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It is important to understand HDL heterogeneity because various subspecies possess different functionalities. To understand the origins of HDL heterogeneity arising from the existence of particles containing only apoA-I (LpA-I) and particles containing both apoA-I and apoA-II (LpA-I+A-II), we compared the abilities of both proteins to promote ABCA1-mediated efflux of cholesterol from HepG2 cells and form nascent HDL particles. When added separately, exogenous apoA-I and apoA-II were equally effective in promoting cholesterol efflux, although the resultant LpA-I and LpA-II particles had different sizes. When apoA-I and apoA-II were mixed together at initial molar ratios ranging from 1:1 to 16:1 to generate nascent LpA-I+A-II HDL particles, the particle size distribution altered, and the two proteins were incorporated into the nascent HDL in proportion to their initial ratio. Both proteins formed nascent HDL particles with equal efficiency, and the relative amounts of apoA-I and apoA-II incorporation were driven by mass action. The ratio of lipid-free apoA-I and apoA-II available at the surface of ABCA1-expressing cells is a major factor in determining the contents of these proteins in nascent HDL. Manipulation of this ratio provides a means of altering the relative distribution of LpA-I and LpA-I+A-II HDL particles.

Monocyte chemoattractant protein-1 (MCP-1) plays crucial roles at multiple stages of atherosclerosis. We hypothesized that MCP-1 might impair the reverse cholesterol transport (RCT) capacity of HepG2 cells by decreasing the cell-surface protein expression of ATP binding cassette A1 (ABCA1), ATP binding cassette G1 (ABCG1), and scavenger receptor class B type I (SR-BI). MCP-1 reduced the total protein and mRNA levels of ABCA1 and SR-BI, but not of ABCG1. MCP-1 decreased the cell-surface protein expression of ABCA1, ABCG1, and SR-BI in dose-dependent and time-dependent manners, as measured using cell-surface biotinylation. We further studied the phosphoinositide 3-kinase (PI3K)/serine/threonine protein kinase Akt pathway in regulating receptor trafficking. Both the translation and transcription of ABCA1, ABCG1, and SR-BI were not found to be regulated by the PI3K/Akt pathway. However, the cell-surface protein expression of ABCA1, ABCG1, and SR-BI could be regulated by PI3K activity, and PI3K activation corrected the MCP-1-induced decreases in the cell-surface protein expression of ABCA1, ABCG1, and SR-BI. Moreover, we found that MCP-1 decreased the lipid uptake by HepG2 cells and the ABCA1-mediated cholesterol efflux to apoA-I, which could be reversed by PI3K activation. Our data suggest that MCP-1 impairs RCT activity in HepG2 cells by a PI3K/Akt-mediated posttranslational regulation of ABCA1, ABCG1, and SR-BI cell-surface expression.

The liver X receptors (LXRs) have been shown to affect lipoprotein plasma profile, lipid metabolism, and reverse cholesterol transport (RCT). In the present study, we investigated whether a short-term administration of the synthetic LXR agonist T0901317 (T0) to mice may affect RCT by modulating the capacity of plasma to promote cellular lipid efflux. Consistent with previous data, the pharmacological treatment of mice caused a significant increase of macrophage-derived [3H]cholesterol content in plasma, liver, and feces and resulted in improved capacity of plasma to promote cellular cholesterol release through passive diffusion and scavenger receptor class B type I (SR-BI)-mediated mechanisms. Differently, plasma from treated mice possessed similar or reduced capacity to drive lipid efflux via ABCA1. Consistent with these data, the analysis of plasma HDL fractions revealed that T0 caused the formation of larger, lipid-enriched particles. These results suggest that T0 promotes in vivo RCT from macrophages at least in part by inducing an enrichment of those HDL subclasses that increase plasma capacity to promote cholesterol efflux by passive diffusion and SR-BI-mediated mechanisms.

ABCG1, one of the half-type ATP binding cassette (ABC) proteins, mediates the efflux of cholesterol to HDL and functions in the reverse cholesterol transport from peripheral cells to the liver. We have shown that ABCG1 mediates the efflux of not only cholesterol but also sphingomyelin (SM) and phosphatidylcholine. Because SM preferentially associates with cholesterol, we examined whether it plays an important role in the ABCG1-mediated efflux of cholesterol. The efflux of cholesterol and SM mediated by ABCG1 was reduced in a mutant CHO-K1 cell line, LY-A, in which the cellular SM level is reduced because of a mutation of the ceramide transfer protein CERT. In contrast, CHO-K1 cells overexpressing CERT showed an increased efflux of cholesterol and SM mediated by ABCG1. The sensitivity of cells to methyl-β-cyclodextrin suggested that cholesterol in nonraft domains was increased due to the disruption of raft domains in LY-A cells. These results suggest that the ABCG1-mediated efflux of cholesterol and SM is dependent on the cellular SM level and distribution of cholesterol in the plasma membrane.

It is important to understand HDL heterogeneity because various subspecies possess different functionalities. To understand the origins of HDL heterogeneity arising from the existence of particles containing only apoA-I (LpA-I) and particles containing both apoA-I and apoA-II (LpA-I+A-II), we compared the abilities of both proteins to promote ABCA1-mediated efflux of cholesterol from HepG2 cells and form nascent HDL particles. When added separately, exogenous apoA-I and apoA-II were equally effective in promoting cholesterol efflux, although the resultant LpA-I and LpA-II particles had different sizes. When apoA-I and apoA-II were mixed together at initial molar ratios ranging from 1:1 to 16:1 to generate nascent LpA-I+A-II HDL particles, the particle size distribution altered, and the two proteins were incorporated into the nascent HDL in proportion to their initial ratio. Both proteins formed nascent HDL particles with equal efficiency, and the relative amounts of apoA-I and apoA-II incorporation were driven by mass action. The ratio of lipid-free apoA-I and apoA-II available at the surface of ABCA1-expressing cells is a major factor in determining the contents of these proteins in nascent HDL. Manipulation of this ratio provides a means of altering the relative distribution of LpA-I and LpA-I+A-II HDL particles.