Characterization of phospholipids in pre- a HDL: selective phospholipid efflux with apolipoprotein A-I

Previously, we have shown that lipid-free apoA-I, when incubated with fibroblasts, will produce lipoproteins of prea mobility (Asztalos, B. F., et al. 1997. Arterioscler. Thromb. Vasc. Biol. 17: 1630–1636). In order to understand the nature of these prea particles, we further characterized their lipid content. The prea particles are high density lipoproteins, having a median density of 1.08 g/ml. They have a surface charge of 2 18.45 mV. The phospholipid composition of these particles showed that they have 4% each of phosphatidyl ethanolamine and inositol; 69% phosphatidyl choline and 18% sphingomyelin. This phospholipid composition is different from those of plasma HDL (81% phosphatidyl choline, 13% sphingomyelin), plasma membrane on the fibroblasts, and whole fibroblast phospholipid. To demonstrate that the prea mobility resides in the lipids, lipids from prea lipoproteins were reconstituted with lipidfree apoA-I. The resultant particles retained their prea mobility. We conclude that apoA-I may react with specific regions of plasma membrane to acquire this unusual lipid composition and that prea mobility is caused in part by the unusual phospholipid composition.— Zhang, W., B. Asztalos, P. S. Roheim, and L. Wong. Characterization of phospholipids in prea HDL: selective phospholipid efflux with apolipoprotein A-I. J. Lipid Res. 1998. 39: 1601–1607. Supplementary key words high density lipoproteins • pre-alpha lipoproteins • pre-beta lipoproteins • fibroblasts • cholesterol • phospholipid composition • cholesterol efflux • two-dimensional electrophoresis High density lipoproteins (HDL) are a major component of plasma lipoproteins. Unlike their low density lipoprotein counterpart, the function of HDL is not well understood. There is evidence that high levels of HDL are protective against coronary heart disease (CHD). In large scale epidemiological studies such as the Framingham study, subjects with high levels of HDL cholesterol have been shown to have lower incidence of CHD even when they have high levels of LDL (1). Three decades of research into the structure and function of various lipoproteins have shown HDL to be heterogeneous. The particles can be separated by charge, density, and apolipoprotein composition into various fractions (2– 4). There is also evidence that the different HDL particles have different metabolic functions (5). Recently, we developed a quantitative two-dimensional agarose non-denaturing polyacrylamide gradient gel electrophoresis system (2DE) to analyze plasma high density lipoproteins (6). Using this procedure, we have observed free apoA-I-like particles in human plasma (7). We have also shown that free apoA-I is capable of causing accelerated cholesterol efflux from cells, and in the process several prea migrating lipoprotein particles are formed (8). The purpose of the present study is to characterize the prea lipoproteins further. We wish to determine the density distribution of prea lipoproteins and the reason for their prea mobility. In this report, we will demonstrate that the particles appear to be HDL, having a density peak at 1.08 g/ml, and their surface charge is considerably greater than the plasma HDL particles. The phospholipid composition of the prea particles is different from plasma HDL particles. It is also different from the phospholipid composition of apoA-I-free media that has been incubated with cells, plasma membrane, and whole cells. Using the lipid from prea lipoproteins, we reconstituted the lipid with lipid-free apoA-I and were able to demonstrate that the reconstituted particles retained their prea mobility.