Abstract
The effect of high density lipoprotein composition on the rates of unesterified cholesterol exchange between low density lipoproteins (LDL) and well-defined homogeneous discoidal lipoproteins (LpA-I) reconstituted with phosphatidylcholine, cholesterol, and apolipoprotein A-I (apoA-I) has been investigated. LpA-I containing cholesterol and 2, 3, and 4 apoA-I molecules per particle differed in their ability to accept or donate cholesterol. A significant cholesterol exchange occurs between LDL and Lp2A-I (7.8 and 9.6 nm), while there is little or no cholesterol exchange detectable between LDL and Lp3A-I (10.8 and 13.4 nm) and Lp4A-I (17.0 nm) complexes. The cholesterol transfer from LDL to the cholesterol-free Lp2A-I (9.6 nm), Lp3A-I (13.4 nm), and Lp4A-I (17.0 nm) particles also shows significant cholesterol transfer to Lp2A-I, while there is no detectable transfer to Lp3- and 4A-I particles. The rates of cholesterol transfer to cholesterol-free and cholesterol-containing Lp2A-I appear to differ significantly. Cholesterol transfer from LDL to cholesterol-free Lp2A-I is zero order with respect to acceptor concentrations when the Lp2A-I/LDL ratio is above 10. Transfer rates from LDL to cholesterol-free Lp2A-I are faster for the smaller Lp2A-I (8.5 nm) than to the larger Lp2A-I (9.7 nm) and exhibit half-times (t1/2) at 25 degrees C of 4.0 and 5.3 h, respectively. In contrast, cholesterol transfer from LDL to cholesterol-containing Lp2A-I remains dependent upon acceptor concentrations to an acceptor/donor particle ratio of 80. In addition, transfer from LDL to cholesterol-containing Lp2A-I is faster to the 9.6 nm than to 7.8 nm particles, with t1/2 of 1.4 and 2.3 h, respectively. The rates of cholesterol transfer from Lp2A-I to LDL are higher than in the opposite direction, in particular for the small Lp2A-I (7.8 nm), which has a t1/2 of approximately 50 min. The results show that changes in the composition and structure of apoA-I-containing particles have a significant effect on inter-lipoprotein exchange of cholesterol. This suggests that the kinetics of cholesterol transfer to and from reconstituted discoidal LpA-I particles cannot be fully explained by passive aqueous diffusion.
Highlights
The abbreviations used are: HDL, high density lipoprotein(s); apo, apolipoprotein; LDL, low density lipoprotein(s); LpA-I, lipoprotein containing apoA-I; LpA-I particles containing cholesterol (LpA-Ic), lipoprotein containing apoA-I and cholesterol; PC, phosphatidylcholine; POPC, I-palmitoyl-2oleoylphosphatidylcholine; SUV, small unilamellar vesicles
A recent study by another group showed that only a fraction of the cellular cholesterol transferred to the pre-I3-HDL pathway is esterified while the majority recycles through a-HDL, LDL, and pre-l3HDL [10]
Studies done by Phillips and co-workers [13,14,15] have shown that the physical state of cholesterol molecules in the phospholipid! water interface of serum lipoprotein particles differs on LDL, HDL, and small unilamellar vesicles (SUV) in a manner that parallels differences in cholesterol flux between these particles [16]
Summary
Preparation of Plasma LDL, ApoA-I, and LpA-I Particles-Plasma LDL and apoA-I were prepared from pooled plasma from normolipi-. Each assay mixture contained 60 iLl of 2% defatted BSA, 1 iLg of [3Hlcholesterol-labeled LDL in 100 iLl of 1% BSA, different amounts of LpA-I particles diluted in 100 iLl of 1% BSA, and TBS was added to a final volume of 460 iLl. The mixtures were incubated at room temperature for different periods of time. At the end ofthe incubation, the mixtures were adjusted to the final density of 1.063 g/ml and transferred into mini tubes for Beckman TLA-100 rotor with a total volume of 250 iLl per tube and ultracentrifuged at 60,000 rpm for 200 min The contents in these tubes were fractionated every 50 1-'1 from the top to the bottom and counted in a The kinetics of cholesterol transfer between LDL and LpA-I were calculated as described by Lund-Katz and colleagues [12, 31]
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