doi: 10.1161/hh1001.091447
© 2001 American Heart Association, Inc.
Review |
New Tricks for Old Dogs
Nonthrombotic Effects of Thrombin in Vessel Wall Biology
From the Program in Molecular Cardiology and Department of Medicine (C.P., G.A.S., N.M., M.S.R.), Department of Pharmacology (C.P.), and Lineberger Comprehensive Cancer Center and Center for Thrombosis and Hemostasis (C.P., M.S.R.), University of North Carolina at Chapel Hill, Chapel Hill, North Carolina.
Correspondence to Cam Patterson, MD, University of North Carolina at Chapel Hill, 324 Burnett-Womack Building, Chapel Hill, NC 27599-7075. E-mail cpatters{at}med.unc.edu
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Abstract |
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 Top Abstract IntroductionThrombin Activation and...Vascular Thrombin ReceptorsThrombin SignalingThrombin and Vascular FunctionSummaryReferences |
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Abstract—Thrombin is a serine protease that potently activates platelets and catalyzes the conversion of fibrinogen to fibrin. Thrombin also exerts direct effects on vascular cells, such as smooth muscle cells, via interactions with members of the protease-activated receptor family. Evidence in several animal models implicates thrombin-mediated signaling events in the response to injury that typifies vascular lesion formation in atherosclerosis and restenosis. In this review, we examine the activation of protease-activated receptors by thrombin, the downstream signaling events mediated by these receptors, and the physiological role of thrombin in vascular cells and vascular disease.
Key Words: atherosclerosis • thrombin • vascular biology
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Introduction |
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TopAbstractIntroductionThrombin Activation and...Vascular Thrombin ReceptorsThrombin SignalingThrombin and Vascular FunctionSummaryReferences |
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Conceptually, our understanding of the interface between the thrombosis and hemostasis systems, on one hand, and vessel wall biology, on the other hand, is undergoing a gradual evolution. In its earliest formulation, the interface between these two processes was the inert barrier provided by the vascular endothelium; disruption of this barrier was considered the major stimulus to activation of the coagulation cascade. A level of complexity was added by the discovery that platelet proteins (such as platelet-derived growth factor) and hemostatic factors (such as thrombin) have potent activities on vascular biology independently of any coagulation effects. Conversely, we now know that vascular endothelial and smooth muscle cells (SMCs) produce proteins like tissue factor that modulate the coagulation cascade in the absence of endothelial disruption. This situation becomes even more complex when the effects of coagulation factors on cardiovascular development are considered; the recent demonstration that factor V–deficient mice, which cannot generate thrombin, die in utero of vascular defects in the absence of hemorrhage shows how closely these two systems are interrelated.1 Not only is the interface between coagulation and vessel wall biology not as simple as it was once thought, the boundaries between these two systems can now hardly be defined at all. It is not surprising, then, that the central molecule in blood coagulation, thrombin, also has profound effects on virtually every aspect of vascular wall biology, including regulation of vessel tone; SMC proliferation, differentiation, and migration; vascular development; atherogenesis; and angiogenesis (Figure 1
>). The purpose of this review is to take thrombin out of the narrow context of its role in the coagulation cascade and in
platelet function (these aspects of thrombin function have been
reviewed elsewhere2 3 ) and
to focus on the roles played by thrombin and its vascular receptors in
SMC biology and vascular disease. Although to date pharmacologic
inhibition of thrombin has yielded mixed results in the treatment and
prevention of atherogenesis, a more precise understanding of how
thrombin mediates its effects in the vasculature may well lead to more
focused therapeutic approaches.
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Thrombin Activation and Inhibition |
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TopAbstractIntroductionThrombin Activation and...Vascular Thrombin ReceptorsThrombin SignalingThrombin and Vascular FunctionSummaryReferences |
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Thrombin is produced predominantly on the surface of circulating platelets as a result of proteolytic activation of the 72-kDa zymogen prothrombin, which is constitutively synthesized in the liver and released into the circulation.4 Therefore, activation of thrombin can occur at a distance from its place of secretion, and, in particular, activation occurs at sites enriched for enzymes that facilitate the conversion of prothrombin to thrombin. In the circulation, factor Xa and its cofactor Va activate thrombin, and the activation of Xa is controlled by tissue factor, a membrane protein that is expressed at sites of vascular injury. Activated thrombin participates in clot formation via its serine protease activity, which cleaves fibrinogen to fibrin. Thrombosis is also facilitated by the ability of thrombin to activate platelets directly.
Knowledge of the structural determinants of the enzymatic activity of thrombin is important for understanding the mechanisms by which thrombin activity may be regulated. Because thrombin plays such a vital role in hemostasis, it comes as no surprise that its activity can be regulated by a variety of endogenous and exogenous factors and that thrombin is a prime target for pharmacologic intervention, as described in detail later in this review. The activity of thrombin can be regulated indirectly via factors that modulate the activation of thrombin (such as activated protein C, which degrades factors V and VIII and therefore inactivates the coagulation cascade5 ). However, direct inhibitors of thrombin, such as the endogenous proteins antithrombin III and heparin cofactor II and the exogenous inhibitors hirudin and bivalirudin (Hirulog), can also target the activity of this enzyme with specificity. Direct inhibitors of thrombin take advantage of the unique structural elements of the activated thrombin molecule, and the activity of direct inhibitors can best be explained on the basis of structural features that determine their effects.
Fortunately, the structure of thrombin, in isolation and in association with cofactors, has been extensively and elegantly studied.6 7 Thrombin assumes a globular football shape, with a deep cleft that contains the active site catalytic residues of the enzyme. Negatively charged residues that participate in substrate recognition surround the active site. Two additional positively charged patches interact with factors that modulate the activity of thrombin. One such region, the fibrinogen-recognition exosite, sits at the base of the active site cleft and mediates interactions between thrombin and the substrates fibrinogen and protease-activated receptor 1.8 9 The endogenous thrombin inhibitor heparin cofactor II docks with this site in a heparin- or dermatan sulfate–dependent manner that allows heparin cofactor II to interfere with the enzymatic activity of thrombin.10 The exogenous leech–derived inhibitor hirudin also binds to the fibrinogen-recognition exosite and the active site to inhibit thrombin function11 and is used clinically as a potent antithrombin. A second positively charged patch, to the opposite side of the active site cleft from the fibrinogen-recognition exosite, is the heparin-binding site that allows heparin to tether antithrombin III to thrombin. Antithrombin III, in turn, interferes with the negatively charged residues around the active site that facilitate substrate recognition, thus preventing their cleavage by thrombin.12
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Vascular Thrombin Receptors |
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TopAbstractIntroductionThrombin Activation and...Vascular Thrombin ReceptorsThrombin SignalingThrombin and Vascular FunctionSummaryReferences |
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Initial observations that thrombin exerts direct effects on platelets, endothelial cells, and SMCs indicated that receptor-mediated events must exist to initiate signaling responses to thrombin. The discovery of the first thrombin receptor (now referred to as protease-activated receptor-1, or PAR1) and the recognition that this receptor was activated by the proteolytic cleavage of its extracellular domain provided a new paradigm for receptor activation.13 The protease-activated receptors are a family of G protein–coupled receptors that contain protease recognition sequences in their extreme amino-terminal extracellular sequences. Proteolytic cleavage at these sites releases a small peptide and unmasks a new amino-terminal domain that can activate the receptor through intramolecular interactions. Proof of such a mechanism is demonstrated by the fact that synthetic peptides analogous to these unmasked sequences (the tethered ligands) can activate these receptors with an efficiency equivalent to that of the proteases themselves. For example, the synthetic peptide SFLLRN, which is exposed by thrombin-mediated cleavage of the extracellular domain of PAR1, efficiently activates signaling responses by this receptor in thrombin-sensitive cell types.13
Both the inability of the PAR1-activating peptide SFLLRN to
completely recapitulate thrombin’s effects in other cell types and the
likelihood that other receptors of this type existed led to the search
for other protease-activated receptors. Presently, 4
members of the protease-activated receptor family have been
identified (Table 1).13 14 15 16 17 Of these, PAR1, PAR3, and PAR4 can be cleaved and activated by thrombin,13 14 15
although thrombin-mediated cleavage of PAR4 occurs at higher
concentrations of thrombin than are required for activation of PAR1 and
PAR3, raising the possibility that activation of this receptor may
occur in an atypical fashion, as described below. PAR2, in contrast, is
not activated directly by thrombin but can be activated
under experimental conditions by trypsin and factor
VIIa.16 18 The existence of multiple thrombin receptors indicates that the cellular responses to thrombin may be determined at least in part by the presence of different combinations of thrombin receptors on different
cell types.
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Thrombin Signaling |
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TopAbstractIntroductionThrombin Activation and...Vascular Thrombin ReceptorsThrombin SignalingThrombin and Vascular FunctionSummaryReferences |
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The classic activation sequence of thrombin receptors (ie, proteolytic cleavage to unmask a tethered ligand) is best illustrated for PAR1, a thrombin receptor that is activated at low thrombin concentrations. Surprisingly, a pathway for receptor activation that involves intermolecular interactions also exists.19 When other thrombin receptors are absent, PAR4 requires high concentrations of thrombin for direct activation, presumably because of the lack of high-affinity thrombin interaction sites in the extracellular domain of this receptor. Activation of PAR4 by low thrombin concentrations occurs only when PAR3 is also present. It seems that PAR4 activation under these circumstances is initiated via binding of thrombin first to the PAR3 exosite. The thrombin-bound PAR3 complex then serves as a cofactor that allows trans-cleavage of the PAR4 exosite to unmask the tethered ligand of PAR4 (Figure 2
).
Trans-activation of PAR2 by
PAR1 may also occur, and PAR2 activation may be required for some
aspects of thrombin signaling, even though it is not itself a thrombin
receptor.20 The advantages
to be gained by such a complicated activation series are not yet
explained, although such events probably contribute to the diversity of
thrombin-mediated responses and allow for cell type–specific
activation sequences to occur. In addition, as described below, the
recruitment of multiple receptor types by thrombin extends beyond the G
protein–coupled receptor family.
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Ongoing work from several laboratories indicates that
signaling downstream from the thrombin receptors is surprisingly
complex. The events that occur consequent to receptor binding and G
protein activation involve broadly dissimilar, parallel signaling
pathways that frequently intersect and even activate signaling
via other, nonthrombin-activated receptors (Figure 1
). For these reasons, assigning specific effects of
thrombin to each pathway has been a challenge. Several of these
pathways have been characterized in a variety of cell types and are
described briefly in this review. More recently, generation of reactive
oxygen species (ROS) as signaling intermediates and crosstalk between
thrombin and other receptor families have been implicated in events
closely linked to vascular dysfunction. These recently elucidated
thrombin-activated signaling events are therefore described in
greater detail.
G Protein–Coupled Receptor Signaling
Although each of the protease-activated
receptors that respond to thrombin undoubtedly mediate different
thrombin responses, most of what is known about thrombin signaling
downstream of the receptors themselves has derived from studies of
PAR1. PAR1 couples with at least 3 G protein families,
G12/13, Gq, and
Gi, to elicit diverse downstream signaling
events.21 22 23
The well-characterized pathways activated via these G
protein–dependent events include Rho activation by
G13 that causes cytoskeletal changes affecting
vascular cell
migration.24 25
Gq-dependent signaling activates
phospholipase C,26 which in
turn leads to mitogen-activated protein kinase (MAPK)
phosphorylation and receptor tyrosine kinase
trans-activation, both
necessary events in thrombin-mediated proliferation. Gß
interactions activate phosphoinositide
3-kinase, which promotes Ca2+ release that
is required for SMC growth in response to
thrombin.27 It is clear that
diverse cellular responses to thrombin depend on this rich signaling
network and that ample opportunity exists for integration of signaling
events elicited by thrombin and by other activation pathways to
modulate the cellular phenotype.
Thrombin and ROS
ROS induce activation of growth-related signaling
pathways in a manner similar to that of endogenous growth
factors, indicating that ROS may lie downstream of some growth factors
in the course of intracellular
signaling.28 Growth factors
known to play a role in atherogenesis, such as platelet-derived
growth factor29 and angiotensin
II,30 generate intracellular
ROS, and ROS in turn can elicit specific growth responses in
SMCs.29 31 It is
therefore quite interesting that ROS production by a unique SMC
oxidase is also required for mitogenic signaling events
elicited by thrombin.32 This
oxidase is similar but not identical to the neutrophil NAD(P)H oxidase.
SMCs express p22phox,
a membrane-associated component of the oxidase, and
p47phox,
a cytoplasmic component that translocates to the activated
membrane–anchored oxidase on
stimulation.32 33
Characterization of the NAD(P)H oxidase in SMCs has indicated that in
these cells, the function of the catalytic membrane–associated
gp91phox
subunit is likely replaced by a homologue (a member of the
nox family) that has functional
and structural similarity to the neutrophil oxidase component
gp91phox.34
A recent review by Griendling et al35 presents a more
complete discussion of the components of the vascular NAD(P)H
oxidase.
Functional studies indicate that both p22phox and nox1 are necessary for ROS generation in SMCs,33 34 and recent data from our laboratories indicate that p47phox is also required for oxidase activity in SMCs (P. Barrylane, C. Patterson, M.S. Runge, unpublished data, 2001). The p47phox component of the oxidase becomes phosphorylated and is recruited to the cell membrane after thrombin stimulation, and this same oxidase component is upregulated after vascular injury in the rat carotid injury model, suggesting that the oxidase in general, and p47phox in particular, plays a central role in signaling events leading to vascular lesion formation. Of interest, angiotensin II activates the SMC oxidase to generate intracellular ROS and initiate cell growth,33 indicating that this oxidase may mediate a central pathway that is activated in vascular pathologies. However, the precise mechanisms that lead from thrombin-mediated receptor activation to oxidase assembly and ROS generation remain obscure. In neutrophils, several different signaling pathways have been implicated in p47phox phosphorylation and receptor activation, but the proximal kinases that activate the ROS-producing capacity of this oxidase continue to be obscure.36 It remains to be determined whether the pathways for thrombin-mediated oxidase activation will be different or similar to the pathways that activate the neutrophil NAD(P)H oxidase.
Although ROS generation via the SMC NAD(P)H oxidase plays a necessary role in thrombin-induced effects in SMCs, the precise signaling events mediated by these ROS are still unclear. It seems that ROS are relatively proximal events in the thrombin-signaling pathway insofar as thrombin-induced MAPK activity and Janus kinase/signal transducers and activators of transcription signaling are ROS-dependent.37 Intriguingly, recent data indicate that ROS may feedback directly on G proteins themselves that then activate signaling via MAPKs and other pathways.38 Clearly, a great deal more needs to be determined about the role of ROS in thrombin-mediated signaling and, by extension, in atherogenesis and other thrombin-associated phenomena. Ascertaining the role of ROS in these events is not a trivial point, because antioxidant therapies have been tested in several formulations for their effectiveness in modulating the progression of vascular lesions, albeit with little success.39 The gap in our understanding of the role of ROS in thrombin-mediated signaling certainly contributes to this oxidative paradox but offers the potential for development of new therapeutic interventions based on growing knowledge of these events.
Crosstalk Between Thrombin-Mediated Signaling
and Other Receptor Types
Trans-activation
of one protease-activated receptor by another, as described
above, provides one means of crosstalk between receptors, albeit within
the same family. G protein–coupled receptors, and in particular the
protease-activated receptors, also interact heterotypically
with other receptor families to elicit appropriate signaling responses.
The first well-characterized example of such an interaction was
described between PAR1 and the epidermal growth factor (EGF)
receptor.40 Thrombin was
found to elicit activation of the EGF receptor, and thrombin signaling
could be inhibited efficiently by a dominant-negative EGF receptor.
Surprisingly, this effect seemed to be independent of EGF itself, which
led to the initial belief that this receptor
trans-activation was
ligand-independent. However, these receptor interactions are in fact
much more complicated, and subsequent studies have shown that EGF
receptor activation by G protein–coupled receptors is dependent not on
EGF but on another ligand that interacts with the EGF receptor,
heparin-binding EGF-like growth factor
(HB-EGF).41 42
HB-EGF is synthesized as an inactive, membrane-bound precursor.
Protease-activated receptor activation by thrombin induces
metalloproteinase activity that sheds the active ectodomain of HB-EGF,
which then can activate the EGF receptor. Such interactions
provide yet another unexpected twist to the thrombin-signaling story,
although some issues, such as the nature of the metalloproteinase
activity that is stimulated by this system, remain to be
clarified.
Receptor tyrosine kinases are not the only heterotypic receptors that interact functionally with the thrombin receptors in vascular SMCs. Interactions between platelet integrins and protease-activated receptors have been well-characterized, and more recent studies indicate that ß3-containing integrins communicate with thrombin receptors and that integrin signaling is necessary for maximal thrombin-mediated proliferative events in SMCs.43 The precise contributions of direct interactions between protease-activated receptors and integrins or of intracellular interactions and ligand-dependent integrin signaling through molecules such as thrombospondin remain to be determined. However, the functional importance of these interactions is likely to be significant, insofar as integrins are required for phenotypic responses in SMCs, such as adhesive and migratory cell-matrix events that are central to the vascular response to injury.44 45
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Thrombin and Vascular Function |
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TopAbstractIntroductionThrombin Activation and...Vascular Thrombin ReceptorsThrombin SignalingThrombin and Vascular FunctionSummaryReferences |
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It is now clear that the effects of thrombin extend beyond its central role in coagulation and platelet function. Receptors for thrombin are present on vascular SMCs and endothelial cells,46 and thrombin stimulation of the endothelium has effects that are prothrombotic47 and proinflammatory.48 Thrombin is mitogenic and chemotactic for inflammatory cells,49 50 such as lymphocytes and macrophages, which may be present in the vessel wall in pathologic conditions. Thrombin is a potent stimulus for endothelium-dependent vasodilatation mediated by activation of nitric oxide production.51 Thrombin also modulates endothelial cell growth responses. Thrombin is among the most potent stimuli for nonhypoxic vascular endothelial growth factor (VEGF) expression52 and also upregulates VEGF receptors in endothelial cells.53 These latter effects may explain, in part, the effects of thrombin on angiogenesis and vascular development, as described below.
Thrombin and Vascular SMCs
Although modulation of endothelial cell
function by thrombin might be easily understandable, in consideration
of the role of thrombin in maintaining hemostasis, a role for thrombin
in SMC biology indicates that its effects are not limited to
coagulation events and are more diverse than originally thought.
Therefore, the importance of initial observations that thrombin
stimulated SMC proliferation cannot be
overestimated.54
Although thrombin-induced proliferation is perhaps the
best-described cellular response of SMCs, the cellular program elicited
by thrombin in this cell type is relatively complex
(Table 2). Thrombin elicits an inflammatory response that
includes secretion of interleukin-6 and monocyte chemotactic
protein-155 and stimulates
SMCs to synthesize
collagen56 that may
contribute to extracellular matrix accumulation. Smooth muscle
migration is also induced in response to thrombin
stimulation.57 The
inflammatory, fibrotic, and chemotactic responses to thrombin may be
major contributors to the deleterious response to injury that
culminates in obstructive vascular lesions. Heat shock proteins such as
Hsp70, Hsp90, and Hsp27 are also activated and upregulated in
SMCs by
thrombin,58 59
and these proteins may contribute to the atherogenic response, because
expression of these proteins is particularly concentrated in SMCs
surrounding the necrotic core of atherosclerotic
plaques.60
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Finally, an interaction between thrombin and smooth muscle that deserves mention is the activation of thrombin that occurs in SMCs undergoing cell death via the apoptotic pathway.61 Phosphatidylserine that is exposed at the membrane of cells undergoing apoptosis facilitates the assembly of a prothrombinase complex that accelerates the generation of thrombin. Apoptosis of SMCs can occur after vascular injury62 and in response to changes in extracellular matrix composition63 64 and may exacerbate the wound-healing response.65 The generation of active thrombin by these apoptotic SMCs may serve as a positive feedback loop to accelerate the atherogenic process and as such may provide one explanation for the uncontrolled proliferation of SMCs after vascular injury.
Thrombin and Vascular Lesion Formation
There has been speculation for some time that thrombin
participates in vascular lesion
formation.66 67
Indeed, the thrombotic effects of thrombin alone would implicate this
protease in vascular lesion formation in the setting of vascular injury
and atherosclerosis. Recent studies indicate that
thrombin does in fact increase the response to injury in these
settings; however, these effects seem to be mediated primarily via
direct effects of thrombin on vascular arterial cells
rather than indirectly through its hemostatic effects.
Concentrations of thrombin are increased at the arterial surface after denuding injury and remain elevated for up to 10 days.68 69 Both activated thrombin and its precursor, prothrombin, can be detected within the neointima of human atherosclerotic lesions.70 Effects of this thrombin on adjacent cells are suggested not only by its presence in activated form but also by upregulation of its cognate receptor PAR1. PAR1 expression is upregulated within 6 hours of injury to the rat carotid artery and remains so during the course of neointima formation.71 Increased expression of PAR1 has also been observed in atherosclerotic plaques from human arteries but not in normal arteries.72 Thus, thrombin and its receptors are in the right place at the right time to contribute to the vasculoproliferative response to injury.
A critical question raised by the presence of active thrombin within the arterial walls is how this thrombin is regulated. The major endogenous inhibitor of thrombin in the intravascular compartment is antithrombin III, yet little antithrombin III is present within vascular lesions and no antithrombin III/thrombin complexes are detectable.70 It has been suggested that plasminogen activator inhibitor 1 (PAI-1) can interact with activated thrombin to inhibit thrombin-mediated activity and that vitronectin is a cofactor in this complex.73 This complex may be converted either to an inactive thrombin/PAI-1 complex or scavenged by the LDL receptor–related protein present on SMCs and macrophages.74 75 Indeed, active thrombin, PAI-1, and vitronectin can be found colocalized within human atherosclerotic lesions,76 suggesting that this is a physiologically relevant mechanism for regulation of thrombin activity within vascular lesions. The existence of a regulatory mechanism involving inhibition of active thrombin by PAI-1 suggests that decreased PAI-1 expression or activity within arteries should enhance the amount of free thrombin available to stimulate vascular cells, with deleterious consequences in the context of lesion formation. Such a scenario is supported by observations that mice lacking PAI-1 have greater neointimal formation after injury than do wild-type mice,77 indicating an inhibitory role for PAI-1 in lesion formation, perhaps via its ability to sequester active thrombin.
Heparin cofactor II is an additional factor that may regulate the activity of thrombin in vascular lesions. Heparin cofactor II binds to and inactivates thrombin and can potently inhibit thrombus formation in models of thrombosis.78 As mentioned above, heparin cofactor II inhibits thrombin by directly interacting with the fibrinogen-recognition exosite of thrombin, and the activity of heparin cofactor II is increased by interactions with dermatan sulfate–containing proteoglycans of the extracellular matrix.79 Interestingly, proteoglycans derived from atherosclerotic vessels, such as biglycan and decorin, are impaired in their ability to activate heparin cofactor II, compared with proteoglycans derived from normal arteries.80 Taken together, these results indicate an additional means by which thrombin activity is regulated within the vasculature and suggest that alterations in dermatan sulfate–containing molecules may accelerate atherosclerotic lesion formation by attenuating the inactivation of thrombin.
The functional role of thrombin in lesion formation has been addressed elegantly in several different models, and hirudin, a potent thrombin inhibitor, has been a potent tool in this regard. Hirudin administration in the periprocedural period reduces neointimal formation after injury in minipig and rabbit models.81 82 This effect may be attributable to prolonged inhibition of thrombin activity within arteries by hirudin after short-term administration.69 Interestingly, studies performed in rats demonstrate that prolonged (but not short-term) administration of hirudin is effective in inhibiting the vasculoproliferative response.83 Likewise, administration of hirudin via adenoviral gene therapy, which produces hirudin over a period of days, is effective in blocking lesion formation in the rat carotid injury model.84 Taken together, these studies demonstrate the importance of thrombin generation in lesion formation but also emphasize the significance of the time course of thrombin inhibition and possibility of species-specific responses to thrombin inhibition in the vasculoproliferative process.
Targeting the receptor PAR1 using inhibitory antibodies has been a second approach to inhibiting thrombin activity experimentally. The feasibility of this approach has been indicated by the inhibition of thrombus formation in Green monkeys85 and by blocking neointimal formation after balloon injury in the rat.86 Because rodent platelets (in contrast with those in primates) do not express PAR1 and are activated by thrombin in a PAR1-independent manner, these experiments provide presumptive evidence that the effects of thrombin on vascular lesion formation are attributable in significant part to direct effects of thrombin on vascular cells.
More precise studies of the role of thrombin signaling via
the PAR1 receptor have been performed using genetically modified mice
that lack the PAR1 receptor. These experiments generally confirm the
preceding studies but also suggest that the role of this pathway may be
more complex than previously
indicated.87
Neointimal and medial areas are decreased in PAR1-deficient
mice after denuding injury, yet in these experiments luminal diameters
also decrease, suggesting a component of adverse remodeling that may
indicate changes in extracellular matrix components in the
PAR1-deficient mice. These studies, taken together with the
species-specific effects of thrombin inhibitors,
demonstrate that the complexity of this system needs to be taken into
account in the translation of this approach to human studies
(Table 3). In addition, doses of hirudin used in the human
trials have been far lower than those used in animal studies, owing to
the risks of bleeding associated with the use of higher doses in
humans. More promising approaches for sustained, local,
high-concentration delivery, including the use of impregnated stents,
may be a useful means to reap the benefits of long-term thrombin
inhibition in the setting of vascular injury without incurring
dose-limiting side effects. Other novel approaches may involve
interrupting specific downstream signals resulting from
thrombin-mediated vascular cell activation.
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Given the preclinical data, it is disappointing that clinical studies using the direct thrombin inhibitors hirudin and bivalirudin have failed to show efficacy in inhibition of restenosis after angioplasty in humans.88 89 However, the disappointment incurred by these studies must be tempered by the realization provided from animal studies that the consequences of thrombin inhibition on lesion formation may depend on the duration of thrombin inhibition and on other factors. In the human studies reported so far, the duration of thrombin inhibition has been relatively short and the doses used were far lower than those used in animal studies. Consideration that thrombin inhibition in these trials was either too brief or incomplete raises the possibility that locally delivered hirudin (or other thrombin inhibitors), perhaps delivered via impregnated stents, may be a more reasonable approach to inhibit thrombin effectively, locally, and for prolonged periods.
Thrombin and Angiogenesis
Thrombosis has long been associated with
angiogenesis-associated diseases such as cancer, and thrombin itself
can be detected in a variety of tumor
types.90 However, an
appreciation that thrombin generation may directly promote angiogenesis
has only emerged recently. Angiogenesis is stimulated by thrombin in
both in vivo and in vitro model
systems.91 92
Several factors apparently contribute to the angiogenic effects of
thrombin. VEGF is a potent angiogenic factor that acts via
endothelial cell–specific receptor tyrosine kinases,
and thrombin is a potent stimulus for release of VEGF stores within
platelets.93 Thrombin
also upregulates expression of the VEGF receptors to potentiate VEGF
activity on endothelial
cells.53 VEGF in turn
accelerates thrombin
generation,94 suggesting
these two factors act as part of a positive feedback loop to accelerate
the angiogenic process. The effects of thrombin in the setting of
angiogenesis are not limited to its modulation of the VEGF axis,
because thrombin also activates matrix metalloproteinases that
are required for tissue remodeling in
angiogenesis.95
An additional and interesting link between thrombin and angiogenesis derives from recent evidence indicating that proteolytic fragments of antithrombin III and prothrombin themselves serve as inhibitors of angiogenesis. Thrombin-cleaved antithrombin III was purified as an antiangiogenic factor derived from small-cell lung cancer cells.96 Likewise, the kringle-2 domain of prothrombin, which is derived from factor Xa–mediated cleavage of prothrombin, was identified in the serum of lipopolysaccharide-treated rabbits as an antiproliferative factor for vascular endothelial cells.97 Although the physiological role of these proteolytic fragments remains to be determined, their identification suggests that the thrombin activation system has a more central role in the angiogenic process than has been previously appreciated. In addition, thrombin-mediated angiogenesis seems to be under both positive and negative feedback controls. A great deal more about this angiogenic pathway, including what role thrombin inhibitors such as hirudin will play in therapeutic modulation of angiogenesis, remains to be determined.
Thrombin and Vascular Development
Although initial reports indicating a role for the
thrombin pathway in vascular development were unanticipated, such an
association is easier to understand given the role of thrombin in
angiogenesis, because angiogenesis is a critical component of the
vascular development program. Deletion of prothrombin by homologous
recombination in mice results in characteristic defects in yolk sac
vasculature.98 99
Similar defects are observed in factor V–deficient mice, which fail to
generate active thrombin.1 It
might be presumed that the developmental effects of thrombin deficiency
are attributable to its effects on hemostasis; however, several lines
of evidence indicate that this is not entirely the case. First,
embryonic hemorrhage was noted in only one of the two reports
of prothrombin deficiency98 ;
similarly, factor V–deficient embryos are devoid of
hemorrhage.1 Second,
deficiency of PAR1 also results in developmental defects (which have
not yet been completely
characterized),100 101
even though this thrombin receptor is not required for platelet
activation or fibrin generation in mice. Third (and quite remarkably),
mice deficient in either
fibrinogen102 or
platelets103 develop
normally, indicating that coagulation per se is not required for the
vascular phenotypes observed in the thrombin pathway during
development. A direct effect of thrombin on vascular cells is plausible
insofar as expression of PAR1 is highest in endothelial
cells at early stages of vascular
development.46
The exact role played by thrombin in vascular development is not yet clear, although it is tempting to speculate that thrombin may be required for local control of VEGF signaling. The developing vascular system is exceptionally sensitive to quantitative changes in VEGF expression, insofar as mice lacking a single VEGF allele die in utero of a vascular developmental catastrophe104 and VEGF receptor expression is required for normal endothelial cell development.105 However, other functions for thrombin on vascular development, instead of or in addition to effects on VEGF signaling, are likely. An important source of information in this regard will be complete descriptions of the developmental phenotypes of mice lacking the different thrombin receptors.
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Summary |
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TopAbstractIntroductionThrombin Activation and...Vascular Thrombin ReceptorsThrombin SignalingThrombin and Vascular FunctionSummaryReferences |
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Thrombin is one of the most well-characterized factors that participates in the vascular response to injury, yet many hypotheses regarding its role in atherogenesis and restenosis have fallen apart on the basis of emerging data indicating direct effects of thrombin on vascular cells, the existence of a novel family of thrombin receptors that are activated in cis and trans via tethered ligands, and trans-activation of receptor tyrosine kinases via metalloproteinase-induced growth factor activation. Despite our expanding knowledge of the role of thrombin in vascular lesion formation in animal models, we still have no effective means to test its role in human disease. Advances in pharmacology and biodelivery or the development of agents that interfere with downstream thrombin-mediated signals may mean that these experiments are on the horizon.
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Acknowledgments |
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The authors’ laboratories are supported by National Institutes of Health Grants HL03658, HL61656, AG10514, and GM61728 (to C.P.) and HL57352 and HL59652 (to M.S.R.). The authors gratefully acknowledge Enid Hatton for assistance with graphics.
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Footnotes |
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Original received March 2, 2001; revision received April 12, 2001; accepted April 12, 2001.
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References |
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TopAbstractIntroductionThrombin Activation and...Vascular Thrombin ReceptorsThrombin SignalingThrombin and Vascular FunctionSummaryReferences |
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1. Cui J, O’Shea KS, Purkayastha A, Saunders TL, Ginsburg D. Fatal haemorrhage and incomplete block to embryogenesis in mice lacking coagulation factor V. Nature. 1996;384:66–68.[Medline] [Order article via Infotrieve]
2. Coughlin SR. Protease-activated receptors and platelet function. Thromb Haemost. 1999;82:353–356.[Medline] [Order article via Infotrieve]
3. Coughlin SR. Thrombin signalling and protease-activated receptors. Nature. 2000;407:258–264.[Medline] [Order article via Infotrieve]
4. Fenton JW. Thrombin. Ann N Y Acad Sci. 1986;485:5–15.[Medline] [Order article via Infotrieve]
5.
Esmon CT. The roles
of protein C and thrombomodulin in the regulation of blood coagulation.
J Biol Chem. 1989;264:4743–4746.
6. Bode W, Mayr I, Baumann U, Huber R, Stone SR, Hofsteenge J. The refined 1.9 Å crystal structure of human a-thrombin: interaction with D-Phe-Pro-Arg chloromethylketone and significance of the Tyr-Pro-Pro-Trp insertion segment. EMBO J. 1989;8:3467–3475.[Medline] [Order article via Infotrieve]
7. Bode W, Turk D, Karshikov A. The refined 1.9 Å x-ray crystal structure of D-Phe-Pro-Arg chloromethylketone inhibited a-thrombin: structure analysis, overall structure, electrostatic properties, detailed active site geometry, structure-function relationships. Protein Sci. 1992;1:426–471.[Medline] [Order article via Infotrieve]
8. Fenton JW 2nd, Zabinski MP, Hsieh K, Wilner GD. Thrombin non-covalent protein binding and fibrin(ogen) recognition. Thromb Haemost. 1981;46:177. Abstract.
9. Mathews I, Padmanabhan KP, Ganesh V, Tulinsky A, Ishii M, Chen J, Turck CW, Coughlin SR, Fenton JW 2nd. Crystallographic structures of thrombin complexed with thrombin receptor peptides: existence of expected and novel binding modes. Biochemistry. 1994;33:3266–3279.[Medline] [Order article via Infotrieve]
10. Church FC, Hoffman MR. Heparin cofactor II and thrombin: heparin-binding proteins linking hemostasis and inflammation. Trends Cardiovasc Med. 1994;4:140–146.
11.
Rydel T,
Ravichandran K, Tulinsky A, Bode W, Huber R, Roitsch C, Fenton JN. The
structure of recombinant hirudin and human 
-thrombin.
Science. 1990;249:277–280.
12. Bourin M-C, Lindahl U. Glycosaminoglycans and the regulation of blood coagulation. Biochem J. 1993;289:313–330.
13. Vu TKH, Hung DT, Wheaton VI, Coughlin SR. Molecular cloning of a functional thrombin receptor reveals a novel proteolytic mechanism of receptor activation. Cell. 1991;64:1057–1068.[Medline] [Order article via Infotrieve]
14. Ishihara H, Connolly AJ, Zeng D, Kahn ML, Zheng YW, Timmons C, Tram T, Coughlin SR. Protease-activated receptor 3 is a second thrombin receptor in humans. Nature. 1997;386:502–506.[Medline] [Order article via Infotrieve]
15. Kahn ML, Zheng YW, Huang W, Bigornia V, Zeng D, Moff S, Farese RV Jr, Tam C, Coughlin SR. A dual thrombin receptor system for platelet activation. Nature. 1998;394:690–694.[Medline] [Order article via Infotrieve]
16.
Nystedt S,
Emilsson K, Wahlestedt C, Sundelin J. Molecular cloning of a potential
proteinase activated receptor.
Proc Natl Acad Sci
U S A. 1994;91:9208–9212.
17.
Xu WF, Andersen
H, Whitmore TE, Presnell SR, Yee DP, Ching A, Gilbert T, Davie EW,
Foster DC. Cloning and characterization of human
protease-activated receptor 4.
Proc Natl Acad Sci
U S A. 1998;95:6642–6646.
18.
Camerer E, Huang
W, Coughlin SR. Tissue factor- and factor X-dependent activation of
protease-activated receptor 2 by factor VIIa.
Proc Natl Acad Sci
U S A. 2000;97:5255–5260.
19. Nakanishi-Matsui M, Zheng YW, Sulciner DJ, Weiss EJ, Ludeman MJ, Coughlin SR. PAR3 is a cofactor for PAR4 activation by thrombin. Nature. 2000;404:609–613.[Medline] [Order article via Infotrieve]
20.
O’Brien PJ,
Prevost N, Molino M, Hollinger MK, Woolkalis MJ, Woulfe DS, Brass LF.
Thrombin responses in human endothelial cells:
contributions from receptors other than PAR1 include the
transactivation of PAR2 by thrombin-cleaved PAR1.
J Biol Chem. 2000;275:13502–13509.
21.
Hung DT, Wong YH,
Vu TK, Coughlin SR. The cloned platelet thrombin receptor couples
to at least two distinct effectors to stimulate
phosphoinositide hydrolysis and inhibit adenylyl
cyclase. J Biol Chem. 1992;267:20831–20834.
22.
Offermanns S,
Laugwitz KL, Spicher K, Schultz G. G proteins of the G12 family are
activated via thromboxane A2 and thrombin receptors
in human platelets. Proc Natl Acad Sci
U S A. 1994;91:504–508.
23.
Barr AJ, Brass
LF, Manning DR. Reconstitution of receptors and GTP-binding regulatory
proteins (G proteins) in Sf9 cells: a direct evaluation of selectivity
in receptor.G protein coupling. J
Biol Chem. 1997;272:2223–2229.
24.
Offermanns S,
Mancino V, Revel JP, Simon MI. Vascular system defects and impaired
cell chemokinesis as a result of G13 deficiency.
Science. 1997;275:533–536.
25.
Seasholtz TM,
Majumdar M, Kaplan DD, Brown JH. Rho and Rho kinase mediate
thrombin-stimulated vascular smooth muscle cell DNA synthesis and
migration. Circ Res. 1999;84:1186–1193.
26.
Taylor SJ, Chae
HZ, Rhee SG, Exton JH. Activation of the ß1
isozyme of phospholipase C by subunits of the Gq class of G
proteins. Nature. 1991;350:516–518.[Medline]
[Order article via Infotrieve]
27.
Krymskaya VP,
Penn RB, Orsini MJ, Scott PH, Plevin RJ, Walker TR, Eszterhas AJ,
Amrani Y, Chilvers ER, Panettieri RA Jr. Phosphatidylinositol 3-kinase
mediates mitogen-induced human airway smooth muscle cell proliferation.
Am J Physiol. 1999;277:L65–L78.
28. Lander HM. An essential role for free radicals and derived species in signal transduction. FASEB J. 1997;11:118–124.[Abstract]
29.
Sundaresan M, Yu
Z-X, Ferrans VJ, Irani K, Finkel T. Requirement for generation of
H2O2 for
platelet-derived growth factor signal transduction.
Science. 1995;270:296–299.
30.
Griendling KK,
Minieri CA, Ollerenshaw JD, Alexander RW. Angiotensin II
stimulates NADH and NADPH oxidase activity in cultured vascular smooth
muscle cells. Circ Res. 1994;74:1141–1148.
31.
Rao GN, Berk BC.
Active oxygen species stimulate vascular smooth muscle cell growth and
proto-oncogene expression. Circ
Res. 1992;70:593–599.
32. Patterson C, Ruef J, Madamanchi NR, Barry-Lane P, Hu Z, Horaist C, Ballinger CA, Bode C, Runge MS. Stimulation of a vascular smooth muscle cell NAD(P)H oxidase by thrombin: evidence that p47phox may participate in forming this oxidase in vitro and in vivo. J Biol Chem. 1999;275:19814–19822.
33.
Ushio-Fukai M,
Zafari A, Fukui T, Ishizaka N, Griendling K,
p22phox
is a critical component of the superoxide-generating NADH/NADPH oxidase
system and regulates angiotensin II-induced
hypertrophy in vascular smooth muscle cells.
J Biol Chem. 1996;271:23317–23321.
34. Suh YA, Arnold RS, Lassegue B, Shi J, Xu X, Sorescu D, Chung AB, Griendling KK, Lambeth JD. Cell transformation by the superoxide-generating oxidase Mox1. Nature. 1999;401:79–82.[Medline] [Order article via Infotrieve]
35.
Griendling KK,
Sorescu D, Ushio-Fukai M. NAD(P)H oxidase: role in
cardiovascular biology and disease.
Circ Res. 2000;86:494–501.
36. Lal AS, Parker PJ, Segal AW. Characterization and partial purification of a novel neutrophil membrane-associated kinase capable of phosphorylating the respiratory burst component p47phox. Biochem J. 1999;338:359–366.
37. Madamanchi NR, Li S, Patterson C, Runge MS. Thrombin regulates vascular smooth muscle cell growth and heat shock proteins via the JAK-STAT pathway. J Biol Chem. March 16, 2001;10.1074/jbc.M008802200. Available at: http://www.jbc.org. Accessed April 18, 2001.
38.
Nishida M,
Maruyama Y, Tanaka R, Kontani K, Nagao T, Kurose H.
Gi and Go are
target proteins of reactive oxygen species.
Nature. 2000;408:492–495.[Medline]
[Order article via Infotrieve]
39.
Patterson C,
Madamanchi NR, Runge MS. The oxidative paradox: another piece in the
puzzle. Circ Res. 2000;87:1074–1076.
40. Daub H, Weiss FU, Wallasch C, Ullrich A. Role of transactivation of the EGF receptor in signalling by G-protein-coupled receptors. Nature. 1996;379:557–560.[Medline] [Order article via Infotrieve]
41.
Kalmes A, Vesti
BR, Daum G, Abraham JA, Clowes AW. Heparin blockade of thrombin-induced
smooth muscle cell migration involves inhibition of epidermal growth
factor (EGF) receptor transactivation by heparin-binding EGF-like
growth factor. Circ Res. 2000;87:92–98.
42. Prenzel N, Zwick E, Daub H, Leserer M, Abraham R, Wallasch C, Ullrich A. EGF receptor transactivation by G-protein-coupled receptors requires metalloproteinase cleavage of proHB-EGF. Nature. 1999;402:884–888.[Medline] [Order article via Infotrieve]
43.
Stouffer GA, Hu
Z, Sajid M, Li H, Jin G, Nakada MT, Hanson SR, Runge MS. ß3 integrins
are upregulated after vascular injury and modulate thrombospondin- and
thrombin-induced proliferation of cultured smooth muscle cells.
Circulation. 1998;97:907–915.
44.
Bilato C, Curto
KA, Monticone RE, Pauly RR, White AJ, Crow MT. The inhibition of
vascular smooth muscle cell migration by peptide and antibody
antagonists of the vß3 integrin complex is reversed by
activated calcium/calmodulin-dependent protein
kinase II. J Clin Invest. 1997;100:693–704.[Medline]
[Order article via Infotrieve]
45.
Srivatsa SS,
Fitzpatrick LA, Tsao PW, Reilly TM, Holmes DR Jr, Schwartz RS, Mousa
SA. Selective vß3 integrin blockade potently limits
neointimal hyperplasia and lumen stenosis following
deep coronary arterial stent injury: evidence for
the functional importance of integrin vß3 and osteopontin
expression during neointima formation.
Cardiovasc Res. 1997;36:408–428.
46. Soifer S, Peters K, O’Keefe J, Coughlin S. Disparate temporal expression of the prothrombin and thrombin receptor genes during mouse development. Am J Pathol. 1994;144:60–69.[Abstract]
47.
Hattori R,
Hamilton KK, Fugate RD, McEver RP, Sims PJ. Stimulated secretion of
endothelial von Willebrand factor is
accompanied by rapid redistribution to the cell surface of the
intracellular granule membrane protein GMP-140.
J Biol Chem. 1989;264:7768–7771.
48.
Lum H, Malik AB.
Regulation of vascular endothelial barrier function.
Am J Physiol. 1994;267:L223–L241.
49.
Bar-Shavit R,
Kahn A, Wilner GD, Fenton JW 2nd. Monocyte chemotaxis: stimulation by
specific exosite region of thrombin.
Science. 1983;220:728–731.
50.
Bar-Shavit R,
Kahn AJ, Mann KG, Wilner GD. Identification of a thrombin sequence with
growth factor activity on macrophages.
Proc Natl Acad Sci
U S A. 1986;83:976–980.
51.
Tesfamariam B,
Allen GT, Normandin D, Antonaccio MJ. Involvement of the
"tethered ligand" receptor in thrombin-induced
endothelium-mediated relaxations.
Am J Physiol. 1993;265:H1744–H1749.
52.
Richard DE, Berra
E, Pouyssegur J. Nonhypoxic pathway mediates the induction of
hypoxia-inducible factor 1 in vascular smooth muscle cells.
J Biol Chem. 2000;275:26765–26771.
53.
Tsopanoglou NE,
Maragoudakis ME. On the mechanism of thrombin-induced angiogenesis:
potentiation of vascular endothelial growth factor
activity on endothelial cells by up-regulation of its
receptors. J Biol Chem. 1999;274:23969–23976.
54. McNamara CA, Sarembock IJ, Gimple LW, Fenton JWI, Coughlin SR, Owens GK. Thrombin stimulates proliferation of cultured rat aortic smooth muscle cells by a proteolytically activated receptor. J Clin Invest. 1993;91:94–98.
55.
Kranzhofer R,
Clinton SK, Ishii K, Coughlin SR, Fenton JW 2nd, Libby P. Thrombin
potently stimulates cytokine production in human
vascular smooth muscle cells but not in mononuclear phagocytes.
Circ Res. 1996;79:286–294.
56. Dabbagh K, Laurent GJ, McAnulty RJ, Chambers RC. Thrombin stimulates smooth muscle cell procollagen synthesis and mRNA levels via a PAR-1 mediated mechanism. Thromb Haemost. 1998;79:405–409.[Medline] [Order article via Infotrieve]
57.
Noda-Heiny H,
Sobel BE. Vascular smooth muscle cell migration mediated by thrombin
and urokinase receptor. Am J
Physiol. 1995;268:C1195–C1201.
58.
Zhou M, Lambert
H, Landry J. Transient activation of a distinct serine protein kinase
is responsible for 27-kDa heat shock protein
phosphorylation in mitogen-stimulated and heat-shocked
cells. J Biol Chem. 1993;268:35–43.
59.
Madamanchi NR, Li
S, Patterson C, Runge MS. Reactive oxygen species regulate heat shock
protein70 via the JAK-STAT pathway.
Arterioscler Thromb Vasc Biol. 2001;21:321–326.
60.
Johnson AD,
Berberian PA, Tytell M, Bond MG. Differential distribution of 70-dD
heat shock protein in atherosclerosis: its potential
role in arterial SMC survival.
Arterioscler Thromb Vasc Biol. 1995;15:27–36.
61.
Flynn PD, Byrne
CD, Baglin TP, Weissberg PL, Bennett MR. Thrombin generation by
apoptotic vascular smooth muscle cells.
Blood. 1997;89:4378–4384.
62.
Perlman H,
Maillard L, Krasinski K, Walsh K. Evidence for the rapid onset of
apoptosis in medial smooth muscle cells after balloon injury.
Circulation. 1997;95:981–987.
63.
Jones PL, Crack
J, Rabinovitch M. Regulation of tenascin-C, a vascular
smooth muscle cell survival factor that interacts with the vß3
integrin to promote epidermal growth factor receptor
phosphorylation and growth.
J Cell Biol. 1997;139:279–293.
64. Baker AH, Zaltsman AB, George SJ, Newby AC. Divergent effects of tissue inhibitor of metalloproteinase-1, -2, or -3 overexpression on rat vascular smooth muscle cell invasion, proliferation, and death in vitro: TIMP-3 promotes apoptosis. J Clin Invest. 1998;101:1478–1487.[Medline] [Order article via Infotrieve]
65.
Walsh K, Smith
RC, Kim HS. Vascular cell apoptosis in remodeling,
restenosis, and plaque rupture.
Circ Res. 2000;87:184–188.
66. Harker LA, Hanson SR, Runge MS. Thrombin hypothesis of thrombus generation and vascular lesion formation. Am J Cardiol. 1995;75:12B–17B.[Medline] [Order article via Infotrieve]
67. Eidt JF, Allison P, Noble S, Ashton J, Golino P, McNatt J, Buja LM, Willerson JT. Thrombin is an important mediator of platelet aggregation in stenosed canine coronary arteries with endothelial injury. J Clin Invest. 1989;84:18–27.
68. Hatton MW, Moar SL, Richardson M. Deendothelialization in vivo initiates a thrombogenic reaction at the rabbit aorta surface: correlation of uptake of fibrinogen and antithrombin III with thrombin generation by the exposed subendothelium. Am J Pathol. 1989;135:499–508.[Abstract]
69.
Barry WL, Gimple
LW, Humphries JE, Powers ER, McCoy KW, Sanders JM, Owens GK, Sarembock
IJ. Arterial thrombin activity after angioplasty in an
atherosclerotic rabbit model: time course and effect of hirudin.
Circulation. 1996;94:88–93.
70. Smith EB, Crosbie L, Carey S. Prothrombin-related antigens in human aortic intima. Semin Thromb Hemost. 1996;22:347–350.[Medline] [Order article via Infotrieve]
71.
Wilcox JN,
Rodriguez J, Subramanian R, Ollerenshaw J, Zhong C, Hayzer DJ, Horaist
C, Hanson SR, Lumsden A, Salam TA, Kelly AB, Harker LA, Runge MS.
Characterization of thrombin receptor expression during vascular lesion
formation. Circ Res. 1994;75:1029–1038.
72. Nelken NA, Soifer SJ, O’Keefe J, Vu T-K, Charo I, Coughlin S. Thrombin receptor expression in normal and atherosclerotic human arteries. J Clin Invest. 1992;90:1614–1621.
73.
van Meijer M,
Smilde A, Tans G, Nesheim ME, Pannekoek H, Horrevoets AJ. The
suicide substrate reaction between plasminogen
activator inhibitor 1 and thrombin is regulated
by the cofactors vitronectin and heparin.
Blood. 1997;90:1874–1882.
74.
Lupu F, Heim D,
Bachmann F, Kruithof EK. Expression of LDL receptor-related
protein/2-macroglobulin receptor in human normal and atherosclerotic
arteries. Arterioscler Thromb. 1994;14:1438–1444.
75.
Stefansson S,
Lawrence DA, Argraves WS. Plasminogen activator
inhibitor-1 and vitronectin promote the
cellular clearance of thrombin by low density lipoprotein
receptor-related proteins 1 and 2. J
Biol Chem. 1996;271:8215–8220.
76.
Stoop AA, Lupu F,
Pannekoek H. Colocalization of thrombin, PAI-1, and
vitronectin in the atherosclerotic vessel wall: a potential
regulatory mechanism of thrombin activity by
PAI-1/vitronectin complexes.
Arterioscler Thromb Vasc Biol. 2000;20:1143–1149.
77.
Carmeliet P,
Moons L, Lijnen R, Janssens S, Lupu F, Collen D, Gerard RD.
Inhibitory role of plasminogen
activator inhibitor-1 in arterial
wound healing and neointima formation: a gene targeting and
gene transfer study in mice.
Circulation. 1997;96:3180–3191.
78. Yamanaga K, Yuuki T, Tsukada M, Koshiba H, Nakajima T, Takechi K, Nakamura N. Heparin cofactor II inhibits thrombus formation in a rat thrombosis model. Thromb Res. 2000;98:95–101.[Medline] [Order article via Infotrieve]
79.
Liaw PC, Austin
RC, Fredenburgh JC, Stafford AR, Weitz JI. Comparison of heparin- and
dermatan sulfate-mediated catalysis of thrombin inactivation by heparin
cofactor II. J Biol Chem. 1999;274:27597–27604.
80.
Shirk RA,
Parthasarathy N, San Antonio JD, Church FC, Wagner WD. Altered dermatan
sulfate structure and reduced heparin cofactor II-stimulating activity
of biglycan and decorin from human atherosclerotic plaque.
J Biol Chem. 2000;275:18085–18092.
81. Abendschein DR, Recchia D, Meng YY, Oltrona L, Wickline SA, Eisenberg PR. Inhibition of thrombin attenuates stenosis after arterial injury in minipigs. J Am Coll Cardiol. 1996;28:1849–1855.[Abstract]
82.
Sarembock IJ,
Gertz SD, Gimple LW, Owen RM, Powers ER, Roberts WC. Effectiveness of
recombinant desulphatohirudin in reducing restenosis after
balloon angioplasty of atherosclerotic femoral arteries in rabbits.
Circulation. 1991;84:232–243.
83.
Gerdes C,
Faber-Steinfeld V, Yalkinoglu O, Wohlfeil S. Comparison of the effects
of the thrombin inhibitor r-hirudin in four animal models
of neointima formation after arterial injury.
Arterioscler Thromb Vasc Biol. 1996;16:1306–1311.
84. Rade JJ, Schulick AH, Virmani R, Dichek DA. Local adenoviral-mediated expression of recombinant hirudin reduces neointima formation after arterial injury. Nature Med. 1996;2:293–298.[Medline] [Order article via Infotrieve]
85.
Cook JJ, Sitko
GR, Bednar B, Condra C, Mellott MJ, Feng DM, Nutt RF, Shafer JA, Gould
RJ, Connolly TM. An antibody against the exosite of the cloned thrombin
receptor inhibits experimental arterial thrombosis in the
African green monkey.
Circulation. 1995;91:2961–2971.
86.
Takada M, Tanaka
H, Yamada T, Ito O, Kogushi M, Yanagimachi M, Kawamura T, Musha T,
Yoshida F, Ito M, Kobayashi H, Yoshitake S, Saito I. Antibody to
thrombin receptor inhibits neointimal smooth muscle cell
accumulation without causing inhibition of platelet aggregation or
altering hemostatic parameters after angioplasty in rat.
Circ Res. 1998;82:980–987.
87.
Cheung WM,
D’Andrea MR, Andrade-Gordon P, Damiano BP. Altered vascular injury
responses in mice deficient in protease-activated receptor-1.
Arterioscler Thromb Vasc Biol. 1999;19:3014–3024.
88. Burchenal JE, Marks DS, Tift Mann J, Schweiger MJ, Rothman MT, Ganz P, Adelman B, Bittl JA. Effect of direct thrombin inhibition with Bivalirudin (Hirulog) on restenosis after coronary angioplasty. Am J Cardiol. 1998;82:511–515.[Medline] [Order article via Infotrieve]
89.
Serruys PW,
Herrman JP, Simon R, Rutsch W, Bode C, Laarman GJ, van Dijk R, van den
Bos AA, Umans VA, Fox KA. A comparison of hirudin with heparin in the
prevention of restenosis after coronary angioplasty:
Helvetica Investigators. N Engl
J Med. 1995;333:757–763.
90. Zacharski LR, Memoli VA, Morain WD, Schlaeppi JM, Rousseau SM. Cellular localization of enzymatically active thrombin in intact human tissues by hirudin binding. Thromb Haemost. 1995;73:793–797.[Medline] [Order article via Infotrieve]
91.
Haralabopoulos
GC, Grant DS, Kleinman HK, Maragoudakis ME. Thrombin promotes
endothelial cell alignment in Matrigel in vitro and
angiogenesis in vivo. Am J
Physiol. 1997;273:C239–C245.
92.
Tsopanoglou NE,
Pipili-Synetos E, Maragoudakis ME. Thrombin promotes angiogenesis by a
mechanism independent of fibrin formation.
Am J Physiol. 1993;264:C1302–C1307.
93.
Mohle R, Green D,
Moore MA, Nachman RL, Rafii S. Constitutive production and
thrombin-induced release of vascular endothelial growth
factor by human megakaryocytes and platelets.
Proc Natl Acad Sci
U S A. 1997;94:663–668.
94. Zucker S, Mirza H, Conner CE, Lorenz AF, Drews MH, Bahou WF, Jesty J. Vascular endothelial growth factor induces tissue factor and matrix metalloproteinase production in endothelial cells: conversion of prothrombin to thrombin results in progelatinase A activation and cell proliferation. Int J Cancer. 1998;75:780–786.[Medline] [Order article via Infotrieve]
95.
Duhamel-Clerin E,
Orvain C, Lanza F, Cazenave JP, Klein-Soyer C. Thrombin
receptor-mediated increase of two matrix metalloproteinases, MMP-1 and
MMP-3, in human endothelial cells.
Arterioscler Thromb Vasc Biol. 1997;17:1931–1938.
96.
O’Reilly MS,
Pirie-Shepherd S, Lane WS, Folkman J. Antiangiogenic activity of the
cleaved conformation of the serpin antithrombin.
Science. 1999;285:1926–1928.
97.
Lee TH, Rhim T,
Kim SS. Prothrombin kringle-2 domain has a growth
inhibitory activity against basic fibroblast growth
factor-stimulated capillary endothelial cells.
J Biol Chem. 1998;273:28805–28812.
98.
Sun WY, Witte DP,
Degen JL, Colbert MC, Burkart MC, Holmback K, Xiao Q, Bugge TH, Degen
SJ. Prothrombin deficiency results in embryonic and neonatal lethality
in mice. Proc Natl Acad Sci
U S A. 1998;95:7597–7602.
99.
Xue J, Wu Q,
Westfield LA, Tuley EA, Lu D, Zhang Q, Shim K, Zheng X, Sadler JE.
Incomplete embryonic lethality and fatal neonatal hemorrhage
caused by prothrombin deficiency in mice.
Proc Natl Acad Sci
U S A. 1998;95:7603–7607.
100. Connolly AJ, Ishihara H, Kahn ML, Farese RV, Coughlin SR. Role of the thrombin receptor in development and evidence for a second receptor. Nature. 1996;381:516–519.[Medline] [Order article via Infotrieve]
101. Darrow AL, Fung-Leung WP, Ye RD, Santulli RJ, Cheung WM, Derian CK, Burns CL, Damiano BP, Zhou L, Keenan CM, Peterson PA, Andrade-Gordon P. Biological consequences of thrombin receptor deficiency in mice. Thromb Haemost. 1996;76:860–866.[Medline] [Order article via Infotrieve]
102.
Suh TT,
Holmback K, Jensen NJ, Daugherty CC, Small K, Simon DI, Potter S, Degen
JL. Resolution of spontaneous bleeding events but failure of pregnancy
in fibrinogen-deficient mice. Genes
Dev. 1995;9:2020–2033.
103. Shivdasani RA, Rosenblatt MF, Zucker-Franklin D, Jackson CW, Hunt P, Saris CJ, Orkin SH. Transcription factor NF-E2 is required for platelet formation independent of the actions of thrombopoietin/MGDF in megakaryocyte development. Cell. 1995;81:695–704.[Medline] [Order article via Infotrieve]
104. Carmeliet P, Ferriera V, Breier G, Pollefeyt S, Nieckens L, Gertsenstein M, Fahrig M, Vandenhoeck A, Harpal K, Eberhardt C, Declercq C, Pawling J, Moons L, Collen D, Risau W, Nagy A. Abnormal blood vessel development and lethality in embryos lacking a single VEGF allele. Nature. 1996;380:435–439.[Medline] [Order article via Infotrieve]
105. Shalaby F, Rossant J, Yamaguchi TP, Gertsenstein M, Wu X-F, Breitman M, Schuh A. Failure of blood-island formation and vasculogenesis in Flk-1-deficient mice. Nature. 1995;376:62–66.[Medline] [Order article via Infotrieve]
106. Damiano BP, D’Andrea MR, de Garavilla L, Cheung WM, Andrade-Gordon P. Increased expression of protease activated receptor-2 (PAR-2) in balloon-injured rat carotid artery. Thromb Haemost. 1999;81:808–814.[Medline] [Order article via Infotrieve]
107.
D’Andrea MR,
Derian CK, Leturcq D, Baker SM, Brunmark A, Ling P, Darrow AL, Santulli
RJ, Brass LF, Andrade-Gordon P. Characterization of
protease-activated receptor-2 immunoreactivity in normal human
tissues. J Histochem
Cytochem. 1998;46:157–164.
108.
Molino M,
Raghunath PN, Kuo A, Ahuja M, Hoxie JA, Brass LF, Barnathan ES.
Differential expression of functional protease-activated
receptor-2 (PAR-2) in human vascular smooth muscle cells.
Arterioscler Thromb Vasc Biol. 1998;18:825–832.
109.
Schmidt VA,
Nierman WC, Maglott DR, Cupit LD, Moskowitz KA, Wainer JA, Bahou WF.
The human proteinase-activated receptor-3 (PAR-3) gene:
identification within a Par gene cluster and characterization in
vascular endothelial cells and platelets.
J Biol Chem. 1998;273:15061–15068.
110. Hollenberg MD, Saifeddine M, Al-Ani B, Gui Y. Proteinase-activated receptor 4 (PAR4): action of PAR4-activating peptides in vascular and gastric tissue and lack of cross-reactivity with PAR1 and PAR2. Can J Physiol Pharmacol. 1999;77:458–464.[Medline] [Order article via Infotrieve]
111. Kanthou C, Parry G, Wijelath E, Kakkar VV, Demoliou-Mason C. Thrombin-induced proliferation and expression of platelet-derived growth factor-A chain gene in human vascular smooth muscle cells. FEBS Lett. 1992;314:143–148.[Medline] [Order article via Infotrieve]
112.
Okazaki H,
Majesky MW, Harker LA, Schwartz SM. Regulation of platelet-derived
growth factor ligand and receptor gene expression by -thrombin in
vascular smooth muscle cells. Circ
Res. 1992;71:1285–1293.
113.
Stouffer GA,
Sarembock IJ, McNamara CA, Gimple LW, Owens GK. Thrombin-induced
mitogenesis of vascular SMC is partially mediated by autocrine
production of PDGF-AA. Am J
Physiol. 1993;265:C806–C811.
114.
Weiss RH,
Maduri M. The mitogenic effect of thrombin in vascular
smooth muscle cells is largely due to basic fibroblast growth factor.
J Biol Chem. 1993;268:5724–5727.
115.
Taylor DS,
Cheng X, Pawlowski JE, Wallace AR, Ferrer P, Molloy CJ. Epiregulin is a
potent vascular smooth muscle cell-derived mitogen induced by
angiotensin II, endothelin-1, and thrombin.
Proc Natl Acad Sci
U S A. 1999;96:1633–1638.
116. Delafontaine P, Anwar A, Lou H, Ku L, G-protein coupled and tyrosine kinase receptors: evidence that activation of the insulin-like growth factor I receptor is required for thrombin-induced mitogenesis of rat aortic smooth muscle cells. J Clin Invest. 1996;97:139–145.[Medline] [Order article via Infotrieve]
117.
Schieffer B,
Drexler H, Ling BN, Marrero MB. G protein-coupled receptors control
vascular smooth muscle cell proliferation via pp60c-src and p21ras.
Am J Physiol. 1997;272:C2019–C2030.
118.
Hoshi S, Goto
M, Koyama N, Nomoto K, Tanaka H. Regulation of vascular smooth muscle
cell proliferation by nuclear factor-
B and its
inhibitor, I-B. J Biol
Chem. 2000;275:883–889.
119.
Peppel K,
Jacobson A, Huang X, Murray JP, Oppermann M, Freedman NJ.
Overexpression of G protein-coupled receptor kinase-2 in smooth muscle
cells attenuates mitogenic signaling via G protein-coupled
and platelet-derived growth factor receptors.
Circulation. 2000;102:793–799.
120.
Rao GN, Runge
MS. Cyclic AMP inhibition of thrombin-induced growth in vascular smooth
muscle cells correlates with decreased JNK1 activity and c-Jun
expression. J Biol Chem. 1996;271:20805–20810.
121. Walters TK, Gorog DA, Wood RF. Thrombin generation following arterial injury is a critical initiating event in the pathogenesis of the proliferative stages of the atherosclerotic process. J Vasc Res. 1994;31:173–177.[Medline] [Order article via Infotrieve]
122.
Bittl JA,
Strony J, Brinker JA, Ahmed WH, Meckel CR, Chaitman BR, Maraganore J,
Deutsch E, Adelman B. Treatment with bivalirudin (Hirulog) as compared
with heparin during coronary angioplasty for unstable or
postinfarction angina: Hirulog Angioplasty Study Investigators.
N Engl J Med. 1995;333:764–769.
123.
Jang Y, Guzman
LA, Lincoff AM, Gottsauner-Wolf M, Forudi F, Hart CE, Courtman DW,
Ezban M, Ellis SG, Topol EJ. Influence of blockade at specific
levels of the coagulation cascade on restenosis in a rabbit
atherosclerotic femoral artery injury model.
Circulation. 1995;92:3041–3050.
124. Sarembock IJ, Gertz SD, Thome LM, McCoy KW, Ragosta M, Powers ER, Maraganore JM, Gimple LW. Effectiveness of hirulog in reducing restenosis after balloon angioplasty of atherosclerotic femoral arteries in rabbits. J Vasc Res. 1996;33:308–3014.[Medline] [Order article via Infotrieve]
125. Buchwald AB, Hammerschmidt S, Stevens J, Goring J, Nebendahl K, Unterberg C. Inhibition of neointimal proliferation after coronary angioplasty by low-molecular-weight heparin (clivarine) and polyethyleneglycol-hirudin. J Cardiovasc Pharmacol. 1996;28:481–487.[Medline] [Order article via Infotrieve]
126. Hadoke PW, Wadsworth RM, Wainwright CL, Butler KD, Giddings MJ. Subcutaneous infusion of r-hirudin does not inhibit neointimal proliferation after angioplasty of the subclavian artery in cholesterol-fed rabbits. Coron Artery Dis. 1996;7:599–608.[Medline] [Order article via Infotrieve]
127. Barry WL, Wiegman PJ, Gimple LW, Gertz SD, Powers ER, Owens GK, Sarembock IJ. A new single-injury model of balloon angioplasty in cholesterol-fed rabbits: beneficial effect of hirudin and comparison with double-injury model. Lab Invest. 1997;77:109–116.[Medline] [Order article via Infotrieve]
128.
Thome LM,
Gimple LW, Bachhuber BG, McNamara CA, Ragosta M, Gertz SD, Powers ER,
Owens GK, Humphries JE, Sarembock IJ. Early plus delayed hirudin
reduces restenosis in the atherosclerotic rabbit more than
early administration alone: potential implications for dosing of
antithrombin agents. Circulation. 1998;98:2301–2306.
129.
Gallo R,
Padurean A, Toschi V, Bichler J, Fallon JT, Chesebro JH, Fuster V,
Badimon JJ. Prolonged thrombin inhibition reduces restenosis
after balloon angioplasty in porcine coronary arteries. Circulation. 1998;97:581–588.
130. Bishop GG, Wiegman P, McNamara C, Din S, Sanders J, Hesselbacher S, Feldman M, McPherson JA, Humphries JE, Hammarskjold ML, Gimple LW, Ragosta M, Powers ER, Dickek D, Owens GK, Sarembock IJ. Local adenovirus-mediated delivery of hirudin in a rabbit arterial injury model. J Vasc Res. 1999;36:343–352.[Medline] [Order article via Infotrieve]
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