Background and Objectives Oral ulceration is one of the most common debilitating condition that affects the oral cavity. In this study, the effect of locally injected platelet rich plasma (PRP) and bone marrow-derived mesenchymal stem cells (BMSCs) on the healing of oral ulcer was investigated. Methods and Results An ulcer was induced in buccal mucosa of rats by using 5mm biopsy punch followed by application of cotton swab soaked with formocresol for 60sec. The ulcer was left untreated in the control group, treated with intralesional injection of PRP, or isolated cultured BMSCs. Data were analyzed clinically, histologically and immunohistologically on day 3, 5, 7 and 10. BMSCs group showed smaller ulcer area throughout the whole experimental period than the other groups with complete resolution of the ulcer on day 10, unlike the control group. However, there was no significant difference with PRP, on day 5, 7 and 10, regarding clinical ulcer size. BMSCs group showed better histological results regarding the rate of epithelial cell migration, the number of inflammatory cells, thickness and organization of collagen fibres and the number of blood vessels, with complete re-epithelization on day 10. BMSCs group showed a greater number of anti-PCNA positive nuclei throughout the whole experimental period than the other groups except on day 5, PRP had higher mean numbers of anti-PCNA positive nuclei in both tissues. Conclusions Both PRP and BMSCs accelerate wound healing and enhance the quality of the healing tissue with the latter being slightly more effective and faster.

Background and ObjectivesRecombinant amelogenin protein (RAP) was reported to induce soft-tissue regeneration in canine infected endodontically treated permanent teeth with open apices. To characterize identities of the cells found in the RAP regenerated tissues compared to authentic pulp by identifying: 1) stem cells by their expression of Sox2; 2) nerve fibers by distribution of the axonal marker peripherin; 3) axons by their expression of calcitonin gene–related peptide (CGRP); 4) the presence of astrocytes expressing glial fibrillary acidic proteins (GFAP).MethodsA total of 240 open-apex root canals in dogs were used. After establishment of oral contamination to the pulp, the canals were cleaned, irrigated, and 120 canals filled with RAP, and the other 120 with calcium hydroxide.ResultsAfter 1, 3, and 6 months, teeth were recovered for immune-detection of protein markers associated with native pulp tissues. Regenerated pulp and apical papilla of RAP group revealed an abundance of stem cells showing intense immunoreactivity to Sox2 antibody, immunoreactivity of peripherin mainly in the A-fibers of the odontoblast layer and immunoreactivity to CGRP fibers in the central pulp region indicative of C-fibres. GFAP immunoreactivity was observed near the odontoblastic, cell-rich regions and throughout the regenerated pulp.ConclusionsRAP induces pulp regeneration following regenerative endodontic procedures with cells identity by gene expression demonstrating a distribution pattern similar to the authentic pulp innervation. A- and C-fibers, as well as GFAP specific to astrocytic differentiation, are recognized. The origin of the regenerated neural networks may be derived from the Sox2 identified stem cells within the apical papilla.

Purpose: To describe and evaluate normal adult dog dentition and temporomandibular joint anatomically and histologically in comparison to humans. Method: Five adult dogs (6-12 months old) were used in this study. The following anatomical structures were histologically evaluated in a qualitative fashion: Teeth and related bony structures mandible, mandibular condyle, disc, Zygomatic arch, temporal bone, glenoid fossa, retrodiscal tissue and synovia. The macroscopical and microscopic study of the human TMJ was based on the current literature. Results: Dogs have three Incisors, one canine, four premolars, and 2 molars in the upper and 3 molars in the lower jaw. The TMJ is surrounded by a thin fibrous tissue capsule, and a synovial lining. The mandibular angle has a prominent shape. The glenoid fossa is flat, with extended mediolaterally with retroarticular process. Histologically, the TMJ is composed of different tissues that comprise the mandibular head, mandibular fossa and fibrocartilaginous disc. A layer of fibrous tissue covers the articulating cortical condyle and temporal bone followed by a layer of hyaline cartilage. Conclusion: Morphologically and numerically dog's teeth are different from humans. Morphologically and histologically, the articular structure of dogs is, on the whole, similar to that of humans. In these animals there is no articular eminence, but they have a retroarticular process.

Receptor activator of NF-κB ligand (RANKL)-binding peptides inhibit bone resorption and were recently shown to activate bone formation. The stimulatory mechanism underlying bone formation associated with these peptides was explained as RANKL-reverse signaling, wherein RANKL molecules on osteoblasts work as receptors to stimulate osteoblast differentiation. However, why RANKL-binding peptides stimulate osteoblast differentiation while osteoprotegerin (OPG), which is well known to bind to RANKL, cannot activate osteoblast differentiation has remained unclear. In this mini-review, we introduce three main issues: (1) The inhibitory effects of two RANKL-binding peptides (W9 and OP3-4) on bone resorption; (2) The stimulatory effects of the RANKL-binding peptides on osteoblast differentiation; and (3) The accumulation and membrane clustering of RANKL molecules at the cell surface of osteoblasts as a potential molecular switch stimulating osteoblast differentiation by RANKL-binding peptides.

Tetracycline is used as a fluorescent reagent to measure bone formation activity in bone histomorphometric analyses. However, there is a possibility to lead a different conclusion when it is used in a bacteria-infected murine model since the tetracycline is considered to work as an antibiotic reagent. There are non-antibiotic fluorescent reagents such as alizarin and calcein for measuring bone formation activity. The purpose of this study was to clarify whether tetracycline could be an appropriate reagent to measure bone formation activity in a murine bacterial model in the same way as a non-antibiotic fluorescent reagent. We used Streptococcus mutans (S. mutans), a normal inhabitant in the oral cavity and tetracycline-sensitive bacteria, for inducing the bacterial model. The murine bacterial model was generated by intravenously inoculating S. mutans to the tail vein, followed immediately by the injection of the first fluorescent reagent, and the second one was injected 2 days prior to euthanization. After one day of inoculation with S. mutans, the subcutaneously injected alizarin had a similar colony count derived from the liver and the bone marrow tissue compared to the phosphate buffered saline (PBS)-injected control group. On the other hand, subcutaneous injection of tetracycline led to a significantly lower colony count from the liver compared to alizarin- or calcein-injected group. However, on day seven, after S. mutans intravenous injections, bone mineral density of distal femurs was significantly reduced by the bacteria inoculation regardless of which fluorescent reagents were injected subcutaneously. Finally, S. mutans inoculation reduced bone-formation-activity indices in both the tetracycline-alizarin double-injected mice and the calcein-alizarin double-injected mice. These results suggested that a one-time injection of tetracycline did not affect bone formation indices in the S. mutans-induced bone loss model. Tetracycline could be used for measuring bone formation activity in the same way as non-antibiotic fluorescent reagent such as calcein and alizarin, even in a tetracycline-sensitive bacterium-infected model.

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Background: Platelet-rich plasma (PRR) was proposed to serve as a possible scaffold model for regenerative endodontic therapies. However, its treatment outcomes are still controversial. Amelogenin protein has been shown to induce stem cell proliferation and differentiation. Thus, the aim of this study was to investigate whether the addition of amelogenin to PRP in non-vital immature permanent teeth with apical periodontitis can improve their treatment outcomes. Methods: Root canals of both maxillary and mandibular premolars in 8 mongrel dogs (n= 128) were instrumented and left open. After 14 days, in a second surgical procedure, canals were cleaned, irrigated, and treated with PRP (n=64) and PRP + amelogenin (n=64). After 1 and 3 months, animals were killed, and treated teeth were evaluated for histological and immune detection for nestin markers. Results: After 1 and 3 months post-surgery, PRP-treated canals showed full closure of opened apexes but minimal cementum, periodontal ligament (PDL), and bone regeneration. Importantly, no pulp regeneration was recognized. In contrast, PRP + amelogenin-treated canals at 1 and 3 months showed full closure of opened apexes, significant deposition of cementum, bone, regeneration of PDL, and pulp-like tissue regeneration compared to PRP-treated groups (p<0.001). By the 3-month period, full regeneration of all the lost dental-pulp complex tissues was seen, including vascular pulp-like tissue. Conclusions: PRP alone did not achieve the desired treatment outcomes, but after addition of amelogenin protein, it induced pulpal regeneration and regenerated the whole attachment apparatus. This combination could serve as a novel approach for regenerative endodontic therapy in non-vital immature permanent teeth with apical periodontitis. However, additional research is warranted to further evaluate the effect of such a combination in different animal settings before administrating this approach in clinical cases.

The receptor activator of NF-κB ligand (RANKL)-binding peptide is known to accelerate bone morphogenetic protein (BMP)-2-induced bone formation. Cholesterol-bearing pullulan (CHP)-OA nanogel-crosslinked PEG gel (CHP-OA nanogel-hydrogel) was shown to release the RANKL-binding peptide sustainably; however, an appropriate scaffold for peptide-accelerated bone formation is not determined yet. This study compares the osteoconductivity of CHP-OA hydrogel and another CHP nanogel, CHP-A nanogel-crosslinked PEG gel (CHP-A nanogel–hydrogel), in the bone formation induced by BMP-2 and the peptide. A calvarial defect model was performed in 5-week-old male mice, and scaffolds were placed in the defect. In vivo μCT was performed every week. Radiological and histological analyses after 4 weeks of scaffold placement revealed that the calcified bone area and the bone formation activity at the defect site in the CHP-OA hydrogel were significantly lower than those in the CHP-A hydrogel when the scaffolds were impregnated with both BMP-2 and the RANKL-binding peptide. The amount of induced bone was similar in both CHP-A and CHP-OA hydrogels when impregnated with BMP-2 alone. In conclusion, CHP-A hydrogel could be an appropriate scaffold compared to the CHP-OA hydrogel when the local bone formation was induced by the combination of RANKL-binding peptide and BMP-2, but not by BMP-2 alone.