Receptor activator of NF-κB ligand (RANKL) increases the release of neutrophil products associated with coronary vulnerability

Summary

The “blood vulnerability”, resulting from the complex balance between serum molecules and inflammatory cell atherosclerotic activities, is a major determinant in the evaluation of the “global patient cardiovascular vulnerability”. In the present study, we focused on the role of the soluble receptor activator of nuclear factor kappa-B (NF-κB) ligand (RANKL, a potential marker of coronary calcification and vulnerability) in the release of neutrophilic proteases. Then, the association between these mediators and the degree of coronary calcification (assessed by coronary calcium score [CCS]) was investigated in 20 subjects (aged ≥65 years) asymptomatic for cardiovascular disease. Results showed that RANKL dose-dependently induced matrix metalloprotease (MMP)-8 and MMP-9 release from human primary neutrophils cultured in Teflon dishes (suspension condition, mimicking cells circulating in the blood stream). Conversely, when adherent to polystyrene, neutrophils became unresponsive to RANKL. RANKL did not influence the release of other neutrophilic products in suspension and adherence cultures as well as neutrophil migration. RANKL-induced release of MMPs was dependent on the activation of defined intracellular signalling pathways (PI3K/Akt and ERK1/2). In asymptomatic subjects, serum levels of RANKL, MMP-8 and MMP-9 positively correlated with CCS, reflecting a potential relationship between circulating RANKL and coronary calcification. In conclusion, RANKL increased the release of neutrophilic products potentially related to the “blood” vulnerability via defined intracellular pathways. Serum levels of RANKL might represent a potential biomarker of coronary calcification and related cardiovascular risk.

Keywords

Atherosclerosis - matrix metalloproteinases - leukocyte function/activation

^* These authors equally contributed as first authors to this work.

^# These authors equally contributed as last authors to this work.

• References

• 1 Montecucco F, Steffens S, Mach F. The immune response is involved in atherosclerotic plaque calcification: could the RANKL/RANK/OPG system be a marker of plaque instability?. Clin Dev Immunol 2007; 2007: 75805.
• 2 Hansson GK, Libby P. The immune response in atherosclerosis: a double-edged sword. Nat Rev Immunol 2006; 06: 508-519.
• 3 Montero I, Orbe J, Varo N. et al. C-reactive protein induces matrix metalloproteinase-1 and -10 in human endothelial cells: implications for clinical and subclinical atherosclerosis. J Am Coll Cardiol 2006; 47: 1369-1378.
• 4 Naghavi M, Libby P, Falk E. et al. From vulnerable plaque to vulnerable patient: a call for new definitions and risk assessment strategies: Part I. Circulation 2003; 108: 1772-1778.
• 5 Pradhan-Palikhe P, Vikatmaa P, Lajunen T. et al. Elevated MMP-8 and decreased myeloperoxidase concentrations associate significantly with the risk for peripheral atherosclerosis disease and abdominal aortic aneurysm. Scand J Immunol 2010; 72: 150-157.
• 6 Brunner S, Kim JO, Methe H. Relation of matrix metalloproteinase-9/tissue inhibitor of metalloproteinase-1 ratio in peripheral circulating CD14+ monocytes to progression of coronary artery disease. Am J Cardiol 2010; 105: 429-434.
• 7 Tuomainen AM, Nyyssönen K, Laukkanen JA. et al. Serum matrix metalloproteinase-8 concentrations are associated with cardiovascular outcome in men. Arterioscler Thromb Vasc Biol 2007; 27: 2722-2728.
• 8 Naghavi M, Libby P, Falk E. et al. From vulnerable plaque to vulnerable patient: a call for new definitions and risk assessment strategies: Part II. Circulation 2003; 108: 1664-1672.
• 9 Khot UN, Khot MB, Bajzer CT. et al. Prevalence of conventional risk factors in patients with coronary heart disease. J Am Med Assoc 2003; 290: 898-904.
• 10 Pearson TA, Mensah GA, Alexander RW. et al. Markers of inflammation and cardiovascular disease: application to clinical and public health practice: A statement for healthcare professionals from the Centers for Disease Control and Prevention and the American Heart Association. Circulation 2003; 107: 499-511.
• 11 Montecucco F, Mach F. Common inflammatory mediators orchestrate patho-physiological processes in rheumatoid arthritis and atherosclerosis. Rheumatology (Oxford) 2009; 48: 11-22.
• 12 Choudhary S, Higgins CL, Chen IY. et al. Quantitation and localization of matrix metalloproteinases and their inhibitors in human carotid endarterectomy tissues. Arterioscler Thromb Vasc Biol 2006; 26: 2351-2358.
• 13 van Oostrom O, Velema E, Schoneveld AH. et al. Age-related changes in plaque composition: a study in patients suffering from carotid artery stenosis. Cardiovasc Pathol 2005; 14: 126-134.
• 14 Montecucco F, Lenglet S, Gayet-Ageron A. et al. Systemic and intraplaque mediators of inflammation are increased in patients symptomatic for ischemic stroke. Stroke 2010; 41: 1394-1404.
• 15 Montecucco F, Vuilleumier N, Pagano S. et al. Anti-Apolipoprotein A-1 auto-antibodies are active mediators of atherosclerotic plaque vulnerability. Eur Heart J 2011; 32: 412-421.
• 16 Zernecke A, Bot I, Djalali-Talab Y. et al. Protective role of CXC receptor 4/CXC ligand 12 unveils the importance of neutrophils in atherosclerosis. Circ Res 2008; 102: 209-217.
• 17 Weber C, Zernecke A, Libby P. The multifaceted contributions of leukocyte subsets to atherosclerosis: lessons from mouse models. Nat Rev Immunol 2008; 08: 802-815.
• 18 De Backer G, Ambrosioni E, Borch-Johnsen K. et al. Third Joint Task Force of European and Other Societies on Cardiovascular Disease Prevention in Clinical Practice. European guidelines on cardiovascular disease prevention in clinical practice. Third Joint Task Force of European and Other Societies on Cardiovascular Disease Prevention in Clinical Practice. Eur Heart J 2003; 24: 1601-1610.
• 19 Kiechl S, Schett G, Schwaiger J. et al. Soluble receptor activator of nuclear factor-kappa B ligand and risk for cardiovascular disease. Circulation 2007; 116: 385-391.
• 20 Quercioli A, Montecucco F, Bertolotto M. et al. Coronary artery calcification and cardiovascular risk: the role of RANKL/OPG signalling. Eur J Clin Invest 2010; 40: 645-654.
• 21 Quercioli A, Luciano Viviani G, Dallegri F. et al. Receptor activator of nuclear factor kappa B ligand/osteoprotegerin pathway is a promising target to reduce atherosclerotic plaque calcification. Crit Pathw Cardiol 2010; 09: 227-230.
• 22 Montecucco F, Steffens S, Burger F. et al. C-reactive protein (CRP) induces chemo-kine secretion via CD11b/ICAM-1 interaction in human adherent monocytes. J Leukoc Biol 2008; 84: 1109-1119.
• 23 DiStasi MR, Ley K. Opening the flood-gates: how neutrophil-endothelial interactions regulate permeability. Trends Immunol 2009; 30: 547-556.
• 24 Montecucco F, Steffens S, Burger F. et al. Tumor necrosis factor-alpha (TNF-alpha) induces integrin CD11b/CD18 (Mac-1) up-regulation and migration to the CC chemokine CCL3 (MIP-1alpha) on human neutrophils through defined signalling pathways. Cell Signal 2008; 20: 557-568.
• 25 Montecucco F, Bertolotto M, Ottonello L. et al. Chlorhexidine prevents hypochlorous acid-induced inactivation of alpha1-antitrypsin. Clin Exp Pharmacol Physiol 2009; 36: e72-77.
• 26 Bruno O, Brullo C, Bondavalli F. et al. Synthesis and biological evaluation of N-pyrazolyl-N'-alkyl/benzyl/phenylureas: a new class of potent inhibitors of interleukin 8-induced neutrophil chemotaxis. J Med Chem 2007; 50: 3618-3626.
• 27 Schindler TH, Facta AD, Prior JO. et al. Structural alterations of the coronary arterial wall are associated with myocardial flow heterogeneity in type 2 diabetes mellitus. Eur J Nucl Med Mol Imaging 2009; 36: 219-229.
• 28 Poubelle PE, Chakravarti A, Fernandes MJ. et al. Differential expression of RANK, RANK-L, and osteoprotegerin by synovial fluid neutrophils from patients with rheumatoid arthritis and by healthy human blood neutrophils. Arthritis Res Ther 2007; 09: R25.
• 29 Borregaard N, Sørensen OE, Theilgaard-Mönch K. Neutrophil granules: a library of innate immunity proteins. Trends Immunol 2007; 28: 340-345.
• 30 Chodniewicz D, Zhelev DV. Novel pathways of F-actin polymerization in the human neutrophil. Blood 2003; 102: 2251-2258.
• 31 Ehrenfeld P, Matus CE, Pavicic F. et al. Kinin B1 receptor activation turns on exocytosis of matrix metalloprotease-9 and myeloperoxidase in human neutrophils: involvement of mitogen-activated protein kinase family. J Leukoc Biol 2009; 86: 1179-1189.
• 32 Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults. Executive Summary of The Third Report of The National Cholesterol Education Program (NCEP) Expert Panel on Detection, Evaluation, And Treatment of High Blood Cholesterol In Adults (Adult Treatment Panel III). J Am Med Assoc 2001; 285: 2486-2497.
• 33 Montecucco F, Bertolotto M, Vuilleumier N. et al. Acipimox reduces circulating levels of insulin and associated neutrophilic inflammation in metabolic syndrome. Am J Physiol Endocrinol Metab 2011; 300: E681-690.
• 34 Catini C, Rontani O, Mayer G. A simple method for the isolation of highly purified neutrophil granulocytes from human peripheral blood. Arch Ital Anat Embriol 1985; 90: 261-266.
• 35 Kim HR, Kim KW, Jung HG. et al. Macrophage migration inhibitory factor enhances osteoclastogenesis through upregulation of RANKL expression from fibroblast-like synoviocytes in patients with rheumatoid arthritis. Arthritis Res Ther 2011; 13: R43.
• 36 Aoki S, Nakagomi A, Asai K. et al. Elevated peripheral blood mononuclear cell count is an independent predictor of left ventricular remodeling in patients with acute myocardial infarction. J Cardiol 2011; 57: 202-207.
• 37 Ress C, Tschoner A, Ciardi C. et al. Influence of significant weight loss on serum matrix metalloproteinase (MMP)-7 levels. Eur Cytokine Netw 2010; 21: 65-70.
• 38 Vik A, Mathiesen EB, Brox J. et al. Serum osteoprotegerin is a predictor for incident cardiovascular disease and mortality in a general population – The Tromsø Study. J Thromb Haemost 2011; 09: 638-644.
• 39 Lieb W, Gona P, Larson MG. et al. Biomarkers of the osteoprotegerin pathway: clinical correlates, subclinical disease, incident cardiovascular disease, and mortality. Arterioscler Thromb Vasc Biol 2010; 30: 1849-1854.
• 40 Asanuma Y, Chung CP, Oeser A. et al. Serum osteoprotegerin is increased and independently associated with coronary-artery atherosclerosis in patients with rheumatoid arthritis. Atherosclerosis 2007; 195: e135-141.
• 41 Mesquita M, Demulder A, Damry N. et al. Plasma osteoprotegerin is an independent risk factor for mortality and an early biomarker of coronary vascular calcification in chronic kidney disease. Clin Chem Lab Med 2009; 47: 339-346.
• 42 Barreto DV, Barreto FC, Carvalho AB. et al. Coronary calcification in hemodialysis patients: the contribution of traditional and uremia-related risk factors. Kidney Int 2005; 67: 1576-1582.
• 43 Huang H, Virmani R, Younis H. et al. The impact of calcification on the biomechanical stability of atherosclerotic plaques. Circulation 2001; 103: 1051-1056.
• 44 Imoto K, Hiro T, Fujii T. et al. Longitudinal structural determinants of atherosclerotic plaque vulnerability: a computational analysis of stress distribution using vessel models and three-dimensional intravascular ultrasound imaging. J Am Coll Cardiol 2005; 46: 1507-1515.