Apolipoprotein E4 Is Deficient in Inducing Macrophage ABCA1 Expression and Stimulating the Sp1 Signaling Pathway

Abstract

ATP binding cassette A1 (ABCA1) is a membrane protein that promotes cellular cholesterol efflux. Using RAW 264.7 macrophages, we studied the relative effects of apolipoprotein (apo) E3 and apoE4 on ABCA1 and on the signaling pathway that regulates its expression. Both lipid-associated and lipid-free apoE4 forms induced ∼30% lower levels of ABCA1 protein and mRNA than apoE3 forms. Phosphorylated levels of phosphoinositol 3-kinase (PI3K), protein kinase Cζ (PKCζ) and specificity protein 1 (Sp1) were also lower when treated with apoE4 compared to apoE3. The reduced ability of apoE4 to induce ABCA1 expression, PKCζ and Sp1 phosphorylation were confirmed in human THP-1 monocytes/macrophages. Sequential phosphorylation of PI3K, PKCζ and Sp1 has been suggested as a mechanism for upregulation of ABCA1 expression. Both apoE3 and apoE4 reduced total cholesterol and cholesterol esters in lipid-laden RAW 264.7 cells, and induced apoAI-mediated cholesterol efflux. However, the cholesterol esters and cholesterol efflux in apoE4-treated cells were ∼50% and ∼24% lower, respectively, compared to apoE3-treated cells. Accumulation of cholesterol esters in macrophages is a mechanism for foam cell formation. Thus the reduced ability of apoE4 to activate the PI3K-PKCζ-Sp1 signaling pathway and induce ABCA1 expression likely impairs cholesterol ester removal, and increases foam cell formation.

APOε4 is associated with enhanced in vivo innate immune responses in human subjects

“…31 Increased cholesterol loading of platelets in mice causes platelet hyperreactivity as well as thrombocytopenia due to increased platelet clearance. 34, 35 Our finding of increased lipid rafts in APOε4 + monocytes, together with past reports that apoE4 protein is defective in promoting cholesterol efflux from cells, 20 raises the interesting possibility that the APOε4 genotype may confer abnormalities in platelet function and number through effects upon platelet cholesterol.…”

“…19 ApoE4, however, is less effective than apoE3 at inducing cholesterol efflux from macrophages. 20 Several TLR cascades are initiated in lipid rafts and are enhanced by raft cholesterol loading, such as occurs with deficient cholesterol efflux. Given this, we next quantified lipid rafts in human peripheral blood monocytes, using fluor-tagged cholera toxin B subunit, a widely used raft probe that recognizes the raft-specific marker GM1.…”

High-density lipoprotein-mediated transcellular cholesterol transport in mouse aortic endothelial cells

“…The lysate, and the apical and basolateral media were each mixed with scintillation fluid to analyze radioactivity. Cholesterol efflux to the apical or basolateral compartment was expressed as the percentage of radioactivity in the apical or basolateral medium compared to the total radioactivity (cells plus media), as described previously [17]. …”

Akt isoform-dependent regulation of ATP-Binding cassette A1 expression by apolipoprotein E

“…Inhibition of PI3K diminished apoE-induced ABCA1 expression. Our studies also suggest that very low-density lipoprotein receptor (VLDLR), apoE receptor 2 (apoER2), and disabled-1 (dab1) are the upstream components, while PKCζ and Sp1 are the downstream components of PI3K in this pathway [3,4,5,6]. …”

“…ApoE is secreted freely by macrophages, which can aid in the removal of excess cholesterol in association with ABCA1 [2]. Recent studies from our laboratory suggest that apoE is able to upregulate ABCA1 expression via activation of a signaling pathway involving phosphatidylinositol 3-kinase (PI3K) [3,4,5,6]. Specifically, treatment of murine macrophages with apoE increased the mRNA and protein levels of ABCA1, and enhanced the phosphorylation of PI3K.…”