Dose-Dependent Osteogenic Effect of Octacalcium Phosphate on Mouse Bone Marrow Stromal Cells

Abstract

Octacalcium phosphate (OCP) has been advocated to be a precursor of biological apatite crystals in bones and teeth. Our previous studies showed that synthetic OCP stimulates bone regeneration, followed by the progressive conversion of OCP into hydroxyapatite (HA), when implanted in bone defects. However, the precise mechanism to induce the osteogenic phenotype in osteoblasts by OCP has not been identified. The present study was designed to investigate whether the physicochemical aspect, specific to and derived from the structural properties of OCP, influences the function of an osteoblastic cell line, mouse bone marrow stromal ST-2 cells. Different amounts of synthetic OCP and synthetic sintered ceramic HA were coated onto 48-well tissue culture plates. The amounts of OCP and HA were controlled to strengthen their intrinsic physicochemical properties, in which the milieu around the crystals will be modified during the culture. The roughness of the OCP coatings was independent of the amount of coating. Chemical analyses of the supernatants of the OCP coatings revealed that the concentration of Ca2+ decreased with increasing amounts of OCP, while the concentration of inorganic phosphate increased markedly, most probably through OCP--apatite conversion. ST-2 cells were cultured on the OCP or HA coatings up to day 21. The OCP coating caused a significant decrease in cell attachment and in the initial stage of proliferation, dependent upon the amount of coating. On the other hand, OCP enhanced the expression of osteogenic markers, including type I collagen, alkaline phosphatase, and osterix. However, HA did not alter the expression of these markers in ST-2 cells cultured on different amounts of HA coating. These results demonstrated that OCP is capable of inducing the differentiation of stromal cells into osteoblastic cells, especially differentiation into early stage osteoblastic cells, prior to reaching the stage of mature osteoblastic cell lineage.

Osteoblastic differentiation of stromal ST-2 cells from octacalcium phosphate exposure via p38 signaling pathway

“…We previously reported that OCP causes changes of Ca 2+ and Pi ion concentrations in acellular culture medium 14) . In the present study, we evaluated the changes of these ion concentrations in cell culture medium caused by OCP-apatite conversion.…”

“…Total RNA (1 μg) was used for cDNA synthesis using the Thermo Script TM RT-PCR system (Invitrogen). The primers and universal probe used in this study were previously reported 14) , except for osteocalcin (OCN) primers, which are as follows:…”

“…The biological responses to OCP have also been investigated from its coating on metallic implants 11,12) , in osteoblastic cell proliferation, or from implantation into subcutaneous tissue or bone defects 8,12) . Recent studies suggested that osteoblastic cell differentiation is enhanced upon contact with OCP, which tends to convert into HA 13) , and differentiation is significantly facilitated in an OCP dose-dependent manner 14) . The chemical route from OCP to HA involves an uptake of calcium ions (Ca 2+ ) and release of inorganic phosphate (Pi) ions 15) .…”

Cranial bone defects: current and future strategies

“…Octacalcium phosphate is a biological precursor of apatite crystals 11 and has been shown to promote differentiation and maturation of osteoblasts in vitro. 6,72,108 Studies have also shown that OCP, compared with HA or b-TCP, has the capacity to increase attachment of osteoblasts to the scaffold. 32 Furthermore, OCP/collagen scaffolds have been demonstrated to direct improved bone regeneration in rat calvarial defects compared with HA or b-TCP alone or HA/collagen or b-TCP/ collagen scaffolds.…”

The impact of octacalcium phosphate on the dynamics of bone matrix formation in experimental bone defects

“…Трансформация ОКФ в ГА -один из факторов, сти-мулирующих дифференциацию остеобластных клеток [3,9,14,16].…”