Endothelial differentiation potential of human dental pulp stem cells under Semaphorin 4D stimulation

Abstract

Objectives: Stem cell-based therapy is a promising approach for regenerating dental pulp/dentin complex tissues. However, the root canal space which is encapsulated by dentin and enamel with only one opening to the host vasculature restricts blood supply to the tissues transplanted into the root canal, and could hamper the process of vascularization. It was reported that Semphorin4D (Sema4D) strongly promotes vascularization in tumor tissue in a manner resembling vascular endothelial growth factor (VEGF). Thus, the objective of this study is to investigate the role of Sema4D on angiogenic and endothelial differentiation of human dental pulp stem cells (DPSCs). Methods: Treated DPSCs with low concentration of Sema4D in α-MEM with fetal bovine serum (10%) for several days, then endothelial differentiation capabilities of DPSCs were evaluated by expression of endothelial cell-specific genes (VEGFR1, VEGFR2, CD31, vWF and ANGPTL4) and angiogenic gene (VEGF) by using real-time polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). Correspondingly, endothelial-related protein (VEGFR2, CD31 and VE-Cadherin) and Sema4D-related downstream signaling (Plexin-B1, Met and ErbB2) were assessed by western blot. Additionally, tube formation ability on Matrigel was performed to evaluate the angiogenic potential of Sema4D treated DPSCs. Results: Sema4D significantly enhanced the genes (VEGFR2, CD31, vWF and ANGPTL4) expression associated with endothelial differentiation. Likewise, endothelial-specific protein (VEGF, VEGFR2, CD31 and VE Cadherin) were notably up-regulated in the presence of Sema4D and protein expression of ErbB2 was increased. In addition, DPSCs pre-treated with Sema4D for 7 days exhibited superior capacity of vessel-like structures formation on Matrigel compared with non-pretreated DPSCs. Conclusions: The findings of this study suggest that Sema4D induces DPSCs endotheliogenesis. Sema4D may be useful for accelerating vasculature formation in dental tissue engineering.