Regulation of Csf1r and Bcl6 in macrophages mediates the stage-specific effects of microRNA-155 on atherosclerosis.

OBJECTIVE: The function of microRNAs is highly context and cell type dependent because of their highly dynamic expression pattern and the regulation of multiple mRNA targets. MicroRNA-155 (miR-155) plays an important role in the innate immune response by regulating macrophage function; however, the effects of miR-155 in macrophages on atherosclerosis are controversial. We hypothesized that the stage-dependent target selection of miR-155 in macrophages determines its effects on atherosclerosis. APPROACH AND RESULTS: The expression of miR-155 increased in lesional macrophages of apolipoprotein E-deficient mice between 12 and 24 weeks of a high-cholesterol diet. Mir155 knockout in apolipoprotein E-deficient mice enhanced lesion formation, increased the lesional macrophage content, and promoted macrophage proliferation after 12 weeks of the high-cholesterol diet. In vitro, miR-155 inhibited macrophage proliferation by suppressing colony-stimulating factor-1 receptor, which was upregulated in lesional macrophages of Mir155(-/-) apolipoprotein E-deficient mice. By contrast, Mir155 deficiency reduced necrotic core formation and the deposition of apoptotic cell debris, thereby preventing the progression of atherosclerosis between 12 and 24 weeks of the high-cholesterol diet. miR-155 inhibited efferocytosis in vitro by targeting B-cell leukemia/lymphoma 6 and thus activating RhoA (ras homolog gene family, member A). Accordingly, B-cell leukemia/lymphoma 6 was upregulated in lesional macrophages of Mir155(-/-) apolipoprotein E-deficient mice after 24 weeks, but not after 12 weeks of the high-cholesterol diet. CONCLUSIONS: Our findings demonstrate a stage-specific role of miR-155 in lesion formation. miR-155 suppressed macrophage proliferation by targeting colony-stimulating factor-1 receptor in early and impaired efferocytosis by downregulating B-cell leukemia/lymphoma 6 in advanced atherosclerosis. Therefore, targeting the interaction between miR-155 and B-cell leukemia/lymphoma 6 may be a promising approach to inhibit the progression of atherosclerosis.

MicroRNA-126-5p promotes endothelial proliferation and limits atherosclerosis by suppressing Dlk1.

Atherosclerosis, a hyperlipidemia-induced chronic inflammatory process of the arterial wall, develops preferentially at sites where disturbed laminar flow compromises endothelial cell (EC) function. Here we show that endothelial miR-126-5p maintains a proliferative reserve in ECs through suppression of the Notch1 inhibitor delta-like 1 homolog (Dlk1) and thereby prevents atherosclerotic lesion formation. Endothelial recovery after denudation was impaired in Mir126(-/-) mice because lack of miR-126-5p, but not miR-126-3p, reduced EC proliferation by derepressing Dlk1. At nonpredilection sites, high miR-126-5p levels in endothelial cells confer a proliferative reserve that compensates for the antiproliferative effects of hyperlipidemia, such that atherosclerosis was exacerbated in Mir126(-/-) mice. In contrast, downregulation of miR-126-5p by disturbed flow abrogated EC proliferation at predilection sites in response to hyperlipidemic stress through upregulation of Dlk1 expression. Administration of miR-126-5p rescued EC proliferation at predilection sites and limited atherosclerosis, introducing a potential therapeutic approach.

Activation of CXCR7 limits atherosclerosis and improves hyperlipidemia by increasing cholesterol uptake in adipose tissue.

BACKGROUND: The aim of this study was to determine the role of the chemokine receptor CXCR7 in atherosclerosis and vascular remodeling. CXCR7 is the alternative receptor of CXCL12, which regulates stem cell-mediated vascular repair and limits atherosclerosis via its receptor, CXCR4. METHODS AND RESULTS: Wire-induced injury of the carotid artery was performed in mice with a ubiquitous, conditional deletion of CXCR7 and in mice treated with the synthetic CXCR7 ligand CCX771. The effect of CCX771 treatment on atherosclerosis was studied in apolipoprotein E-deficient (Apoe(-/-)) mice fed a high-fat diet for 12 weeks. Lipoprotein fractions were quantified in the plasma of Apoe(-/-) mice by fast protein liquid chromatography. Uptake of DiI-labeled very low-density lipoprotein to adipose tissue was determined by 2-photon microscopy. We show that genetic deficiency of Cxcr7 increased neointima formation and lesional macrophage accumulation in hyperlipidemic mice after vascular injury. This was related to increased serum cholesterol levels and subsequent hyperlipidemia-induced monocytosis. Conversely, administration of the CXCR7 ligand CCX771 to Apoe(-/-) mice inhibited lesion formation and ameliorated hyperlipidemia after vascular injury and during atherosclerosis. Treatment with CCX771 reduced circulating very low-density lipoprotein levels but not low-density lipoprotein or high-density lipoprotein levels and increased uptake of very low-density lipoprotein into Cxcr7-expressing white adipose tissue. This effect of CCX771 was associated with an enhanced lipase activity and reduced expression of Angptl4 in adipose tissue. CONCLUSIONS: CXCR7 regulates blood cholesterol by promoting its uptake in adipose tissue. This unexpected cholesterol-lowering effect of CXCR7 is beneficial for atherosclerotic vascular diseases, presumably via amelioration of hyperlipidemia-induced monocytosis, and can be augmented with a synthetic CXCR7 ligand.

The microRNA-342-5p fosters inflammatory macrophage activation through an Akt1- and microRNA-155-dependent pathway during atherosclerosis.

BACKGROUND: Atherosclerosis is a chronic inflammatory vascular disease driven by the subendothelial accumulation of macrophages. The mechanism regulating the inflammatory response in macrophages during atherogenesis remains unclear. Because microRNAs (miRNAs) play a crucial role in cellular signaling by posttranscriptional regulation of gene expression, we studied the miRNA expression profiles during the progression of atherosclerosis. METHODS AND RESULTS: Using an miRNA real-time polymerase chain reaction array, we found that macrophage-derived miR-342-5p and miR-155 are selectively upregulated in early atherosclerotic lesions in Apoe(-/-) mice. miR-342-5p directly targets Akt1 through its 3'-untranslated region. Akt1 suppression by miR-342-5p induces proinflammatory mediators such as Nos2 and II6 in macrophages via the upregulation of miR-155. The local application of an miR-342-5p antagomir inhibits the development of atherosclerosis in partially ligated carotid arteries. In atherosclerotic lesions, the miR-342-5p antagomir upregulated Akt1 expression and suppressed the expression of miR-155 and Nos2. This reduced Nos2 expression was associated with a diminished generation of nitrotyrosine in the plaques. Furthermore, systemic treatment with an inhibitor of miR-342-5p reduced the progression of atherosclerosis in the aorta of Apoe(-/-) mice. CONCLUSIONS: Macrophage-derived miR-342-5p promotes atherosclerosis and enhances the inflammatory stimulation of macrophages by suppressing the Akt1-mediated inhibition of miR-155 expression. Therefore, targeting miR-342-5p may offer a promising strategy to treat atherosclerotic vascular disease.

MicroRNA-155 promotes atherosclerosis by repressing Bcl6 in macrophages.

Macrophages in atherosclerotic plaques drive inflammatory responses, degrade lipoproteins, and phagocytose dead cells. MicroRNAs (miRs) control the differentiation and activity of macrophages by regulating the signaling of key transcription factors. However, the functional role of macrophage-related miRs in the immune response during atherogenesis is unknown. Here, we report that miR-155 is specifically expressed in atherosclerotic plaques and proinflammatory macrophages, where it was induced by treatment with mildly oxidized LDL (moxLDL) and IFN-γ. Leukocyte-specific Mir155 deficiency reduced plaque size and number of lesional macrophages after partial carotid ligation in atherosclerotic (Apoe-/-) mice. In macrophages stimulated with moxLDL/IFN-γ in vitro, and in lesional macrophages, loss of Mir155 reduced the expression of the chemokine CCL2, which promotes the recruitment of monocytes to atherosclerotic plaques. Additionally, we found that miR-155 directly repressed expression of BCL6, a transcription factor that attenuates proinflammatory NF-κB signaling. Silencing of Bcl6 in mice harboring Mir155-/- macrophages enhanced plaque formation and CCL2 expression. Taken together, these data demonstrated that miR-155 plays a key role in atherogenic programming of macrophages to sustain and enhance vascular inflammation.

Lipoprotein-derived lysophosphatidic acid promotes atherosclerosis by releasing CXCL1 from the endothelium.

Oxidatively modified low-density lipoprotein (oxLDL) plays a key role in the initiation of atherosclerosis by increasing monocyte adhesion. The mechanism that is responsible for the oxLDL-induced atherogenic monocyte recruitment in vivo, however, still remains unknown. Oxidation of LDL generates lysophosphatidylcholine, which is the main substrate for the lysophosphatidic acid (LPA) generating enzyme autotaxin. We show that oxLDL requires endothelial LPA receptors and autotaxin to elicit CXCL1-dependent arterial monocyte adhesion. Unsaturated LPA releases endothelial CXCL1, which is subsequently immobilized on the cell surface and mediates LPA-induced monocyte adhesion. Local and systemic application of LPA accelerates the progression of atherosclerosis in mice. Blocking the LPA receptors LPA(1) and LPA(3) reduced hyperlipidemia-induced arterial leukocyte arrest and atherosclerosis in the presence of functional CXCL1. Thus, atherogenic monocyte recruitment mediated by hyperlipidemia and modified LDL crucially depends on LPA, which triggers endothelial deposition of CXCL1, revealing LPA signaling as a target for cardiovascular disease treatments.