Stem Cells from Human Exfoliated Deciduous Teeth (SHEDs) and Dental Pulp Stem Cells (DPSCs) Display a Similar Profile with Pericytes.

BACKGROUND: Pericytes play an important role in forming functional blood vessels and establishing stable and effective microcirculation, which is crucial for vascular tissue engineering. The slow ex vivo expansion rate, limited proliferative capacity, and variability of tissue-specific phenotypes would hinder experimental studies and clinical translation of primary pericytes. In this study, the angiogenic and pericyte functions of stem cells from human exfoliated deciduous teeth (SHEDs) and postnatal human dental pulp stem cells (DPSCs) were investigated. METHODS: Osteogenic and adipogenic induction assays were performed to evaluate the mesenchymal potential of SHEDs, DPSCs, and pericytes. An in vitro Matrigel angiogenesis assay was conducted to reveal the ability of SHEDs, DPSCs, and pericytes to stabilize vascular-like structures. Quantitative real-time polymerase chain reaction (RT-qPCR) was performed to evaluate mRNA expression. Flow cytometry, western blotting, and immunostaining were used to assess the protein expression. Wound healing and transwell assays were performed to evaluate the migration ability of SHEDs, DPSCs, and pericytes. RESULTS: The osteogenic and adipogenic induction assays showed that SHEDs, DPSCs, and pericytes exhibited similar stem cell characteristics. The mRNA expression levels of PDGFR-ß, α-SMA, NG2, and DEMSIN in SHEDs and DPSCs cultured in EC medium were significantly higher than those in the control groups on day 7 (P < 0.05), but significantly higher than those in the pericytes group on day 14 (P < 0.05). Flow cytometry showed that high proportions of SHEDs and DPSCs were positive for various pericyte markers on day 7. The DPSCs, SHEDs, and pericytes displayed strong migration ability; however, there was no significant difference among the groups (P > 0.05). CONCLUSION: The SHEDs and DPSCs display a profile similar to that of pericytes. Our study lays a solid theoretical foundation for the clinical use of dental pulp stem cells as a potential candidate to replace pericytes.

[Effect of transfected EphB4 on osteogenic differentiation in stem cells from human exfoliated deciduous teeth].

PURPOSE: To evaluate the effect of stem cells from human exfoliated deciduous teeth (SHED) transfected with EphB4 gene in regulating osteogenic differentiation. METHODS: Human dental pulp tissue were harvested from extracted deciduous teeth and digested by collagenase and dispase. The SHEDs were transfected with transgenic (hEphB4-GFP) vector or empty vector (GFP-vector). Real time-polymerase chain reaction（real time-PCR） analysis and Western blot were used to detect the expression of EphB4 in SHEDs after transfection. EphB4-SHEDs and GFP-SHEDs were subjected to osteogenic induction and assessed by alkaline phosphatase(ALP) assay and Alizarin-red S staining. SPSS 16.0 software package was used for statistical analysis. RESULTS: Real time-PCR revealed that the expression of EphB4 was significantly enhanced in EphB4-SHEDs compared to GFP-SHEDs (P<0.05). The expression of EphB4 protein was significantly higher (P<0.05) in EphB4-SHEDs compared to GFP-SHEDs. ALP assay and Alizarin-red S staining demonstrated higher ALP activity and increased mineralization with EphB4-SHEDs. CONCLUSIONS: The results indicate that transgenic expression of EphB4 in SHEDs could promote osteogenic differentiation.

[The mechanism of corticotomy accelerating orthodontic tooth movement in SD rats].

PURPOSE: To study the influences of corticotomy on orthodontic tooth movement (OTM) and the underlying mechanism in rats. METHODS: Forty-eight female Sprague-Dawley rats were randomly assigned to corticotomy group (Cort, 24 rats) or sham-corticotomy group (Sham, 24 rats). All rats were subjected OTM after corticotomy or sham surgery. 6 rats of each group were sacrificed at 0, 1, 3 and 7 day of OTM. OTM were measured with an electronic digital caliper. Osteoclasts were counted in pressure side with TRAP. RANKL were measure by IHC at pressure side. The data were analyzed with SPSS 16.0 software package. RESULTS: OTM at the 1 and 7 day in Cort group increased compared with sham group. Number of TRAP positive osteoclasts in pressure side increased in Cort group at the 3 and 7 day. Expression of RANKL in pressure side also increased in Cort group at the 3 and 7 day. CONCLUSIONS: Corticotomy accelerates OTM in rats and it may result from promoted bone resorption via increased RANKL expression in periodontal tissue.

TGF-ß1-induced differentiation of SHED into functional smooth muscle cells.

BACKGROUND: Adequate vascularization is crucial for supplying nutrition and discharging metabolic waste in freshly transplanted tissue-engineered constructs. Obtaining the appropriate building blocks for vascular tissue engineering (i.e. endothelial and mural cells) is a challenging task for tissue neovascularization. Hence, we investigated whether stem cells from human exfoliated deciduous teeth (SHED) could be induced to differentiate into functional vascular smooth muscle cells (vSMCs). METHODS: We utilized two cytokines of the TGF-ß family, transforming growth factor beta 1 (TGF-ß1) and bone morphogenetic protein 4 (BMP4), to induce SHED differentiation into SMCs. Quantitative real-time polymerase chain reaction (RT-qPCR) was used to assess mRNA expression, and protein expression was analyzed using flow cytometry, western blot and immunostaining. Additionally, to examine whether these SHED-derived SMCs possess the same function as primary SMCs, in vitro Matrigel angiogenesis assay, fibrin gel bead assay, and functional contraction study were used here. RESULTS: By analyzing the expression of specific markers of SMCs (α-SMA, SM22α, Calponin, and SM-MHC), we confirmed that TGF-ß1, and not BMP4, could induce SHED differentiation into SMCs. The differentiation efficiency was relatively high (α-SMA+ 86.1%, SM22α+ 93.9%, Calponin+ 56.8%, and SM-MHC+ 88.2%) as assessed by flow cytometry. In vitro Matrigel angiogenesis assay showed that the vascular structures generated by SHED-derived SMCs and human umbilical vein endothelial cells (HUVECs) were comparable to primary SMCs and HUVECs in terms of vessel stability. Fibrin gel bead assay showed that SHED-derived SMCs had a stronger capacity for promoting vessel formation compared with primary SMCs. Further analyses of protein expression in fibrin gel showed that cultures containing SHED-derived SMCs exhibited higher expression levels of Fibronectin than the primary SMCs group. Additionally, it was also confirmed that SHED-derived SMCs exhibited functional contractility. When SB-431542, a specific inhibitor of ALK5 was administered, TGF-ß1 stimulation could not induce SHED into SMCs, indicating that the differentiation of SHED into SMCs is somehow related to the TGF-ß1-ALK5 signaling pathway. CONCLUSIONS: SHED could be successfully induced into functional SMCs for vascular tissue engineering, and this course could be regulated through the ALK5 signaling pathway. Hence, SHED appear to be a promising candidate cell type for vascular tissue engineering.