Molecular Cues from Extracellular Matrix Proteins Drive the Differentiation of Dental Pulp Stem Cells into Endothelial Cells in the Presence of Dentin Matrix Protein 1.

INTRODUCTION

Dental pulp stem cells (DPSCs) have the potential to differentiate into endothelial cells (ECs) because they express the key cell surface markers like VEGFR1 and VE-cadherin and can be stimulated by various growth factors. We have previously demonstrated that DPSCs, when stimulated by DMP1 combined with cues from human umbilical vein endothelial cells matrix, promoted their differentiation into ECs. In this study, we explored the response of DPSCs to specific extracellular matrix components such as laminin and Matrigel combined with DMP1 stimulation. The extracellular matrix plays a vital role in tissue development by providing structural support, anchorage for cell adhesion, mechanical cues, and the capacity to sequester soluble factors.

METHODS

DPSCs were seeded onto plates coated with laminin and Matrigel, treated with recombinant DMP1 (500 ng/mL), and cultured for 7 days. Various biological assays, such as gene expression (quantitative real-reverse transcription PCR) and protein expression assessments (flow cytometry), alongside functional characteristics as assessed by tubule assays, were performed to evaluate the response of DPSCs to DMP1 stimulation in the presence of laminin or Matrigel.

RESULTS

An analysis of the data from this study demonstrates that DMP1 significantly enhances the differentiation of DPSCs to the endothelial lineage. This enhancement is evidenced by the upregulation of angiogenic genes, comprising CD31, VE-cadherin, vascular endothelial growth factor A, Angiopoietin 1, and Endoglin, under both laminin-coated and Matrigel-coated conditions in response to DMP1. At the gene expression level, all angiogenic markers, except CD31, exhibited higher expression in Matrigel conditions relative to laminin with DMP1 stimulation. However, CD31, CD144, and CD146 expression were elevated at the protein level in laminin-coated conditions. Several members of the integrin family, such as α5, αV, β1, β3, and β4, were upregulated in the presence of laminin. The binding of the integrin receptors with the laminin matrix plays a pivotal role in vascular differentiation. Thus, DMP1-mediated integrin expression on the cell surface of DPSCs would facilitate cell anchorage and promote their differentiation into ECs. Finally, the functional characterization of the de novo ECs shows a strong propensity to form tubules under laminin-coated conditions compared to DPSCs cultured on Matrigel matrix.

CONCLUSIONS

Overall, findings reveal that DMP1 directs the DPSCs to endothelial-like cells in the presence of laminin and Matrigel.