Integrated scRNA-seq and spatial transcriptomics identify caveolin-1 as a regulator of endothelial progenitor cell function associated with PI3K/AKT signalling and DVT recanalization

Abstract Background Deep vein thrombosis (DVT), a prevalent vascular disease, frequently manifests in conjunction with post-thrombotic syndrome (PTS), a condition that can markedly compromise prognoses. The objective of this study is to investigate the internal mechanisms and pathways associated with DVT recanalization, with a view to establishing a foundation for the development of relevant drug targets. Methods The SD rat inferior vena cava (IVC) ligation model was constructed, and thrombus ( n = 3) and normal IVC tissues ( n = 3) were obtained for standard single-cell RNA sequencing (scRNA-seq) analysis. The aim was to characterise the DVT-associated single-cell transcriptional profiles and localise key cellular subpopulations. In combination with lentiviral knockdown/overexpression (KD/OE) assays, the present study explored the functional regulatory role of caveolin-1 (CAV-1) on target cells and the corresponding key signalling pathways, and illustrated its biological effects on DVT at the animal model level. Results Based on the analysis of the scRNA-seq standard process, the heterogeneity of each cell population related to the DVT microenvironment was demonstrated after cell dimensionality reduction, population size and annotation, and neutrophils and fibroblasts were the most obvious. In addition, it was speculated that the presence of endothelial progenitor cells (EPCs) in DVT plays a regulatory role through subpopulation annotation, cell communication analysis and pseudotime analysis. Through multi-angle experiments of cell counting kit-8 (CCK-8), flow cytometry, immunofluorescence (IF) and transwell assay proved that up-regulation of CAV-1 can promote EPC proliferation and migration ability. Subsequently, the utilisation of animal models was employed to demonstrate that up-regulation of CAV-1 promotes angiogenesis and enhances EPC survival. Furthermore, the PI3K-AKT signalling pathway was discovered and validated as a key pathway in the aforementioned process. Furthermore, the spatial transcriptome provided further evidence for alterations in CAV-1 levels and key inflammatory pathways. Conclusions The combination of scRNA-seq and functional experiments has led to the proposal that CAV-1 is a regulator of EPC function associated with PI3K/AKT signalling and DVT recanalization.