Elsevier

Atherosclerosis

Volume 192, Issue 1, May 2007, Pages 75-84
Atherosclerosis

Sequential patterns of chemokine- and chemokine receptor-synthesis following vessel wall injury in porcine coronary arteries

https://doi.org/10.1016/j.atherosclerosis.2006.05.050Get rights and content

Abstract

Inflammation plays a central role in vascular repair, and spreads into perivascular tissue (PVT) following angioplasty. Chemokines (CK) and chemokine receptors (CKR) are key determinants of inflammatory chemotaxis. We sought to assess the arterial and perivascular expression of the CK CCL2 and CXCL2, and the CKR CCR2, CCR5, and CXCR4 in balloon-injured porcine coronary arteries. Vascular cells that express specific CK and CKR mRNA during post-angioplasty time course were detected by in situ hybridization (ISH), and expression was quantified by real time RT-PCR in PVT. CCL2 was maximal in PVT from 2 to 24 h post injury, coincident with local macrophage-activation. Expression was upregulated in media and adventitia from 24 h to 3 days, and in neointima at 7 days. CXCL2 was detected in media at 2 and 4 h, and also in some neointimal cells. CCR2 and CCR5 were maximal in PVT at 24 h and 3 days, respectively. Expression shifted to media and adventitia at 2 and 3 days, and to neointima and adventitia at 7 days, and was low at 14 days. CXCR4 was low in PVT, but was upregulated in media and adventitia at 2 and 3 days, as well as in neointima and adventitia at 7 days.

In conclusion, PVT is the primary source of inflammatory CK and CKR early post-angioplasty. Specific sequential patterns of CK- and CKR-synthesis are identified that may regulate phase-specific chemotaxis by spatio-temporally differential expression during coronary response to injury.

Introduction

Inflammation plays a key role in vascular repair, neointima formation and restenosis [1], [2]. Systemic leukocyte depletion with attenuated mononuclear cell recruitment reduces neointimal volume [3], [4]. As to specific leukocyte subtypes, work from our group previously showed neutrophil granulocyte and macrophage infiltration throughout extensive perivascular coronary tissue post-angioplasty [5], and vascular dendritic cells were identified as a novel cell type in rodent and human neointimal lesions [6], [7]. Although increasing evidence suggests a coordinated recruitment of inflammatory cell populations as a prerequisite for vascular healing, the determinants that induce and sustain this process are incompletely understood. Analyzing mechanisms responsible for the phase-specific and spatially differential chemotaxis of leukocytes and other vascular cells is essential for understanding coronary healing and neointima formation.

Inflammatory cell extravasation and tissue homing depend on chemoattractants such as chemokines (CK), cytokines that induce directed basal and inflammatory leukocyte chemotaxis, gene transcription, proliferation, and apoptosis [8], [9]. Chemokines interact with seven-transmembrane G protein-coupled chemokine receptors (CKR) in a complex, partly overlapping system of >50 ligands and >20 receptors identified to date [9]. Endothelial cells (ECs), smooth muscle cells (SMCs), stromal cells, and epithelial cells can also express chemokines and their receptors, indicating that these cell types are sensitive to specific chemokine signaling [8], [9], [10], [11]. Chemokines like CCL2 and -11, CXCL8, -9, -10, -11, and -12, as well as chemokine receptors like CCR2 and -3, and CXCR2 have been located in atherosclerotic plaques [12], [13], [14], [15], [16], [17]. CCL2 and -11, CXCL8, CCR2 and -3 and others were described following vascular injury [18], [19], [20]. However, the exact time course and transmural arterial localization of chemokine and chemokine receptor expression after angioplasty has not been described in detail.

In the present study, we hypothesized that distinct transmural and perivascular expression patterns of selected chemokines and chemokine receptors implicated in granulocyte, macrophage, and progenitor cell recruitment should be distinguishable after porcine coronary balloon angioplasty. Our findings provide novel insights into coronary inflammatory cell chemotaxis that is essential to ensure a coordinated inflammatory process, and thereby initiates and modulates vascular repair.

Section snippets

Porcine coronary angioplasty

Balloon overstretch injury was performed in left anterior descending and circumflex coronary arteries of female domestic pigs (n = 28; 25–35 kg; Clemson University, Clemson, SC) [5], [21]. Animals were sedated by an intramuscular injection of ketamine (20 mg/kg), xylazine (2 mg/kg), and atropine (0.05 mg/kg). Thiopental (10 mg/kg) was administered intravenously, and general anesthesia was maintained with 1.0–1.5% isoflurane. At 2, 4, and 24 h, 2, 3, 7, or 14 days, animals were sacrificed with an

Results

Porcine coronary arteries were examined at 2, 4, and 24 h, 2, 3, 7, and 14 days after balloon overstretch injury (Table 2). Inflammatory cell aggregation was maximal adjacent to medial injury, and occurred in perivascular tissue (Fig. 1, Table 3). MPO immunostaining was seen in cells with typical neutrophil granulocyte appearance, predominantly located in adventitial and adjacent perivascular areas at 2, 4, and 24 h (Fig. 1A, Table 3). Macrophages, identified by HB141/142 immunostaining and by c-

Discussion

This study demonstrates the transmural and perivascular expression patterns of the chemokines CCL2 and CXCL2 and the chemokine receptors CCR2, CCR5, and CXCR4 following porcine coronary angioplasty. Perivascular tissue is identified as the primary source of the inflammatory leukocyte chemoattractant CCL2 and abundantly contains cells that express chemokine receptors early post-angioplasty. Inflammatory cytokine expression predominantly occurs in granulocytes, macrophages, and myofibroblasts (

Acknowledgements

Supported, in part, by NIH (National Institutes of Health, Bethesda, MD, USA) grant HL57908 (to J.N.W.) and a German Cardiac Society (Duesseldorf, Germany) postdoctoral scholarship (to A.J.). The authors would like to thank Giji Joseph and Tracey Couse for their expert technical assistance.

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    Present address: Department of Medicine II, Director: Prof. Dr. T. Munzel, Johannes Gutenberg-University, Mainz, Germany

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