What is hemostasis?
Hemostasis is a balance of two opposing forces: clot formation
and dissolution. Appropriate clotting is essential at the site
of a wound in order to maintain hemostasis, however clotting away
from the site of a wound must be minimized in order to prevent
life-threatening thrombotic events. Regulation of clotting involves
a series of zymogen conversions with extensive use of co-factors.
Each stage involves the conversion of a precursor to an active
form, which then is involved in the activation of a factor in
the next stage of the cascade. The ultimate goal is the activation
of the protease thrombin with the subsequent conversion of fibrinogen
to fibrin for use in clot formation.
Effects Of Venoms Upon Hemostasis
Venoms often have profound effects upon blood coagulation,
acting directly upon important clotting factors either by inappropriate
activation or through prevention of activation. The same net effect
(i.e. inability to stop bleeding) may be produced by dramatically
different mechanisms allowing for the selective use of venom or
venom components to address a specific deficiency in blood chemistry.
As blood coagulation therapies or diagnostic tools, the most important
snake venom components have been homeostatic or antithrombotic
agents.
Russell's Viper Venom
The use of snake venoms as homeostatic agents is based upon
early observations of the potent coagulative properties of snake
venoms, the Russell's viper (Daboia russelli) in particular
gaining early use as a treatment of hemophilia. The mechanism
of action of D. russelli venom is in the activation of
the factors V, X, IX, as well as Protein C. While D. russelli
venom is no longer used as a therapeutic agent, it still is used
as a diagnostic agent to determine deficiencies in clotting factor
X.
Prothrombin Activators
In addition to measurement of factor X by D. russelli venom,
other blood chemistry research or diagnostic uses of snake venoms
utilizes the enzymes affecting the coagulation cascade, such as
prothrombin activators to measure factor V levels in patients'
blood. Prothrombin activators found in snake venoms can be divided
into four groups:
Group I convert prothrombin to meizothrombin with activity insensitive
to the presence of the non-enzymatic prothrombinase complex cofactors
(CaCL2, factor V and phospholipid)
Group II and III activators are able to cleave both peptide bonds
in prothrombin essential for the conversion of prothrombin to
thrombin, the difference between the II and III being that the
converting activity of II is strongly stimulated by phospholipids
and factor Va in the presence of calcium while III is only stimulated
by CaCl2 and phospholipid
Group IV activators are proteases that cleave prothrombin
into non-active precursor forms of thrombin rather than converting
prothrombin into the enzymatically active products.
Group I prothrombin activators such as ecarin from the saw-scaled viper (Echis carinatus) are excellent hospital laboratory tools in the analysis of blood from patients with liver diseases or being treated with anticoagulants such as vitamin K antagonists. The usefulness of this enzyme lies in its ability, unlike factor Xa, to act independently of any co-factor, including factor V and to catalyze intermediates of both carboxylated and acarboxylated prothrombin, with the levels of acarboxylated prothrombin being abnormally present in patients with disease states such as liver disease. Thus, even if factor V levels are also affected, the amount of prothrombin present can still be measured.
Since factor V is a necessary co-factor in the conversion of prothrombin to thrombin, the amount of thrombin generated upon the addition of Group II venom prothrombin activating enzymes is directly proportional to the level of factor V present in the blood. Therefore, Group II prothrombin activators are useful for accurate measurement of factor V. Ecarin has another important diagnostic use in conjunction with textarin, a group II prothrombin activator from the brown snake Pseudonaja textilis.
Together these components are a specific and highly sensitive
test for the lupus anticoagulant based on the relative presence
or absence of specific lipids and other co-factors. Since ecarin
is a co-factor independent prothrombin activator while textarin
is co-factor dependent, the relative of clotting time ratios are
useful as diagnostic evidence of the presence of the anti-phospholipid
lupus anticoagulant antibodies. As the level of lupus antibodies
increase, the amount of circulating phospholipid proportionally
decreases resulting in greatly increased clotting times. The co-factor
independent ecarin is used to provide a base-line measure of the
amount of prothrombin present in the serum, thus increasing the
diagnostic evidence.
Thrombin-Like Enzymes
Thrombin-like enzymes, such as those abundantly found in viper
venoms, are used to determine the levels of fibrinogen in a plasma
sample by converting fibrinogen to fibrin and the levels of fibrin
subsequently measured against a baseline. One of these serine
proteases, batroxobin from the Central American pit viper Bothrops
moojeni, is quite useful in its ability, unlike circulating
prothrombin, to form clots even in the presence of heparin. This
allows for a patient's plasma to be monitored for levels of fibrinogen
even whilst undergoing heparin therapy. Consequently, these studies
concluded that batroxobin is quite useful not only as a diagnostic
agent but also possibly as a direct therapeutic agent.
