Abstract Body: Background: In-stent restenosis (ISR) and re-thrombosis (RT) are pathological processes that develop after venous stenting, leading to late stent failure. Currently, post-stenting medical management primarily targets the coagulation cascade, but neglects to address associated inflammation subsequent vascular fibrosis, which are linked to the interactions between venous endothelial cells and immune cells.
Objective: To determine cell-specific gene expression profiles that contribute to venous stent failure.
Methods: We performed single-cell RNA sequencing (scRNA-seq) analysis of native femoral veins and stent acute RT and ISR, characterized by old thrombus/diffuse intimal thickening (DIT). Selected specific cell-expression profiles were confirmed using immunohistochemistry (IHC).
Results: Using scRNA-seq, we identified the cell landscape in human deep veins and RT and ISR processes. Clustering analysis of transcriptional profiles identified 12 cell lineages using SingleR, Further sub-clustering of the main cell clusters identified 6 endothelial (mature venous endothelial cells/EC: EPHB4+ NR2F2+, Inflammatory EC: SELE+ ICAM1+ VCAM1+), 3 vascular smooth muscle (VSMC) (Contractile: SPEG+ SMTN+, Synthetic/Transitioning VSMC: ACTA2+ TAGLN+, and Fibroblast-like: SERPINE1+, DUSP6+), 4 fibroblast (Inflammatory: NLRP3+, MRC1+, Matrix-producing: MMP2+ COL1A1+, Senescent: DEPP1+ CCL2+), and 5 monocyte/macrophage (Mo/MΦ) subpopulations. The cellular composition reverted to primarily vascular structural cells in ISR compared to circulating cells in RT. Gene enrichment analysis on VECs and VSMCs subsets identified inflammation, hypoxia, and PI3K-Akt-mTOR as common upregulated pathways in both RT and ISR. Figure 1. A-F.
Conclusions: A single-cell analysis of native deep veins, RT, and ISR reveal how the gene expression landscape is altered in these pathologic processes, providing a cell specific transcriptomic map to nominate causal target genes.