Liposome-mediated calcium phosphate formation in metastable solutions

The present study examined the effect of anionic liposomes on precipitate formation in supersaturated calcium phosphate solutions. The liposomes were prepared by dispersing 7:2:1 molar mixtures of phosphatidylcholine, dicetyl phosphate, and cholesterol in buffered aqueous solutions containing 0 or 50 mM inorganic phosphate (PI). Unencapsulated PI was removed by gel filtration. The liposomes were then suspended in reaction solutions containing 2.25 mM Ca2+ and either 0, 1.0, or 1.5 mM PI. All experiments were carried out at 22 degrees C, pH 7.4, and 240 mOsm. External solution Ca2+ and PI losses were found to be appreciable only when the membranes of liposomes containing entrapped PI were made permeable to Ca2+ with the addition to the suspension of the ionophore X-537A. The Ca2+ losses, moreover, were up to 3 times as great (1.5 vs 0.5 mM) when accompanied by external PI losses than in PI-free external solutions where Ca2+ alone was involved. Previous studies showed that in this latter situation, decline in external Ca2+ concentration was the result of precipitate formation in the aqueous interiors of the liposomes. The present findings suggest that when the external solution phase was metastable, the apparent coupling of large additional Ca2+ losses with intraliposomal precipitation was due to secondary precipitation brought about by the seeding action of intraliposomal crystals penetrating into the external solution. The results may explain in part the origin of extravesicular mineral deposits in matrix vesicle calcification.