ATP1A1-Driven Intercellular Contact Between Dental Pulp Stem Cell and Endothelial Cell Enhances Vasculogenic Activity.

AIM: The interaction between dental pulp stem cells (DPSCs) and vascular endothelial cells (ECs) is crucial to the speedy establishment of functional blood circulation within the transplanted pulp tissue. It is a complex process involving direct cell contact and paracrine signalling. The transmembrane domains of α1-Na(+)/K(+)-ATPase (ATP1A1) have been shown to influence tumour angiogenesis. Its role in regulating DPSCs/ECs interaction in vascular formation remains unknown. This study aimed to explore ATP1A1 on DPSCs/ECs communication, vascular network formation, and underlying mechanisms. METHODS: The formation of vessel structures within different culture systems was examined. The expression of pericyte-like markers and Na(+)/K(+)-ATPase-related genes and proteins were systematically analysed. Immunofluorescence staining was performed to examine the localisation of ATP1A1. Total and phosphorylated proteins were evaluated to identify and explore the signalling pathways activated under cocultured conditions. Downstream signalling was also investigated after the inhibition of ATP1A1. RESULTS: Direct coculture accelerated vessel network formation and prolonged its stability compared to indirect systems. ATP1A1 expression and SMC-specific marker (α-SMA) levels significantly increased in direct coculture systems, with nuclear α-SMA localisation and ATP1A1 enrichment at cell-contact sites. Protein assay revealed activated Src/AKT pathways and upregulated FGF-2/activin A secretion in coculture supernatants. ATP1A1 inhibition reduced α-SMA expression, impairing SMC differentiation. CONCLUSION: Direct DPSCs-HUVECs contact stabilises vessel networks via ATP1A1-mediated Src/AKT activation, driving FGF-2/activin A secretion and initiating SMC differentiation. This highlights that ATP1A1 may be critical for pericyte-like transition and vascular microenvironment optimisation in pulp angiogenesis. CLINICAL SIGNIFICANCE: This research informed strategies aimed at pulp tissue regeneration. The findings hold significant implications for enabling the biological restoration of tooth vitality and function in the field of clinical regenerative treatment.