Association of Methotrexate Encapsulated in Negatively Charged Liposomes with Cells at Nanomolar Lipid Concentrations

Abstract

AbstractAn in vitro liposome-cell association system has been developed that will allow the study of uptake and metabolism of liposomes by cultured cells at nanomolar lipid concentrations. The fate of cell associated liposomes is followed through the liposome encapsulated marker, methotrexate. Detection is based on the inhibition of dihydrofolate reductase by methotrexate, after its release from cells through boiling. Methotrexate in phospha-tidylglycerol (PG) liposomes is taken up by cells and then subsequently lost from the cells. Uptake is partially blocked by monensin. Loss from the cells is blocked by metabolic inhibitors, monensin, ammonium chloride, and chloroquine. Methotrexate in distearoylphosphatidylglycerol (DSPG) liposomes is taken up by cells slowly, and there is minimal lost of methotrexate after uptake. Pulse studies show that metabolism of PG liposomes after endocytosis is occurring at a much higher rate than that of DSPG liposomes, and substantial retention of encapsulated methotrexate o...