Macrophage Specific IGF‐1 Overexpression Decreases Atherosclerosis, CXCL12 Chemokine, And Increases Cholesterol Efflux In ApoE Deficient Mice

Abstract

Increased bioavailability of Insulin Like Growth Factor (IGF-1) has been shown to decrease cardiovascular event risk. We have shown that systemic IGF-1 administration reduced atherosclerosis in ApoE deficient (Apoe-/-) mice fed a high fat diet and that those animals have reduced plaque macrophages. Thus, we have shown that IGF-1 is atheroprotective, but cell-specific mechanism(s) mediating IGF-1's effect need to be identified. IGF-1 interacts with major cell types in atherosclerotic plaque: macrophages, smooth muscle cells, and endothelial cells. Results of our previous in vitro experiments suggest that macrophages play a predominant role in mediating IGF-1 effects. We hypothesized that increasing IGF-1 levels strictly in macrophages will prevent atherosclerosis. Using a novel macrophage-specific IGF-1 overexpressing transgenic mouse bred to an Apoe-/- background (MF-IGF1 mice), we assessed atherosclerotic plaque burden, stability, and monocyte recruitment. We isolated plaque using laser capture microdissection and we analyzed monocyte recruitment via intravenous injection of fluorescent red beads among other traditional techniques. Macrophage IGF-1 overexpression downregulated plaque burden by 30% (P<0.01), reduced plaque macrophages by 47% (P<0.001) and promoted a stable plaque phenotype. Monocyte recruitment was reduced by 70% (P<0.05) in MF-IGF1 mice and was associated with a decrease in circulating levels of CXC Chemokine Ligand 12 (CXCL12) (27% reduction compared to control, P<0.05). CXCL12 protein levels were reduced in plaque and peritoneal macrophages in MF-IGF1 mice. IGF-1 completely blocked oxidized low-density lipoprotein (oxLDL)-dependent increase of CXCL12 mRNA transcription (98% reduction vs. control, P<0.01) and IGF-1 treatment reduced CXCL12 protein (56% decrease vs. control, P<0.001)in vitro. ATP-binding cassette transporter A1 (ABCA1) is the key cholesterol transporter mediating macrophage cholesterol efflux. We found that peritoneal macrophages isolated from MF-IGF1 mice have a 2-fold increase in ABCA1 protein levels. To study whether IGF-1 changes macrophage cholesterol efflux, we loaded THP-1 cells with oxLDL and then attached a fluorescent label to measure changes in efflux. We found a 27% increase in cholesterol efflux in IGF-1 (100ng/mL)-treated cells (P<0.01) when using Apolipoprotein A I as a cholesterol acceptor. Overall, our results indicate that macrophage IGF-1 reduces atherosclerosis and decreases CXCL12, a chemokine newly implicated in atheroprogression. CXCL12 downregulates ABCA1 expression. IGF-1 potentially exerts its atheroprotective effect via this reduction in CXCL12 by reducing monocyte recruitment and by increasing ABCA1 and thereby increasing cholesterol efflux capacity.