Interaction of French-pressed liposomes with isolated bovine adrenal chromaffin cells. Characterization of the cell-liposome interactions.
Small unilamellar liposomes with an average external diameter of approximately 550 A were prepared by high pressure extrusion in a French press. Liposomes, composed of phosphatidylcholine, phosphatidylserine, and cholesterol at a molar ratio of 7:1:2, were incubated with suspensions of bovine adrenal chromaffin cells. The cell-liposome interactions were characterized using fluorescence and radiotracer techniques. Transfer of the liposomal contents into the cytoplasm was visualized by fluorescence microscopy, using fluorescence-labeled macromolecules, and further documented by flow cytometry with liposome-entrapped 5,6-carboxy-fluorescein. The dose dependence, time course, and temperature dependence of the cell-liposome association, as determined by radioactive labeling both the liposomal membranes and their contents, indicate saturable interaction of the cells with intact liposomes (KappM approximately 5 X 10(-7) M lipid/10(6) cells at 37 degrees C). Using nonexchangeable fluorescent phospholipid analogs, the cell-liposome interactions were characterized by fluorescence resonance energy transfer and by fluorescence recovery after photobleaching. From these latter experiments we conclude that after 1-h incubation of 10(6) cells with 1 microM lipid at 37 degrees C, 30% of the cell-associated liposomes will have fused with the plasma membranes, resulting in the delivery of the contents of approximately 1.25 X 10(5) liposomes into each cell. Thus, liposomal delivery is an effective means to gain access to the cytoplasm and can be exploited to modulate physiological responses from within intact chromaffin cells.
D000311:Adrenal Glands / Q000166:cytology*; D000818:Animals; D002232:Carbocyanines / Q000378:metabolism; D002417:Cattle; D002838:Chromaffin System / Q000166:cytology*; D003593:Cytoplasm / Q000378:metabolism; D004735:Energy Transfer; D005434:Flow Cytometry; D005456:Fluorescent Dyes; D007700:Kinetics; D008081:Liposomes / Q000378:metabolism*; D008561:Membrane Fusion; D008856:Microscopy, Fluorescence; D010743:Phospholipids / Q000378:metabolism; D011312:Pressure; D013050:Spectrometry, Fluorescence
D002232:Carbocyanines; D005456:Fluorescent Dyes; D008081:Liposomes; D010743:Phospholipids; C024286:3,3'-dioctadecylindocarbocyanine