Characterization of inorganic phosphate transport in osteoclast-like cells

Abstract

Osteoclasts possess inorganic phosphate (Pi) transport systems to take up external Piduring bone resorption. In the present study, we characterized Pitransport in mouse osteoclast-like cells that were obtained by differentiation of macrophage RAW264.7 cells with receptor activator of NF-κB ligand (RANKL). In undifferentiated RAW264.7 cells, Pitransport into the cells was Na+dependent, but after treatment with RANKL, Na+-independent Pitransport was significantly increased. In addition, compared with neutral pH, the activity of the Na+-independent Pitransport system in the osteoclast-like cells was markedly enhanced at pH 5.5. The Na+-independent system consisted of two components with Kmof 0.35 mM and 7.5 mM. The inhibitors of Pitransport, phosphonoformic acid, and arsenate substantially decreased Pitransport. The proton ionophores nigericin and carbonyl cyanide p-trifluoromethoxyphenylhydrazone as well as a K+ionophore, valinomycin, significantly suppressed Pitransport activity. Analysis of BCECF fluorescence indicated that Pitransport in osteoclast-like cells is coupled to a proton transport system. In addition, elevation of extracellular K+ion stimulated Pitransport, suggesting that membrane voltage is involved in the regulation of Pitransport activity. Finally, bone particles significantly increased Na+-independent Pitransport activity in osteoclast-like cells. Thus, osteoclast-like cells have a Pitransport system with characteristics that are different from those of other Na+-dependent Pitransporters. We conclude that stimulation of Pitransport at acidic pH is necessary for bone resorption or for production of the large amounts of energy necessary for acidification of the extracellular environment.