Accelerating Socket Repair via WNT3A Curtails Alveolar Ridge Resorption.

Tooth extraction triggers alveolar ridge resorption, and when this resorption is extensive, it can complicate subsequent reconstructive procedures that use dental implants. Clinical data demonstrate that the most significant dimensional changes in the ridge occur soon after tooth extraction. Here, we sought to understand whether a correlation existed between the rate at which an extraction socket heals and the extent of alveolar ridge resorption. Maxillary molars were extracted from young and osteoporotic rodents, and quantitative micro-computed tomographic imaging, histology, and immunohistochemistry were used to simultaneously follow socket repair and alveolar ridge resorption. Extraction sockets rapidly filled with new bone via the proliferation and differentiation of Wnt-responsive osteoprogenitor cells and their progeny. At the same time that new bone was being deposited in the socket, tartrate-resistant acid phosphatase-expressing osteoclasts were resorbing the ridge. Significantly faster socket repair in young animals was associated with significantly more Wnt-responsive osteoprogenitor cells and their progeny as compared with osteoporotic animals. Delivery of WNT3A to the extraction sockets of osteoporotic animals restored the number of Wnt-responsive cells and their progeny back to levels seen in young healthy animals and accelerated socket repair in osteoporotic animals back to rates seen in the young. In cases where the extraction socket was treated with WNT3A, alveolar ridge resorption was significantly reduced. These data demonstrate a causal link between enhancing socket repair via WNT3A and preserving alveolar ridge dimensions following tooth extraction.

Interspecies Comparison of Alveolar Bone Biology, Part I: Morphology and Physiology of Pristine Bone.

INTRODUCTION: Few interspecies comparisons of alveolar bone have been documented, and this knowledge gap raises questions about which animal models most accurately represent human dental conditions or responses to surgical interventions. OBJECTIVES: The objective of this study was to employ state-of-the-art quantitative metrics to directly assess and compare the structural and functional characteristics of alveolar bone among humans, mini pigs, rats, and mice. METHODS: The same anatomic location (i.e., the posterior maxillae) was analyzed in all species via micro-computed tomographic imaging, followed by quantitative analyses, coupled with histology and immunohistochemistry. Bone remodeling was evaluated with alkaline phosphatase activity and tartrate-resistant acid phosphatase staining to identify osteoblast and osteoclast activities. In vivo fluorochrome labeling was used as a means to assess mineral apposition rates. RESULTS: Collectively, these analyses demonstrated that bone volume differed among the species, while bone mineral density was equal. All species showed a similar density of alveolar osteocytes, with a highly conserved pattern of collagen organization. Collagen maturation was equal among mouse, rat, and mini pig. Bone remodeling was a shared feature among the species, with morphologically indistinguishable hemiosteonal appearances, osteocytic perilacunar remodeling, and similar mineral apposition rates in alveolar bone. CONCLUSIONS: Our analyses demonstrated equivalencies among the 4 species in a plurality of the biological features of alveolar bone. Despite contradictory results from older studies, we found no evidence for the superiority of pig models over rodent models in representing human bone biology. KNOWLEDGE TRANSFER STATEMENT: Animal models are extensively used to evaluate bone tissue engineering strategies, yet there are few state-of-the-art studies that rigorously compare and quantify the factors influencing selection of a given animal model. Consequently, there is an urgent need to assess preclinical animal models for their predictive value to dental research. Our article addresses this knowledge gap and, in doing so, provides a foundation for more effective standardization among animal models commonly used in dentistry.

Defective Mineralization in X-Linked Hypophosphatemia Dental Pulp Cell Cultures.

X-linked hypophosphatemia (XLH) is a skeletal disease caused by inactivating mutations in the PHEX gene. Mutated or absent PHEX protein/enzyme leads to a decreased serum phosphate level, which cause mineralization defects in the skeleton and teeth (osteomalacia/odontomalacia). It is not yet altogether clear whether these manifestations are caused solely by insufficient circulating phosphate availability for mineralization or also by a direct, local intrinsic effect caused by impaired PHEX activity. Here, we evaluated the local role of PHEX in a 3-dimensional model of extracellular matrix (ECM) mineralization. Dense collagen hydrogels were seeded either with human dental pulp cells from patients with characterized PHEX mutations or with sex- and age-matched healthy controls and cultured up to 24 d using osteogenic medium with standard phosphate concentration. Calcium quantification, micro-computed tomography, and histology with von Kossa staining for mineral showed significantly lower mineralization in XLH cell-seeded scaffolds, using nonparametric statistical tests. While apatitic mineralization was observed along collagen fibrils by electron microscopy in both groups, Raman microspectrometry indicated that XLH cells harboring the PHEX mutation produced less mineralized scaffolds having impaired mineral quality with less carbonate substitution and lower crystallinity. In the XLH cultures, immunoblotting revealed more abundant osteopontin (OPN), dentin matrix protein 1 (DMP1), and matrix extracellular phosphoglycoprotein (MEPE) than controls, as well as the presence of fragments of these proteins not found in controls, suggesting a role for PHEX in SIBLING protein degradation. Immunohistochemistry revealed altered OPN and DMP1 associated with an increased alkaline phosphatase staining in the XLH cultures. These results are consistent with impaired PHEX activity having local ECM effects in XLH. Future treatments for XLH should target both systemic and local manifestations.

Phosphate and Vitamin D Prevent Periodontitis in X-Linked Hypophosphatemia.

X-linked hypophosphatemia (XLH) is a rare genetic skeletal disease where increased phosphate wasting in the kidney leads to hypophosphatemia and prevents normal mineralization of bone and dentin. Here, we examined the periodontal status of 34 adults with XLH and separated them according to the treatment they received for hypophosphatemia. We observed that periodontitis frequency and severity were increased in adults with XLH and that the severity varied according to the hypophosphatemia treatment. Patients who benefited from an early and continuous vitamin D and phosphate supplementation during their childhood presented less periodontal attachment loss than patients with late or incomplete supplementation. Continued hypophosphatemia treatment during adulthood further improved the periodontal health. Extracted teeth from patients with late or incomplete supplementation showed a strong acellular cementum hypoplasia when compared with age-matched healthy controls. These results show that XLH disturbs not only bone and dentin formation but also cementum and that the constitutional defect of the attachment apparatus is associated with attachment loss.

Abnormal osteopontin and matrix extracellular phosphoglycoprotein localization, and odontoblast differentiation, in X-linked hypophosphatemic teeth.

Mutations in phosphate-regulating gene (PHEX) lead to X-linked hypophosphatemic rickets (XLH), a genetic disease characterized by impaired mineralization in bones and teeth. In human XLH tooth dentin, calcospherites that would normally merge as part of the mineralization process are separated by unmineralized interglobular spaces where fragments of matrix proteins accumulate. Here, we immunolocalized osteopontin (OPN) in human XLH teeth, in a three-dimensional XLH human dental pulp stem cell-collagen scaffold culture model and in a rat tooth injury repair model treated with acidic serine- and aspartate-rich motif peptides (ASARM). In parallel, matrix extracellular phosphoglycoprotein (MEPE) immunolocalization and alkaline phosphatase (ALP) activity were assessed in XLH teeth. OPN was expressed by odontoblasts in the XLH models, and localized to the abnormal calcospherites of XLH tooth dentin. In addition, ALP activity and MEPE localization were abnormal in human XLH teeth, with MEPE showing an accumulation in the unmineralized interglobular spaces in dentin. Furthermore, XLH odontoblasts failed to form a well-polarized odontoblast layer. These data suggest that both MEPE and OPN are involved in impaired tooth mineralization associated with XLH, possibly through different effects on the mineralization process.

Mineralization of dense collagen hydrogel scaffolds by human pulp cells.

While advances in biomineralization have been made in recent years, unanswered questions persist on bone- and tooth-cell differentiation, on outside-in signaling from the extracellular matrix, and on the link between protein expression and mineral deposition. In the present study, we validate the use of a bioengineered three-dimensional (3D) dense collagen hydrogel scaffold as a cell-culture model to explore these questions. Dental pulp progenitor/stem cells from human exfoliated deciduous teeth (SHEDs) were seeded into an extracellular matrix-like collagen gel whose fibrillar density was increased through plastic compression. SHED viability, morphology, and metabolic activity, as well as scaffold mineralization, were investigated over 24 days in culture. Additionally, measurements of alkaline phosphatase enzymatic activity, together with immunoblotting for mineralized tissue cell markers ALPL (tissue-non-specific alkaline phosphatase), DMP1 (dentin matrix protein 1), and OPN (osteopontin), demonstrated osteo/odontogenic cell differentiation in the dense collagen scaffolds coincident with mineralization. Analyses of the mineral phase by electron microscopy, including electron diffraction and energy-dispersive x-ray spectroscopy, combined with Fourier-transform infrared spectroscopy and biochemical analyses, were consistent with the formation of apatitic mineral that was frequently aligned along collagen fibrils. In conclusion, use of a 3D dense collagen scaffold promoted SHED osteo/odontogenic cell differentiation and mineralization.