High biocompatibility and improved osteogenic potential of amorphous calcium carbonate/vaterite

In human bone, amorphous calcium carbonate (ACC) is formed as a precursor of the crystalline carbonated apatite/hydroxyapatite (HA). Here we describe that the metastable ACC phase can be stabilized by inorganic polyphosphate (polyP) that is also used as a phosphate source for the non-enzymatic carbonate/phosphate exchange during HA formation. This polymer was found to suppress the transformation of ACC into crystalline CaCO₃ at a percentage of 5% [w/w] ("CCP5") with respect to CaCO₃ and almost completely at 10% [w/w] ("CCP10"). Both preparations (CaCO₃/polyP) are amorphous, but also contain small amounts of vaterite, as revealed by XRD, FTIR and SEM analyses. They did not affect the growth/viability of SaOS-2 cells. Cell culture and Ca²⁺ release experiments revealed that the CaCO₃ particles formed in the presence of polyP (CaCO₃/polyP) are degradable and, unlike calcite, become disintegrated with time during the cell culture incubation. Again in contrast to calcite, "CCP5" and "CCP10" were found to exhibit osteogenic activity and induce the expression of alkaline phosphatase gene in SaOS-2 cells as well as in human mesenchymal stem cells (MSC). In vivo studies in rats, using PLGA microspheres inserted in the muscles of the back of the animals, revealed that the encapsulated "CCP10" is not only biocompatible but also supports the regeneration at the implant region. We conclude that ACC containing small amounts of vaterite has osteogenic potential and offers superior properties compared to the biologically inert calcite with respect to a potential application as a scaffold material for bone implants.