THROMBOSIS RESEARCH Printed in the
INHIBITION
OF
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States
C OX31UNI
BRIEF PURIFIED
PLASMA AND
Pp. 223-225,
7,
Vol.
United
CAT1
KALLIKREIN
Press,
197j
Inc.
OX
BY ANTITHROMBIN
III
HEPARIN
Anne Marie Vennertjd and Knut LaakeX Department
of Pharmacology, University
(Received
to Yin et al.
identical
with the heparin of heparin
(1) antithrombin cofactor
antithrombin
factor Xa and the factor XIa heparin
forms a complex
serine proteinases
antithrombin
importance trinsic (5).
III of human plasma
III inhibits
(2). Rosenberg
in and related
thrombin,
III which
is rapidly
of heparin
is
Xa. In the the
(3) has proposed
that
inactivates
to the coagulation
that plasmin
III in the presence
Recently,
form 9.5.1974. Godal)
and the antifactor
with antithrombin
and it has been observed
of Pharmacy,
of Oslo, Norway
in revised 29.4.1975; Accepted by Editor H.C.
According
presence
Institute
cascade,
inactivated
by
(4).
plasma
kallikrein
has been reported
in the initial
activation
of the factor XII in the in-
blood coagulation
Plasma
kallikrein
and the factor link between
IX
also activates
(7). Plasma
the initial
XPresent
system by negatively
Department
kallikrein
Haematological
IX, University
surfaces
plasminogen
may also represent
parts of the intrinsic
address:
Oslo, Norway.
purified
charged
to be of
a
and the extrinsic
Research
Clinic,
(6)
Laboratory,
Ullevaal
Hospital,
blood
coagulation
systems
arin and antithrombin factor VII purified
III somehow
(9). The present
plasma
(a), and it has been observed
kallikrein
interfere
experiments
that hep-
with the activated
were undertaken
is inactivated
to see if
by antithrombin
III and
heparin. Partially activated viously
purified
with purified
human plasma
factor XII fragments
shown that the activity
a-N-benzoyl-L-arginine
prekallikrein
was
(11). We have pre-
of such preparations
ethyl ester
(10)
towards
(BAEe) is due to the activat-
05-
Fig. 1 Inhibition of plasma kallikrein by antithrombin III and heparin. -c: Plasma kallikrein, antithrombin III and heparin _ : Plasma kallikrein and antithrombin III & : Plasma kallikrein and heparin + : Plasma kallikrein alone The final concentrations in the incubations were: Plasma kallikrein 0.7 BAEe U/ml, antithrombin III 0.4 U/ml (defining 1 U as the amount present in 1 ml titrated plasma), and heparin 0.85 U/ml (obtained dry from A/S Apothekernes Laboratorium for Specialpreparatgr, Oslo). All incubations were carried out at 25 C in 0.05 M Tris/HCl pH 7.4. The BAEe esterase activity in the incubations was determined at intervals with NAD and alcohol dehydrogenase (10).
IMLLIKREIN
t’cJl.~,lio.l
ion of plasma
L AXTITHROMBIS
Antithrombin
16).
prekallikrein
from human titrated
plasma by a two-step
inity chromatography
on heparin-coupled
DEAE-Sephadex
A-50 chromatography
No inhibition observed
ever, the kallikrein and antithrombin
plasma
antithrombin
plasma
kallikrein
a2-macroglobulin that plasma vation
kallikrein
of the initial
the results impaired heparin
probably
intrinsic
sustaining
the results
Plasma pathogenesis
ological
ment with heparin
system,
of factor XII is also during
of factor VII by the plasma
kallikrein
and antithrombin
angioneurotic of plasma
III,
(13).
edema which
(14). The rapid III and heparin
concentrations
might be of benefit
acti-
to play a role in the
by antithrombin
and pharmacological
of
and the
coagulation
experiments
is supposed
of the hereditary
kallikrein
inhibitor
for the normal
through
by heparin
of previous
kallikrein
blood and
in vitro and possibly
system mediated
to lack of the Ci-inactivator ion of plasma
is necessary
in vivo. The activation
also counteracted
(Fig. 1).
As it is now accepted
mean that the activation
coagulation
is probably
acting
stages of the intrinsic
plasma
III. How-
in the incubations
of proteinases.
was
when both heparin
to the Ci-inactivator
activity
in heparinized treatment
or antithrombin
III may be a rapidly
inhibitor
aff-
gel and
activity
that in heparinized
in addition
employing
polyacrylamide
inactivated
III were present indicate
procedure
kallikrein
with heparin
was rapidly
The results
III was purified
(12).
of the plasma
upon incubation
225
III
indicates
is due inactivatin physi-
that treat-
to such patients.
REFERENCES 1. YIN, E.T., WESSLER, S., and STOLL, P.J. Identity of plasma-activated factor X inhibitor with antithrombin III and heparin cofactor. J. biol. Chem.: 246, 3712-3719, 1971.
2. DAWS, P.S., HICKS, M., and ROSENBERG, R.D. Anticoagulant action of heparin. Nature: 246, 355-357, 1973. 3.
ROSENBERG, R.D. Actions and interactions of antithrombin and heparin. N. Eng. J. Med.: 292, 146-151, 1975.
4. HIGHSMITH, R.F. and ROSENBERG, R.D. The inhibition of human plasmin by antithrombin-heparin cofactor. J. biol. Chem.: 249, 4335-4338, 1974. -5.
COCHRANE, C.G., REVAK, S.D., and WJEPPER, K.D. Activation of Hageman factor in solid and fluid phases. J. exp. : 138, 1564-1538, 1973. Med. -6. LAAXE, K. and VENNERgD, A.M. Factor XII-induced fibrinolysis: Studies on the separation of prekallikrein, plasminogen proactivator, and factor XI in human plasma. Thromb. Res.: Q, 285-302, 1974. B., LAAKE, 7. %TERUD, factor IX. In press
K., and PRYDZ, H. Activation in: Thromb. Diath. haemorrh.
Of
8. GJijNNAESS, H. Cold promoted activation of factor VII. II. Identification of the activator. Thromb. Diath. haemorrh.: 28, 169-181, 1972. 9.
GODAL, H-C., RYGH, M., and LAAKE, K. Progressive inactivation of purified factor VII by heparin and antithrombin 111. Thromb. Res.: 2, 773-775, 1974.
10.
LAAKE, K. and VENNERGD, A.M. Determination of factor XII in human plasma with arginine proesterase (prekallikrein). I. Preparation and properties of the substrate. Thromb. Res.: 2_, 393-407, 1973.
11. VENNERijD, A.M. and LAAKE, K. Isolation and characterization of a prealbumin activator of prekallikrein from acetone-activated human plasma. Thromb. Res.: 4, 103-108, 1974. 12. MILLER-ANDERSON, M., BORG, H., and ANDERSSON, L.-O. Purification of antithrombin III by affinity chromatography. Thromb. Res.: 2, 439-452, 1974. 13. GODAL, H-C., GRAVEM, K., and LAAKE, K. Progressive inactivation of factor VII in the cold by heparin. Thromb. Res.: fi, 707-710, 1974. 14. DENNEHY, J.J. Hereditary angioneurotic edema. Report of a large kindred with defect in C'l esterase inhibitor and review of the literature. Ann. intern. Med.: 73, 55-59, 1970.