Distribution of Matrix Metalloproteinases in Human Atherosclerotic Carotid Plaques and Their Production by Smooth Muscle Cells and Macrophage Subsets

Molecular Imaging and Biology, Sep 2015

Purpose In this study, the potential of matrix metalloproteinase (MMP) sense for detection of atherosclerotic plaque instability was explored. Secondly, expression of MMPs by macrophage subtypes and smooth muscle cells (SMCs) was investigated. Procedures Twenty-three consecutive plaques removed during carotid endarterectomy were incubated in MMPSense™ 680 and imaged with IVIS® Spectrum. mRNA levels of MMPs, macrophage markers, and SMCs were determined in plaque specimens, and in in vitro differentiated M1 and M2 macrophages. Results There was a significant difference between autofluorescence signals and MMPSense signals, both on the intraluminal and extraluminal sides of plaques. MMP-9 and CD68 messenger RNA (mRNA) expression was higher in hot spots, whereas MMP-2 and αSMA expression was higher in cold spots. In vitro M2 macrophages had higher mRNA expression of MMP-1, MMP-9, MMP-12, and TIMP-1 compared to M1 macrophages. Conclusion MMP-9 is most dominantly MMP present in atherosclerotic plaques and is produced by M2 rather than M1 macrophages.

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Distribution of Matrix Metalloproteinases in Human Atherosclerotic Carotid Plaques and Their Production by Smooth Muscle Cells and Macrophage Subsets

Distribution of Matrix Metalloproteinases in Human Atherosclerotic Carotid Plaques and Their Production by Smooth Muscle Cells and Macrophage Subsets Nynke A. Jager 2 Bastiaan M. Wallis de Vries 1 Jan-Luuk Hillebrands 0 Niels J. Harlaar 1 René A. Tio 5 Riemer H. J. A. Slart 4 Gooitzen M. van Dam 1 Hendrikus H. Boersma 3 4 Clark J. Zeebregts 1 Johanna Westra 2 0 Departments of Pathology and Medical Biology, University Medical Center Groningen, University of Groningen , PB 30.0019700 RB, Groningen , The Netherlands 1 Department of Surgery, Division of Vascular Surgery, University Medical Center Groningen, University of Groningen , Groningen, PB 30.0019700 RB, Groningen , The Netherlands 2 Departments of Rheumatology and Clinical Immunology, University Medical Center Groningen, University of Groningen , PB 30.0019700 RB, Groningen , The Netherlands 3 Departments of Clinical Pharmacy and Pharmacology, University Medical Center Groningen, University of Groningen , PB 30.0019700 RB, Groningen , The Netherlands 4 Departments of Nuclear Medicine and Molecular Imaging, University Medical Center Groningen, University of Groningen , PB 30.0019700 RB, Groningen , The Netherlands 5 Department of Cardiology, University Medical Center Groningen, University of Groningen , PB 30.0019700 RB, Groningen , The Netherlands Purpose: In this study, the potential of matrix metalloproteinase (MMP) sense for detection of atherosclerotic plaque instability was explored. Secondly, expression of MMPs by macrophage subtypes and smooth muscle cells (SMCs) was investigated. Procedures: Twenty-three consecutive plaques removed during carotid endarterectomy were incubated in MMPSense™ 680 and imaged with IVIS® Spectrum. mRNA levels of MMPs, macrophage markers, and SMCs were determined in plaque specimens, and in in vitro differentiated M1 and M2 macrophages. Results: There was a significant difference between autofluorescence signals and MMPSense signals, both on the intraluminal and extraluminal sides of plaques. MMP-9 and CD68 messenger RNA (mRNA) expression was higher in hot spots, whereas MMP-2 and αSMA expression was higher in cold spots. In vitro M2 macrophages had higher mRNA expression of MMP-1, MMP-9, MMP-12, and TIMP-1 compared to M1 macrophages. Conclusion: MMP-9 is most dominantly MMP present in atherosclerotic plaques and is produced by M2 rather than M1 macrophages. Matrix metalloproteinase; Macrophage; Atherosclerotic plaque; Smooth muscle cell; MMPSense - Abbreviations: BMI, Body mass index; CCD, Charged-coupled device; CEA, Carotid endarterectomy; Ct, Threshold cycle; CVA, Cerebrovascular accident; CXCL4, Chemokine ligand 4; ECM, Extracellular matrix; ELISA, Enzyme-linked immunosorbent assay; FCS, Fetal calf serum; GAPDH, Glyceraldehyde-3-phosphate dehydrogenase; HRP, Horseradish peroxidase; IFN-γ, Interferon-gamma; IL, Interleukin; IRB, International Review Board; LPS, Lipopolysacharride; MCSF, Macrophage colony-stimulating factor; MMP, Matrix metalloproteinases; mRNA, Messenger ribonucleic acid; PBMCs, Peripheral blood mononuclear cells; SMCs, Smooth muscle cells; TBR, Target to background; TIA, Transient ischemic attack; TIMP, Tissue inhibitors of metalloproteinases Atherosclerosis is a progressive inflammatory disease characterized by the accumulation of lipid-filled macrophages within the arterial intima. Continued inflammation may promote rupture of the atherosclerotic plaque’s protective fibrous cap causing subsequent clinical ischemic events [1, 2]. The fibrous cap covering an advanced atherosclerotic plaque is typically composed of smooth muscle cells (SMCs) and extracellular matrix (ECM) [3]. Activated monocyte-derived macrophages and SMCs are critically involved in the development of high-risk vulnerable plaques by producing matrix metalloproteinases (MMPs) [1, 4, 5]. A heterogeneous population of macrophages exists including a classically activated macrophage type (M1) as well as an alternatively activated macrophage population (M2) [6]. The M1 macrophage is thought to have proinflammatory properties, and polarization in vitro is driven by low concentration lipopolysacharride (LPS) and interferon-gamma (IFN-γ). Defined as classically, however, M2 macrophages are anti-inflammatory and immune regulatory. Upon cytokine stimulation, they modify development of atherosclerotic plaques. The M2 macrophage population can be divided in M2a, M2b, and M2c subtypes, depending on the cytokine environment (IL-4, immune complexes, and IL-10, respectively)[7]. In contrast to its classical M2 properties, the function of M2a macrophages is type II inflammation, and the function of type M2c is matrix deposition and tissue remodeling; this last mentioned type might be most important in atherosclerosis [8]. Recently, Wolfs et al. suggested that additional circumstances in the local microenvironment makes macrophage polarization in the atherosclerotic tissue even more complex than the typically described M1 and M2 macrophage d (...truncated)


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Nynke A. Jager, Bastiaan M. Wallis de Vries, Jan-Luuk Hillebrands, Niels J. Harlaar, René A. Tio, Riemer H. J. A. Slart, Gooitzen M. van Dam, Hendrikus H. Boersma, Clark J. Zeebregts, Johanna Westra. Distribution of Matrix Metalloproteinases in Human Atherosclerotic Carotid Plaques and Their Production by Smooth Muscle Cells and Macrophage Subsets, Molecular Imaging and Biology, 2015, pp. 283-291, Volume 18, Issue 2, DOI: 10.1007/s11307-015-0882-0