Modulation of calcium phosphate formation by phosphatidate-containing anionic liposomes

Summary

Liposomes prepared from 7∶2∶1 molar mixtures of phosphatidylcholine, dicetyl phosphate, and cholesterol to which 1–20 mole % dioleoylphosphatidic acid (DOPA) was added were used to examine the effect of membrane-bound monoester phosphatidate phosphate anions on calcium phosphate formation in aqueous solutions at 22°C, pH 7.4, and 240 mOsm. Results showed that up to 20 mole % DOPA in the liposomal envelope did not initiate mineralization in solutions made meta-stable with 2.25 mM CaCl₂ and 1.50 mM KH₂PO₄. Results also revealed that precipitation induced in metastable solutions by the seeding action of intraliposomally formed mineral was measurably reduced with as little as 1 mole % DOPA and completely suppressed with 5 mole % DOPA. In contrast, 10 mole % concentrations of diester phosphate lipids either had no effect on extraliposomal precipitation (e.g., phosphatidylglycerol and phosphatidylinositol) or, as reported previously for phosphatidylserine, only partially reduced the amount of precipitate formed. Transmission electron microscopical analysis suggests that DOPA-containing lipid bilayers adhering to the seed crystals inhibited extraliposomal mineralization by encapsulating the crystals within the liposomes and/or by blocking potential growth sites on the crystal faces. The polar head group of DOPA, being more negatively charged and sterically less encumbered than diester phosphate ligands, most probably was responsible for this adherence of the lipid bilayers to the crystal surfaces.