Delivery of Alkaline Phosphatase Promotes Periodontal Regeneration in Mice

Factors regulating the ratio of pyrophosphate (PP_i) to phosphate (P_i) modulate biomineralization. Tissue-nonspecific alkaline phosphatase (TNAP) is a key promineralization enzyme that hydrolyzes the potent mineralization inhibitor PP_i. The goal of this study was to determine whether TNAP could promote periodontal regeneration in bone sialoprotein knockout mice (Ibsp^-/- mice), which are known to have a periodontal disease phenotype. Delivery of TNAP was accomplished either systemically (through a lentiviral construct expressing a mineral-targeted TNAP-D₁₀ protein) or locally (through addition of recombinant human TNAP to a fenestration defect model). Systemic TNAP-D₁₀ delivered by intramuscular injection at 5 d postnatal (dpn) increased circulating alkaline phosphatase (ALP) levels in Ibsp^-/- mice by 5-fold at 30 dpn, with levels returning to normal by 60 dpn when tissues were evaluated by micro-computed tomography and histology. Local delivery of recombinant human TNAP to fenestration defects in 5-wk-old wild type (WT) and Ibsp^-/- mice did not alter long-term circulating ALP levels, and tissues were evaluated by micro-computed tomography and histology at postoperative day 45. Systemic and local delivery of TNAP significantly increased alveolar bone volume (20% and 37%, respectively) and cementum thickness (3- and 42-fold) in Ibsp^-/- mice, with evidence for periodontal ligament attachment and bone/cementum marker localization. Local delivery significantly increased regenerated cementum and bone in WT mice. Addition of 100-μg/mL bovine intestinal ALP to culture media to increase ALP in vitro increased media P_i concentration, mineralization, and Spp1 and Dmp1 marker gene expression in WT and Ibsp^-/- OCCM.30 cementoblasts. Use of phosphonoformic acid, a nonspecific inhibitor of sodium P_i cotransport, indicated that effects of bovine intestinal ALP on mineralization and marker gene expression were in part through P_i transport. These findings show for the first time through multiple in vivo and in vitro approaches that pharmacologic modulation of P_i/PP_i metabolism can overcome periodontal breakdown and accomplish regeneration.