Thromb Haemost 2012; 107(01): 140-149
DOI: 10.1160/TH11-05-0368
Cardiovascular Biology and Cell Signalling
Schattauer GmbH

Increased secretion of Gas6 by smooth muscle cells in human atherosclerotic carotid plaques

Sylvain Clauser
1   EA 4531, Université Paris-Sud, Châtenay-Malabry, France
2   AP-HP, Hôpital Ambroise Paré, Boulogne-Billancourt, France
3   Faculté de Médecine, Université de Versailles-Saint Quentin, Guyancourt, France
,
Olivier Meilhac
4   INSERM U698, Paris, France
5   AP-HP, CHU X-Bichat, Paris, France
6   Université Paris Diderot, Paris, France
,
Ivan Bièche
7   INSERM UMR S745, Faculté de Pharmacie, Paris, France
8   Université Paris Descartes, Paris, France
,
Pierre Raynal
9   Centre Hospitalier de Versailles, Le Chesnay, France
,
Patrick Bruneval
8   Université Paris Descartes, Paris, France
10   INSERM U970, Paris, France
11   AP-HP, Hôpital Européen Georges-Pompidou, Paris, France
,
Jean-Baptiste Michel
4   INSERM U698, Paris, France
5   AP-HP, CHU X-Bichat, Paris, France
6   Université Paris Diderot, Paris, France
,
Delphine Borgel
1   EA 4531, Université Paris-Sud, Châtenay-Malabry, France
2   AP-HP, Hôpital Ambroise Paré, Boulogne-Billancourt, France
› Author Affiliations
Financial support: This work was funded by the Leducq foundation: Leducq Transatlantic Network of Excellence on Atherothrombosis Research (LENA), and Leducq International Network Against Thrombosis (LINAT).
Further Information

Publication History

Received: 31 May 2011

Accepted after major revision: 04 October 2011

Publication Date:
29 November 2017 (online)

Summary

Vitamin K-dependent protein Gas6 (growth-arrest specific gene 6) plays a role in vascular smooth muscle cell (VSMC) survival and migration, as well as in endothelium and leukocyte activation, and could therefore be involved in atherosclerosis. However, the study of mouse models has led to contradictory results regarding the pro- or anti-atherogenic properties of Gas6, and relatively few data are available in human pathophysiology. To better understand the implication of Gas6 in human atherosclerosis, we studied Gas6 expression and secretion in vitro in human VSMC, and analysed the effect of Gas6 on inflammatory gene expression in these cells. We show that Gas6 secretion in VSMC is strongly induced by the anti-inflammatory cytokine transforming growth factor (TGF)β, and that VSMC stimulation by recombinant Gas6 decreases the expression of inflammatory genes tumour necrosis factor (TNF)α and intracellular adhesion molecule (ICAM)-1. The study of Gas6 expression in human carotid endarterectomy samples revealed that Gas6 is mainly expressed by VSMC at all stages of human atherosclerosis, but is not detected in normal vessel wall. Analysis of plaque secretomes showed that Gas6 secretion is markedly higher in non-complicated plaques than in complicated plaques, and that TGFβ secretion pattern mirrors that of Gas6. We conclude that Gas6 is secreted in human atherosclerotic plaques by VSMC following stimulation by TGFβ, and that Gas6 secretion decreases with plaque complication. Therefore, we propose that Gas6 acts as a protective factor, in part by reducing the pro-inflammatory phenotype of VSMC.

 
  • References

  • 1 Manfioletti G, Brancolini C, Avanzi G. et al. The protein encoded by a growth arrest-specific gene (gas6) is a new member of the vitamin K-dependent proteins related to protein S, a negative coregulator in the blood coagulation cascade. Mol Cell Biol 1993; 13: 4976-4985.
  • 2 Schneider C, King RM, Philipson L. Genes specifically expressed at growth arrest of mammalian cells. Cell 1988; 54: 787-793.
  • 3 Nakano T, Higashino K, Kikuchi N. et al. Vascular smooth muscle cell-derived, Gla-containing growth-potentiating factor for Ca(2+)-mobilizing growth factors. J Biol Chem 1995; 270: 5702-5705.
  • 4 O’Donnell K, Harkes IC, Dougherty L. et al. Expression of receptor tyrosine kinase Axl and its ligand Gas6 in rheumatoid arthritis: evidence for a novel endothelial cell survival pathway. Am J Pathol 1999; 154: 1171-1180.
  • 5 Avanzi GC, Gallicchio M, Cavalloni G. et al. GAS6, the ligand of Axl and Rse receptors, is expressed in hematopoietic tissue but lacks mitogenic activity. Exp Hema-tol 1997; 25: 1219-1226.
  • 6 Angelillo-Scherrer A, de Frutos P, Aparicio C. et al. Deficiency or inhibition of Gas6 causes platelet dysfunction and protects mice against thrombosis. Nat Med 2001; 07: 215-221.
  • 7 Hafizi S, Dahlback B. Gas6 and protein S. Vitamin K-dependent ligands for the Axl receptor tyrosine kinase subfamily. Febs J 2006; 273: 5231-5244.
  • 8 Goruppi S, Ruaro E, Schneider C. Gas6, the ligand of Axl tyrosine kinase receptor, has mitogenic and survival activities for serum starved NIH3T3 fibroblasts. On-cogene 1996; 12: 471-480.
  • 9 Fridell YW, Villa Jr. J, Attar EC. et al. GAS6 induces Axl-mediated chemotaxis of vascular smooth muscle cells. J Biol Chem 1998; 273: 7123-7126.
  • 10 Ishimoto Y, Ohashi K, Mizuno K. et al. Promotion of the uptake of PS liposomes and apoptotic cells by a product of growth arrest-specific gene, gas6. J Biochem 2000; 127: 411-417.
  • 11 O’Bryan JP, Fridell YW, Koski R. et al. The transforming receptor tyrosine kinase, Axl, is post-translationally regulated by proteolytic cleavage. J Biol Chem 1995; 270: 551-557.
  • 12 Nakano T, Kawamoto K, Higashino K. et al. Prevention of growth arrest-induced cell death of vascular smooth muscle cells by a product of growth arrest-specific gene, gas6. FEBS Lett 1996; 387: 78-80.
  • 13 Melaragno MG, Wuthrich DA, Poppa V. et al. Increased expression of Axl tyrosine kinase after vascular injury and regulation by G protein-coupled receptor agonists in rats. Circ Res 1998; 83: 697-704.
  • 14 Korshunov VA, Mohan AM, Georger MA. et al. Axl, a receptor tyrosine kinase, mediates flow-induced vascular remodeling. Circ Res 2006; 98: 1446-1452.
  • 15 Li Y, Gerbod-Giannone MC, Seitz H. et al. Cholesterol-induced apoptotic macrophages elicit an inflammatory response in phagocytes, which is partially attenuated by the Mer receptor. J Biol Chem 2006; 281: 6707-6717.
  • 16 Ait-Oufella H, Pouresmail V, Simon T. et al. Defective mer receptor tyrosine ki-nase signaling in bone marrow cells promotes apoptotic cell accumulation and accelerates atherosclerosis. Arterioscler Thromb Vasc Biol 2008; 28: 1429-1431.
  • 17 Thorp E, Cui D, Schrijvers DM. et al. Mertk receptor mutation reduces efferocytosis efficiency and promotes apoptotic cell accumulation and plaque necrosis in atherosclerotic lesions of apoe-/- mice. Arterioscler Thromb Vasc Biol 2008; 28: 1421-1428.
  • 18 Seimon T, Tabas I. Mechanisms and consequences of macrophage apoptosis in atherosclerosis. J Lipid Res 2009; 50 (Suppl) S382-387.
  • 19 Tjwa M, Bellido-Martin L, Lin Y. et al. Gas6 promotes inflammation by enhancing interactions between endothelial cells, platelets, and leukocytes. Blood 2008; 111: 4096-4105.
  • 20 Lutgens E, Tjwa M, Garcia de Frutos P. et al. Genetic loss of Gas6 induces plaque stability in experimental atherosclerosis. J Pathol 2008; 216: 55-63.
  • 21 Cavet ME, Smolock EM, Ozturk OH. et al. Gas6-axl receptor signaling is regulated by glucose in vascular smooth muscle cells. Arterioscler Thromb Vasc Biol 2008; 28: 886-891.
  • 22 Alciato F, Sainaghi PP, Sola D. et al. TNF-alpha, IL-6, and IL-1 expression is inhibited by GAS6 in monocytes/macrophages. J Leukoc Biol 2010; 87: 869-875.
  • 23 Murao K, Imachi H, Sayo Y. et al. A product of growth arrest-specific gene 6 modulates scavenger receptor expression in human vascular smooth muscle cells. FEBS Lett 1999; 459: 363-366.
  • 24 Avanzi GC, Gallicchio M, Bottarel F. et al. GAS6 inhibits granulocyte adhesion to endothelial cells. Blood 1998; 91: 2334-2340.
  • 25 Munoz X, Sumoy L, Ramirez-Lorca R. et al. Human vitamin K-dependent GAS6: gene structure, allelic variation, and association with stroke. Hum Mutat 2004; 23: 506-512.
  • 26 Hurtado B, Abasolo N, Munoz X. et al. Association study between polymorphims in GAS6-TAM genes and carotid atherosclerosis. Thromb Haemost 2010; 104: 592-598.
  • 27 Balogh I, Hafizi S, Stenhoff J. et al. Analysis of Gas6 in human platelets and plasma. Arterioscler Thromb Vasc Biol 2005; 25: 1280-1286.
  • 28 Clauser S, Bachelot-Lozat C, Fontana P. et al. Physiological plasma Gas6 levels do not influence platelet aggregation. Arterioscler Thromb Vasc Biol 2006; 26: e22.
  • 29 Cosemans JM, Van Kruchten R, Olieslagers S. et al. Potentiating role of Gas6 and Tyro3, Axl and Mer (TAM) receptors in human and murine platelet activation and thrombus stabilization. J Thromb Haemost 2010; 08: 1797-1808.
  • 30 Clauser S, Peyrard S, Gaussem P. et al. Development of a novel immunoassay for the assessment of plasma Gas6 concentrations and their variation with hormonal status. Clin Chem 2007; 53: 1808-1813.
  • 31 Saposnik B, Borgel D, Aiach M. et al. Functional properties of the sex-hormone-binding globulin (SHBG)-like domain of the anticoagulant protein S. Eur J Bio-chem 2003; 270: 545-555.
  • 32 Sambrook J, Fritsch EF, Maniatis T. Molecular Cloning: A Laboratory Manual. 2nd ed.. Cold Spring Harbor Laboratory Press, Cold Spring Harbor; 1989
  • 33 Bieche I, Parfait B, Le Doussal V. et al. Identification of CGA as a novel estrogen receptor-responsive gene in breast cancer: an outstanding candidate marker to predict the response to endocrine therapy. Cancer Res 2001; 61: 1652-1658.
  • 34 Stary HC, Chandler AB, Dinsmore RE. et al. A definition of advanced types of atherosclerotic lesions and a histological classification of atherosclerosis. A report from the Committee on Vascular Lesions of the Council on Arteriosclerosis, American Heart Association. Circulation 1995; 92: 1355-1374.
  • 35 Duran MC, Mas S, Martin-Ventura JL. et al. Proteomic analysis of human vessels: application to atherosclerotic plaques. Proteomics 2003; 03: 973-978.
  • 36 Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 1976; 72: 248-254.
  • 37 Tedgui A, Mallat Z. Cytokines in atherosclerosis: pathogenic and regulatory pathways. Physiol Rev 2006; 86: 515-581.
  • 38 Owens GK, Kumar MS, Wamhoff BR. Molecular regulation of vascular smooth muscle cell differentiation in development and disease. Physiol Rev 2004; 84: 767-801.
  • 39 Orr AW, Hastings NE, Blackman BR. et al. Complex regulation and function of the inflammatory smooth muscle cell phenotype in atherosclerosis. J Vasc Res 2010; 47: 168-180.
  • 40 Geng YJ, Libby P. Evidence for apoptosis in advanced human atheroma. Colocaliz-ation with interleukin-1 beta-converting enzyme. Am J Pathol 1995; 147: 251-266.
  • 41 Clarke MC, Figg N, Maguire JJ. et al. Apoptosis of vascular smooth muscle cells induces features of plaque vulnerability in atherosclerosis. Nat Med 2006; 12: 1075-1080.
  • 42 Angelillo -Scherrer A, Burnier L, Lambrechts D. et al. Role of Gas6 in erythropoie-sis and anemia in mice. J Clin Invest 2008; 118: 583-596.
  • 43 Grainger DJ. TGF-beta and atherosclerosis in man. Cardiovasc Res 2007; 74: 213-222.
  • 44 Gamble JR, Khew-Goodall Y, Vadas MA. Transforming growth factor-beta inhibits E-selectin expression on human endothelial cells. J Immunol 1993; 150: 4494-4503.
  • 45 Hurtado B, Munoz X, Recarte-Pelz P. et al. Expression of the vitamin K-depend-ent proteins GAS6 and protein S and the TAM receptor tyrosine kinases in human atherosclerotic carotid plaques. Thromb Haemost 2011; 105: 873-882.
  • 46 Meir KS, Leitersdorf E. Atherosclerosis in the apolipoprotein-E-deficient mouse: a decade of progress. Arterioscler Thromb Vasc Biol 2004; 24: 1006-1014.
  • 47 Miyoshi T, Tian J, Matsumoto AH. et al. Differential response of vascular smooth muscle cells to oxidized LDL in mouse strains with different atherosclerosis susceptibility. Atherosclerosis 2006; 189: 99-105.