Page 2156 - Hematology_ Basic Principles and Practice ( PDFDrive )

Chapter 127 Regulatory Mechanisms in Hemostasis 1909

26
of the human orthologue. 18,19 The components of the tenase and a conformational change (snap) that crushes the protease. AT
prothrombinase complexes are similar in sequence and structure, circulates at about 2.3 µM and acts to mop up lumenal thrombin
and it has always been assumed that they assemble in an analogous and factor Xa. A fraction of AT is associated with heparan sulfate on
manner. However, how the very different substrates of factor X vascular endothelial cells, where it helps to confer an anticoagulant
and prothrombin bind to their respective complexes has yet to be environment to intact vessels. Heparan sulfate binding also activates
resolved. AT roughly 1000-fold toward its main targets, thrombin, factor Xa,
and factor IXa. This is indeed how the structurally related GAG
heparin asserts its therapeutic anticoagulant effect. About one-third
THE REGULATORS OF COAGULATION of the chains of medicinal heparin (much less for heparan sulfate)
contain a pentasaccharide sequence that binds AT with high affinity
ADAMTS13 and induces a conformational change in AT. Low-molecular-weight
heparins, including the synthetic pentasaccharide fondaparinux,
Platelet adhesion is an early event in hemostasis and is initiated pri- accelerate inhibition of factor IXa and factor Xa, but have almost no
marily through vWF, which acts as a molecular bridge between effect on thrombin inhibition. This is because thrombin is insensitive
exposed collagen and the GPIb/IX/V receptor on the platelet mem- to the conformational change in AT conferred by heparin binding,
20
brane. vWF, a multidomain protein that forms large disulfide-linked and instead requires long chains composed of at least 18 saccharide
27
multimers, is secreted from endothelial cells in a latent form that is units to “bridge” AT to thrombin. Factors IXa and Xa can also be
unable to bind platelets. Binding to exposed collagen via the A3 bridged, but this is a secondary effect to the allosteric activation by
domain in environments with high shear stress causes vWF to par- heparins, and requires longer heparin chains than those needed for
28
tially unfold, exposing the binding site for platelet glycoprotein thrombin (36 or more saccharide units in length). Other serpins
21
receptor GPIbα, which is located in the A1 domain (Fig. 127.3A). play relatively minor roles in regulating clotting proteases, including
The ease of unfolding, and therefore the procoagulant activity, is heparin cofactor II (HCII or SERPIND1), which inhibits thrombin
related to the size of the multimers, and this in turn is regulated by and is activated by dermatan sulfate and heparin; protease nexin-1
the multidomain metalloprotease ADAMTS13 (vWF-cleaving pro- (PN-1 or SERPINE2), which is a specific thrombin inhibitor and is
tease). Although ADAMTS13 circulates in a constitutively active only found on cell surfaces bound to GAGs; and protein C inhibitor
state, its ability to cleave vWF is dependent on the unfolding of the (PCI or SERPINA5), which is a promiscuous inhibitor of coagulation
A2 domain that harbors the scissile bond, Tyr 1605 –Met 1606 . Cleavage proteases and can be activated by GAGs to inhibit thrombin, factor
29
of ultralarge multimers of vWF by ADAMTS13 reduces the capacity Xa, factor XIa and the TF–factor VIIa complex. The structures
of vWF to initiate platelet adhesion and spontaneous platelet aggrega- of most of these recognition complexes have been solved by x-ray
tion. The importance of this regulatory mechanism is highlighted by crystallography.
the microvascular thrombosis that characterizes thrombotic throm-
bocytopenic purpura (TTP), a disorder associated with deficiency of
ADAMTS13. 22 Protein C Pathway
Protein C is activated by thrombin bound to thrombomodulin on
Tissue Factor Pathway Inhibitor the surface of endothelial cells. Activation is more efficient when
30
protein C binds to the endothelial cell protein C receptor (EPCR)
23
31
TFPI is an endogenous inhibitor of the extrinsic Xase complex. It via its Gla domain. Activated protein C (APC) is a physiologic
is expressed by endothelial cells in α and β isoforms generated by anticoagulant that down-regulates thrombin generation by cleaving
alternative splicing of its premessenger RNA (Fig. 127.3B). The α and inactivating factor Va and factor VIIIa, the cofactors of the
form is composed of three Kunitz domains (K1, K2, K3) followed prothrombinase and intrinsic Xase complexes, respectively; protein
by an unstructured basic C-terminal region. The first and second S acts as a cofactor for APC-mediated inactivation of factor Va
Kunitz domains inhibit factor VIIa and factor Xa, respectively. and factor VIIIa by helping target APC to the negatively charged
32
Inhibition is thought to be a two-step process with binding of factor surface of activated platelets. Deficiency in protein C results in
Xa to K2 followed by binding of K1 to factor VIIa-TF. The third thrombophilia and its complete absence is associated with purpura
33
Kunitz domain does not possess inhibitory activity but binds to fulminans. The most common thrombophilic mutation is the
protein S, and the basic C-terminus binds to cell surface GAGs. Leiden mutation in factor V that reduces the rate of factor Va inacti-
34
The β-isoform consists of the first and second Kunitz domains and vation by APC. Conversely, excessive APC activity is associated with
a glycophosphatidylinositol (GPI)-anchor on the C-terminus that bleeding. 35
tethers it to the surface of the endothelial cell that produced it.
Endothelial cells secrete both α- and β-isoforms, but platelets only
produce the soluble α isoform. The basic C-terminal tail of the α Fibrinolysis
isoform has been shown to bind to an acidic stretch on factor Va
that has been activated by factor Xa and on the partially activated Fibrin is a key component of the hemostatic clot and is the primary
24
factor V that is released from platelets. However, that acidic stretch target for plasmin, the effector protease of the fibrinolytic system.
is missing in thrombin-activated factor Va, so the relevance of this During healing of an injured vessel, the thrombus is lysed by plasmin.
interaction in normal clotting is unclear. Protein S binding to the Plasmin is generated by plasminogen activators, principally tissue
K3 domain helps to localize TFPIα to cell surfaces and thus effects plasminogen activator (tPA). It is essential that tPA and plasmin
36
a cofactor activity, enhancing the rate of inhibition of factor Xa by activity are regulated to prevent hyperfibrinolysis. The principle
90-fold. 25 inhibitors of these enzymes are the serpins plasminogen activator
inhibitor 1 (PAI-1) and antiplasmin. In addition, during hemostasis
thrombin down-regulates fibrinolytic activity through activation of
Serpins TAFI. However, hyperfibrinolysis may occur if there is excessive tPA
or plasmin. Dysregulation and consequent hyperfibrinolysis occur in
The principal inhibitor of the coagulation proteases is the serpin, AT, disseminated intravascular coagulation (DIC), liver disease, nephrotic
11
also known as SERPINC1 and previously as ATIII. Like all other syndrome and with some metastatic tumors. Very rarely, heritable
members of the serpin family of protease inhibitors, AT utilizes a deficiencies of PAI-1 or antiplasmin produce hyperfibrinolysis and a
mousetrap-like mechanism whereby the protease (mouse) takes a bleeding tendency. In addition, normal fibrinolysis may contribute
bite of the reactive center loop (cheese), and before the protease to bleeding when there is inadequate thrombin generation, as in
can disengage, the serpin (trap) releases its stored energy through hemophilia. 36