SEARCH

SEARCH BY CITATION

Keywords:

  • coagulation;
  • microparticles;
  • P-selectin;
  • P-selectin glycoprotein ligand 1;
  • thrombosis;
  • tissue factor

Abstract

  1. Top of page
  2. Abstract
  3. P-selectin – link between inflammation and hemostasis
  4. Tissue factor in blood coagulation and beyond
  5. P-selectin/PSGL-1 binding induces procoagulant microparticle generation from leukocytes
  6. P-selectin and PSGL-1 in therapeutic strategies to modulate hemostasis
  7. Future directions
  8. Acknowledgements
  9. References

Summary.  The primary importance of tissue factor (TF) in blood coagulation and thrombus propagation has been recognized for many years. Nevertheless, our view about the origin of TF activity, necessary for normal hemostasis and found in pathologic conditions, needs to be revised in the light of recent observations. Pioneering work by Yale Nemerson's group showed that circulating TF on microparticles (MPs), could promote thrombus growth. The origin and characteristics of this ‘blood-borne’ TF are targets of intense research as well as intense debate. Surprising observations now implicate the adhesion receptor P-selectin (P-sel), known for its role in inflammation, in these MPs’ generation. P-sel, translocated from granules to the cell surfaces of activated platelets and endothelial cells, was recently found to play multiple roles in hemostasis. Expressed on endothelium, it can mediate platelet rolling. Signaling by P-sel through its receptor on leukocytes, P-selectin glycoprotein ligand 1 (PSGL-1), induces the generation of TF-positive, highly procoagulant MPs. In addition, P-sel on activated platelets helps to recruit these MPs specifically to thrombi. In this review, we discuss the roles of P-sel and TF-positive MPs and highlight strategies to modulate hemostasis by modulating the P-sel, TF, coagulation triad.


P-selectin – link between inflammation and hemostasis

  1. Top of page
  2. Abstract
  3. P-selectin – link between inflammation and hemostasis
  4. Tissue factor in blood coagulation and beyond
  5. P-selectin/PSGL-1 binding induces procoagulant microparticle generation from leukocytes
  6. P-selectin and PSGL-1 in therapeutic strategies to modulate hemostasis
  7. Future directions
  8. Acknowledgements
  9. References

P-selectin (P-sel) is a member of the selectin family of cell adhesion receptors, which mediate binding to specific carbohydrate-containing ligands [1]. Selectins mediate adhesion among leukocytes, platelets, and endothelium. P-sel, the largest member of the selectin family, is localized in the membranes of platelet α-granules [2] and in storage granules of endothelial cells called Weibel-Palade bodies [3,4] (Fig. 1). The major component of Weibel-Palade bodies is von Willebrand factor (VWF) [5]. VWF plays two main functions in hemostasis: it mediates platelet adhesion to the injured vessel wall, and it carries coagulation factor VIII. VWF directs the formation of the Weibel-Palade bodies and, in its absence, P-sel is mislocalized in the endothelial cell [6]. Weibel-Palade body exocytosis is triggered by secretagogues such as thrombin and histamine and occurs within seconds to minutes after stimulation. Similarly in platelets, upon activation, P-sel translocates with the α-granule membrane to the cell surface. Once present on the cell surface, the receptor can mediate cellular adhesion.

image

Figure 1. Electron micrographs of Weibel-Palade bodies (arrows) in sections of human umbilical vein endothelial cells. (A) Immunogold labeling for P-selectin. The distribution of the gold particles is close to the limiting membrane (arrowheads) of the organelle. (B) Immunogold labeling for von Willebrand factor (VWF). The gold particles (arrowheads) co-localize with the tubular structures present in the matrix of the Weibel-Palade body (bar: 0.1 μm). The photographs were generously provided by Dr Elisabeth Martin-Cramer, adapted from Ref. [70].

Download figure to PowerPoint

A key role of P-sel is in mediating leukocyte interactions in inflammation [2]. P-sel has long been known to support leukocyte rolling, as shown in vitro [7] and in vivo [8]. The most clearly defined ligand for P-sel is P-selectin glycoprotein ligand 1 (PSGL-1). It is a homodimeric mucin expressed on the majority of leukocytes [2], concentrated at the tips of microvilli, a location favorable for mediating leukocyte tethering and rolling (Fig. 2). PSGL-1 is a signaling receptor and dimeric or oligomeric forms of P-sel may be needed for cross-linking of PSGL-1 molecules on the leukocyte and subsequent signal transduction [9,10]. Indeed, it was found that the transmembrane form of P-sel on platelets and endothelial cells is dimeric [11,12].

image

Figure 2. Model of tissue factor (TF)- and P-selectin action in thrombus formation. (A) After vessel wall injury limited to a few cells or stimulation of endothelial cells (left), subendothelial TF or TF expressed on EC initiates clotting. Platelets adhering to the site of injury with deposited fibrin promptly seal vessel wall TF from the blood stream. TF activity required for thrombus growth and stabilization is provided from blood-borne TF (orange). When large-scale damage to the vessel wall occurs (right) a significant amount of TF of subendothelial origin, in the membrane of cell debris or microparticles (pink), enters the blood stream lessening the importance of blood-borne TF in thrombus propagation. (B) The P-selectin, tissue factor, coagulation triad. P-selectin (P) on activated platelets and endothelial cells and P-selectin shed from these cells (sP) binds to P-selectin glycoprotein ligand 1 (PSGL-1; v) on monocytes and this induces TF-positive MPs generation. P-selectin on the activated platelets in thrombi helps in the recruitment of these MPs to the thrombus, by binding to PSGL-1 on the MP. This ultimately leads to increased thrombin generation at the site of injury.

Download figure to PowerPoint

A soluble form of P-selectin (sP-sel) has been detected in human [13] and mouse [14] plasma. In humans, at least part of plasma P-sel is an alternatively spliced, secreted molecule that lacks the transmembrane domain, and at least part of the P-sel in mouse plasma results from proteolytic shedding of the extracellular domain [14,15]. Elevated levels of sP-sel are found in thrombotic consumptive disorders, such as disseminated intravascular coagulation, thrombotic thrombocytopenic purpura, and heparin-induced thrombocytopenia [16,17]. Furthermore, increased levels of sP-sel in the plasma have been documented in a variety of diseases, including coronary artery disease, diabetes, hypertension, and atrial fibrillation (reviewed in Ref. [18]). Moreover, elevated sP-sel might also serve as a predictive marker for future cardiovascular events, such as myocardial infarction and stroke [19].

Platelets, like leukocytes, were found to roll on activated endothelium in mesenteric venules and this process, dependent on shear rate, is supported by either VWF or P-sel [20,21]. Small amounts of PSGL-1 are expressed on platelets, and it can mediate P-sel-dependent platelet rolling [22]. The VWF receptor, glycoprotein (GP)Ibα, has also been shown to be a counter-receptor for P-sel [23] and may thus promote platelet rolling on P-sel as well. The striking parallel between platelet and leukocyte behavior indicates that both cell types need to slow down (roll) before they firmly adhere to the site of injury/inflammation. The fact that the adhesion molecules responsible for initial adhesion of both leukocytes and platelets are stored in the same organelle (Fig. 1) and are, therefore, always released together, shows how closely hemostatic and inflammatory responses are intertwined.

A role for P-sel in hemostasis and/or thrombosis became evident when P-sel-deficient mice were evaluated. The animals have a 40% prolongation in bleeding time, resulting from amputation of the tip of the tail, and a greater hemorrhage (extravascular red blood cells) in the Shwartzman-like reaction induced in the skin by lipopolysaccharide (LPS) and tumor necrosis factor (TNF)-α administration [24]. The contribution of P-sel to thrombus formation is indicated by several observations. P-sel blockade affected the stability of platelet aggregates in vitro, a phenomenon not seen with a PSGL-1 antagonist, supporting a role for GPIbα as a P-sel receptor on platelets [25]. P-sel-deficient thrombi formed in a flow chamber were thinner and taller [26]. Moreover, in an in vivo model of transluminal endothelial injury of the femoral artery, Smyth et al. reported that platelet deposits were less compact in P-sel-deficient mice than those observed in wild-type mice [27]. How platelet P-sel might contribute to platelet aggregate structure is not clear. P-sel might support platelet–platelet interactions either directly or indirectly by promoting fibrin deposition that is essential for thrombus stability [28]. The role of P-sel in fibrin formation will be discussed.

Tissue factor in blood coagulation and beyond

  1. Top of page
  2. Abstract
  3. P-selectin – link between inflammation and hemostasis
  4. Tissue factor in blood coagulation and beyond
  5. P-selectin/PSGL-1 binding induces procoagulant microparticle generation from leukocytes
  6. P-selectin and PSGL-1 in therapeutic strategies to modulate hemostasis
  7. Future directions
  8. Acknowledgements
  9. References

Research on tissue factor (TF) stretches back to 1834 when De Blaineville reported that i.v. injection of a suspension of brain tissue in a vein led to formation of clots and immediate death of the animal (history of TF is reviewed in Ref. [29]). TF, also known as thrombokinase, thromboplastin, CD142, or FIII of blood coagulation, is a 47 kDa membrane glycoprotein that consists of a large extracellular domain with two fibronectin type III modules joined by a hinge region, single transmembrane domain, and a short cytoplasmic tail [30–32]. The two intramolecular disulphide bonds in the extracellular domain are important for TF function in coagulation [30]. As a cofactor, TF binds coagulation FVII/FVIIa and enhances the catalytic activity of FVIIa more than one million-fold. TF is the principal initiator of the coagulation cascade and it is this function that makes its absence incompatible with life [33].

Tissue factor is highly expressed in brain, heart, kidneys, lung, uterus, and placenta [34,35]. Blood vessels are surrounded by cells constitutively expressing TF and the above organs exhibit defects in low TF mice [36]. When the integrity of the vascular system is disrupted, blood coagulation is immediately activated by TF now in direct contact with blood. Peripheral blood monocytes and endothelial cells can also express TF but only after stimulation. For many years it was believed that the low levels of TF antigen detectable in circulating blood represented inactive TF. However, work by Nemerson's group clearly demonstrated that circulating TF either is active or can become activated (‘blood-borne TF’) [37]. Human blood flowing over collagen-coated slides or pig arterial media not containing TF, promoted thrombi growth. Immunoelectron microscopy of the thrombi revealed the presence of TF-positive microparticles (MPs) [37]. In these studies, as well as in a rabbit venous thrombosis model in which fibrin deposition was quantified on collagen-coated threads inserted in the jugular vein [38], anti-TF antibodies inhibited thrombus formation.

What is ‘blood-borne’ TF and how does it become active when it is needed? It is likely that MPs originally derived from monocytes are the major carriers for blood-borne TF [39]. MPs are submicron vesicles that are released from cells after activation or during apoptosis [40]. MPs are procoagulant because of exposure of negatively charged phospholipids [mainly phosphatidyl serine (PS)] on their surface. A small portion of circulating blood MPs are highly procoagulant by expressing not only PS, but TF as well. It is not yet known whether TF on MPs is constitutively active or needs to be decrypted from an inactive form or location for binding FVII/FVIIa [41]. It is possible that TF becomes active upon MP adhesion to fibrin or activated platelets. A subpopulation of platelets was found to be positive for TF by electron microscopy [42] and transfer of leukocyte-derived TF-positive MPs to platelets was shown to be dependent on CD15 (epitope also found on PSGL-1), PSGL-1, P-sel, and TF [39,43]. Recently, Chou et al. demonstrated that hematopoietic cell-derived TF contributed to thrombus formation after laser injury of the microvessels in the cremaster muscle [44]. In contrast, Day et al. found that hematopoietic cell-derived TF did not promote thrombus formation in a carotid artery injury, induced by ferric chloride, in the same mouse model [45]. This discrepancy is probably caused by the significant differences between the injury inflicted, the later exposing large areas positive for TF in the vessel wall. It is likely that blood-borne TF plays a major role in the development and growth of thrombi in situations where the damage to the vessel is just to a few cells, as is likely the case in the laser injury [44] or venous thrombosis when thrombosis is induced by blood stasis with no or minimal damage to the vessel wall (Fig. 2A). In this model, both P- and E-selectins were found to play an important role in thrombus growth [46,47], likely in part by recruiting TF-positive MPs to the activated vessel wall. Once TF-expressing subendothelial cells or stimulated endothelial cells (EC) expressing TF are covered by a layer of platelets and fibrin, the plaque becomes impermeable to clotting factors [48]. In such situations, the TF requirement for thrombus growth and stabilization will rely on ‘blood-borne’ TF. When there is extensive damage, the significance of the original pool of circulating TF may be lessened by TF in the membrane of cell debris or microvesicles entering the blood at the site of injury (Fig. 2A).

The TF is the only clotting factor for which a congenital disorder has not been described. Low-TF mice (1% TF levels) are viable but they have a high incidence of fatal hemorrhages [33,36]. The procoagulant effect of TF is balanced by tissue factor pathway inhibitor (TFPI), a major anticoagulant protein in vertebrates [49]. TFPI gene disruption produces embryonic lethality [50], demonstrating the utmost importance of TF regulation. TF also functions in vascular development, angiogenesis, and tumor cell metastasis. The molecular mechanisms of TF functions in these processes are still poorly defined. Recent results showed a key role for TF cytoplasmic domain in angiogenesis by regulating signaling through protease-activated receptor-2 [51]. Recent data also revealed cross-talk of integrin α3β1 and TF in cell migration [52].

P-selectin/PSGL-1 binding induces procoagulant microparticle generation from leukocytes

  1. Top of page
  2. Abstract
  3. P-selectin – link between inflammation and hemostasis
  4. Tissue factor in blood coagulation and beyond
  5. P-selectin/PSGL-1 binding induces procoagulant microparticle generation from leukocytes
  6. P-selectin and PSGL-1 in therapeutic strategies to modulate hemostasis
  7. Future directions
  8. Acknowledgements
  9. References

P-selectin and PSGL-1 are no doubt key players in the recruitment of leukocytes in acute and chronic inflammation [1]. Recently, these molecules began to be considered important in hemostasis and thrombosis as well [53]. Early observations indicated that inhibition of P-sel influences fibrin deposition simultaneously with a reduction of leukocyte adhesion to platelets deposited on a thrombogenic graft in baboons [54]. Additional evidence indicating potential links between P-sel and coagulation came from mice with elevated levels of circulating sP-sel (ΔCT mice) [14]. In these mice, the wild-type gene of P-sel was replaced by one lacking the sequence encoding the cytoplasmic domain of P-sel. This P-sel was not correctly targeted to the Weibel-Palade bodies and was proteolytically shed from the plasma membrane into blood [14]. Surprisingly, these mice showed a shortening in plasma clotting time [55]. In addition, a propensity to deposit fibrin was demonstrated in the ΔCT mice: first, in the Shwartzman-like reaction where rapid fibrin deposition resulted in 50% less hemorrhage than seen in wild-type mice [55]; secondly, on platelet-rich thrombi in a collagen-coated capillary linked directly to vena cava [55]; and thirdly, in a model of deep vein thrombosis [46]. This is in contrast with P-sel-deficient animals that usually presented less fibrin formation than wild-type controls in these animal models [47,56]. These experiments lead to the conclusion that P-sel expression modulates fibrin deposition and the resulting thrombus size.

How can P-sel potentiate coagulation and ultimately fibrin formation? It appears that the primary reason is the role of P-sel in the formation and recruitment of leukocyte-derived MPs containing TF. The ΔCT mice, which shed high levels of sP-sel, have a large population of these MPs, which is not found in wild-type mice [55]. The depletion of these MPs from the plasma of the ΔCT mice prolongs its clotting time [55]. The TF-positive MPs were also positive for Mac-1 (αMβ2 integrin), demonstrating that they were derived from leukocytes. Because activated monocytes can express TF and P-sel was shown to induce TF synthesis in monocytes in vitro [57], it is most likely that the TF-positive MPs originated from monocytes. To further demonstrate the role of P-sel in this process, infusion of a P-sel–immunoglobulin chimera (P-sel–Ig) into wild-type mice was performed. Ultimately, this induced a similar procoagulant state to that seen in the ΔCT mice (Fig. 3) [55]. Thus, P-sel induces formation of procoagulant MPs some of which contain TF. In addition to TF, MPs carry procoagulant activity through the expression of prothrombinase activity on their membrane [58]. Leukocyte-derived MPs were also shown to induce TF expression on endothelial cells in vitro [59] and thus the procoagulant state induced by P-sel could be propagated further.

image

Figure 3. Effect of P-selectin–immunoglobulin chimera (P-sel–Ig) on hemorrhage and fibrin distribution at the Shwartzman reaction lesion site. Micrographs showing fibrin deposition (F) in paraffin section from the Shwartzman lesion site. Tissue sections from wild-type (WT) mice treated with control IgG1 or P-sel–Ig were immunostained with antibodies to fibrinogen (the brown reaction product). Note the presence of hemorrhage (H) in the section from the IgG1-treated animals, and a diffuse staining for fibrin inside and outside the vessel. A strong fibrin staining (F) is found on the luminal face of the vessel wall in the P-sel–Ig-treated mice, without detectable fibrin deposition in the surrounding tissue. White arrowheads point to the vessel wall (bar: 40 μm). Reproduced with permission from Andre et al. [55].

Download figure to PowerPoint

Recently, PSGL-1 was found to play a crucial role in TF-containing MP formation (Fig. 2B). The formation of procoagulant activity induced by P-sel–Ig in human blood was fully inhibited by an antibody to PSGL-1 [60] and the generation of procoagulant MPs by the infusion of P-sel–Ig in mice was not observed in PSGL-1-deficient animals [60]. Thus, the P-sel–PSGL-1 interactions could be particularly important after vessel injury (Fig. 2). The larger the injury, the more granular exocytosis of P-sel will occur from both platelets and endothelium and the more TF-containing MPs will be produced to help generate thrombin and fibrin. The specific recruitment of the MPs to the platelet plug [39,60] serves to prevent disseminated vascular coagulation while stabilizing the platelet plug at the site of injury.

P-selectin and PSGL-1 in therapeutic strategies to modulate hemostasis

  1. Top of page
  2. Abstract
  3. P-selectin – link between inflammation and hemostasis
  4. Tissue factor in blood coagulation and beyond
  5. P-selectin/PSGL-1 binding induces procoagulant microparticle generation from leukocytes
  6. P-selectin and PSGL-1 in therapeutic strategies to modulate hemostasis
  7. Future directions
  8. Acknowledgements
  9. References

Abnormalities in hemostasis, which push the delicate balance either to bleeding or thrombosis, are life-threatening conditions. Based on the prominent role of TF in initiating blood coagulation as well as in thrombus propagation, it is logical to assume that interfering with the TF pathway would significantly down-regulate blood clotting potential while in contrast stimulating the pathway would promote blood clotting and reduce bleeding. Approaches directly targeting TF involve administration of anti-TF antibodies, recombinant TFPI, or inhibitory synthetic small molecules (reviewed in Ref. [32]). The interaction between P-sel and PSGL-1 represents an important newly uncovered mechanism by which TF-positive procoagulant MPs are produced and recruited to thrombi (Fig. 2B). Accordingly, antithrombotic strategies are currently aimed at inhibiting this interaction (reviewed in Ref. [53]) while recent results also suggest that the procoagulant potential of P-sel could be used to treat bleeding disorders such as hemophilia [60].

Infusion of anti-P-sel antibodies, recombinant PSGL-1-immunoglobulin chimera (rPSGL–Ig), or small molecule inhibitors of P-sel binding to PSGL-1, has shown beneficial effects in various experimental models of arterial and deep vein thrombosis, thrombotic glomerulonephritis, and stroke [61–65]. Several studies also document that P-sel inhibitors promote thrombolysis of preformed thrombi and prevent vascular reocclusion [61,63,65]. Treatments with rPSGL–Ig probably reduce both MP generation and recruitment, and therefore limit the extent of new fibrin deposition in thrombi. One illustration of this effect is provided by the P-sel ΔCT mice, in which the procoagulant MP count is reduced and clotting time is extended following rPSGL–Ig treatment [46,55]. On the contrary, infusion of P-sel–Ig in FVIII-deficient mice, a model for hemophilia A, shortened their bleeding time and with increasing time postinfusion corrected the clotting parameters of their blood [60] (Table 1). These results show the viability of strategies based on the procoagulant effect of P-sel–Ig by inducing the production of TF-positive MPs.

Table 1.  P-selectin–immunoglobulin (P-sel–Ig) treatment of factor VIII-deficient mice
F8−/− mice infused with6 h72 h
Clotting time (T; min)*Clotting rate (R)Clotting time (T; min)**Clotting rate (R)**
  1. P-sel–Ig treatment of hemophilia A mice accelerates the onset of blood clotting and increases the rate of clotting. In mice treated with control immunoglobulin (control Ig), T and R (% peak amplitude min−1) were not significantly different from that of untreated hemophilia A mice (data not shown). For comparison, the values in wild-type mice were T = 3.1 ± 0.3 min and R = 26.2 ± 5.1. Although T in the mice treated with P-sel–Ig is still slightly higher than the wild-type value, the difference in R was not significant (n = 4 in each group).

  2. *P < 0.02, **P < 0.001 (adapted from Ref. [60]).

P-sel–Ig8.5 ± 1.14.4 ± 3.34.5 ± 0.518.0 ± 2.8
Control Ig19.2 ± 2.50.4 ± 0.017.9 ± 2.00.6 ± 0.1

Because of the prominent role of the TF pathway in blood clotting, the emerging success story of recombinant FVIIa (rFVIIa) as a ‘general hemostatic agent’ in a wide variety of conditions with bleeding tendency [66] is not surprising. In vitro data raise the question whether the pharmacologic doses of rFVIIa currently used for therapy in humans may directly activate FIX and FX on the surface of activated platelets [67]. Nevertheless, it is likely that direct activation by FVIIa is marginal in vivo and the TF/FVIIa complex is the major constituent in the action mechanism during rFVIIa therapy. It is important to note that rFVIIa is not a magic bullet as it is ineffective or only partially effective in more than 10% of hemophilia patients with inhibitors [68,69]. It is possible that insufficiently low levels of circulating TF or the short half-life of infused FVIIa are behind the ineffectiveness of rFVIIa in these cases. Using P-sel–Ig to boost the level of circulating TF-positive MPs, either alone or in combination with rFVIIa, is a promising new strategy for controlling bleeding in hemophilia.

Future directions

  1. Top of page
  2. Abstract
  3. P-selectin – link between inflammation and hemostasis
  4. Tissue factor in blood coagulation and beyond
  5. P-selectin/PSGL-1 binding induces procoagulant microparticle generation from leukocytes
  6. P-selectin and PSGL-1 in therapeutic strategies to modulate hemostasis
  7. Future directions
  8. Acknowledgements
  9. References

Despite the impressive progress in this field during the last few years, many questions remain open. In our view, some of the most exiting are the following.

  • 1
    How is P-selectin expression (Weibel-Palade bodies and platelet α-granule secretion) regulated?
  • 2
    What induces P-sel shedding and which is the enzyme(s) responsible?
  • 3
    Which cellular and sP-sel can induce TF expression and generation of TF-positive MPs from monocytes?
  • 4
    What is the signaling pathway from PSGL-1 engagement to TF-positive MPs generation?
  • 5
    Which receptors on monocytes could act in synergy with PSGL-1 to produce MPs?
  • 6
    It is assumed that TF-positive MPs are shed from the plasma membrane, and are not exosomes secreted by the cells – is it really so?
  • 7
    Do the other two selectins have similar procoagulant activity because they can both bind PSGL-1?
  • 8
    Which receptors on MPs and on platelets produce stable adhesion between the two entities? Do MPs bind to fibrin?
  • 9
    Do other cells (endothelial cells, smooth muscle cells, adventitial fibroblasts) contribute to the blood-borne TF pool and in which situations?
  • 10
    Can platelets take up TF-positive MPs and thus form a reservoir of TF activity to be used later after subsequent platelet activation?
  • 11
    Is TF on MPs latent/encrypted or is it active, ready to bind and stimulate FVII/FVIIa? If it is encrypted, what initiates its decryption?

One can try to predict the answers to these questions but only one prediction is accurate: frequent updates will be required on the ‘P-selectin, tissue factor, coagulation triad’ for some time to come.

Acknowledgements

  1. Top of page
  2. Abstract
  3. P-selectin – link between inflammation and hemostasis
  4. Tissue factor in blood coagulation and beyond
  5. P-selectin/PSGL-1 binding induces procoagulant microparticle generation from leukocytes
  6. P-selectin and PSGL-1 in therapeutic strategies to modulate hemostasis
  7. Future directions
  8. Acknowledgements
  9. References

The authors thank Lesley Cowan for help preparing the manuscript and Jaime Lazarte for artwork. Our laboratory's work was supported by NIH NHLBI grants R37 HL41002, P01 HL56949, and R01 HL53756 (to D.D.W.).

References

  1. Top of page
  2. Abstract
  3. P-selectin – link between inflammation and hemostasis
  4. Tissue factor in blood coagulation and beyond
  5. P-selectin/PSGL-1 binding induces procoagulant microparticle generation from leukocytes
  6. P-selectin and PSGL-1 in therapeutic strategies to modulate hemostasis
  7. Future directions
  8. Acknowledgements
  9. References
  • 1
    Ley K. The role of selectins in inflammation and disease. Trends Mol Med 2003; 9: 2638.
  • 2
    McEver RP. Adhesive interactions of leukocytes, platelets, and the vessel wall during hemostasis and inflammation. Thromb Haemost 2001; 86: 74656.
  • 3
    Bonfanti R, Furie BC, Furie B, Wagner DD. PADGEM (GMP140) is a component of Weibel-Palade bodies of human endothelial cells. Blood 1989; 73: 110912.
  • 4
    McEver RP, Beckstead JH, Moore KL, Marshall-Carlson L, Bainton DF. GMP-140, a platelet alpha-granule membrane protein, is also synthesized by vascular endothelial cells and is localized in Weibel-Palade bodies. J Clin Invest 1989; 84: 929.
  • 5
    Wagner DD, Olmsted JB, Marder VJ. Immunolocalization of von Willebrand protein in Weibel-Palade bodies of human endothelial cells. J Cell Biol 1982; 95: 35560.
  • 6
    Denis CV, Andre P, Saffaripour S, Wagner DD. Defect in regulated secretion of P-selectin affects leukocyte recruitment in von Willebrand factor-deficient mice. Proc Natl Acad Sci U S A 2001; 98: 40727.
  • 7
    Lawrence MB, Springer TA. Leukocytes roll on a selectin at physiologic flow rates: distinction from and prerequisite for adhesion through integrins. Cell 1991; 65: 85973.
  • 8
    Mayadas TN, Johnson RC, Rayburn H, Hynes RO, Wagner DD. Leukocyte rolling and extravasation are severely compromised in P selectin-deficient mice. Cell 1993; 74: 54154.
  • 9
    Hidari KI, Weyrich AS, Zimmerman GA, McEver RP. Engagement of P-selectin glycoprotein ligand-1 enhances tyrosine phosphorylation and activates mitogen-activated protein kinases in human neutrophils. J Biol Chem 1997; 272: 287506.
  • 10
    Piccardoni P, Sideri R, Manarini S, Piccoli A, Martelli N, de Gaetano G, Cerletti C, Evangelista V. Platelet/polymorphonuclear leukocyte adhesion: a new role for SRC kinases in Mac-1 adhesive function triggered by P-selectin. Blood 2001; 98: 10816.
  • 11
    Ushiyama S, Laue TM, Moore KL, Erickson HP, McEver RP. Structural and functional characterization of monomeric soluble P-selectin and comparison with membrane P-selectin. J Biol Chem 1993; 268: 1522937.
  • 12
    Barkalow FJ, Barkalow KL, Mayadas TN. Dimerization of P-selectin in platelets and endothelial cells. Blood 2000; 96: 30707.
  • 13
    Dunlop LC, Skinner MP, Bendall LJ, Favaloro EJ, Castaldi PA, Gorman JJ, Gamble JR, Vadas MA, Berndt MC. Characterization of GMP-140 (P-selectin) as a circulating plasma protein. J Exp Med 1992; 175: 114750.
  • 14
    Hartwell DW, Mayadas TN, Berger G, Frenette PS, Rayburn H, Hynes RO, Wagner DD. Role of P-selectin cytoplasmic domain in granular targeting in vivo and in early inflammatory responses. J Cell Biol 1998; 143: 112941.
  • 15
    Ishiwata N, Takio K, Katayama M, Watanabe K, Titani K, Ikeda Y, Handa M. Alternatively spliced isoform of P-selectin is present in vivo as a soluble molecule. J Biol Chem 1994; 269: 2370815.
  • 16
    Chong BH, Murray B, Berndt MC, Dunlop LC, Brighton T, Chesterman CN. Plasma P-selectin is increased in thrombotic consumptive platelet disorders. Blood 1994; 83: 153541.
  • 17
    Katayama M, Handa M, Araki Y, Ambo H, Kawai Y, Watanabe K, Ikeda Y. Soluble P-selectin is present in normal circulation and its plasma level is elevated in patients with thrombotic thrombocytopenic purpura and haemolytic uraemic syndrome. Br J Haematol 1993; 84: 70210.
  • 18
    Kappelmayer J, Nagy B Jr, Miszti-Blasius K, Hevessy Z, Setiadi H. The emerging value of P-selectin as a disease marker. Clin Chem Lab Med 2004; 42: 47586.
  • 19
    Ridker PM, Buring JE, Rifai N. Soluble P-selectin and the risk of future cardiovascular events. Circulation 2001; 103: 4915.
  • 20
    Frenette PS, Johnson RC, Hynes RO, Wagner DD. Platelets roll on stimulated endothelium in vivo: an interaction mediated by endothelial P-selectin. Proc Natl Acad Sci U S A 1995; 92: 74504.
  • 21
    Andre P, Denis CV, Ware J, Saffaripour S, Hynes RO, Ruggeri ZM, Wagner DD. Platelets adhere to and translocate on von Willebrand factor presented by endothelium in stimulated veins. Blood 2000; 96: 33228.
  • 22
    Frenette PS, Denis CV, Weiss L, Jurk K, Subbarao S, Kehrel B, Hartwig JH, Vestweber D, Wagner DD. P-Selectin glycoprotein ligand 1 (PSGL-1) is expressed on platelets and can mediate platelet-endothelial interactions in vivo. J Exp Med 2000; 191: 141322.
  • 23
    Romo GM, Dong JF, Schade AJ, Gardiner EE, Kansas GS, Li CQ, McIntire LV, Berndt MC, Lopez JA. The glycoprotein Ib-IX-V complex is a platelet counter-receptor for P-selectin. J Exp Med 1999; 190: 80314.
  • 24
    Subramaniam M, Frenette PS, Saffaripour S, Johnson RC, Hynes RO, Wagner DD. Defects in hemostasis in P-selectin-deficient mice. Blood 1996; 4: 123842.
  • 25
    Merten M, Thiagarajan P. P-selectin expression on platelets determines size and stability of platelet aggregates. Circulation 2000; 102: 19316.
  • 26
    Ruggeri ZM, Subramaniam M, Dent JA, Wagner DD, Saldivar E. P-selectin and the three-dimensional structure of platelet thrombi. Blood 2000; 96: 812a.
  • 27
    Smyth SS, Reis ED, Zhang W, Fallon JT, Gordon RE, Coller BS. Beta(3)-integrin-deficient mice but not P-selectin-deficient mice develop intimal hyperplasia after vascular injury: correlation with leukocyte recruitment to adherent platelets 1 hour after injury. Circulation 2001; 103: 25017.
  • 28
    Ni H, Denis CV, Subbarao S, Degen JL, Sato TN, Hynes RO, Wagner DD. Persistence of platelet thrombus formation in arterioles of mice lacking both von Willebrand factor and fibrinogen. J Clin Invest 2000; 106: 38592.
  • 29
    Bachli E. History of tissue factor. Br J Haematol 2000; 110: 24855.
  • 30
    Ruf W, Edgington TS. Structural biology of tissue factor, the initiator of thrombogenesis in vivo. FASEB J 1994; 8: 38590.
  • 31
    Konigsberg W, Kirchhofer D, Riederer MA, Nemerson Y. The TF:VIIa complex: clinical significance, structure-function relationships and its role in signaling and metastasis. Thromb Haemost 2001; 86: 75771.
  • 32
    Eilertsen KE, Osterud B. Tissue factor: (patho)physiology and cellular biology. Blood Coagul Fibrinolysis 2004; 15: 52138.
  • 33
    Mackman N. Role of tissue factor in hemostasis, thrombosis, and vascular development. Arterioscler Thromb Vasc Biol 2004; 24: 101522.
  • 34
    Drake TA, Morrissey JH, Edgington TS. Selective cellular expression of tissue factor in human tissues. Implications for disorders of hemostasis and thrombosis. Am J Pathol 1989; 134: 108797.
  • 35
    Fleck RA, Rao LV, Rapaport SI, Varki N. Localization of human tissue factor antigen by immunostaining with monospecific, polyclonal anti-human tissue factor antibody. Thromb Res 1990; 59: 42137.
  • 36
    Pawlinski R, Pedersen B, Erlich J, Mackman N. Role of tissue factor in haemostasis, thrombosis, angiogenesis and inflammation: lessons from low tissue factor mice. Thromb Haemost 2004; 92: 44450.
  • 37
    Giesen PL, Rauch U, Bohrmann B, Kling D, Roque M, Fallon JT, Badimon JJ, Himber J, Riederer MA, Nemerson Y. Blood-borne tissue factor: another view of thrombosis. Proc Natl Acad Sci U S A 1999; 96: 23115.
  • 38
    Himber J, Wohlgensinger C, Roux S, Damico LA, Fallon JT, Kirchhofer D, Nemerson Y, Riederer MA. Inhibition of tissue factor limits the growth of venous thrombus in the rabbit. J Thromb Haemost 2003; 1: 88995.
  • 39
    Falati S, Liu Q, Gross P, Merrill-Skoloff G, Chou J, Vandendries E, Celi A, Croce K, Furie BC, Furie B. Accumulation of tissue factor into developing thrombi in vivo is dependent upon microparticle P-selectin glycoprotein ligand 1 and platelet P-selectin. J Exp Med 2003; 197: 158598.
  • 40
    Morel O, Toti F, Hugel B, Freyssinet JM. Cellular microparticles: a disseminated storage pool of bioactive vascular effectors. Curr Opin Hematol 2004; 11: 15664.
  • 41
    Maynard JR, Heckman CA, Pitlick FA, Nemerson Y. Association of tissue factor activity with the surface of cultured cells. J Clin Invest 1975; 55: 81424.
  • 42
    Muller I, Klocke A, Alex M, Kotzsch M, Luther T, Morgenstern E, Zieseniss S, Zahler S, Preissner K, Engelmann B. Intravascular tissue factor initiates coagulation via circulating microvesicles and platelets. FASEB J 2003; 17: 4768.
  • 43
    Rauch U, Bonderman D, Bohrmann B, Badimon JJ, Himber J, Riederer MA, Nemerson Y. Transfer of tissue factor from leukocytes to platelets is mediated by CD15 and tissue factor. Blood 2000; 96: 1705.
  • 44
    Chou J, Mackman N, Merrill-Skoloff G, Pedersen B, Furie BC, Furie B. Hematopoietic cell-derived microparticle tissue factor contributes to fibrin formation during thrombus propagation. Blood 2004; 104: 31907.
  • 45
    Day SM, Reeve JL, Pedersen B, Farris D, Myers DD, Im M, Wakefield TW, Mackman N, Fay WP. Macrovascular thrombosis is driven by tissue factor derived primarily from the blood vessel wall. Blood 2005; 105: 19298.
  • 46
    Myers DD, Hawley AE, Farris DM, Wrobleski SK, Thanaporn P, Schaub RG, Wagner DD, Kumar A, Wakefield TW. P-selectin and leukocyte microparticles are associated with venous thrombogenesis. J Vasc Surg 2003; 38: 107589.
  • 47
    Sullivan VV, Hawley AE, Farris DM, Knipp BS, Varga AJ, Wrobleski SK, Thanaporn P, Eagleton MJ, Myers DD, Fowlkes JB, Wakefield TW. Decrease in fibrin content of venous thrombi in selectin-deficient mice. J Surg Res 2003; 109: 17.
  • 48
    Hathcock JJ, Nemerson Y. Platelet deposition inhibits tissue factor activity: in vitro clots are impermeable to factor Xa. Blood 2004; 104: 1237.
  • 49
    Broze GJ Jr. Tissue factor pathway inhibitor and the revised theory of coagulation. Annu Rev Med 1995; 46: 10312.
  • 50
    Huang ZF, Higuchi D, Lasky N, Broze GJ Jr. Tissue factor pathway inhibitor gene disruption produces intrauterine lethality in mice. Blood 1997; 90: 94451.
  • 51
    Belting M, Dorrell MI, Sandgren S, Aguilar E, Ahamed J, Dorfleutner A, Carmeliet P, Mueller BM, Friedlander M, Ruf W. Regulation of angiogenesis by tissue factor cytoplasmic domain signaling. Nat Med 2004; 10: 5029.
  • 52
    Dorfleutner A, Hintermann E, Tarui T, Takada Y, Ruf W. Cross-talk of integrin alpha3beta1 and tissue factor in cell migration. Mol Biol Cell 2004; 15: 441625.
  • 53
    Cambien B, Wagner DD. A new role in hemostasis for the adhesion receptor P-selectin. Trends Mol Med 2004; 10: 17986.
  • 54
    Palabrica T, Lobb R, Furie BC, Aronovitz M, Benjamin C, Hsu YM, Sajer SA, Furie B. Leukocyte accumulation promoting fibrin deposition is mediated in vivo by P-selectin on adherent platelets. Nature 1992; 359: 84851.
  • 55
    Andre P, Hartwell D, Hrachovinova I, Saffaripour S, Wagner DD. Pro-coagulant state resulting from high levels of soluble P-selectin in blood. Proc Natl Acad Sci U S A 2000; 97: 1383540.
  • 56
    Subramaniam M, Frenette PS, Saffaripour S, Johnson RC, Hynes RO, Wagner DD. Defects in hemostasis in P-selectin-deficient mice. Blood 1996; 87: 123842.
  • 57
    Celi A, Pellegrini G, Lorenzet R, De Blasi A, Ready N, Furie BC, Furie B. P-selectin induces the expression of tissue factor on monocytes. Proc Natl Acad Sci U S A 1994; 91: 876771.
  • 58
    Satta N, Toti F, Feugeas O, Bohbot A, Dachary-Prigent J, Eschwege V, Hedman H, Freyssinet JM. Monocyte vesiculation is a possible mechanism for dissemination of membrane-associated procoagulant activities and adhesion molecules after stimulation by lipopolysaccharide. J Immunol 1994; 153: 324555.
  • 59
    Mesri M, Altieri DC. Leukocyte microparticles stimulate endothelial cell cytokine release and tissue factor induction in a JNK1 signaling pathway. J Biol Chem 1999; 274: 231118.
  • 60
    Hrachovinova I, Cambien B, Hafezi-Moghadam A, Kappelmayer J, Camphausen RT, Widom A, Xia L, Kazazian HH Jr, Schaub RG, McEver RP, Wagner DD. Interaction of P-selectin and PSGL-1 generates microparticles that correct hemostasis in a mouse model of hemophilia A. Nat Med 2003; 9: 10205.
  • 61
    Toombs CF, DeGraaf GL, Martin JP, Geng JG, Anderson DC, Shebuski RJ. Pretreatment with a blocking monoclonal antibody to P-selectin accelerates pharmacological thrombolysis in a primate model of arterial thrombosis. J Pharmacol Exp Ther 1995; 275: 9419.
  • 62
    Connolly ES Jr, Winfree CJ, Prestigiacomo CJ, Kim SC, Choudhri TF, Hoh BL, Naka Y, Solomon RA, Pinsky DJ. Exacerbation of cerebral injury in mice that express the P-selectin gene: identification of P-selectin blockade as a new target for the treatment of stroke. Circ Res 1997; 81: 30410.
  • 63
    Kumar A, Villani MP, Patel UK, Keith JC Jr, Schaub RG. Recombinant soluble form of PSGL-1 accelerates thrombolysis and prevents reocclusion in a porcine model. Circulation 1999; 99: 13639.
  • 64
    Ito I, Yuzawa Y, Mizuno M, Nishikawa K, Tashita A, Jomori T, Hotta N, Matsuo S. Effects of a new synthetic selectin blocker in an acute rat thrombotic glomerulonephritis. Am J Kidney Dis 2001; 38: 26573.
  • 65
    Myers D, Wrobleski S, Londy F, Fex B, Hawley A, Schaub R, Greenfield L, Wakefield T. New and effective treatment of experimentally induced venous thrombosis with anti-inflammatory rPSGL-Ig. Thromb Haemost 2002; 87: 37482.
  • 66
    Uhlmann EJ, Eby CS. Recombinant activated factor VII for non-hemophiliac bleeding patients. Curr Opin Hematol 2004; 11: 198204.
  • 67
    Hoffman M, Monroe DM III. The action of high-dose factor VIIa (FVIIa) in a cell-based model of hemostasis. Semin Hematol 2001; 38: 69.
  • 68
    Negrier C, Hay CR. The treatment of bleeding in hemophilic patients with inhibitors with recombinant factor VIIa. Semin Thromb Hemost 2000; 26: 40712.
  • 69
    Arkin F, Blei F, Fetten J, Foulke R, Gilchrist GS, Heisel MA, Key N, Kisker CT, Kitchen C, Shafer FE, Shah PC, Strickland D. Human coagulation factor FVIIa (recombinant) in the management of limb-threatening bleeds unresponsive to alternative therapies: results from the NovoSeven emergency-use programme in patients with severe haemophilia or with acquired inhibitors. Blood Coagul Fibrinolysis 2000; 11: 25559.
  • 70
    Wagner DD. The Weibel-Palade body: the storage granule for von Willebrand factor and P-selectin. Thromb Haemost 1993; 70: 10510.