Microparticles- Signaling in Atherothrombosis Agneta Siegbahn, MD, PhD, FESC Professor in Coagulation Science

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1 Microparticles- Signaling in Atherothrombosis Agneta Siegbahn, MD, PhD, FESC Professor in Coagulation Science Department of Medical Sciences and Uppsala Clinical Research Center, Uppsala University Uppsala, Sweden

2 No disclosure Disclosure

3 Microparticles (MPs) in hemostasis MP MP MP 0.5 µm MPs are vesicles that bud off from cells, lack a nucleus, contain a membrane skeleton; considered as ectosomes Originate from platelets, leukocytes, erythrocytes,endothelial cells and SMC MP formation is a physiological phenomenon Harbor a large repertoire of cell surface receptors, proteins, mrna and micrornas Vectors for exchange of biological signals and information between cells Elevated levels of MPs in atherosclerosis, thrombosis, inflammation, metabolic diseases and malignancies Image adapted from Tilley et al. Thromb Res 2008; 122: 604-9

4 Leroyer AS. et al, J Am Coll Cardiol Feb 20;49(7):772-7 AnnexinV-Positive Microparticles in Atherosclerotic Plaques

5 Leroyer AS. et al, J Am Coll Cardiol Feb 20;49(7):772-7 Cellular Origin of Human Plaque Microparticles

6 Thrombin Generation by Plasma, Plaque and Cell-Derived Microparticles Leroyer AS. et al, J Am Coll Cardiol Feb 20;49(7):772-7

7 Rautou PE. et al, Circ Res Feb 4;108(3): ICAM-1 levels on Endothelial Cells are Increased by Plaque MPs

8 ICAM-1 Transferred to Endothelial Cells by Plaque MPs is Functionally Active Rautou PE. et al, Circ Res Feb 4;108(3):335-43

9 ICAM-1 expressing HCAEC induce Tissue Factor in Human Monocytes Lindmark E. & Siegbahn A., Thromb Res Oct 1;108(1):77-84.

10 Effect of EMPs on TF protein level in THP-1 cells Sabatier F. et al, Blood Jun 1;99(11):

11 Role of adhesion molecules in THP-1 cell procoagulant activity induced by EMPs Sabatier F. et al, Blood Jun 1;99(11):

12 Platelet derived MPs in whole blood Christersson, Johnell, Siegbahn; Submitted 2012

13 P-selectin expressing MPs in whole blood Christersson, Johnell, Siegbahn Submitted 2012

14 Membrane and Cytoplasmic Proteins Harbored by Platelet-Derived Microparticles Receptors; GP1a, GP1b, GPIIb/IIIa, vwf, chemokine receptors (CXCR4), P-selectin, CD40L and tissue factor (TF) Intraparticular proteins; RANTES/CCL5, VEGF, PDGF, bfgf, PPAR-g, fibrinogen, Caspase 3

15 Production of tissue factor and IL8 in monocytes in TRAP-stimulated whole blood Christersson et al. J Thromb Haemost (6)

16 Phosphorylation of the Scr-family member Lyn in monocytes fromtrap-stimulated whole blood Christersson et al..j Thromb Haemost (6)

17 COAGULATION Fibrinogen receptor PDMP GPIIb- IIIa Laminin receptor TF CD40L P-selectin Cytokines FVIIa TFPI PSGL CD40 PSGL TF EPCR ICAM-1 TM PAR2 MDMP SIGNALING

18 TF/FVIIa induces cytokine production in human monocytes * * LPS 3h LPS 3h + FVIIa 3h Johnell &Siegbahn, Acta Universitas Uppsaliensis, 2003

19 TF/FVIIa induce hyperchemotaxis in monocytes towards PDGF-BB Potentiation also in fibroblasts, VSMC, PAEC, malignant cells Selective towards PDGF-BB; not to 5 other chemotactic factors Dependent on the cytoplasmic domain Src-family, PLC, and PAR-2 dependent TF/FVIIa induces transactivation of the PDGFRβ PDGF-BB SU6656 = Src-family inhibitor SLIGKV = PAR-2 agonist Siegbahn, A. et al. Blood 2000;96:10. Siegbahn, A. et al Thrombosis and Haemostasis 2005;93:1. Siegbahn, A. et al. ATVB 2008;28:135.

20 FVIIa FVIIa PAR 2 PDGFRb TF TF P P ABP- 280 Src family P P FAK activation Signal transduction [Ca 2+ ] MAPK PI3-kinase / AKT EC adhesion and migration Gene expression VEGF, upar, CCN 1&2 IL-8, Egr-1, MMPs Apoptosis inhibition Potentiation of cell migration Angiogenesis, Cell migration, Cell survival Arteriosclerosis, Thrombosis, Tumor proliferation and Metastasis

21 Accumulation of TF into Developing Thrombi is Dependent upon PSGL-1 and Platelet P-selectin Falati S. et al, J Exp Med Jun 2;197(11):

22 Accumulation of monocyte-derived MPs expressing TF and PSGL-1 in thrombus is dependent on P-selectin expression Falati S. et al, J Exp Med Jun 2;197(11):

23 Conclusions MPs released into the bloodstream act as messengers delivering biologically active molecules to distal cells contributing to inflammation and thrombosis. Binding of MPs to monocytes induces the expression of proinflammatory and procoagulant molecules. Elevated numbers of monocyte-derived MP in blood may trigger thrombosis. TF and P-selectin/PSGL-1 positive MPs may prove to be a useful biomarker to identify patients at risk of thrombosis. Targeted inhibition of the generation and release of MPs may represent a novel therapeutic strategy for treatment of thrombotic diseases.

24 Research Group of Coagulation and Inflammation Science, Uppsala University Christina Christersson Mikael Åberg Oskar Eriksson Dariush Mokhtari Jenny Alfredsson Lena Kask Åsa Thulin Helena Vretman Former fellows Matilda Johnell Eva Lindmark Ristoff Teet Velling Anders Mälarstig