Abstract 833: An ApoE Mimetic Peptide Modulates HDL Subspeciation, Increases ApoE Secretion and Decreases MCP-1 mRNA Levels in Cells
Background: An apoE mimetic peptide, Ac-hE18A-NH2, has been shown to:
enhance the uptake of atherogenic lipoproteins in HepG2 cells,
dramatically lower plasma cholesterol associated with apoB-containing lipoproteins, without affecting high density lipoprotein (HDL) levels, in dyslipidemic animal models and
improve HDL- and endothelial function in Watanabe Heritable Hyperlipidemic rabbits.
Objective: In this study we investigated the mechanism by which Ac-hE18A-NH2 exerts its anti-atherogenic and anti-inflammatory effects at the cellular level with particular reference to apoA-I and apoE synthesis and secretion and monocyte chemotaxis protein-1 (MCP-1) expression.
Methods and Results: Metabolic labeling with 35S-methionine and immunoprecipitation techniques in presence and absence of heparin demonstrated that Ac-hE18A-NH2 causes a 4-fold increase over controls in the secretion of cell-surface apoE in HepG2 cells. It enhances the level of apoA-I associated with preβ-HDL particles in the medium in a time- and concentration- dependent manner, although it does not cause an increase in the total mass of apoA-I secreted by HepG2 cells. An 18h incubation with 20 μg/ml of peptide produces a 2.5 fold increase over controls in secreted preβ-HDL levels. This effect appears to be at the post translational level as it is inhibited by cycloheximide and bafilomycin A. These data suggest that the peptide remodels newly synthesized apoA-I-containing particles. Sucrose density ultracentrifugation demonstrated that both the apolipopro-teins form separate particles: i.e. an apoE-rich particle (d=1.05g/ml) with apoE:apoA-I = 9:0.5 and an apoA-I rich particle (d = 1.12g/ml) with apoE:apoA-I = 1:8. Increased secretion of cell-surface apoE by Ac-hE18A-NH2 was also observed in THP-1 monocyte-derived macrophages. In these cells, real time PCR indicated that Ac-hE18A-NH2 completely abolished the 18-fold increase in LPS-induced mRNA levels for MCP-1.
Conclusions: These results suggest that Ac-hE18A-NH2 exerts its anti-atherogenic and anti-inflammatory activities by:
promoting the formation of preβ-HDL particles
stimulating the secretion of cell-surface-derived apo E and
inhibiting the formation of LPS-induced MCP-1.