Integrin-Associated Signalling Regulates Jagged1-Induced Osteogenic Differentiation of Human Dental Pulp Stem Cells

To investigate the role of integrin-associated adhesion signalling in Jagged1-driven osteogenic differentiation and to determine the specific contribution of integrin β3 (ITGβ3).

RNA sequencing data from Jagged1-activated human dental pulp cells were analysed to identify changes in integrin-associated signalling. Jagged1-induced Notch activation and ITGβ3 expression were validated by quantitative real-time polymerase chain reaction and immunofluorescence. Functional inhibition of integrin activity was performed using the αvβ3 antagonist cilengitide, and the specific role of ITGβ3 was examined using ITGβ3-targeting siRNA. Apoptosis was evaluated by flow cytometry, migration by a wound-healing assay, and osteogenic differentiation by Alizarin Red S staining and osteogenic gene expression.

RNA sequencing analysis revealed enrichment of extracellular matrix- and integrin-associated pathways among Jagged1-responsive genes in human dental pulp stem cells (hDPSCs), with ITGβ3 identified as a prominently upregulated integrin subunit. Jagged1 induced ITGB3 mRNA expression in parallel with dose- and time-dependent increases in the Notch target genes HES1 and HEY1, and increased ITGβ3 protein expression was further confirmed by immunofluorescence. Pharmacological inhibition with cilengitide did not affect viability, proliferation, or migration, but significantly suppressed basal and Jagged1-induced mineralisation and reduced RUNX2 expression. High-dose cilengitide (1000 nM) increased late-stage apoptosis under Jagged1 stimulation. In contrast, ITGB3 silencing attenuated basal osteogenic differentiation but did not impair Jagged1-induced mineralisation.

Jagged1-induced osteogenic differentiation of hDPSCs requires integrin-dependent adhesion signalling but is not exclusively dependent on ITGB3 transcription. These findings support integrin-Notch crosstalk as a regulatory mechanism in hDPSC osteogenesis and provide a rationale for designing regenerative dental biomaterials that combine Notch-activating and adhesion-supportive cues.