Intravenous administration of apolipoprotein (apo) A

Both increased (mice study) and decreased (hamster study) CETP activity could result in enhanced RCT.

Rationale: Apolipoprotein (apoA)-I mimetic peptides are a promising type of antiatherosclerosis therapy, but how the structural features of these peptides relate to the multiple antiatherogenic functions of HDL is poorly understood. Objective: To establish structure/function relationships of apoA-I mimetic peptides with their antiatherogenic functions. Methods and Results: Twenty-two bihelical apoA-I mimetic peptides were investigated in vitro for the capacity and specificity of cholesterol efflux, inhibition of inflammatory response of monocytes and endothelial cells, and inhibition of low-density lipoprotein (LDL) oxidation. It was found that mean hydrophobicity, charge, size of hydrophobic face, and angle of the link between the helices are the major factors determining the efficiency and specificity of cholesterol efflux. The peptide with optimal parameters was more effective and specific toward cholesterol efflux than human apoA-I. Charge and size of hydrophobic face were also the major factors affecting antiinflammatory properties, and the presence of cysteine and histidine residues was the main factor determining antioxidant properties. There was no significant correlation between capacities of the peptides to support individual functions; each function had its own optimal set of features.

ApoA-I contains a tandem array of amphipathic helices with varying lipid affinity, which are critical in its ability to bind and remove lipids from cells by the ABCA1 transporter. In this study, the effect of asymmetry in the lipid affinity of amphipathic helices in a bihelical apoA-I mimetic peptide, 37pA, on lipid efflux by the ABCA1 transporter was examined. Seven peptide variants of 37pA were produced by substituting a varying number of hydrophobic amino acids for alanine on either one or both helices. The 5A peptide with five alanine substitutions in the second helix had decreased helical content compared with 37pA (5A, 12 ؎ 1% helicity; 37pA, 28 ؎ 2% helicity) and showed less self-association but, similar to the parent peptide, was able to readily solubilize phospholipid vesicles. Furthermore, 5A, unlike the parent peptide 37pA, was not hemolytic (37pA, 27 ؎ 2% RBC lysis, 2 h, 18 M). Finally, the 5A peptide stimulated cholesterol and phospholipid efflux by the ABCA1 transporter with higher specificity (ABCA1-transfected versus untransfected cells) than 37pA (5A, 9.7 ؎ 0.77%, 18 h, 18 M versus 1.5 ؎ 0.27%, 18 h, 18 M (p < 0.0001); 37pA, 7.4 ؎ 0.85%, 18 h, 18 M versus 5.8 ؎ 0.20%, 18 h, 18 M (p ‫؍‬ 0.03)). In summary, we describe a novel bihelical peptide with asymmetry in the lipid affinity of its helices and properties similar to apoA-I in terms of specificity for cholesterol efflux by the ABCA1 transporter and low cytotoxicity.ApoA-I, the predominant protein constituent of HDL 5 (1), can promote the efflux of excess cholesterol from cells by the ABCA1 transporter (2), which is one of the main antiatherogenic functions of HDL (3). ApoA-I contains a tandem array of amphipathic helical domains (4), but the amino and carboxyl terminal helices have the highest lipid affinity and are critical for the physical and biological properties of apoA-I (5-7). When synthesized as individual peptides, helices 1 and 2 and helices 9 and 10 exhibit good detergent-like properties, as assessed by dimyristoylphosphatidylcholine (DMPC) vesicle solubilization (5,7,8). The lipid affinity and detergent properties of the amphipathic helices of apoA-I are also important in how it facilitates cholesterol efflux from cells. In particular, the high lipid affinity COOH-terminal helix of apoA-I is essential to lipid efflux by the ABCA1 transporter (1,6,8,9,11,12). Recently, it has been proposed that the ABCA1 transporter promotes lipid efflux from cells by creating a lipid microdomain that is solubilized and removed by apoA-I by a microsolubilization process dependent upon the detergent-like properties of its amphipathic helices (13-17).ApoA-I and other apolipoproteins (18), including some relatively short peptides (13,17,19,20), can promote cholesterol efflux from cells if they contain an amphipathic helix. ApoA-I mimetic peptides, containing amphipathic helices, are currently being investigated as an alternative to apoA-I for acute HDL therapy of patients with acute coronary syndromes (21,22). One of these peptides, 37pA, is a dimer of...

Eight proteins potentially involved in cholesterol efflux [ABCA1, ABCG1, CYP27A1, phospholipid transfer protein (PLTP), scavenger receptor type BI (SR-BI), caveolin-1, cholesteryl ester transfer protein, and apolipoprotein A-I (apoA-I)] were overexpressed alone or in combination in RAW 264.7 macrophages. When apoA-I was used as an acceptor, overexpression of the combination of ABCA1, CYP27A1, PLTP, and SR-BI (Combination I) enhanced the efflux by 4.3-fold. It was established that the stimulation of efflux was due to increased abundance of ABCA1 and increased apoA-I binding to non-ABCA1 sites on macrophages. This combination caused only a small increase of the efflux to isolated HDL. When HDL was used as an acceptor, overexpression of caveolin-1 or a combination of caveolin-1 and SR-BI (Combination II) was the most active, doubling the efflux to HDL, without affecting the efflux to apoA-I. When tested in the in vivo mouse model of cholesterol efflux, overexpression of ABCA1 and Combination I elevated cholesterol export from macrophages to plasma, liver, and feces, whereas overexpression of caveolin-1 or Combination II did not have an effect. We conclude that pathways of cholesterol efflux using apoA-I as an acceptor make a predominant contribution to cholesterol export from macrophages in vivo.