AJP - Lung Cellular and Molecular Physiology, Vol 269, Issue 1 11-L19, Copyright © 1995 by American Physiological Society
Pathways for uptake of fluorescently labeled liposomes by alveolar type II cells in culture
W. J. Muller, K. Zen, A. B. Fisher and H. Shuman
Institute for Environmental Medicine, University of Pennsylvania Medical Center, Philadelphia 19104-6068, USA.
Isolated alveolar type II cells are shown in real time to internalize
1-palmitoyl-2-[12-[(7 nitro-2-1,3-benzoxadiazol-4-yl)amino]dodecanoyl]
phosphatidyl choline (C12-NBD-PC)-labeled liposomes in an intact manner
with the use of fluorescence microscopy. Cells isolated from rat lungs,
cultured on plastic for 24 h, are exposed at 37 degrees C to liposomes on
the microscope stage, using a microperfusion system. Liposomes composed of
a self-quenching concentration of fluorophore (25 mol% C12-NBD-PC) do not
dequench at the level of the plasma membrane; rather, they appear to fuse
with intracellular lipids primarily in vesicular structures consistent with
lamellar bodies. Temperature and energy dependence of this uptake mechanism
is demonstrated by the lack of uptake of maximally fluorescent liposomes
(15 mol% C12-NBD-PC) at 4 degrees C or when depleted of ATP. Inhibition of
the coated-pit endocytic pathway by hypertonic media reduces fluorescent
lipid uptake by 50%, suggesting a separate, clathrin-independent endocytic
pathway for intact internalization of liposomes by type II cells. Uptake is
inhibited by cytochalasin D by 49%, suggesting a dependence on action, and
by 88% with combined cytochalasin D and hypertonicity, or nearly the same
level as ATP depletion. The evidence is consistent with the existence of
two separate endocytic pathways for lipids in type II cells: one clathrin
dependent and the other actin dependent.
