We used a novel approach of cytostatically induced leucocyte depletion and subsequent reconstitution with leucocytes deprived of classical (inflammatory/Gr1hi) or non-classical (resident/Gr1lo) monocytes to dissect their differential role in atheroprogression under high-fat diet (HFD). Apolipoprotein E-deficient (Apoe−/−) mice lacking classical but not non-classical monocytes displayed reduced lesion size and macrophage and apoptotic cell content. Conversely, HFD induced a selective expansion of classical monocytes in blood and bone marrow. Increased CXCL1 levels accompanied by higher expression of its receptor CXCR2 on classical monocytes and inhibition of monocytosis by CXCL1-neutralization indicated a preferential role for the CXCL1/CXCR2 axis in mobilizing classical monocytes during hypercholesterolemia. Studies correlating circulating and lesional classical monocytes in gene-deficient Apoe−/− mice, adoptive transfer of gene-deficient cells and pharmacological modulation during intravital microscopy of the carotid artery revealed a crucial function of CCR1 and CCR5 but not CCR2 or CX3CR1 in classical monocyte recruitment to atherosclerotic vessels. Collectively, these data establish the impact of classical monocytes on atheroprogression, identify a sequential role of CXCL1 in their mobilization and CCR1/CCR5 in their recruitment.

Methods and Results: Compared to Apoe؊/؊ mice, Cramp ؊/؊ Apoe ؊/؊ mice exhibit reduced lesion sizes with lower macrophage numbers. In atherosclerotic aortas, we could detect CRAMP specifically in neutrophils, but not in monocytes or macrophages. By use of intravital microscopy, CRAMP was found to be deposited by activated neutrophils on inflamed endothelium of large arteries. In this location cathelicidins promote adhesion of classical monocytes and neutrophils, but not nonclassical monocytes in a formyl-peptide receptor-dependent manner. Key Words: atherosclerosis Ⅲ monocyte recruitment Ⅲ neutrophil A therosclerosis is a chronic inflammation of the arterial vessel wall with relatively well-defined roles for leukocytes such as macrophages and lymphocytes. 1,2 Recent studies, however, have revealed that neutrophils infiltrate atherosclerotic lesions at various time points, 3-5 and depletion studies provide evidence for a proatherogenic role of neutrophils. 5,6 Nevertheless, mechanistic insights into how neutrophils promote early atherosclerotic lesion formation remain elusive. Neutrophils contain granules with more than 300 different proteins that undergo limited exocytosis on neutrophil extravasation. 7 Some of these proteins are able to activate and recruit immune cells and thus have been coined alarmins. 8 Cathelicidins (CRAMP in mouse, LL37 in humans) residing in neutrophil secondary granules were shown to potently activate and recruit monocytes and macrophages, 9,10 thus fulfilling alarmin criteria. Because cathelicidins were identified in atherosclerotic lesions, 11 we investigated their role in a mouse model of atherosclerosis. Conclusions: Editorial, see p 1036 In This Issue, see p 1035 MethodsDetailed Methods are provided in the Online Supplement. Plaque StudiesCramp Ϫ/Ϫ mice 12 were crossed with Apoe Ϫ/Ϫ mice. Atherosclerotic lesion size as well as lesional neutrophil and macrophage content were assessed by histology and immunohistochemistry. Intravital MicroscopyLeukocyte adhesion to the carotid artery was studied by intravital fluorescence microscopy as described previously. 5 ResultsTo investigate the role of CRAMP in early atherosclerotic lesion formation, we fed Apoe Ϫ/Ϫ and CrampOriginal received January 29, 2012; revision received February 15, 2012; accepted February 27, 2012. In January 2012, the average time from submission to first decision for all original research papers submitted to Circulation Research was 13.88 days.Brief UltraRapid Communications are designed to be a format for manuscripts that are of outstanding interest to the readership, report definitive observations, but have a relatively narrow scope.

Note: The editorial process for this article was fully handled by Prof. G. Y. H. Lip, Editor-in-Chief.

Percutaneous transluminal angioplasty with stent implantation is used to dilate of arteries narrowed by atherosclerotic plaques and to revascularize coronary arteries occluded by atherothrombosis in myocardial infarction. Commonly applied drug-eluting stents release antiproliferative or anti-inflammatory agents to reduce the incidence of in-stent stenosis. However, these stents may lead to in-stent stenosis and increase the rate late stent thrombosis, an obstacle to Correspondence: Oliver Soehnlein, MD, PhD or Christian Weber, MD, Institute for Cardiovascular Prevention, Pettenkoferstr. 9, 80336 Munich, Phone +49-(0)89-5160-4350, Fax +49-(0)89-5160-4352, oliver.soehnlein@med.uni-muenchen.de or christian.weber@med.uni-muenchen.de. * These authors contributed equally. Competing interests:The authors do not declare any competing financial interests. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript optimal revascularization possibly related to endothelial recovery. Here we examined the contribution of neutrophils and neutrophilic granule proteins to arterial healing after injury. We found that neutrophil-born cathelicidin (mouse CRAMP, human LL-37) promoted reendothelization and thereby limited neointima formation after stent implantation. We then translated these findings, generating a neutrophil-instructing biofunctionalized miniaturized Nitinol stent coated with LL-37. This stent reduced in-stent stenosis in a mouse model of atherosclerosis, suggesting that LL-37 may promote vascular healing after interventional therapy.

Chronic hepatitis leads to liver fibrosis and cirrhosis. Cirrhosis is a major cause of worldwide morbidity and mortality. Macrophages play a key role in fibrosis progression and reversal. However, the signals that determine fibrogenic vs fibrolytic macrophage function remain ill defined. We studied the role of interleukin-4 receptor α (IL-4Rα), a potential central switch of macrophage polarization, in liver fibrosis progression and reversal. We demonstrate that inflammatory monocyte infiltration and liver fibrogenesis were suppressed in general IL-4Rα−/− as well as in macrophage-specific IL-4Rα−/− (IL-4RαΔLysM) mice. However, with deletion of IL-4RαΔLysM spontaneous fibrosis reversal was retarded. Results were replicated by pharmacological intervention using IL-4Rα-specific antisense oligonucleotides. Retarded resolution was linked to the loss of M2-type resident macrophages, which secreted MMP-12 through IL-4 and IL-13-mediated phospho-STAT6 activation. We conclude that IL-4Rα signaling regulates macrophage functional polarization in a context-dependent manner. Pharmacological targeting of macrophage polarization therefore requires disease stage-specific treatment strategies.Research in ContextAlternative (M2-type) macrophage activation through IL-4Rα promotes liver inflammation and fibrosis progression but speeds up fibrosis reversal. This demonstrates context dependent, opposing roles of M2-type macrophages. During reversal IL-4Rα induces fibrolytic MMPs, especially MMP-12, through STAT6. Liver-specific antisense oligonucleotides efficiently block IL-4Rα expression and attenuate fibrosis progression.