An analysis of the role of a retroendocytosis pathway in ABCA1-mediated cholesterol efflux from macrophages.
The ATP binding cassette transporter A-1 (ABCA1) is critical for apolipoprotein-mediated cholesterol efflux, an important mechanism employed by macrophages to avoid becoming lipid-laden foam cells, the hallmark of early atherosclerotic lesions. It has been proposed that lipid-free apolipoprotein A-I (apoA-I) enters the cell and is resecreted as a lipidated particle via a retroendocytosis pathway during ABCA1-mediated cholesterol efflux from macrophages. To determine the functional importance of such a pathway, confocal microscopy was used to characterize the internalization of a fully functional apoA-I cysteine mutant containing a thiol-reactive fluorescent probe in cultured macrophages. ApoA-I was also endogenously labeled with (35)S-methionine to quantify cellular uptake and to determine the metabolic fate of the internalized protein. It was found that apoA-I was specifically taken inside macrophages and that a small amount of intact apoA-I was resecreted from the cells. However, a majority of the label that reappeared in the media was degraded. We estimate that the mass of apoA-I retroendocytosed is not sufficient to account for the HDL produced by the cholesterol efflux reaction. Furthermore, we have demonstrated that lipid-free apoA-I-mediated cholesterol efflux from macrophages can be pharmacologically uncoupled from apoA-I internalization into cells. On the basis these findings, we present a model in which the ABCA1-mediated lipid transfer process occurs primarily at the membrane surface in macrophages, but still accounts for the observed specific internalization of apoA-I.
Faulkner
Loren E
LE
Department of Pathology and Laboratory Medicine, University of Cincinnati, Cincinnati OH, 45237, USA.
Panagotopulos
Stacey E
SE
Johnson
Jacob D
JD
Woollett
Laura A
LA
Hui
David Y
DY
Witting
Scott R
SR
Maiorano
J Nicholas
JN
Davidson
W Sean
WS
D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters / Q000502:physiology*; D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D002784:Cholesterol / Q000378:metabolism*; D004705:Endocytosis; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D018613:Microscopy, Confocal
D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters; D002784:Cholesterol