Role of free apolipoprotein A-I in cholesterol efflux. Formation of pre-alpha-migrating high-density lipoprotein particles.
This article characterizes products formed by the interaction of purified apolipoprotein (apo) A-I and human fibroblasts. Fibroblasts were incubated with different concentrations of purified apoA-I (1 to 30 micrograms/mL) in tissue culture medium for different periods of time (0 to 24 hours). The medium was then characterized by one- (agarose) and two-dimensional (agarose: polyacrylamide nondenaturing gradient gel) electrophoresis. At any given concentration of apoA-I, the rate of cellular cholesterol efflux appeared linear over 24 hours. Incubating purified apoA-I with fibroblasts for 4 hours, we detected five pre-alpha lipoproteins with particle sizes between 114 and 684 kDa. Formation of pre-alpha lipoproteins was concentration-dependent. At low concentrations (below 5 micrograms/mL apoA-I), all purified apoA-I (with pre-beta mobility) was converted to pre-alpha lipoproteins. At higher concentrations (greater than 5 micrograms/mL apoA-I), more apoA-I remained with pre-beta mobility. The pre-alpha lipoproteins were characterized by colocalization of apoA-I particles with 14C-cholesterol and 32P-phospholipids. Results showed that the pre-alpha particle of lowest molecular weight contained phospholipid and apoA-I but no cholesterol. The remaining pre-alpha particles contained all three substances. When pre-alpha particles were subjected to ultracentrifugation, all particles floated at d < 1.21 g/mL with some of the smallest phospholipid apoA-I only particles being present in the d > 1.21 g/mL fraction. Based on these results, we postulated that in the first stages of reverse cholesterol transport, pre-alpha lipoproteins are formed by the interaction of lipid free apoA-I and peripheral cells.
Asztalos
B
B
Department of Physiology, Louisiana State University Medical Center, New Orleans 70112, USA.
Zhang
W
W
Roheim
P S
PS
Wong
L
L
D016632:Apolipoprotein A-I / Q000378:metabolism*; D002478:Cells, Cultured; D002784:Cholesterol / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D008075:Lipoproteins, HDL / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism
D016632:Apolipoprotein A-I; D008075:Lipoproteins, HDL; D010446:Peptide Fragments; D010743:Phospholipids; D002784:Cholesterol