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J. Biol. Chem., Vol. 258, Issue 20, 12221-12227, Oct, 1983
JF Nagelkerke, KP Barto and TJ van Berkel
Isolation and separation of rat liver cells into endothelial, Kupffer, and
parenchymal cell fractions were performed at different times after
injection of human 125I-acetyl low density lipoproteins (LDL). In order to
minimize degradation and redistribution of the injected lipoprotein during
cell isolation, a low temperature (8 degrees C) procedure was applied. Ten
min after injection, isolated endothelial cells contained 5 times more
acetyl-LDL apoprotein per mg of cell protein than the Kupffer cells and 31
times more than the hepatocytes. A similar relative importance of the
different cell types in the uptake of acetyl- LDL was observed 30 min after
injection. For studies on the in vitro interaction of endothelial and
Kupffer cells with acetyl-LDL, the cells were isolated with a collagenase
perfusion at 37 degrees C. Pure endothelial (greater than 95%) and purified
Kupffer cells (greater than 70%) were obtained by a two-step elutriation
method. It is demonstrated that the rat liver endothelial cell possesses a
high affinity receptor specific for the acetyl-LDL because a 35-fold excess
of unlabeled acetyl-LDL inhibits association of the labeled compound for
70%, whereas unlabeled native human LDL is ineffective. Binding to the
acetyl-LDL receptor is coupled to rapid uptake and degradation of the
apolipoprotein. Addition of the lysosomotropic agents chloroquine (50
microM) or NH4Cl (10 mM) resulted in more than 90% inhibition of the high
affinity degradation, indicating that this occurs in the lysosomes. With
the purified Kupffer cell fraction, the cell association and degradation of
acetyl-LDL was at least 4 times less per mg of cell protein than with the
pure endothelial cells. Although cells isolated with the cold pronase
technique are also still able to bind and degrade acetyl-LDL, it appeared
that 40-60% of the receptors are destroyed or inactivated during the
isolation procedure. It is concluded that the rat liver endothelial cell is
the main cell type responsible for acetyl-LDL uptake.
In vivo and in vitro uptake and degradation of acetylated low density lipoprotein by rat liver endothelial, Kupffer, and parenchymal cells
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