Objectives Apolipoprotein E (apoE) exerts potent anti-inflammatory effects. We here investigated the effect of apoE on the functional phenotype of macrophages. Methods and Results Human apoE receptors VLDL-R or apoER2 were stably expressed in RAW264.7 mouse macrophages. In these cells apoE downregulated markers of the pro-inflammatory M1 phenotype (iNOS, IL-12, MIP-1α), but upregulated markers of the anti-inflammatory M2 phenotype (arginase-I, SOCS3, IL-1RA). In addition, M1 macrophage responses (migration, generation of reactive oxygen species, antibody-dependent cell cytotoxicity, phagocytosis) as well as poly(I:C)- and/or IFN-γ-induced production of pro-inflammatory cytokines, COX-2 expression, and activation of NF-κB, IκB and STAT1 were suppressed in VLDL-R- or apoER2-expressing cells. Conversely, the suppression of M2 phenotype and the enhanced response to poly(I:C) were observed in apoE-producing bone marrow macrophages derived from VLDL-R-deficient mice, but not wild type or LDL receptor-deficient mice. The modulatory effects of apoE on macrophage polarization were inhibited in apoE receptor-expressing RAW264.7 cells exposed to SB220025, a p38MAP kinase inhibitor, and PP1, a tyrosine kinase inhibitor. Accordingly, apoE induced tyrosine kinase-dependent activation of p38MAP kinase in VLDL-R- or apoER2-expressing macrophages. Under in vivo conditions, apoE−/− mice transplanted with apoE-producing wild-type bone marrow showed increased plasma IL-1RA levels and peritoneal macrophages of transplanted animals were shifted to the M2 phenotype (increased IL-1RA production and CD206 expression). Conclusion ApoE signaling via VLDL-R or apoER2 promotes macrophage conversion from the pro-inflammatory M1 to the anti-inflammatory M2 phenotype. This effect may represent a novel anti-inflammatory activity of apoE.

Background-Numerous in vitro studies suggest that sphingosine 1-phosphate (S1P), a bioactive lysosphingolipid associated with high-density lipoproteins, accounts at least partly for the potent antiinflammatory properties of high-density lipoprotein and, thereby, contributes to the antiatherogenic potential attributed to high-density lipoproteins. The present study was undertaken to investigate whether modulation of S1P signaling would affect atherosclerosis in a murine model of disease. Methods and Results-Low-density lipoprotein receptor-deficient mice on a cholesterol-rich diet were given FTY720, a synthetic S1P analogue, at low (0.04 mg/kg per day) or high (0.4 mg/kg per day) doses for 16 weeks. FTY720 dose-dependently reduced atherosclerotic lesion formation, both in the aortic root and brachiocephalic artery, and almost completely blunted necrotic core formation. Plasma lipids remained unchanged during the course of FTY720 treatment. However, FTY720 lowered blood lymphocyte count (at a high dose) and significantly interfered with lymphocyte function, as evidenced by reduced splenocyte proliferation and interferon-␥ levels in plasma. Plasma concentrations of proinflammatory cytokines such as tumor necrosis factor-␣, interleukin (IL)-6, IL-12, and regulated on activation normal T cell expressed and secreted were reduced by FTY720 administration. Moreover, lipopolysaccharide-elicited generation of nitrite/nitrate and IL-6 -two markers of classical (M1) macrophage activation-was inhibited, whereas IL-4 -induced production of IL-1-receptor antagonist, a marker of alternative (M2) macrophage activation, was augmented in peritoneal macrophages from FTY720-treated low-density lipoprotein receptor-deficient mice. Conclusions-The present results demonstrate that an S1P analogue inhibits atherosclerosis by modulating lymphocyte and macrophage function, and these results are consistent with the notion that S1P contributes to the antiatherogenic potential of high-density lipoprotein. (Circulation. 2007;115:501-508.)