Inorganic polyphosphate accelerates bone mineralization

Objectives: Inorganic polyphosphates [Poly(P)] are orthophosphate polymers found in many mammalian species and in many cell types. It has been suggested that mammalian Poly(P) stimulates the growth of fibroblasts on periodontal ligament and activates osteogenetic cells. Thus Poly(P) is a possible stimulant of bone formation. However mechanisms of osteogenesis and calcification of the bone matrix have not yet been clarified. In this study, we investigated the effects of Poly(P) on bone mineralization using mouse osteoblast-like cells and chondrogenic precurser cells as in vitro cell models.

Methods: The cells used were the mouse osteoblast-like cell line MC3T3-E1 and the chondrogenic precurser cell line ATDC5. Poly(P), with an average chain length 60-75, was used at a final concentration of 1 mM. Reverse transcription polymerase chain reaction (RT-PCR) and Real Time PCR were performed to examine gene expression. Calcification of the cell matrix was examined by alizarin red staining. To visualize polyphosphatase activity, thin column chromatography analyses using [³²P] labeled Poly(P) were performed.

Results: Both Poly(P) treated MC3T3-E1 and ATDC5 cells showed increased calcification of the cell matrix. The expression of the osteogenic marker genes including osteopontin, osteocalcin, collagen type I, alkaline phosphatase and the progressive ankylosis gene were also increased in Poly(P) treated cells. In addition, column chromatographic analysis showed that degradation of Poly(P) into phosphates, pyrophosphates, and Poly(P) oligomers was activated by a Poly(P) treatment.

Conclusion: These results suggested that Poly(P) modulates phosphate metabolism and promotes calcification by the induction of bone matrix protein and phosphatase activity.