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Distribution
of Phospholipids,
Fatty
Acids,
and Platelet
Factor
3 Activity
Among
Subcellular
Fractions
of Human
Platelets
By M. Johan
As compared
cently
with
other
reported
of
for
human
separation
of
Robert
methods,
method
fractionation
the
Broekman,
I. Handin,
our
re-
subcellular
platelets
improves
mitochondria,
pertaining
pholipids
were
alpha
granule
phatidyl
zone,
In
plasmalogen
at
membrane
suggested
lysophosphatidyl
that
it
LATELETS
vascular
may
have
been
participate
injury
and
85%,
tent
of
more
are
active
platelets
clot-promoting
make
polyenoic
were
acids
esterified
The
most
related
to
non-
in
formed
planations
the
the
new
clot-promoting
which,
content
basis
are
VIII
centers
for this
protein
fractions.
on
novel
both
necessary
.2 and
for the
possible
ex-
finding.
at the
for the
activation
site of
to prooptimal
of pro-
systems
in vitro,
phospholipids
fatty acids are both
unsaturated
coagulation:4
however,
is not established.
platelet
factor
of
nonmembranous
discussion
model
or whose
activity
other
the
important
to
on
acids,
of 20:4,
was
highest
in the membrane
but on the basis of phospholipid
The
In
fatty
72%
the
relevance
The clot-promoting
3 (PF-3).
The mechanism
available
is not
well
of
these
properties
of
by which
understood:
how-
with
the release
reaction.9
PF-3
since other
blood
cells,’0
as
origin,
can also
furnish
clot-
activity.
the Department
From
Peter
the
18:2
18:0,
higher
choline
72%,
total
con-
94%,
content,
fractions,
ever, its appearance
is closely
associated
in vitro
is not a unique
property
of platelet
phospholipids,
well as cells or subcellular
fractions
of diverse
promoting
of
in hemostasis
by forming
aggregates
by providing
a source
of phospholipoprotein
in promoting
processes
been
called
of
group.
choline
factors
Xa and
V.’3
are negatively
charged
data to in vivo
platelets
have
held
and
the
mem-
activity,
mote
clotting.
Phospholipid
and calcium
activation
of factor
X by factors
IXa and
thrombin
by
whose
micelles
of
the
versus
latter
18:1,
20:4
of
asymmetrical
respectively,
16:0,
information
the
phospholipase
acids
noncholine
The
67%
Cohen
platelet
fatty
in
choline
the
Phin
a markedly
55%
than
phos-
peaked
of
The
showed
and
and
and
action
whereas
of diacyl
phosphatidyl
ethanol(2) The fatty acid composition
of
amine.
P
(1)
activity.
phospholipids.
of phos-
new.
ethanolamine
expense
the
entirely
the
A2
distribution
alpha
to distribution
by
branes
granules,
and Iysosomal
enzyme
activities.
The relative
purity
of these fractions
has
led us to undertake
the present
study to
compare
the subcellular
distribution
of
phospholipids,
fatty
acids,
and
platelet
factor
3 (clot-promoting)
activity.
Two
findings
Derksen,
Arie
Bent
Brigham
of Nutrition,
Hospital;
Harvard
and
the
School
Department
of
of
Public
health;
Medicine,
the
Harvard
Division
of hematology.
Medical
School.
Boston,
Mass.
Submitted
June
Supported
in part
0083
the
from
Hospital.
Dr.
Cancer
United
by
accepted
USPHS
States
February
Grants
A rmy;
Handin
is a Cancer
reprint
requests:
ilL
/4.
/976.
/3584.
13802.
and
/7513;
and
the Hematology
Research
Fund
Research
Scholar
of the
Massachusetts
M.D..
Department
of
Contract
of the
DADA
Peter
Division
of
Bent
the
/ 7-70-CBrigham
American
Society.
Address
Boston,
for
Mass.
/976
Blood,
4. 1975;
hi
Phin
Cohen,
Nutrition,
665
Huntington
Ave.,
021/5.
Grune
& Stratton,
Vol. 47, No. 6 (June). 1976
Inc.
963
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964
BROEKMAN
After
platelet
ity that
organelles,
method
aggregation
and
the
release
reaction,
was
incomplete.
tion
of human
Recently
that
of
among
procurement
(Fenwal
decompression
of
one.
tamed,
and
Phospholipid
was
and
25 sec and
fractions
for
an
excess
several
days
Chemicals.
fore
use.
of
was
45,
22.5,
so
of platelet
at 4’C
substrates
and
-
and
from
subjected
217,500
that
(clot-
three
Nine
pack
the
sys-
nitrogen
decompressions
fractions
were
ob-
reported.’3
reported
The
was
a plastic
of
to
gmax’
methods
RF
a gift
of
of
were
used
to
cx-
bis-(monoacylglyceryl)
Dr.
phospholipids,’4
Cintron21
1:2,
9.0,
the
20’C
was
1:5,
and
that
J.
fatty
4.5
did
in
.ng per
shorter
not
change
F.
Wherrett,
acid
their
each
M
To-
methyl
no
platelet
NaCI
to
esters
7 sec.
Sigma
at
protein
of
platelet
con-
Russell’s
material
Storage
clot-promoting
and
fraction
give
concentration
added
than
Methanol
from
assay
The
with
never
obtained
to
0.15
assay.
time
was
grade.
were
used
1:10,
clotting
of reagent
RVV
PF-3
and
a modification
as previously
of
material
or
were
was
at
sample
diluted
adjusted
All chemicals
Enzyme
and
availwhich
described.20
of Spaet
undiluted,
The
study,
platelets.
derived
phospholipids.
authentic
as recently
method
were
Previously
quantify’4
activities.’3
present
acids,
by
monitored
transesterification
quantified
(RVV)
with
an
mm
studies.
and
with
respectively,
venom
were
composition
After
The
four concentrations:
viper
acid
90
the
afrac-
METHODS
platelets
of platelets
for
a
ac-
fraction
mitochondrial
of human
homogenized
batch
activities
localize,14
determined
activity.
centrations,
each
and
was
enzyme
Canada.19
separated
PF-3
and
and fatty
Ont.,
were
in that
separate,’4’8
phosphate
Ill.)
Ultracentrifugation
protein
tract,14’15
AND
subcellular
and
fatty
fractions
Washed
Deerfield,
technique’3
instead
for
granule
phospholipids,
subcellular
and fractionation.
Laboratories,
a method
alpha
MATERIALS
Platelet
ronto,
activ-
that,
in addition
to the memcontain
PF-3
activity.
Howfraction
into its components
and lysosomal
enzyme
fractions
has promoted
distributions
activity
reported
separates
to microsomal
purified
platelet
the
promoting)
we have
platelets
in addition
of more
compares
tem
clot-promoting
is made
available
could
reside
in plasma
membranes
or intracellular
or both.
Subcellular
fractionation
of platelets
should
provide
to examine
various
platelet
constituents
for their
clot-promoting
tivity.
Marcus
et al.” and Day et al.’2 have shown
branes,
various
subcellular
fractions
of platelets
ever, in those
studies
the resolution
of the granular
tions
ability
the
ET AL.
was
of subcellular
activities.
chloroform
Chemical
were
Co.
(St.
redistilled
Louis,
be-
Mo.).
RESULTS
Phospholipid
Composition
Fifty-eight
per
membranous
rather
evenly
phospholipid
and
lowest
Choline
myelin
which
sitide;
of the
of
total
alpha
PC
a reciprocity
been
pointed
different
I)
platelet
(zones
2 and
among
the
in particulate
in zone 7, where
phospholipids.
cells
of
the
phospholipid
was
recovered
from
the
The
remaining
phospholipid
was
other
particulate
fractions.
The ratio
of
fractions
was highest
in the membranes
3))3
granules
decreased
predominated
only
in those
between
the major
out in comparisons
same
species’4
or
the
overwhelmingly.’3
zones
where
sphingo-
choline
phospholipids
of phospholipid
comsame
cell
in
different
22
*Abbreviations
phatidyl
cent
fractions
distributed
to protein
was increased,
has previously
position
species
(Table
in
ethanolamine;
DPG,
text:
PC,
PS,
diphosphatidylglycerol.
phosphatidyl
phosphatidyl
choline;
serine;
LPC,
P1,
lysophosphatidyl
phosphatidyl
inositol:
choline;
DPI,
PE,
diphosphoino-
phos-
From www.bloodjournal.org by guest on June 17, 2017. For personal use only.
DISTRIBUTION
Table
OF
PHOSPHOLIPIDS
1. Distribution
of Phospholipid
Phosphorus
in Subcellular
Fractions
of Human
Platelets
Zone
.
Phospholupud
Class
Choline
965
1
H
2
3
4
5
6
7
8
9
1.0
31.8
23.6
1.3
34.5
22.8
1.3
35.4
21.2
1.4
15.0
16.1
10.3
13.0
PL
PC
SM
37.8
15.1
41.8
39.5
38.6
39.3
38.4
31.6
13.9
16.4
12.1
15.8
16.4
21.6
1.0
1.3
0.9
0.6
0.5
17.4
17.8
15.5
15.4
19.4
19.2
11.4
15.7
13.4
12.9
11.4
9.0
LPC
Ethonolamine
Plasmalogen
PE
17.9
DiacylPE
25.2
13.6
Plasmalogen
PE/
1.32
DiacylPE
Other
1.1
PL
1.70
1.19
1.16
1.13
1.53
-
2.13
1.46
1.24
7.4
P1
PS
9.0
10.5
8.9
9.6
8.8
7.9
7.9
9.6
9.4
P1
3.6
4.6
4.4
4.6
3.8
3.5
3.8
2.8
4.6
3.5
DPI*
0.6
0.6
-
tr
tr
tr
0.7
0.9
tr
tr
tr
-
tr
0.7
2.3
2.5
1.9
0.9
1.1
1.1
1.0
0.6
0.7
1.2
1.0
0.9
1.0
0.6
DPG
0.8
Origin
1.6
P1 phosphorust
nm/mg
protein
Per cent of total
Data
shown
recovered
for
from
analysis,
For most
(< 0.5%);-,
*The
only
13.3
492.2
546.8
288.6
267.5
185.0
171.2
181.1
257.6
4.4
43.0
15.4
6.5
4.1
4.5
6.7
4.4
10.9
each
whole
nique of Broekman
pholipid
164.6
100.0
phospholipid
are
homogenate
the
(H) and
et al.13 For preparation
see Materials
means
the
of
nine
of
four
analyses
subcellular
platelet
as
fractions
fractions,
per
cent
that
can
be
of
phospholipids,
separation
phospholipid
phosphorus
obtained
by
the
and
techphos-
and Methods.
abbreviations,
see
not determined.
member
of the
text.
Other
abbreviations,
polyphosphoinositides
PL,
that
was
phospholipid;
found
SM,
in more
than
trace
sphingomyelin;
tr,
trace
amounts.
of which
was re-
The abrupt
change
in the sphingomyelin
content
between
the upper
lower
halves
of the gradient,
as defined
by zones
5 and 6, respectively,
was
striking.
Marcus
et al.23 found
a similar
trend,
but not of such
sharpness
and
most
or
tPL
phosphorus
covered
from
magnitude.
blood
for
mem
shown
the
as nm/mg
and
per
cent
of
the
total
P1 phosphorus,
96%
5%
±
gradient.
By
platelet
contrast,
Kaulen
and
Gross,24
procurement,
found
the highest
using
content
EDTA-anticoagulated
of sphingomyelin
The
fatty
1-acyl
LPC
acid
of the
ratio
membranous
of
configuration.
zones,
approximately
Ethanolamine
in their
high
diacyl
PE,
By contrast,
phospholipids.
content
of plasmalogen
in zones
6 and
2 and
0.2,
and
a value
in human
plasma
LPC25
reflecting
derivatives.
Thus,
we suggest
that
plasma
but probably
derived
from
of
3, had
an
thus
was
I was
reported
unsaturated/saturated
predominantly
Minorphospholipids.
as well
an equal
distribution
of 1-acyl
and
the membrane
LPC was not adsorbed
endogenous
phospholipase
A2 activity.20
2-acyl
from
7, where
The
as the fractions
was
data
on lipid
extracts
the
ratio
PE.
resemble
The latter
alpha
granules
brain26
peaked
combined
less than
derived
harvest
one
from
might
intact
and
erythrocytes26
at the expense
predominate
of
DPI
from
the
of
same
Platelets
for
relationship,
if any,
between
plasmalogen
PE or sphingomyelin
other
constituents
of alpha
granules27
remains
to be elucidated.
vious
in
bra nes.
the
of
heavily.
The
content
and
homogenate
have
expected
from
our
platelets.’5
This
finding
presug-
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966
BROEKMAN
Table 2.
Distributio
Acids (F A) Among
n of Fatty
ET AL.
of Zone 2 (Membranes)
the Ph ospholipids
FA Types’
Other
Phospholipid
Class
Total
160
18,0
18,1
182
204
Polyenoict
Otherst
FA
U/S
PC
73.2
38.6
71.4
83.9
27.9
33.2
23.5
48.4
1.3
SM
19.8
5.5
0.7
1.4
0
0
73.7
10.1
1.2
1.4
0.2
0.1
0.1
0
Choline
LPC
0.1
0.6
Noncholine
PPE
0.7
2.1
1.9
2.0
24.3
38.7
0
10.7
DPE
4.0
20.0
7.7
9.1
23.9
18.6
0
14.0
PS
0.4
22.4
15.5
2.2
12.9
9.4
2.7
10.9
1.8
P1
0.7
10.0
2.5
1.3
10.9
0
0
5.4
2.1
Total
FA
On the
major
20.1
left
and
FA types.
tical
listing);
that
are
Given
(2)
all
esterified
derived
For
diacyl
of
FA
for
to
human
the
all
7.0
platelet’s
percentage
(total
FA, vertical
diacyl
6.8
seven
content
phospholipids
(total
the
29.1
the
are
phospholipid
FA,
The
(1)
horizontal
On
the
raw
100.1
phospholipids
among
listing).
phospholipids.
major
all
listing);
far
data
right
from
and
seven
phospholipids
are
and
(ver-
(3)
given
which
all
FA
the
un-
these
results
request.
membranous
and
4.7
listed
esterified
FA ratios
fractions,
separation
of phospholipids,
Other
abbreviations:
and
phospholipid
and
fatty
acid
Methods.
abbreviations,
see
text.
SM, sphingomyelin;
PPE, plasmalogen
PE; DPE,
PE.
‘FA are designated
by the number
t 203,
205,
161,
18,3,20,0,22,0,22,1,24:0,24:1.
224,
22,5.
22,6,
gested
that
the
zation
and
fractionation.’5’28
DPG
Pertinently,
Marinetti
platelets
for
No
found
Fatty
C atoms,number
with
ofendogenous
fractions
5-7,
was the first phospholipid
showed
its association
et al. found
an increase
nonmembranous
fractions,
DPG
in platelets.
bis-(monoacylglyceryl)
in any subcellular
our
fraction,
of double
the
same
bonds.
retention
phosphatase
in
where
was
as 24, 1.
enhanced
mitochondria
were
that
qualified
with
mitochondria.29
in DPG
in a granular
particles.23
data
are
phosphate,
even
time
the
Because
first
to
a putative
in concentrated
by homogenimost
as
of the
show
abundant.
a marker
since
In studies
with
fraction
that was
greater
clearly
lysosomal
samples.
purity
of
a marker
marker,’9
was
A cid Composition
As previously
acid
shown
and
whole
DPG
sphingomyelin
PE (20:4,
(18:1,
As shown
in Table
*Fatty
are
acids
for
fractions
obtained
predominance
was
PC (16:0,
l8:I),*
20:4),
plasmalogen
20:4),
of
an unknown
this
et al.
Marcus
as platelet
fatty
activity
and
peaked
a mixture
of all
our mitochondrial
role
are
of the
are
(U/S)
see Materials
most
each
be sent upon
For preparation
analysis,
for
to a given
will
16.2
respectively,
FA that
saturated/saturated
were
16.2
top,
2.4
designated
human
erythrocytes3#{176}
by the Teflon-pestle
associated
with
(16:0,
20:0,
higher
polyenoic
and
platelets,’4’23
technique,23
each
phospholipid
as well
a characteristic
in all zones:
22:0,
24:0,
24:1),
diacyl
acids),
PS (18:0,
20:4),
PE (18:0,
P1 (18:0,
18:2).
2 the
platelet’s
by the
number
fatty
acids
ofC-atoms:number
were
asymmetrically
ofdouble
bonds.
distributed
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DISTRIBUTION
OF
PHOSPHOLIPIDS
967
-
-
-
=
HOMOGENATE
-
20
‘5
‘44
ZONES
l0
K
4
d
.
5
45
90
22.5
jig
Fig. 1 . PF-3
let fractionation
activity
in human
and PF-3 assay.
seconds; on the abscissa,
groupings
homogenate;
among
,
are
the
choline
versus
choline
phospholipids
tent of 16:0, 18:1, and
noncholine
sociated
was
the
held
18:2.
18:0
esterified
polyenoic
subcellular
fractions.
See Methods
for details
ordinate
is the Russell’s
viper
venom
clotting
than
are
group.
20:4.
PF-3 assay.
the
clotting:
of
of the
which
The
latter
also
disproportionately
Figure
Table
The
Clot-promoting
results
with
tivity
clot-promoting
the
activity
nine
4 was
1 and
much
remaining
zones
Table
3 compares
per
activity
phosphorus
was
zones
67
of
the
plasmalogens
the
latter
assay
fell into four
the membranous
higher
in
which
were
more
the diacyl
phospho-
7 was
more
to
protein
content
of
the
fractions:
content.
tested
at four
fell into three
at 9 j.g protein/assay.
of zones
3, and
tent
1 relates
The
(PF-3)
3 to phospholipid
delineated
the
A clivity
At this
considerably
difficult
concentrations
groupings
which
concentration,
diminished,
to attenuate.
the
of protein
(Fig.
I).
were most
clearly
clot-promoting
whereas
The
that
clot-promoting
ac-
of zones
2,
activity
of
clustered
about
the values
for whole
homogenate.
the clotting
time
with
the phospholipid
phosphorus
of ten
groupings.
zones
and
origins,3’
abundant
in PC than the noncholine
fractions.
Also, considering
only
species,
the unsaturated/saturated
fatty acid ratios
of the noncholine
lipids
were considerably
higher
than that of PC.
Clot-promoting
three
9,
as-
various
held
represented,
8,
of the total
con18:0 was found
in
of
excluding
6,
is preferentially
phospholipids
However,
5,
membrane.
respectively,
16:0, 55
of the
of 20:4,
of platetime in
At 9 g,
2-4;
85#{176},
72,
species
noncholine
in
of
phospholipids
Pertinently,
other
polyenoics
of platelet
protein
least
acceleration
noncholine
in molecular
to the
acids
/4SSAY
94,
72#{176}c,
and
By contrast
with
phospholipids.
with
higher
platelet
On the
the concentration
from greatest
to
apparent
1 7.
450
PROTEIN
selected
points
in Fig.
1. The
At similar
concentrations
gave considerably
longer
results
with
of phospholipid
RVV clotting
the
times
nine
conzones
phosphorus,
than
zones
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968
BROEKMAN
Table
3.
Relationship
of Phospholipid
Phosphorus
of Subcellular
Fractions
Zone’
No.
Content
and
of Human
Clot-promoting
Activity
Platelets
nmoles Phospholipid
Phosphorus/Assay
Content
ET AL.
Russell’s Viper Venom
Clotting Time (sec)
2
Membranes
2.21
19.3
3
Membranes
2.46
23.3
Microsomes
4
Mitochondria
2.60
13.0
5
Acid hydrolases
2.41
13.9
9
Debris
2.32
15.3
1.67
14.8
Storage
granules
Mitochondria
6
Alpha
granules
7
Alpha
granules
1.54
17.0
8
Alpha
granules
1.63
14.4
Debris
1
Supernatant
0.30
14.8
H
Homogenate
1.49
14.4
Data
were
derived
‘Numbers
arated
using
from
Table
1 and
characterizations
of the
our
recently
method.13
reported
pholipid
phosphorus
where
mitochondria,
granule
of the
zones
had the lowest
particulate
zones,
yet
from
any
zones,
lipid
selected
and
points
nine
on Fig.
subcellular
The
1.
fractions
zones
are
of
arranged
human
platelets
in four
that
groupings
can
be
according
sep-
to phos-
content.
acid
particulate
whereas
with
the
content,
shortened
collected.
of the
RVV
The
concentrations
activity
grouped
lowest
the
organellar
granules
phosphorus
clot-promoting
homogenate
supernatant,
or any
or storage
phospholipid
held
as much
zoh#{235}.Whole
phosphorus
as homogenate
hydrolases,
with
tested
clotting
the
as
alpha
values
time
for
to the
alpha
of any
material
granule
phospho-
same
degree
fraction.
DISCUSSION
Our
data
show
that
(1) there
is a marked
asymmetry
in distribution
of some
phospholipids
and fatty acids
among
subcellular
fractions
of human
platelets,
and (2) on the basis
of the RVV
time,
clot-promoting
activity
of the fractions
does not correlate
with their phospholipid
phosphorus
content.
When
the clot-promoting
activity
of the fractions
was assayed
at equivalent
protein
concentrations,
organelles
(Fig.
the
1). This
membranes
finding
and Day et al.’2 However,
protein
mark
for assessing
clot-promoting
considered
lipid
to
to be dependent
on
protein
varies
widely
highest,
and
alpha
granules
(Table
1). Yet, on the
hold more
clot-promoting
do not
activity
by
fit with the
of platelets
differences
basis
prevailing
to their
in phospholipid
were
agrees
more
with
concentration
activity
active
previous
the
lowest
view that
membranes.
or
fatty
the
by
intracellular
Marcus
may be an inappropriate
by the RVV time,
since
the
phospholipid.
Pertinently,
over
the gradient,
with
ratio
of phospholipid
activity
than
the
than
studies
among
concentration,
membranes
the ratio
membranes
the
attributes
most
of
The findings
cannot
the
acid
Thus,
composition.
benchlatter
is
of phosphohaving
the
particulate
the
(Table
et al.”
fractions
alpha
granules
3). These
data
clot-promoting
be accounted
it seems
for
ap-
From www.bloodjournal.org by guest on June 17, 2017. For personal use only.
DISTRIBUTION
OF
propriate
whole
PHOSPHOLIPIDS
to
reconsider
platelet
as well
969
both
the
nature
and
as its subcellular
availability
PF-3 activity
may be defined
as a clot-promoting
mammalian
tissues
and is universally
unmasked
of the
cells.
in the
case
Intact
cells
with
keep
their
erythrocytes
platelets,
activity
is made
the cell’s physiologic
role.
with platelet
aggregation,33
The
source
of the
matter
of
(Table
1), should
pholipid
stimulated
in the
is not
However,
than
material
from
release
reaction
siderations
(1) The
makes
those
subcellular
(zones
of the
it is
effects,
4-9)
membrane
useful
to
PF-3
or
(zone
review
come to mind.
composition
of
from
aspects
simply
of
total
of
phos-
this
major
reorientation
of molecular
In
of
Three
themselves
PF-3
participants
to be determined.
PF-3
activity.
this
concon-
is important.
With
micelles
and liposomes
made
from
synthetic
phospholipids,
clot-promoting
tivity
is enhanced
as the negative
charge58
or the unsaturation
of the
acids4-7 increases.
However,
none
of our subcellular
fractions
contained,
percentage
basis,
an excess
of negatively
rated
molecular
species,
as compared
with
(2) In the natural
state
influenced
by interactions
the
clot-promoting
with
other
,
Niemetz
and
Marcus
have
recently
charged
phospholipids
other
fractions.
ability
membrane
demonstrated
promoters
by showing
that the procoagulant
in the presence
of whole
platelets.33
Possibly,
sented
a favorable
(3) We believe
tivity
as related
inner
acfatty
on a
diunsatucould
Pertinently,
platelets
as
be
clot
is enhanced
granules
pre-
surface
for the clotting
reactions
of the PF-3 assay.
that more
attention
should
be paid to the latency
of PF-3
to newly
tribution
of membrane
more
thromboplastic,
the
for
of leukocytes
intact
alpha
developing
concepts
of membrane
PF-3 activity
is normally
not accessible
to the plasma
reacts.
It is therefore
unlikely
that
the PF-3
reactive
surface
of the cell. In this context,
recent
information
on
or
of phospholipids
constituents.
a role
activity
our
a
the
The intact,
unrelease
reaction,
Whether
are
of
in parallel
concentration
that
a parallel
phospholipids
58%
available.
2) remains
enactment
If it were
with
the
erythrocytes
in that their
in the
known.
as
hypotonic
develops
reaction.9
its availability.
hidden.
The
activity
certain
the
point
constituents
unless,
to
Otherwise,
are unique
membranes,
role.
components
nection,
the
important
reactive
to other
in the
that resides
in all
total
disruption
exposed
in vitro
usually
upon
the release
a dominant
derives
activity
quiescent
are
at a critical
activity
phospholipid,
play
cells
would
appear
to be less
platelet
keeps
its PF-3
in addition
activity
available
PF-3
activation
but may depend
or
activity
the
nonelectrolytes.’#{176}’32
activity.
Platelets
clot-promoting
furnishing
capacity
partial
by
clot-promoting
and
media’#{176} or certain
penetrating
must be lysed to show clot-promoting
clot-promoting
of PF-3
fractions.
side
phospholipids
noncholine
of
the
cell
contact
with the plasma.
These
are clot promoting
in several
abundance
of more
negatively
lecular
species4’7
in their
mixed
man erythrocyte
various
combinations,
36
structure.
cofactors
material
on the
with
which
it
is on the outer
asymmetric
dis-
is of interest.
In human
erythrocytes,
phospholipids
are almost
exclusively
membrane’36
where
they
would
ac-
Clearly
have
no
the
located
direct
noncholine
phospholipids-diacyl
PE, PS, P1combinations,4’37
probably
owing
to the greater
charged
polar
groups5t
and diunsaturated
momicelles.
By contrast,
the outer
layer
of the hu-
contains
the
particularly
phospholipids,
those
with
PC and sphingomyelin,
the highest
PC content,
that in
have
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970
BROEKMAN
anticoagulant
properties.38
A preliminary
is almost
exclusively
located
on the
a basis
for the groupings
in Table
acid
composition
The
of the
foregoing
the basis
fractions
an
indicates
surface
show
of the membrane,39
providing
the remarkably
different
fatty
and
noncholine
explanation
for
phospho
the
fact
activity,
(Table
PF-3 activity
to its reaction
in association
3), requires
of intact
partners
be.’#{176}’2’
This
alpha
than
observation
activity,27
with
some
the
suggests
alpha
a relatively
explanation.
granules
(see Fig.
the PF-3
activity
that,
granules
5 of
of
membranes
also
be
PE
are,
ref.
intact
on
of the particulate
rather
vigorous
low phospholipid
It would
appear
in addition
can
platelet’s
lipids.
that
of phospholipid
phosphorus
content,
the least active
(Table
3). In the same
context,
the alpha
granules’
clot-promoting
phorus
content
PF-4
choline
seeks
the
AL.
report
inner
2 that
that
El
that
phosthe
13) is more
accessible
platelets
is known
to
to
holding
important
fibrinogen
and
contributors
to
PF-3
activity.
ACKNOWLEDGMENT
The
authors
Willcox,
acknowledge
and
Vera
platelet
suspensions
General
Hospital.
the
Martin.
were
highly
capable
technical
Through
the
cooperation
derived
from
blood
assistance
of
of
drawn
in
Dr.
the
Alice
Raynesford,
Charles
Blood
Bank
of
Eileen
E.
Huggins,
the
Massachusetts
the
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1976 47: 963-971
Distribution of phospholipids, fatty acids, and platelet factor 3 activity
among subcellular fractions of human platelets
MJ Broekman, RI Handin, A Derksen and P Cohen
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