Dental pulp stem cells (DPSCs) invoke wide interest due to their roles in in dentin regeneration and repair, and more research is required to illustrate the molecular basis. In this study, we induced osteogenic differentiation in human DPSCs to investigate the role of interferon-induced transmembrane (IFITM) family in osteogenic differentiation. Our results demonstrated that genes of the IFITM family, IFITM1, IFITM3, and IFITM5 were upregulated during the osteogenic differentiation of DPSCs. Loss-of-function and gain-of-function assays indicated that IFITM3 knockdown suppressed osteogenic differentiation, assessed by the reduction of alkaline phosphatase (ALP) activity, nodule mineralization, and the expressions of several osteogenic differentiation-associated genes; in contrast, IFITM3 overexpression increased ALP activity, nodule mineralization, and the expressions of osteogenic differentiation-associated genes. Further investigation revealed that IFITM3 was involved in the regulation of mitogen-activated protein kinase (MAPK) signaling during osteogenic differentiation, and pharmacological inhibition of both ERK and p38 suppressed the effects of IFITM3 overexpression in osteogenic differentiation. These results suggest that IFITM3 promotes osteogenic differentiation of DPSCs by regulating MAPK signaling. Our findings enhance the existing knowledge on the role of IFITM family in osteogenic differentiation and the molecular basis of IFITM3.

We generated podoplanin-conditional knockout mice where the floxed podoplanin exon3 was deleted by the Dmp1-driven Cre (Dmp1-Cre;Pdpn^Δ/Δ) and investigated the cell process elongation of podoplanin-deficient mouse osteocyte in vitro and in vivo. The expression of podoplanin is found in odontoblasts while not observed in odontoblasts of Dmp1-Cre;Pdpn^Δ/Δ mice, indicating that the conditional knockout of podoplanin in Dmp1-expressing cells in Dmp1-Cre;Pdpn^Δ/Δ mice is successful. There were no differences in the growth of wild-type and Dmp1-Cre;Pdpn^Δ/Δ mice, and no differences in calcification and alkaline phosphatase activity in cultured calvarial osteoblasts of the wild-type and Dmp1-Cre;Pdpn^Δ/Δ mice, in total this suggests that the podoplanin-cKO has no effect on generation of the bone. The cell process elongation was suppressed in cultured calvarial osteoblasts of Dmp1-Cre;Pdpn^Δ/Δ mice compared with wild-type mice. In the electron microscopic study, there were no morphological differences in bone matrix formation and osteocyte distribution in Dmp1-Cre;Pdpn^Δ/Δ and wild-type mice, whereas the cell process formation was sparser and the network with neighboring cells was more deficient in Dmp1-Cre;Pdpn^Δ/Δ mice than in wild-type mice. In the quantitative analysis, the number and thickness of the cell processes were significantly smaller and thinner in Dmp1-Cre;Pdpn^Δ/Δ mice than in wild-type mice. This could suggest that podoplanin plays a role in the formation of the osteocyte network created by the cell process elongation.

Treatment of tumors in the head and neck region using anticancer agents or radiotherapy often causes oral mucositis, which is associated with severe mucosal pain. Hyaluronic acid (HA) is a moisturizing agent used clinically to treat oral mucositis; however, HA does not remain long-term in the mucositis region due to its liquid nature. We recently developed the original sheet form HA using a freeze-drying method. In this study, we evaluated the effect of topical application of the newly developed HA sheets on oral mucositis healing using an animal model. Experimental oral mucositis was induced in 6-week-old male Syrian hamsters by applying filter paper soaked with acetic acid to the cheek pouches. After determining the appropriate conditions for obtaining uniform mucositis region useful for evaluating the healing process, the effect of HA sheets on oral mucositis healing were evaluated macroscopically and histopathologically comparing three groups: an HA group, in which HA sheets were applied to the mucositis region from the second day after mucositis induction; a carboxymethylcellulose (CMC) control group treated with CMC sheets; and a control group in which no sheets were applied. Compared to the control group, the HA group exhibited significantly smaller mucositis region on Days 2-4. Histopathology analysis on Day 2 showed mild inflammatory cell infiltration, reduced edematous changes, partial regeneration of the oral mucosal epithelium, and parallel thick bundles of collagen fibers in the superficial layer in the HA group compared with the Control and CMC groups. In conclusion, a hamster oral mucositis model was established that allows for convenient evaluation of the healing process. Daily topical application of freeze-dried sheets of HA accelerated the healing of oral mucositis in this hamster model. These results suggest that HA sheets could be an effective material in the treatment of oral mucositis in cancer patients.

Regenerative medicine has limitations of stem cell amplification and immune rejection that make it difficult to apply in emergency treatment, such as that required for trauma. We have transplanted DPSCs or conditioned medium (CM; secretome accumulation) in animal models and found evidence for pulp regenerative potential, with these findings caused by various trophic effects of DPSCs and CM. Trauma affects various tissues, all of which require regeneration, and there is a need to evaluate trophic effects in trauma models. To examine this issue, we made a canine teeth complete dislocation model, removed the dental pulp and periodontal ligament, performed DPSC transplantation and retransplantation, and observed the regenerative response of periodontal tissues. These results suggest that pulp regeneration is important for regeneration of periodontal ligament in a complete dislocation model. In short, these results indicate the need to focus on cross talk between dental pulp and periodontal tissues. Thus, we added DPSC-CM to periodontal ligament cells and investigated whether a trophic effect would occur in regenerated tissues. The results of this study suggest that migration and proliferation were promoted by trophic effects of DPSC-CM, suggesting regeneration of dental pulp. Next, we performed coculture of dental pulp and periodontal ligament to observe changes in trophic factors by cross talk between these tissues. The results of the current study showed that both VEGF and HGF were highly expressed, suggesting a synergistic effect. Tissue regeneration after trauma was found to be caused by the trophic effect of CM, promotion of proliferation, migration and antiresorptive activity. This effect may be enhanced by crosstalk between the pulp and the periodontal ligament, and VEGF and HGF are candidates for this effect.

Periosteal distraction is promising for augmenting new bone. This study aimed to develop a protocol for achieving stable bone augmentation. We applied mandibular alveolar ridge augmentation in beagles to test a trial distraction device. Premolars and molars were extracted bilaterally, and adjacent buccal cortical bone (3–4-mm thick) was removed to mimic atrophy. A 0.9-mm-diameter titanium screw was placed under the periosteum for later use to induce distraction. After healing (16 weeks), a titanium periosteal distraction device was applied on both sides in three beagles (six sides), and the periosteum was distracted laterally (0.5 mm/day for 6 days). One beagle was a control (two sides). No infection or displacement of the device was observed. All beagles were euthanized 10 weeks after distraction. Specimens were harvested and the mandibles were sectioned in half side was decalcified with formic acid and hematoxylin-eosin stained. Histological examination showed dense bone formation with a lamellar structure, with a bone width increase of 0.79 ± 0.11 mm compared with the control specimens. Bone formation can be induced by a periosteal distraction device, suggesting potential clinical utility.