IER3 Paucity Inhibits oxLDL-induced Foam Cell Formation in ApoE-deficient macrophages via Promoting an Alternative Phenotype

Abstract ID 100168 Poster Board 270 Introduction: Atherosclerosis is a chronic inflammatory disorder and the central cause of ischemic heart disease and stroke. It is characterized by the narrowing of large arteries due to the accumulation of fatty and fibrous plaque. The early stages are defined by subendothelial aggregation of foam cells–lipid-filled macrophages, lacking the ability to migrate as a result of cholesterol engorgement. The accumulation of these foam cells ultimately leads to the buildup of cholesterol, and atherosclerotic plaque formation. The Immediate early response gene 3 (IER3) is a stress inducible gene that is highly expressed in macrophages and regulates their phenotype. We hypothesize that IER3 in macrophages is required for foam cell formation and thereby contributing to atherosclerosis pathogenesis. Methods: To investigate a cell autonomous role of IER 3 in foam cell formation, we harvested bone marrow (BM) cells from mice lacking Apolipoprotein-E (ApoE-/-) or ApoE and IER3 (ApoE-/-IER3-/-). These cells were first treated with macrophage-colony stimulating factor (M-CSF) to differentiate into mature macrophages and next with LPS (10 ng/ml) or IL-4 (10 nM) for 24 hours to transform them into pro- or anti-inflammatory phenotypes. Macrophage phenotypes were determined by quantifying the mRNA levels of pro- and anti-inflammatory marker genes using qRT-PCR and flow cytometry. In another experiment, macrophages were treated with oxLDL (10 μg/ml) for 24 hours to induce foam cell formation, which was subsequently quantified using Oil-Red-O staining. Results: LPS increased mRNA levels of TNF- α (15±3-fold, p<0.05) and iNOS (2.4±0.2-fold, p<0.05) in ApoE−/− cells, an effect which was inhibited in ApoE−/−IER3−/− cells. Conversely, IL-4-induced increases in mRNA levels of Arg-1 (675±131-fold) and MRC-1 (3.2±0.2-fold) were markedly potentiated in ApoE−/−IER3−/− cells as compared to ApoE−/− control. Likewise, LPS-induced activation of proinflammatory state as determined by flow cytometry was attenuated in ApoE−/−IER3−/− cells as compared to ApoE−/− control. Moreover, ox-LDL-induced foam cell formation and accumulation of intracellular oil droplets were also suppressed in ApoE−/−IER3−/− macrophages as compared to ApoE−/− cells. Collectively, this data suggests that IER3 paucity hinders foam cell formation by inhibiting the proinflammatory state of macrophages. Conclusion: IER3 contributes to foam cell formation in macrophages and may promote atherogenesis.