Inflammation and thrombosis


  • C. T. Esmon

    1. Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation; Departments of Pathology, and Biochemistry & Molecular Biology, University of Oklahoma Health Sciences Center; and Howard Hughes Medical Institute, Oklahoma City, Oklahoma, USA
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Dr Charles T. Esmon, Oklahoma Medical Research Foundation, Cardiovascular Biology Research Program, 825 NE 13th Street, Oklahoma City, OK 73104, USA.
Tel.: +405 271 6474; fax: +405 271 3137; e-mail:


Summary.  Systemic inflammation is a potent prothrombotic stimulus. Inflammatory mechanisms upregulate procoagulant factors, downregulate natural anticoagulants and inhibit fibrinolytic activity. In addition to modulating plasma coagulation mechanisms, inflammatory mediators appear to increase platelet reactivity. In vivo, however, natural anticoagulants not only prevent thrombosis, but they also dampen inflammatory activity. Some insights into the evolution and linkages between inflammatory mechanisms and the coagulation/anticoagulation mechanisms have become evident from recent structural studies. This review will summarize the interactions between inflammation and coagulation.

The impact of inflammation on procoagulant reactions

A simplified blood coagulation scheme with multiple control mechanisms is shown in Fig. 1. Tissue factor is normally present in the circulation at very low levels [1]. An increase in tissue factor expression caused by inflammation tends to shift the hemostatic balance in favor of coagulation/thrombosis. Normal hemostasis is initiated when the blood vessels are breached allowing blood to contact extravascular cells bearing tissue factor [2]. Inflammatory mediators promote coagulation by elevating tissue factor. Endotoxin, tumor necrosis factor α (TNFα) and interleukin-1α (IL-1α) induce tissue factor expression, primarily on monocytes/macrophages [3,4], and probably play a role in inducing tissue factor in atherosclerotic plaques as well [5]. As denoted in Fig. 1, all of the reactions involved in coagulation initiation and amplification require a membrane surface. This surface needs to contain negatively charged phospholipids such as phosphatidylserine. Normally, these negatively charged lipids are not present on the cell surface in sufficient concentrations to support effective initiation or propagation of the coagulation responses, but require potent cell agonists such as a combination of collagen and thrombin [6] or the complement C5b9 complex to optimally express the procoagulant lipid surface [7]. In vivo, the limiting nature of the accessible lipid surface is readily demonstrated by the observation that factor (F)Xa infusion alone is not very thrombogenic unless negatively charged phospholipid vesicles are coinfused [8]. Inflammation will also elevate fibrinogen synthesis. Fibrinogen levels rise under these conditions unless a consumptive coagulopathy occurs [9].

Figure 1.

Regulation of blood clotting. Inhibitors that control coagulation are shown in gray boxes above the complex or factor they regulate (AT, antithrombin; ZPI, protein Z-dependent protease inhibitor; PCI, protein C inhibitor). Top panel: FVIIa binds to tissue factor (TF) to activate FX, generating FXa. FXa then binds to FVa. The complex of FXa–FV converts prothrombin (pro) to thrombin (T). Thrombin can then either bind to TM or carry out procoagulant reactions like fibrin formation or platelet activation. When bound to TM, thrombin can activate protein C (PC) to APC. This process is enhanced when protein C is bound to the EPCR. APC bound to EPCR cleaves substrates other than FVa. APC dissociates from EPCR and can then interact with protein S to inactivate FVa. Bottom panel: The FIXa–FVIIIa complex is inactivated by APC. In this case, FV participates with APC and protein S in the inactivation of FVIIIa. For simplicity, the activation of FVII, FV and FVIII are not shown.

Thrombin has a variety of activities on cells that result in augmentation of the inflammatory response (Fig. 2). While thrombin is thought to be involved primarily in coagulation, its ability to augment leukocyte adhesion and activation likely contributes to amplification of the inflammatory response. Thrombin activation of the endothelium results in high levels of platelet-activating factor formation [10], which works as a potent neutrophil agonist, especially when neutrophils are tethered to selectins [11]. Furthermore, P-selectin seems to play an important role in thrombosis since inhibition of P-selectin decreases thrombus formation under both flow [12] and stasis conditions [13]. In addition, thrombin activation of platelets releases CD40 ligand which in turn can induce tissue factor formation [14,15] and increase inflammatory cytokines, including IL-6 and IL-8 [16,17]. Increased levels of IL-6 have been shown in vivo to increase platelet reactivity increasing their thrombogenic potential [18], thus further linking inflammation and thrombosis. Fortunately, natural anticoagulant mechanisms dampen this inflammatory–coagulation interface. Any effective anticoagulant should decrease the coagulant-mediated increase in inflammatory potential. Some of these natural anticoagulant mechanisms are concentrated on the microvascular endothelium and because of this location, may have advantages in inhibiting thrombin activation of the endothelium.

Figure 2.

Thrombin is a multifunctional enzyme. Thrombin generates procoagulant, anticoagulant, inflammatory and mitogenic responses. These responses serve to shift the hemostatic balance. EC, endothelial cell; PMNs, polymorphonucleocytes; PAF, platelet activating factor; PDGF, platelet-derived growth factor; TGF-β, transforming growth factor-β; CD40L, CD40 ligand; MCP-1, macrophage chemotactic protein-1; IL-6, interleukin-6; IL-8, interleukin-8. (Modified with permission from Esmon et al. Annual Review of Cell Biology 1993; 9: 1–26,with permission from Esmon CT. Cell mediated events that control blood coagulation and vascular injury. Annu Rev Cell Biol 1993; 91: 1–26 [77], © Annual Reviews.)

Natural anticoagulant mechanisms and their downregulation by inflammatory mediators

Tissue factor–FVIIa activity is controlled by tissue factor pathway inhibitor (TFPI) [19–22] and heparin-antithrombin [23]. The relative importance of TFPI in controlling thrombosis is documented by the severe thrombotic complications observed upon TFPI gene deletion in mice that leads to an embryonic lethal phenotype [19]. The importance of tissue factor–FVIIa inhibition by antithrombin remains uncertain. It is difficult to assess the role of inflammation in controlling TFPI since the majority of TFPI is vessel associated [20] and a significant percentage of this seems to be stored in endothelial cell granules that can be released by thrombin treatment [24].

Factor Xa (FXa) is inhibited by antithrombin and by a complex between vitamin K-dependent protein Z and the protein Z-dependent protease inhibitor (ZPI), a plasma serpin [25]. The protein Z–ZPI complex binds to negatively charged phospholipids to inhibit FXa [25]. Inflammation impacts on this reaction since protein Z appears to be a negative acute phase reactant [26]. In murine models, protein Z deficiency has been shown to augment the risk of thrombosis [27], at least in the case of mice with FV Leiden. In addition, low levels of protein Z have been associated with an increased risk of stroke [28]. Acute inflammatory events can also decrease vascular heparin proteoglycans [29] thought to contribute to inhibition of the tissue factor–FVIIa complex, FXa and thrombin.

Of the natural anticoagulants, the protein C pathway appears to be the most negatively influenced by inflammation. Thrombomodulin (TM) and the endothelial cell protein C receptor (EPCR) are both downregulated by inflammatory cytokines like TNFα[30,31]. Neutrophil elastase readily cleaves TM from the endothelial cell surface [32,33], generating a form with reduced protein C activation potential. Furthermore, TM is very sensitive to oxidation of an exposed methionine [34], and hence can be damaged by oxidants emerging from endothelial cell-associated leukocytes. Both EPCR and TM are severely downregulated over atherosclerotic plaque [35], potentially contributing to both thrombosis and plaque rupture through localized thrombin generation and subsequent metalloproteinase activation. In addition, TM is downregulated in models of diabetes [36], possibly contributing to thrombotic complications associated with that disease. In acute inflammatory events leading to severe sepsis, the levels of protein C and protein S drop markedly and the extent of this decrease is associated with an increased risk of death [37]. Analysis of circulating activated protein C (APC) levels in patients with severe sepsis indicates that the protein C activation complex is indeed compromised in these patients, but that the extent of the compromise varies widely among patients [38]. Consistent with these observations, patients with severe meningococcemia can have markedly depressed levels of TM and EPCR with the anticipated decrease in protein C activation potential [39]. To put this in context, however, several clinical reports have shown rapid improvement in septic patients following protein C infusion [40].

Regulation of inflammation by natural anticoagulant pathways

Natural anticoagulants appear to possess unique, anti-inflammatory activities distinct from their anticoagulant activities. Antithrombin, TFPI and APC have all been shown to protect baboons from E. coli sepsis when given prior to the challenge. In this and several rodent models [41] either synthetic FXa inhibitors [41] or active site-blocked FXa, a potent inhibitor of prothrombin activation in vivo[42], failed to protect the animals or modulate cytokine elaboration. Thus current data suggest that the anti-inflammatory activities of the natural anticoagulants may be very important to their physiological functions.


In experimental animal settings, antithrombin has been shown to protect from septic shock [43]. Heparin appears to prevent protection despite increasing the antithrombotic activity of antithrombin, and a similar negative effect of heparin in combination with antithrombin was observed in clinical trials [44]. In vitro, supra-physiological levels of antithrombin have been shown to have anti-inflammatory activity [43]. Antithrombin has been found to inhibit endotoxin-induced interleukin-6 formation by mononuclear cells and endothelium [45]. These activities appear to be mediated through interaction of antithrombin with proteoglycans, like syndecan-4 [45]. Antithrombin interaction with the cell surfaces appears to be able to block NFκB nuclear translocation [46] and the subsequent release of cytokines and induction of adhesion molecules. In addition, antithrombin stimulates prostacyclin release from endothelial cells in culture [47], which appears to contribute to the protective effects of antithrombin in lung injury models [48].


TFPI has been shown to reduce leukocyte activation and decrease TNFα[41]. Inhibition of cytokine elaboration appears to be independent of blood coagulation, but the mechanism responsible for TFPI-mediated cellular effects remains unknown.

Protein C pathway

APC has been shown to protect non-human primates from E. coli-induced sepsis, whether given before or after the E. coli challenge. APC binding to monocytic cells has been shown to block agonist-induced calcium transients [49] and to inhibit NFκB-mediated signaling [50–53] and mRNA levels [54]. In endothelium, APC reduces the expression of cell surface adhesion molecules and cytokine formation, and elevates molecules involved in preventing apoptosis [54]. APC has also been shown to dampen both basal levels and the phorbol-induced expression of tissue factor on U937 cells [55]. The latter activities are EPCR dependent. Under appropriate conditions, APC can also cleave protease-activated receptors (PARs) [56]. It remains unclear to the author how this form of cell surface activation can elicit anti-inflammatory functions, since most of the downstream events following activation of these receptors enhance inflammation [57]. It is possible that activation of the PARs is a negative side reaction occurring under the in vitro conditions.

TM has also been found to exhibit anti-inflammatory properties. In part, this is mediated by protein C activation, but recent studies have identified additional pathways. TM also accelerates thrombin activation of a plasma procarboxypeptidase B, sometimes referred to as thrombin activatable fibrinolysis inhibitor (TAFI) [58]. TAFI was named for its ability to remove terminal Lys residues in fibrin. Since these lysine residues are important in binding plasminogen/plasmin and tissue plasminogen activator (TPA), removal of the lysines slows, but does not eliminate, clot lysis [58]. Originally, these activities were thought to stabilize the clot and therefore this activity of TM appeared to be thrombogenic.

Carboxypeptidase B enzymes remove C terminal Arg residues from vasoactive peptides like the complement anaphylatoxin C5a. C5a is generated during complement activation. Recent studies have found that TAFI is the major enzyme responsible for the inactivation of C5a [59,60]. In this context, the thrombin–TM complex protects the microvasculature by accelerating the clearance by activated TAFI and possibly other vasoactive peptides. Further linking this pathway to inflammation, C reactive protein, a strong acute phase reactant, has been found to initiate complement activation [61]. Inflammation leading to downregulation of TM, decreases in TAFI activation and increased complement activation could work together to injure the vessel wall and expose coagulant phospholipid surfaces.

Very recently, an additional potent anti-inflammatory activity of TM has been revealed by Conway et al. [62]. TM is composed of several domains. The N terminal domain has homology to the C type lectins. Conway et al. found that this domain dampened activation of the MAP kinase and NFκB signaling systems. Interestingly, TM had these activities whether on the cell surface or in solution.

This finding is potentially quite important as a contributing factor to arterial, venous and microvascular thrombosis. TM appears to be downregulated on endothelium overlying atherosclerotic plaques, vein bypass grafts, in diabetes, and by acute inflammatory insults like bacterial infection [63]. Loss of the TM would not only reduce protein C activation, but would also increase the sensitivity of the endothelium to phenotype modulation by inflammatory mediators. The net effect would be to promote leukocyte adhesion, increase permeability and reduce the natural antithrombotic surface. Consistent with this prediction, overexpression of TM has been shown to reduce thrombosis, restenosis and leukocyte infiltration in rabbits with deep arterial injury [64,65].

In addition to the anti-inflammatory activities of TM, EPCR has recently been found to exhibit anti-inflammatory activity. Inhibition of EPCR-protein C binding increased the coagulant and cytokine responses in animals challenged with low dose E. coli[66]. In addition, leukocyte migration into the tissues was increased substantially. A potential mechanism for the increased leukocyte migration came from the observations that soluble EPCR, released by a metalloproteinase in endothelium [67], can bind to selectively activated neutrophils and that the binding appears to involve interaction with proteinase 3 bound to Mac-1 (CD11b/CD18) [68]. In vivo data suggest that this interaction reduces tight binding of neutrophils to activated endothelium.

It has been recognized since EPCR was cloned [30] and the gene structure determined [69] that EPCR was closely related to the MHC class 1/CD1 family of proteins. The crystal structure confirmed and expanded this view by showing that EPCR has a tightly bound phospholipid in the ‘antigen-presenting groove’[70]. CD1 family members are antigen-presenting molecules, but unlike the MHC class 1 family, they present lipid antigens. For instance in the case of CD1c, this is a lipid derived from tuberculosis [71]. The CD1 series of proteins then instruct T cells and modulate the cellular and humoral response to inflammation. Furthermore, they appear likely candidates to be involved in autoimmunity [72]. Whether EPCR plays similar roles should become clear through analysis of genetically modified mice [73].

Structures linking the coagulation pathway and inflammation

Several factors within the coagulation system share structural similarity to components of the inflammatory pathway. For instance, tissue factor has structural homology to the cytokine receptors [74], the lectin domain of TM has homology to the selectins involved in leukocyte adhesion [75], and the structure of EPCR is almost superimposable over the structure of the corresponding regions of the MHC class 1/CD1 family of molecules [70]. Furthermore, linkages to the complement system can be seen in the interaction of complement regulatory protein, C4-binding protein, with protein S, an interaction that allows C4-binding protein to interact with membrane at the expense of protein S anticoagulant function [76]. Together these structural observations and interactions suggest that the coagulation and inflammatory pathways interact, but that there was very likely even tighter evolutionary linkage.


In the past few years, structural and functional studies have demonstrated increasingly tight interplay between the inflammatory and coagulation systems (Fig. 3). The mechanisms responsible for some of the hypercoaguable states fostered by inflammation are beginning to be clarified. These studies support a role for inflammation in arterial, venous and microvascular thrombosis. Rapidly evolving insights into the role of coagulation inhibitors in controlling inflammation may further clarify previously unrecognized risk factors and aid in assessing the transition to frank thrombosis. We now recognize that even the statins, thought originally to function by lowering cholesterol, have important anti-inflammatory effects as well. Further delineation of the links between inflammation and thrombosis should provide novel approaches to new diagnostics and therapeutics.

Figure 3.

Inflammation can have a prothrombotic effect.