-specific molecular mechanisms that distinguish adaptive from negative remodeling remain incompletely understood. We aimed to investigate the vascular microenvironment of patent and stenotic grafts using spatial transcriptomics. Methods Vein specimens were obtained from three patients undergoing revision surgery. For each patient, a matched set of three samples was collected: unused saphenous vein (Denovo), non-stenotic portion of the graft (Non-stenosed), and stenosed vein graft (Stenosed) (n = 3 patients). GeoMx Digital Spatial Profiling with the Human Whole Transcriptome Atlas was used to map gene expression across intimal, medial, and adventitial layers in 80 regions of interest; Denovo sections were evaluable in two of the three patients. Differential expression, gene ontology, spatial deconvolution, and immunofluorescence were integrated for analysis, with sensitivity analyses accounting for the nesting of regions of interest within patients. Results Non-stenosed and Stenosed grafts shared transcriptional features distinct from Denovo veins, particularly in pathways related to cell proliferation. Non-stenosed grafts showed increased expression of CDKN1A across all vascular layers, whereas Stenosed grafts showed higher mitogen-activated protein kinase (MAPK) scores, lower DUSP1 expression, and greater adventitial macrophage marker expression. Extracellular matrix (ECM) remodeling showed layer-specific organization, with VCAN and ACAN enriched in the intima and DCN in the adventitia, while Stenosed grafts showed a trend toward collagen-dominant remodeling. Cell deconvolution suggested a shift toward vascular smooth muscle cell (VSMC)-dominant architecture after arterialization, with modest enrichment of synthetic VSMC signatures in stenotic regions. Differences between Non-stenosed and Stenosed segments in DUSP1 and the Net MAPK score were concordant in direction across all three patients in every layer. Conclusions In this exploratory study, vein graft stenosis was associated with layer-specific differences in cell cycle regulation, inflammatory signaling, ECM remodeling, and VSMC phenotype. Spatial transcriptomic analysis resolved molecular heterogeneity not captured by bulk approaches. Because only three patients were studied, these findings are hypothesis-generating and require confirmation in larger cohorts.