From www.bloodjournal.org by guest on June 17, 2017. For personal use only.
Polybrene-Induced
Electrophoretic
Mobility:
Platelet
Agglutination
Enhancement
by von
Inhibition
by Vancomycin
By
It has
platelets
recently
more
proposed
that
between
low
been
reported
extensively
in
that
the
polycation
Polybrene
the
presence
of normal
plasma
ristocetin
initiate
agglutination
by
induce
agglutination
and
may
Polybrene’s
ability
concentrations
of
concentrations
were
concentrations
produced
reductions
and
platelet
produced
to
surface
Willebrand
produce
more
in mobility
with
These
charge
data
may
platelet
were
are
much
and
larger
with
(but
perhaps
aggregates
reversed
our
with
both
ANTIBIOTIC
ristocetin
platelets
if von
This
agglutination
ent.”2
antibiotic,
dependence,5
agglutinates
Willebrand
factor
is
vancomycin.”4
the effect
the two antibiotics,4
von
Willebrand
(vWf)
inhibited
by
Based
on
of chemical
and
factor
sufficient)
normal
Log
is pres-
bars.
normal
support
the
interaction
the
The
plasma.
the
that
between
Both
that
and
view
same
plasma.
an antibiotic
agglutination
Higher
these
Willebrand
Vancomycin,
and
studies
of
modification
(Havertown,
described6
pH
of
Rank
mobility
horizontal
Bros.,
Bottisham,
Measurements
a model
that
stressed
the
positive
charge
and its abilcharge
of platelets
in initiat-
inhibits
reductions
reductions
platelets
for each
Based
on its
process
does
molecular
patient
with
George
King
assess
agglutination,
I have
platelet
electrophoretic
vancomycin
to inhibit
agglutination
and
centrifuged
electrophoretic
The
volume
40%
at
g for
3 mm
PRP
buffer(0.l5
was then
at
effect
on
ability
of
platelet
changes.
22#{176}Cfor
2-4
whole
of
and
von
Tenon
stir
at
times
pH
in a dual
was
Co. (Fain
Vlll/vWf
14 ml of
7.4).
channel
and
N.J.).
of
4
severe
von
from
Miles
Ind.)
and hepanin
ries,
Chnono-
Health
Division
Sciences
Stony
Supported
and
Center,
N.Y.
in part
by
Blood
Hematology,
Hematology,
State
Brook.
Department
University
of
of
New
Medicine.
York
(96%
19278
from
the
at Stony
National
and
I 980 by Grune
accepted
requests
SUNY-Stony
to
Brook,
t4 Stratton,
0006-4971/80/5502-0026$O1.OO/O
was negative
Heart,
by
October
Dr.
Barry
Stony
Inc.
18, 1979.
S.
Brook,
Coller,
N. Y. I I 794.
of
weeks
bled
at
method
37#{176}Cfor
Q,
a
plasma
factor
studies,
blood
severe
von
concentra-
Becton-Dickinson,
by centni-
albumin
was obtained
Fraction
V.
from
the first
Elkhardt,
Abbott
2 hr,
of
Laborato-
0.1 5
injection
was
Willebrand
was
mg
purified
La Jolla,
of0.4
mg without
weekly
thereafter,
The
blood
was allowed
then
the
serum
was
Vol. 55,
clottable
antigen
(at
rabbit
fibninogen
Calif.)
the
in
followed
adjuvant.
Four
rabbit
to clot
separated
By immunodiffusion,
Blood,
Cutter
-96%
in an albino
and
30 mm.
purified
from
factor
raised
of
254B),
obtained
material
(Calbiochem,
25 ml.
and
This
PR
was
VIlI/von
injection
at 56#{176}C
for
number
antibody
adjuvant
of approximately
inactivated
this
with
a final
the
from
from
was prepared
Pentex
(lot
Calif.
injection
by intradermal
after
from
no detectable
patient
to give
U/mI)
of Mosesson,9
for factor
Freund’s
I wk later
Division
as a gift
Scientific
obtained
Kans.);
and
from
Plasma
In some
pure,
10,000
Antifibninogen
intradermal
complete
18. 1979;
reprint
Park,
3204
mean
Ill.
Berkeley,
10 mg/mI).
Grant
was obtained
at 4#{176}C.
Human
(Panhepnin
and
individuals.
assay5
its
Fisher
was
x
VllI/vWf
N.J.)
plasma
fibninogen
to the
anti-factor
plasma
(Vacutainen
for 30 mm
of
from
(Overland
4.8
was obtained
disease
platelet-poor
it has
manufactun-
citrate
8 healthy
by nistocetin
and
Chicago,
Laboratories,
Institute.
SubmittedJuly
Address
of
0A456)
(Somerville,
normal
(Milwaukee,
ofthe
rabbit
Willebrand
Laboratories
North
according
the
Ind.),
least
EDTA
at 2,500g
at least
direction)
pH,
assessment
(lot
by electnoimmunoassay.8
N.J.),
Co.
nature
accurate
HCI
Inc.
vWf
mM
I,
V/cm.
and
in each
at neutral
tnisodium
Bio-Medical,
Rutherford,
(Mark
plane,
Chemical
The
an
Pooled
ofat
Plasminogen-free
From
Aldrich
Diagnostics
blood
antigen
previously
of 3.82
going
structure,
(Indianapolis,
had no detectable
gradient
measured
from
permit
Hultin,
Lawn,
as
apparatus
in the stationary
Vancomycin
Behning
Mae
fugation
plasma
22#{176}C
with
M EDTA,
was monitored
Dr.
measured
sample.
not
Co.
matched
capillary
from
a normal
individual
and/on
the
Willebrand
disease was collected
in tubes
blood
platelet-rich
and
from
from
tion
citrate)
MTnis-0.OOl
agglutination
of this
METHODS
sodium
washed
M NaCI-0.OI
Platelet
mobility
AND
(1/100
750
mechanism
tested
Polybrene’s
mobility
and
the
Polybrene-induced
MATERIALS
Citrated
the
Lilly
was
per microgram.
weight.
whole
To
Eli
only
molecular
charges
using
at a voltage
platelets
was obtained
1014 positive
ing
made
(10
averaged
Polybrene
Wisc.).
U.K.)
were
20 measurements
were
aggregometen
electrophonetic
citrated
plasma.
Pa.)
in a cylindrical,
Willebrand
276
of
von
model
serum
©
than
of plasma.
mobility.
plasma;
presence
normal
citrate.
ing agglutination.4
Recently,
Rosborough
and Swaim’
reported
that the polycation
hexadimethrine
bromide
(Polybrene),
like
ristocetin,
agglutinated
platelets
more
extensively
in the presence
of normal
than
von
Lung
Willebrand
to initiate
Platelet
a similar
the effect of ristocetin
and/or
on platelet
electrophoretic
mobility,6
We proposed
importance
of ristocetin’s
ity to reduce
the surface
Brook,
of von
correlation
absence
factor.
HE
(PRP).
electrophoretic
in the
the
In the
the
Polybrene-induced
electrostatic
not
I investigated
mobility.
presence
of
like
ristocetin.
agglutinates
Since
we
have
previously
charge.
reduced
presence
inhibited
consistent
surface
electrophoretic
in the
in the
be partially
specificity.
be necessary
platelet
platelet
aggregates
greater
could
(hexadimethrine
bromide).
than
von Willebrand
plasma.
aggregates
and
also
some
alter
and
Collen
reducing
platelet
agglutination
mobility
reduction
small
small
rapid
agglutination
mobility.
platelet
T
Polybrene
needed
the
ristocetin-induced
in
to
in electrophoretic
agglutination
S.
Barry
and Reduction
in
Willebrand
Factor
was
in glass
and
the
No. 2 (February),
heat
resulting
1980
From www.bloodjournal.org by guest on June 17, 2017. For personal use only.
POLYBRENE-INDUCED
antisera
gave
plasma
AGGLUTINATION
a single
and
line
purified
277
of identity
fibrinogen;
there
when
tested
against
normal
was
no line
against
normal
Po/ybrene
serum.
Fibronectin
affinity
was
described
by
was eluted
450,000
220,000
was
and
0.4
weight
depleted
column),
against
normal
the
en) gave
a line
5-200
recently
descnibed.
Method
IV
by the
modiflcations.#{176}
void
brene
cofacton
imidazole,
6.5)
antiserum
(kindly
identity
of
of
material
from
10%
factor
over
the
tube
(A280
vWf
the column
was
buffer
IgG
=
tested
(0.25
an AO
70
as
O.D.
60
minor
0.075)
plasma
50
Poly0.02
azide,
1 5 for
Norrnc/
.
of
for
M NaCI,
=
80
1-lyland
only
0.027
of calculation,
normal
from
with
90
the factor
and Sephaof
purified
peak
I 100%
charMosh-
against
et al.
Sero
line
Dean
preparations
was
50
the gelatin
well
by Dr.
tested
60
with
a single
a previously
methyl-a-D-glucopynanoside
For purposes
Willebrand
and
O.D.
serum
unadsonbed
only
supplied
To
weight
absorption
passage
Marchesi
containing
with
The
these
antisera
and
by pepsin digestion
VlII/vWf
activity,
of approxito fibronectin
After
gave
purified
protein
antifibninogen
when
from
prepared
Factor
peak
as a control.
VIll/von
This
technique
The
volume
repeated
antiserum
chromatography
80
Antiserum
the
90
as
fibninectin
of molecular
plasma.
antiserum
fragments
were
the
purified
respectively.
(by
of complete
plasma.
F(ab’)2
VIIl/vWfantiserum
that
by
10)
gel analysis
gels.
as for
normal
resulting
plasma.
fibronectin
SDS
injections,
of fibninectin
actenized
cryl
mg
gave 4 lines against
affinity
SDS
rabbit
plasma
(Affi-gel
and a doublet
on reduced
in an albino
plasma
citnated
of 8 M urea. The
on unreduced
0.24
drawn
Ruoslahti,’#{176} except
instead
band
mg and
antiserum
freshly
gelatin-agarose
molecular
200,000
prepared
using
and
4 M urea
a single
mately
M
from
on
Engvall
with
revealed
the
purified
chromatography
‘4
control
serum
40
I mm
30
pH
factor
was used.
20
RESULTS
Polybrene
consistently
agglutinated
washed
platel-
ets in the absence
of plasma
at concentrations
equal
to
or greater
than
50 j.g/ml.
The aggregation
tracings
were notable
for a lack of increase
in the amplitude
of
the
deflections
that
aggregation
and
usually
accompanies
is attributable
to
large
platelet
aggregates
entering
and leaving
the light path.
Microscopic
analysis
confirmed
the
presence
of agglutination,
but
virtually
all the aggregates
were small,
consisting
of
only several
platelets.
When
citrated
was added
to the washed
platelets
higher
concentrations
of Polybrene
were
required
In the
to induce
presence
agglutination
but
some
amplitude
strong
normal
plasma
(25%-50%
v/v).
(0.3-0.6
mg/mI)
agglutination
(Fig.
I).
of vWf-deficient
citrated
plasma,
the
was both less rapid
and less extensive,
agglutination
of deflections
normal
than
inspection
of much
plasma.
confirmed
that
larger
aggregates
The preincubation
vW
citrated
was
was
always
much
plasma,
present.
greater
and
The
with
microscopic
this was due to the presence
in the samples
with normal
of anti-factor
VIII/vWf
serum
with normal
plasma
inhibited
the agglutination
and
the amplitude
of deflections,
whereas
control
serum
did not (Fig.
I). Because
citrate
is known
to
reverse
the effect of Polybrene
in red blood cell agglutination,’3
and both normal
and vW citrated
plasmas
inhibited
low-dose
Polybrene-induced
agglutination,
Fig. 1 .
(Above)
Aggregometen
tracings
of Polybrene-induced
platelet
agglutination
of washed
platelets
in the presence
of
normal
or severe
von Willebrand
factor
(vWf)
deficient
plasma.
Citrated
platelet-rich
plasma
(PRP) was washed
3 times in 0.1 5 M
NaCI, 0.01 M Tnis-HCI,
1 mM EDTA.
pH 7.4. and resuspended
in
the same buffer
to a platelet
count of 4.2 x 10’/ml.
A 0.2-mI
aliquot
of the platelet-rich
buffer
(PRB) was then added
to an
aggnegometer
cuvette
along
with
0.2 ml of normal
on vWfdeficient
plasma.
After a baseline
was established.
Polybrene
(0.6
mg/mI)
was added. To increase
the sensitivity
of the tracings.
the
“O” OD reading
was set with a mixture
of 1 part PRB and 3 parts
buffer.
(Below)
Effect of anti-factor
Vlll/vWf
serum on Polybreneinduced
agglutination.
Platelets
were prepared
as above.
Control
unimmunized
serum
or anti-factor
Vlll/vWf
serum
(10 .tl) was
added to the cuvette
1 mm before the Polybrene
(0.6 mg/mI).
the effect
of citrate
was
tested.
addition
ofcitrate
(34 mM)
of platelets
in the presence
and decreased
sample
with
the
normal
As shown
in Fig.
2, the
reversed
the agglutination
of normal
or vW plasma
amplitude
plasma.
of the
In other
deflections
experiments,
in the
the
addition
of 10 mM citrate
resulted
in more than a 60%
inhibition
of the
initial
slope
of Polybrene
(0.06
mg/mI)
induced
agglutination
of washed
platelets
in
the absence
of plasma,
and even a fourfold
increase
in
Polybrene
nation
mg/ml
concentration
completely.
and albumin
tion of agglutination
than that produced
U/ml)
was
also
failed
to restore
The addition
at 2 mg/mI
by Polybrene
(60
by 10 mM citrate.
able
to
reverse
the
of fibrinogen
caused
some
the
.tg/ml),
Heparin
agglutiat I .6
inhibibut
agglutination
less
(12.5
From www.bloodjournal.org by guest on June 17, 2017. For personal use only.
BARRY
278
Fig. 2.
90
80
0.
D.
After
brene
50
PLASMA
induced
citrate.
by Polybrene
When
plasma
in a manner
similar
to that for
from
a single
individual
was
collected
washed
into both EDTA
and citrate
platelets
and Polybrene,
the
gave consistently
greater
50, 100, and 200 tg/ml
rich buffer
and
I .3 versus
0.4,
optical
density
ence
tions
sample
plasma
did not
response
slopes
were
11.4 and
respectively).
Analysis
platelets
in the presence
revealed
tion
responses
(at
3 parts
platelet-
I part plasma,
the initial
slopes
were
4.6 versus
2.0, and
1 1 .0 versus
4.6
units/mm,
respectively).
This
differ-
could
be minimized
of Polybrene,
since
plasma
citrated
agglutination
employing
and tested
with
EDTA
plasma
variable
results
of Polybrene
used.
by use of higher
concentrathe response
of the EDTA
continue
did (at
to increase
0.6 mg/ml
while the
the initial
10 optical
density
units/mm,
of agglutination
responses
of
of buffer
or EDTA
plasma
depending
on the
concentra-
At low concentrations
of Poly-
a baseline
was established,
(1 .2 mg/mI)
was added.
At
Polythe
90
indicated
times. tnisodium
citrate
(34 mM
final
concentration)
was
added.
To
increase
the sensitivity
of the aggregom-
80
eter. the “0’S OD reading
was set with
mixture
of 1 part PRB and 2 parts buffer.
40
50 %
on Polybrene-
aliquot of the PRB was added to an aggregometer
cuvette
along
with
0.2 ml of
normal
or severe
vWf-deficient
plasma.
70
I mm
of citrate
induced
platelet
agglutination.
Citrated
PRP was washed
twice
in 0.1 5 M NaCI.
0.01
M Tnis-HC1
. 1 mM
EDTA.
pH 7.4.
and resuspended
in the same buffer to a
8
platelet
count of 3.5 x 10 /ml. A 0.2-mI
100
60
Effect
S. COLLER
of agglutination
washed
platelets
a
(50 versus
30 optical
density
units)
of
when
Polybrene
(0.6
mg/mI)
was
added.
It also enhanced
the amplitude
ofdefiections
the aggregometer
tracing.
Since
vancomycin
is known
to inhibit
ristocetininduced
agglutination,
its effect on Polybrene-induced
of
agglutination
was
tested.
With
washed
platelets
suspended
in buffer
alone,
vancomycin
I .2 mg/ml)
had a small
and variable
effect
on the initial
rate of
(
agglutination,
of agglutination
concentrations
mg/ml,
the
but
consistently
reduced
the final extent
after
2 mm over a wide
range
of
of Polybrene
(at 0.05, 0. 1 , 0.3, and 0.6
final
extents
of agglutination
in the
absence
and
10, 20 versus
density
presence
of vancomycin
were
I 8, 29 versus
I 8, and 25 versus
units,
respectively).
plasma
(25 v/v)
the initial
rate
When
was present,
of Polybrene
citrated
decreased
induced
agglutination
I I mg/ml
the response
present.
This
inhibited
the
small
amount
of agglutination
that
occurred
in the presence
of von Willebrand
plasma.
When
platelets
were agglutinated
by Polybrene
(0.6
Polybrene
plasma,
response
was greater
when
EDTA
finding
resulted
from
the
to induce
regardless
continued
normal
plasma
was increased
citrated
in the
large aggregates
in the absence
of the concentration,
whereas
to get stronger
in the presence
when
the concentration
up to 0.6-1.2
mg/mI.
plasmas,
presence
plasma
inability
the agglutination
of normal
EDTA
EDTA
plasma.
Moreover,
the
of normal
EDTA
plasma
was
VIII/vWf
F(ab’)2
fragments
was
of
of
the
of
of Polybrene
As with
the
was much
greater
plasma
than
vW
response
inhibited
(---30
in the presence
by anti-factor
j.tg/ml
final
concentrations
for
2 mm
at
37#{176}C;initial
slope
decreased
by 48% and total extent
of agglutination
at
4 mm by 70%) but not by similar
fragments
of normal
rabbit
antifibrinogen
The
presence
concentration
U/mI)
versus
of
of
significantly
I 5 optical
-
and antifibronectin
purified
factor
13
g/ml
serum.
VllI/vWf
containing
enhanced
the
density
units/mm)
275
(final
vWf
initial
rate
(54
and total
extent
I .2 mg/mI.
mg/ml)
and
Vancomycin
in the
presence
and the
virtually
normal
vancomycin
(0.6
mg/mI)
brene,
the response
of platelets
in buffer
was greater
than that in the presence
of EDTA
plasma.
However,
at higher
concentrations
of Polybrene
(> 0.3 mg/ml),
.
by 29%
caused
I 2 versus
I 6 optical
total
extent
by 53%
complete
inhibition
( I .2 mg/mg)
of normal
citrated
addition
of vancomycin,
like
citrate
reversed
the agglutination.
In the absence
of plasma,
Polybrene
also
at
at
completely
plasma,
and
the
heparin,
decreased
the
electrophoretic
mobility
of washed
platelets
at concentrations
as low as 2 g/ml
(Fig. 3). The decrease
was
dose-dependent,
mobility
occurring
with
almost
a 50%
reduction
in
with 6 zg/ml.
The decrease
could
be reversed
by washing;
34% reversal
after
a single
wash,
50%
after
2 washes,
and
80%
after
3. The
decrease
in mobility
could
not be accounted
for by
nonspecific
changes
in the suspending
solution
(such
as viscosity
or bulk ionic strength)
produced
by Polybrene,
since its effect on erythrocytes
was significantly
less (Fig. 3; p < 0.01)
than that on platelets.
Preincubating
the platelets
with citrate
or vancomycin
inhibited the decrease
in mobility
caused
by Polybrene.
In
From www.bloodjournal.org by guest on June 17, 2017. For personal use only.
POLYBRENE-INDUCED
AGGLUTINATION
279
20
mM
cl/role
presence
of
versus
27%
27%
0
±
normal
I .4%,
l.4%,p
±
than
p
<
0.001,
<
These
70
-j
CD
0
60
Ui
50
Ui
40
on the effect
agglutination
was the first
of the polycation
and electrophoretic
to report
that
weight
(but
that
of
required
that
not
2000)
EDTA
at
to
(g/ml)
Fig. 3.
Effect of Polybrene
on the electrophoretic
mobility
of
platelets
(.) and erythrocytes
()
in the absence
of plasma.
Platelets
were washed
3 times and resuspended
in 0.1 5 M NaCl.
0.01 M Tnis-HC1
. pH 7.0.
In some experiments.
citrate
or vancomycin was added as indicated.
the pH readjusted
to 7.00 ± 0.05.
and the measurements
taken.
The technique
for determining
electrophoretic
mobility
is described
in the Materials
and Methods
section.
presence
of
citrated
concentrations
of
plasma
(25%-33%
Polybrene
EDTA
did
were
v/v),
required
serum
supported
responsible
also
observed
the
that
present
the reduction
in mobility
than vW plasma.
and anti-factor
fresh
plasma
(25%-33%
electrophoretic
v/v) produced
mobility
than
a greater
decrease
that
found
in
amplitude
of the defiections
the presence
of vW plasma,
vW
plasma
±
(
(74.7%
SEM;
p
suspension
<
±
8.0% versus
0.05).
When
was preincubated
the
of the
with vancomycin
I .2 mg/ml),
the
ity produced
by 0.6 mg/mI
Polybrene
decrease
in mobilwas only 32%
3.2% (p < 0.001).
Similar
inhibition
in mobility
produced
by Polybrene
citrate,
with the mobility
decreasing
of the reduction
was observed
with
by 56.2%
± I .2%
(p < 0.05)
in the
42.0%
± 1.1%
(p
EDTA
citrated
Polybrene
normal
plasma,
(100
presence
0.001)
<
and
vW
of 20 mM citrate
with 40 mMcitrate.
plasmas
were
used
and
±
only
When
instead
of
the reduction
in mobility
produced
sg/ml)
was
again
greater
in
by
the
that
and
the
vW
IgG
In the presence
VlII/vWf,
high
of this
was
serum
They
must
agglutina-
platelets
were
of plasma,
at relatively
of normal
plasma
concentrations
of
of much
and
of the recorder
these
larger
and preincubation
VIII/vWfserum
fraction
vWf
plasma
washed
resulted
in the development
as judged
both microscopically
failed
to appear,
with anti-factor
or
Additional
VIII/vWf
Polybrene
in the absence
were small in size even
Polybrene
aggregates,
mean
plasma
that
noted
in agglutination.
normal
mg/mI
Polybrene
to
of normal
citrated
of citrated
the
No studies
were reported
on
disease.
Rosborough
and
platelets,
found
that Polyproceeded
more
rapidly
addition
in the
53.5%
± 2.6%,
platelet-normal
Grottum
of Poly-
by approximately
in an aggregom-
contention
study,
agglutinated
by
but the aggregates
in
one-eighth
sg/mI)
He
experiments,
the
washed
platelets
in
the
platelets
doses
(12.5-25
a 30% decrease.
high concentrations.
or purified
factor
of 0.6
presence
clump
mobility
changes
difference
both
Swaim’
evidence
of agglutinating
(8.3 1ug/ml)
reduced
produce
reductions
in mobility
comparable
to those
produced
in the absence
of plasma,
with a 50% reduction requiring
approximately
300 tg/ml.
In each of 6
presence
could
contain
some inhibitory
substance(s),
since
tion was fastest
in the absence
of plasma.
In the
to
their
for
and
erythrocytes.
concentration
not inhibit
in the presence
of normal
studies
with cryoprecipitate
observations
Polybrene
on platelet
mobility.
Polybrene
of molecular
clump
minimum
the
the
concentrations
the platelet
agglutination.
patients
with von Willebrand
Swaim,
using formalinized
brene-induced
agglutination
4
POLYBRENE
higher
6000
presence
eter
tracing
required
produced
approximately
30
3
the
extend
Rosborough
platelets’
electrophoretic
15%. To achieve
significant
I-
-J
and
and
brene
that gave microscopic
platelets
in normal
plasma
I
a:
confirm
Grottum,’5
those
reported
0/one
studies
of Lalezari,’4
the
C)
I-
in 2 experiments).
DISCUSSION
80
>-
.
90
C
0
U
1-
vW plasma
(34%
±
I I%
0.001 , and 39% ± I .7% versus
larger
by the
tracing.
aggregates
In
of normal
plasma
or F(ab’)2
fragments
also
prevented
the
development
of these
larger
aggregates.
F(ab’),
fragments
of control,
antifibrinogen,
and antifibronectin
IgG did not inhibit
the reaction,
reinforcing
the specificity for vWf.
whole
serum
The
was
nonspecific
IgG
ristocetin-induced
has been
shown
to
platelet
agglutination
16
was significantly
is approximately
use of these
fragments
necessary,
since
the
The
Polybrene-induced
inhibited
what
by citrate
the
final
Fe
rather
than
portion
of
interfere
under
with
certain
agglutination
at 10 mM,
concentration
which
of
From www.bloodjournal.org by guest on June 17, 2017. For personal use only.
280
BARRY
citrate
would
be
if citrated
plasma
and
the
vancomycin
platelet
inhibited
the
decrease
S. COLLER
in
mobility
suspension
were added
in equal
volumes.
In addition,
EDTA-anticoagulated
plasma
produced
significantly
less inhibition
than citrated
plasma
at low concentra-
produced
by Polybrene.
As contrasted
with ristocetin,
Polybrene
produced
a greater
reduction
in mobility
at
concentrations
that were sufficient
to induce
aggluti-
tions
nation.
of
Polybrene
and
enhanced
more
at high
concentrations,
citrate
in plasma
accounted
inhibition
produced
by citrated
(2 mg/ml)
and
minor
inhibition,
inhibition,
ough
known
fibrinogen
so they
as has
already
formation,
been
an
peptide-binding
to ristocetin,
suggested
also
able
but
will
tion.3’4
This
inhibition
antibiotic
Rosbor-
inhibit
the
similar
in
mechanism
platelets
is
by
reduction
in mobility
by Polybrene
suggesting
result
the
expression
ristoce-
vWf-specific
that histone
at limiting
concentrations
of ADP.3
of vancomycin
to inhibit
Polybreneagglutination
is strong
evidence
that
is operating
by a mechanism
similar
Since
it was previously
hypothesized
to that
that
vancomycin
competes
with ristocetin
for binding
sites
on the platelet
membrane
and,
by virtue
of its less
positive
change,
prevents
the
decrease
in surface
change
caused
by ristocetin,
the effects
of vancomycin
and
Polybrene
on platelet
electrophoretic
mobility
were tested.
Polybrene
produced
decreases
in platelet
electrophoretic
mobility,
and
preincubation
than
presence
its
in the
EDTA),
that
the
produced
of vWf-deficient
plasma,
of vWf to the platelet
may
in mobility
in
with ristocetin.6
polycation
with
no
to induce
platelet
of
ristocetin
charge
of
or
of the decrease
the
that
of
positive
to platelets
greater
in the presence
the binding
in
factor
induced
aggregation3’18”9
and only mildly
adenosine
diphosphate
(ADP)
induced
pla-
Polybrene
of ristocetin.
was
in an enhancement
hypothesis.
that simple
telet aggregation
Thus,
the ability
induced
platelet
that
added
(citrated
a manner
similar
to that observed
The
ability
of Polybrene,
a
structural
similarity
to ristocetin,
does not
von Wille-
vancomycin
or bovine
was
plasma
vWf
in
a manner
supports
the view that
plays an important
role in
agglutinating
activity.
Rosbor-
ough’s
recent
preliminary
report
that other
unrelated
polycations
(polylysine
and polyornithine)
also have
similar
activity20
is additional
evidence
in favor of this
agglutinafor
Polybrene
of normal
similar
to
ristocetin’s
of action
as ristocetin
specific
When
presence
agglutination
agglu-
structure,
ristocetin-induced
is relatively
tin-induced
agglutination,
since
inhibit
thrombin,
epinephrine,
brand
inhibits
by
to inhibit
properties,
and
does not aggregate
does,17
some
to the
expected,
heparin,
which
by Polybrene,
presumably
was
tination.
Vancomycin,
agglutination
(1 .6 mg/mI)
produced
may also contribute
and
Swaim.7
As
to be inactivated
complex
the
indicating
that
the
for at least
part
of the
plasmas.
Both albumin
with
However,
acidification
agglutination,4
and
also failed
to induce
nation.2#{176}Thus,
sary,
but
not
a positive
in itself
phenomenon.
teristic
in unpublished
studies,
and lysozyme
did
I favor
of these
mycin’s)
surface
ability
where
Rosborough
vWf-specific
charge
appears
sufficient,
to
the view
agents
we found
not induce
that
accounts
to be necesproduce
the
some
other
for their
to bind to specific
the positive
charge
reported
aggluti-
charac-
(and
vanco-
sites on the
can then
platelet
initiate
agglutination.
ACKNOWLEDGMENT
I would
like to thank
excellent
technical
for typing
Marilyn
assistance
Banszczowski
and
Vesta
and
Carucci
Dorene
Turi
and Jennifer
for
Floyd
the manuscript.
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1980 55: 276-281
Polybrene-induced platelet agglutination and reduction in electrophoretic
mobility: enhancement by von Willebrand factor and inhibition by
vancomycin
BS Coller
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