Endocytotic uptake of small unilamellar liposomes by human trophoblast cells in culture

Abstract

We evaluated the mechanism of uptake of carboxyfluorescein-containing small unilamellar liposomes of different surface charge by trophoblast cells in culture. Carboxyfluorescein-encapsulated neutral liposomes were prepared by using equimolar concentrations of lecithin and cholesterol. Anionic and cationic liposomes were prepared by adding dicetylphosphate and stearylamine. Trophoblast cells from human term placenta were cultured and incubated on the first day at 37°C with liposome-encapsulated carboxyfluorescein or 500 nM of free carboxyfluorescein. The mechanism of uptake was determined by pre-treating the cells with metabolic inhibitors: 2 mM of sodium azide and 25 mM of deoxyglucose for 30 min. The uptake of liposomes was also evaluated both qualitatively under fluorescent microscope and quantitatively by measuring carboxyfluorescein fluorometrically. The uptake of free carboxyfluorescein and cationic liposomes was comparable. The anionic liposomes were taken up by the trophoblast cells more avidly than the neutral (13.2 ± 1.6 versus 9.5 ± 1.4%; P < 0.01), cationic (2.9 ± 0.4%; P < 0.001) and the free carboxyfluorescein (2.1 ± 0.9%; P < 0.01). When cells were pre-treated with metabolic inhibitors, the uptake of anionic (5.9 ± 1.8%; P < 0.001) and neutral liposomes (4.0 ± 0.8%; P < 0.01) was significantly reduced,whereas uptake of cationic and free carboxyfluorescein remained unaltered. This study indicates that small unilamellar liposomes are internalized by the trophoblast cells in culture by an energy-dependent pathway; most probably by endocytosis. The neutral and anionic liposomes are internalized more avidly than cationic liposomes.