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.

Chemokines mediate monocyte adhesion to dysfunctional endothelial cells (ECs) and promote arterial inflammation during atherosclerosis. Hypoxia-inducible factor (HIF)-1α is expressed in various cell types of atherosclerotic lesions and is associated with lesional inflammation. However, the impact of endothelial HIF-1α in atherosclerosis is unclear. HIF-1α was detectable in the nucleus of ECs covering murine and human atherosclerotic lesions. To study the role of endothelial HIF-1α in atherosclerosis, deletion of the Hif1 a gene was induced in ECs from apolipoprotein E knockout mice (EC- Hif1a −/− ) by Tamoxifen injection. The formation of atherosclerotic lesions, the lesional macrophage accumulation, and the expression of CXCL1 in ECs were reduced after partial carotid ligation in EC- Hif1a −/− compared with control mice. Moreover, the lesion area and the lesional macrophage accumulation were decreased in the aortas of EC- Hif1a −/− mice compared with control mice during diet-induced atherosclerosis. In vitro, mildly oxidized low-density lipoprotein or lysophosphatidic acid 20:4 increased endothelial CXCL1 expression and monocyte adhesion by inducing HIF-1α expression. Moreover, endothelial Hif1a deficiency resulted in downregulation of miR-19a in atherosclerotic arteries determined by microRNA profiling. In vitro, HIF-1α–induced miR-19a expression mediated the upregulation of CXCL1 in mildly oxidized low-density lipoprotein–stimulated ECs. These results indicate that hyperlipidemia upregulates HIF-1α expression in ECs by mildly oxidized low-density lipoprotein–derived unsaturated lysophosphatidic acid. Endothelial HIF-1α promoted atherosclerosis by triggering miR-19a–mediated CXCL1 expression and monocyte adhesion, indicating that inhibition of the endothelial HIF-1α/miR-19a pathway may be a therapeutic option against atherosclerosis.

Objective-Hypoxia-inducible factor (HIF)-1␣ is the regulatory subunit of a transcriptional complex, which controls the recruitment of multipotent progenitor cells and tissue repair in ischemic tissue by inducing stromal cell-derived factor (SDF)-1␣ expression. Because HIF-1␣ can be activated under normoxic conditions in smooth muscle cells (SMCs) by platelet products, we investigated the role of HIF-1␣ in SDF-1␣-mediated neointima formation after vascular injury. Methods and Results-Wire-induced injury of the left carotid artery was performed in apolipoprotein E-deficient mice.HIF-1␣ expression was increased in the media as early as 1 day after injury, predominantly in SMCs. Nuclear translocation of HIF-1␣ and colocalization with SDF-1␣ was detected in neointimal cells after 2 weeks. HIF-1␣ mRNA expression was induced at 6 hours after injury as determined by real-time RT-PCR. Inhibition of HIF-1␣ expression by local application of HIF-1␣-siRNA reduced the neointimal area by 49% and significantly decreased the neointimal SMCs content compared with control-siRNA. HIF-1␣ and SDF-1␣ expression were clearly diminished in neointimal cells of HIF-1␣-siRNA treated arteries. Key Words: atherosclerosis Ⅲ restenosis Ⅲ vascular biology Ⅲ chemokines Ⅲ progenitor cells Ⅲ smooth muscle cells N eointima formation and inward remodeling are the basic mechanisms of the vascular response to mechanical injury, eg, by balloon angioplasty or stent implantation in obstructive coronary artery disease. 1,2 This can result in renarrowing of the target vessel, which is termed restenosis and limits therapeutic revascularization. 3 Initial endothelial denudation leads to an exposure of subendothelial matrix, which precipitates the adhesion of activated platelets and fibrin deposition, supporting the inflammatory recruitment of leukocytes. 4 The progress of neointimal hyperplasia is attributable to the accumulation of dedifferentiated smooth muscle cells (SMCs), which by far constitute the majority of neointimal cells. 5 The traditional idea of neointima formation focused on the concept that neointimal SMCs originate from medial cells within the vessel wall, which migrate into the intima and start to proliferate. 5,6 In recent years, evidence has accumulated that bone marrow-derived vascular progenitor cells participate significantly to neointima formation after selective recruitment to the injury site, thereby directing the vascular wound healing. 7 Smooth muscle progenitor cells (SPCs) have been detected in the peripheral blood of humans 8 and appear to be the critical vascular stem cell subtype for vascular repair after mechanical injury. Neointimal lesions after endothelial denudation consist of approximately 50% to 60% bone marrow-derived SMCs. 7 Among the multiple chemokines directing vascular cell trafficking, the CXC chemokine stromal cell-derived factor (SDF)-1␣ and its receptor CXCR4 represent an important molecular signaling axis for the mobilization and recruitment of SPCs. 9,10 In a "remote control" function, upregulation of SDF...

Key Words: vascular remodeling Ⅲ vascular smooth muscle Ⅲ neointima Ⅲ restenosis Ⅲ chemokines T he response to mechanical vessel injury clinically encountered as restenosis after percutaneous interventions is characterized by the formation of neointimal tissue comprising primarily smooth muscle cells (SMCs) and leukocytes. 1,2 Although drug-eluting stents have significantly reduced the need for repeated target vessel revascularization, long-term treatment with inhibitors of platelet aggregation after drug-eluting stent implantation to avoid late stent thrombosis is still a significant constraint. 3 Impaired endothelial recovery of drug-eluting stents is caused by the unspecific antimigratory and antiproliferative activities of drugs like Rapamycin or Paclitaxel and has been identified as a major risk factor for late stent thrombosis. 4 Therefore, development of alternative drugs which inhibit specifically neointimal SMC accumulation would be preferable. Apart from proliferation of neointimal SMCs, recruitment of circulating smooth muscle progenitor cells (SPCs) contributes decisively to the accumulation of SMCs in the neointima. [5][6][7] The chemokine CXCL12 (CXC motif ligand 12) and its receptor CXC motif receptor (CXCR)4 regulate the mobilization and recruitment of SPCs after vascular injury, 5,6,8 without affecting re-endothelialization. 9 Early apoptosis of medial SMCs and the nonhypoxic activation of the transcription factor hypoxia- Original received November 10, 2009; revision received March 22, 2010; accepted March 24, 2010. In March 2010, the average time from submission to first decision for all original research papers submitted to Circulation Research was 13.3 days.From the Institute for Molecular Cardiovascular Research (P.S., E.K., T.A., R.T.A.M., Z.Z., S.A., U.S., X.L., M.v.Z., C.W., A.S.) and Interdisciplinary Center for Clinical Research BIOMAT within the Faculty of Medicine (P.S., X.L., R.T.A.M.), RWTH Aachen University, Germany; Cardiology Unit (E.K., P.R.), Medical Policlinic-City Center Campus, University of Munich, Germany; Polyphor Ltd (C.L.), Allschwill, Switzerland; and Cardiovascular Research Institute Maastricht (CARIM) 20,21 Among a variety of unsaturated LPA species, LPA20:4 (1-arachidonoyl-2-lyso-sn-glycero-3-phosphate) and the alkyl-ether analog 1-AGP18:1 (1-oleyl-2-lyso-sn-glycero-3-phosphate) have been shown to stimulate neointimal growth in rats most efficiently. 20 Although LPA serves as a mitogenic growth factor for SMCs in vitro, 22 the exact mechanism of LPA-induced neointima formation remains unclear. Interestingly, in contrast to mice with combined deficiency of LPA 1 and LPA 2 , neointimal hyperplasia after carotid ligation was enhanced in LPA 1 Ϫ/Ϫ mice. This finding suggests a major role of Edg family LPA receptors in vascular repair. 23 In vitro, unsaturated LPA induces CXCL12 expression, which is inhibited by the LPA 1 and LPA 3 receptor antagonist Ki16425. 24 We therefore hypothesize that any blocking of the LPA receptors LPA 1 and LPA 3 reduces CXCL12 expres...

Rationale: Besides their essential role in hemostasis, platelets also have functions in inflammation. In platelets, junctional adhesion molecule (JAM)-A was previously identified as an inhibitor of integrin α IIb β 3 -mediated outside-in signaling and its genetic knockdown resulted in hyperreactivity. Objective: This gain-of-function was specifically exploited to investigate the role of platelet hyperreactivity in plaque development. Methods and Results: JAM-A–deficient platelets showed increased aggregation and cellular and sarcoma tyrosine-protein kinase activation. On α IIb β 3 ligation, JAM-A was shown to be dephosphorylated, which could be prevented by protein tyrosine phosphatase nonreceptor type 1 inhibition. Mice with or without platelet-specific (tr)JAM-A-deficiency in an apolipoprotein e ( apoe –/– ) background were fed a high-fat diet. After ≤12 weeks of diet, trJAM-A –/– apoe–/– mice showed increased aortic plaque formation when compared with trJAM-A +/+ apoe –/– controls, and these differences were most evident at early time points. At 2 weeks, the plaques of the trJAM-A –/– apoe –/– animals revealed increased macrophage, T cell, and smooth muscle cell content. Interestingly, plasma levels of chemokines CC chemokine ligand 5 and CXC-chemokine ligand 4 were increased in the trJAM-A –/– apoe –/– mice, and JAM-A–deficient platelets showed increased binding to monocytes and neutrophils. Whole-blood perfusion experiments and intravital microscopy revealed increased recruitment of platelets and monocytes to the inflamed endothelium in blood of trJAM-A –/– apoe –/– mice. Notably, these proinflammatory effects of JAM-A–deficient platelets could be abolished by the inhibition of α IIb β 3 signaling in vitro. Conclusions: Deletion of JAM-A causes a gain-of-function in platelets, with lower activation thresholds and increased inflammatory activities. This leads to an increase of plaque formation, particularly in early stages of the disease.

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SummaryIn recent years, it has become clear that platelets and platelet-derived chemokines, beyond their role in thrombosis and haemostasis, are important mediators affecting a broad spectrum of (patho)physiological conditions. These biologically active proteins are released from α-granules upon platelet activation, most probably even during physiological conditions. In this review, we give a concise overview and an update on the current understanding of platelet-derived chemokines in a context of health and disease.

Objective: Inflammatory activation changes the mitochondrial function of macrophages from oxidative phosphorylation to reactive oxygen species production, which may promote necrotic core formation in atherosclerotic lesions. In hypoxic and cancer cells, HIF-1α (hypoxia-inducible factor) promotes oxygen-independent energy production by microRNAs. Therefore, we studied the role of HIF-1α in the regulation of macrophage energy metabolism in the context of atherosclerosis. Approach and Results: Myeloid cell–specific deletion of Hif1a reduced atherosclerosis and necrotic core formation by limiting macrophage necroptosis in apolipoprotein E-deficient mice. In inflammatory bone marrow–derived macrophages, deletion of Hif1a increased oxidative phosphorylation, ATP levels, and the expression of genes encoding mitochondrial proteins and reduced reactive oxygen species production and necroptosis. microRNA expression profiling showed that HIF-1α upregulates miR-210 and downregulates miR-383 levels in lesional macrophages and inflammatory bone marrow–derived macrophages. In contrast to miR-210 , which inhibited oxidative phosphorylation and enhanced mitochondrial reactive oxygen species production, miR-383 increased ATP levels and inhibited necroptosis. The effect of miR-210 was due to targeting 2,4-dienoyl-CoA reductase, which is essential in the β oxidation of unsaturated fatty acids. miR-383 affected the DNA damage repair pathway in bone marrow–derived macrophages by targeting poly(ADP-ribose)-glycohydrolase (Parg), which reduced energy consumption and increased cell survival. Blocking the targeting of Parg by miR-383 prevented the protective effect of Hif1a deletion in macrophages on atherosclerosis and necrotic core formation in mice. Conclusions: Our findings unveil a new mechanism by which activation of HIF-1α in inflammatory macrophages increases necroptosis through microRNA-mediated ATP depletion, thus increasing atherosclerosis by necrotic core formation.