Clinical Biochemistry of Domestic Animals (Sixth Edition) - UMK ...

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Chapter | 10 Hemostasis
However, when the vessel wall is damaged, thrombin is so
inefficiently inhibited by AT, the conversion of fibrinogen
to fibrin and hence clot formation can proceed ( Pike et al .,
2005 ).
Heparin cofactor II (HCII) is a specific thrombin inhibitor
that has a similar mechanism of action as AT, but it
is dissimilar both antigenically and in its cofactor requirements
( Johnstone, 2000 ). HCII appears to be synthesized
exclusively by the liver as a 65.6-kDa glycoprotein with
a unique amino-terminal extension that contains two tandem
repeats rich in acidic amino acids with two sulfated
tyrosines ( Inhorn and Tollefsen, 1986 ). These repeats are
homologous to the carboxyl-terminal sequence of hirudin,
the specific thrombin inhibitor from the medicinal
leech. Although heparin, heparan sulfate, and dermatan
sulfate are all physiological activators of HCII, many different
polyanions, including polyphosphates, polysulfates,
and polycarboxylates, are also able to accelerate HCII
inhibition of thrombin up to 1000-fold ( Tollefsen, 2004 ).
Although the physiological function of HCII has not been
fully elucidated, it has been suggested that it may function
as a thrombin-inhibitor reserve when AT levels become
subnormal such as occurs in DIC (see Section IV.B.4;
Tran and Duckert, 1984). Experimental animal studies and
human clinical studies indicate that HCII may have a protective
antithrombotic effect following arterial oxidative
damage ( Tollefsen, 2004 ).
As summarized in Table 10-6 , a number of other serpins
participate in modulating hemostasis ( Silverman et al .,
2001 ). The broad spectrum protease inhibitor known
both as α 1 -antitrypsin and α 1 -protease inhibitor ( α 1 -PI)
can inhibit thrombin, FXa and FXIa, APC, and plasmin,
although its main physiological function is as a circulating
antitryspin inhibitor. However, α 1 -PI appears to be a more
effective inhibitor of FXIa than AT ( Scott et al ., 1982 ). It
circulates as a 50-kDa glycoprotein and contains the characteristic
reactive site loop of a serpin ( Long et al ., 1984 ).
α 2 -Macroglobulin circulates as a large 650-kDa glycoprotein
composed of four identical subunits. This protein
can bind one or two molecules of a protease depending on
its size. Small proteases such as trypsin maximally form
2:1 complexes with α 2 -M, whereas larger proteases such
as plasmin form 1:1 complexes. It has been suggested
that α 2 -M is one of the major physiological inhibitors of
FXa in vivo ( Fuchs and Pizzo, 1983 ). α 2 -M-protease complexes
are rapidly removed from the circulation by the liver
through interaction with the lipoprotein receptor-related
protein that serves as a large multifunctional endocytosis
receptor ( Narita et al. , 1998 ). This receptor protein is
also responsible for the removal of other proteases such
as tissue-type plasminogen activator and thrombin activatable
fibrinolysis inhibitor (TAFI) ( Orth et al. , 1992 ;
Warshawsky et al. , 1994 ).
C1 inhibitor is a 104-kDa single-chain glycoprotein
that exhibits structural homology with other members of
the serpin superfamily. It appears to be the main circulating
inhibitor of FXIa, although it can also suppress the activity
of FXIIa and kallikrein but not FXa or plasmin ( Harpel
et al ., 1985 ; Pixley et al ., 1985 ). As its name suggests, the
main physiological role of C1 inhibitor is to modulate the
complement system.
Protein C inhibitor (PCI), also known as plasminogen
activator inhibitor-3, inactivates the proteolytic activity of
numerous serine proteases including thrombin, FXa, FXIa,
APC, tissue-type plasminogen (tPA) activator, and urokinase-type
plasminogen activator (uPA) ( Meijers et al .,
2002 ). The mature PCI protein is 57 kDa and is found in
several body fluids as well as plasma ( Pike et al ., 2005 ).
Like HCII, several GAGs and polyanions can enhance
the activity of PCI. In the presence of heparin, a ternary
complex is formed between PCI and the active protease
through a GAG binding region localized on the H-helix
region of PCI ( Pratt and Church, 1992 ). However, even in
the presence of heparin, the inhibitory effects of PCI for
thrombin and FXa are still modest compared to those of
AT and HCII ( Pike et al ., 2005 ). Although PCI can inhibit
APC in the presence of heparin and calcium ions, the
more physiologically relevant role of PCI in modulating
the protein C anticoagulant system is the result of its ability
to inhibit the activity of the thrombin-thrombomodulin
(T-TM) complex ( Yang et al ., 2003 ). As described later, this
complex is key to the conversion of the inactive zymogen
protein C to its active form.
c. Protein C Anticoagulant System
The protein C anticoagulant system regulates thrombin formation
by modulating the activity of the cofactors FVIIIa
and FVa ( Dahlback, 1995 ). This inhibitory system is composed
of several components: protein C (PC), thrombomodulin
(TM), endothelial cell protein C receptor (EPCR),
and protein S ( Table 10-6 ). Protein C, the key component,
is a vitamin K-dependent, 62-kDa glycoprotein that is synthesized
by the liver and circulates as a biologically inactive
molecule consisting of a 40-kDa heavy chain and a
20-kDa light chain joined by a disulfide bond ( Stenflo and
Fernlund, 1982 ). As occurs with other vitamin K-dependent
proteins, PC lacks biological activity unless it has
undergone vitamin K-dependent posttranslational carboxylation
of glutamine residues in the N-terminal region of
the molecule. Thrombin cleavage of this N-terminal region
converts PC to its proteolytic active form, APC ( Esmon,
2003 ). For thrombin to activate PC at a physiologically relevant
rate, it requires the presence of the cofactor thrombomodulin
(TM) so that T-TM complexes can form. TM is
a 58.7-kDa transmembrane glycoprotein that is present on
vascular endothelial cells with the highest concentrations
being present in microvascular beds ( Esmon, 2003 ). In
the presence of TM, the rate of PC activation is increased
by more than 1000-fold ( Van de Wouwer et al ., 2004 ).
The TM molecule is organized into five distinct domains.