The bidirectional flux of cholesterol between cells and lipoproteins. Effects of phospholipid depletion of high density lipoprotein.
The bidirectional surface transfer of free cholesterol (FC) between Fu5AH rat hepatoma cells and human high density lipoprotein (HDL) was studied. Cells and HDL were prelabeled with [4-14C]FC and [7-3H]FC, respectively. Influx and efflux of FC were measured simultaneously from the appearance of 3H counts in cells and 14C counts in medium. Results were analyzed by a computerized procedure which fitted sets of kinetic data to a model assuming that cell and HDL FC populations each formed a single homogeneous pool and that together the pools formed a closed system. This analysis yielded values for the first-order rate constants of FC influx and efflux (ki and ke), from which influx and efflux of FC mass (Fi and Fe) could be calculated. With normal HDL, the uptake and release of FC tracers conformed well to the above-described model; Fi and Fe were approximately equal, suggesting an exchange of FC between cells and HDL. HDL was depleted of phospholipid (PL) by treatment with either phospholipase A2 or heparin-releasable rat hepatic lipase, followed by incubation with bovine serum albumin. PL depletion of HDL had little or no effect on ki, but reduced ke, indicating that PL-deficient HDL is a relatively poor acceptor of cell cholesterol. The reduction in ke resulted in initial Fi greater than Fe and, thus, in net uptake of FC by the cells. This result explained previous results demonstrating net uptake of FC from PL-depleted HDL. In the presence of an inhibitor of acyl coenzyme A:cholesterol acyltransferase, the steady state distribution of FC mass between cells and HDL was accurately predicted by the ratio of rate constants for FC flux. This result provided additional validation for describing FC flux in terms of first-order rate constants and homogeneous cell and HDL FC pools.
W J
WJ
Johnson
M J
MJ
Bamberger
R A
RA
Latta
P E
PE
Rapp
M C
MC
Phillips
G H
GH
Rothblat
D000818:Animals; D001692:Biological Transport; D002250:Carbon Radioisotopes; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism*; D008076:Cholesterol, HDL / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008049:Lipase / Q000378:metabolism; D008075:Lipoproteins, HDL / Q000097:blood*; D008099:Liver / Q000201:enzymology; D008114:Liver Neoplasms, Experimental / Q000378:metabolism*; D008563:Membrane Lipids / Q000502:physiology*; D010743:Phospholipids / Q000502:physiology*; D051381:Rats; D014316:Tritium
D002250:Carbon Radioisotopes; D008076:Cholesterol, HDL; D008075:Lipoproteins, HDL; D008563:Membrane Lipids; D010743:Phospholipids; D014316:Tritium; D002784:Cholesterol; D008049:Lipase