Clodronate-Loaded Liposome Treatment Has Site-Specific Skeletal Effects.

Ineffective oral wound healing is detrimental to patients' oral health-related quality of life. Delineating the cellular mechanisms involved in optimal healing will elicit better approaches to treating patients with compromised healing. Osteal macrophages have recently emerged as important positive regulators of bone turnover. The contributions of macrophages to long bone healing have been studied, but their role in oral osseous wound healing following tooth extraction is less clear. Clodronate-loaded liposomes were used as a tool to deplete macrophages in C57BL/6J mice and assess oral osseous bone fill after extraction. In addition to macrophage ablation, osteoclast ablation occurred. Interestingly, depletion of macrophages and osteoclasts via clodronate treatment had differential effects based on skeletal location. In the nonwounded tibiae, clodronate treatment significantly increased CD68+ cells and decreased F4/80+ cells in the marrow, which correlated with increased trabecular bone volume fraction after 7 and 14 d. Serum formation and resorptive markers P1NP and TRAcP 5b were decreased as were tibial TRAP+ osteoclasts. In healing extraction sockets, clodronate treatment increased extraction socket trabecular bone thickness at 14 d, which correlated with decreased TRAP+ osteoclasts and F4/80+ macrophages. Conversely, nonwounded maxillary interseptal bone was unaffected by clodronate treatment. Furthermore, the increase in extraction socket bone fill with clodronate was less than the large increase in trabecular bone observed in a nonwounded long bone. These data suggest a temporal and spatial specificity in the roles of macrophages and osteoclasts in normal turnover and healing.

Role of PHOSPHO1 in Periodontal Development and Function.

The tooth root and periodontal apparatus, including the acellular and cellular cementum, periodontal ligament (PDL), and alveolar bone, are critical for tooth function. Cementum and bone mineralization is regulated by factors including enzymes and extracellular matrix proteins that promote or inhibit hydroxyapatite crystal growth. Orphan Phosphatase 1 (Phospho1, PHOSPHO1) is a phosphatase expressed by chondrocytes, osteoblasts, and odontoblasts that functions in skeletal and dentin mineralization by initiating deposition of hydroxyapatite inside membrane-limited matrix vesicles. The role of PHOSPHO1 in periodontal formation remains unknown and we aimed to determine its functional importance in these tissues. We hypothesized that the enzyme would regulate proper mineralization of the periodontal apparatus. Spatiotemporal expression of PHOSPHO1 was mapped during periodontal development, and Phospho1(-/-) mice were analyzed using histology, immunohistochemistry, in situ hybridization, radiography, and micro-computed tomography. The Phospho1 gene and PHOSPHO1 protein were expressed by active alveolar bone osteoblasts and cementoblasts during cellular cementum formation. In Phospho1(-/-) mice, acellular cementum formation and mineralization were unaffected, whereas cellular cementum deposition increased although it displayed delayed mineralization and cementoid. Phospho1(-/-) mice featured disturbances in alveolar bone mineralization, shown by accumulation of unmineralized osteoid matrix and interglobular patterns of protein deposition. Parallel to other skeletal sites, deposition of mineral-regulating protein osteopontin (OPN) was increased in alveolar bone in Phospho1(-/-) mice. In contrast to the skeleton, genetic ablation of Spp1, the gene encoding OPN, did not ameliorate dentoalveolar defects in Phospho1(-/-) mice. Despite alveolar bone mineralization defects, periodontal attachment and function appeared undisturbed in Phospho1(-/-) mice, with normal PDL architecture and no evidence of bone loss over time. This study highlights the role of PHOSPHO1 in mineralization of alveolar bone and cellular cementum, further revealing that acellular cementum formation is not substantially regulated by PHOSPHO1 and likely does not rely on matrix vesicle-mediated initiation of mineralization.