From www.bloodjournal.org by guest on June 18, 2017. For personal use only.
Activation
of
Glycogen
Phosphorylase
in Blood
By
\Vith
B
LOOD
the
platelets
l)y
platelets,
the
must
be
mechanisms
relatively
metabolism.
lytic
nance
until
In muscle,
5’-AMP,
the
serves
form
as
an
activation
phosphorvlase
1) form.
Liver
the
Li forni
only,
although
electrical
which
is active
a form
in that
its
of its
activity
stimulation
of
either
the
active
molecular
muscle
through
without
or by
this
another
to
the
is more
of the
like
pres-
Epinephrine,
influence
it.7
also
other
the
and
Still
is independent
deactivate
from
a form
weight
contractio&#{176}
AMP
utilized.
contraction.
and
activate
enzyme
b kinase,
reduce
level.
active
Transformation
muscular
to
the
influences
on
the
phos-
1) kinase.
the
present
study
human
platelets
hence,
phosphorvlase
determine
From
time
1/mis
Decenmher
First
RoBER1’
activity
of
vitro
under
in
activation.
synthesis
been
favoring
of
were
during
Medicine,
measured
in
glycogenolysis-
experiments
continued
De,)artmermt
Dwzsio,m,
has
conditions
Additional
glvcogen
hematology
phosphorylase
performed
active
Medical
to
glycogenolysis
College
of
Virginia,
from
time
Natioimal
imrginw.
Supported
Cancer
the
incubated
whether
Ricimmond,
Ricimmond,
be
serves
majority
glycogen
could
phosphorylase
enzymes
of phosphorviase,
phorvlase
In
the
similar
enzyme
mechanisms
enzyme
differs
The
to
5’-AMP.
during
mainte-
of functioning
a constant
requires
phosphatase,
and
and
activity
the
mechanism
I)llosphorylase
of 5’-A\IP
glucagon.)
by
proteo-
the
unavailable
in a tetrameric
which
is iiiediated
enzyme,
ence
exists
balance
is a
from
for
energy.
The
the
platelet
of glycogen
protected
maintain
be
of these
b form
a dimeric
(1
may
combination
some
phosphorvlase
and
may
substrates
the
mechanisms
used.
mole-
control
for needed
to be
phosphorylase
or
or
waiting
gram
possesses
reason
of the
possible
for
of porcine
decreased.4
longer
this
somehow
of
its degradation
form
and
co-factors
needed,
and
inactive
synthetase
Necessary
1) to
reserve
in a totally
enzymes
a gram
phosphorylase
no
aspects
example,
clotting
methods,
platelet
to study
a number
on
glycogen
For
as dem-
For
in vitro
chemical
by
synthesis.
of
are
muscle
anucleate
a complement
of glycogen,
microscopy.3
during
preexisting
enzyme
Cooper
amounts
skeletal
measured
the
\V.
electron
that
in which
there
of a glvcogcn
glycogen
as
new
system
Given
be
as
reported
since
inducing
degradation,
may
been
content,
simplified
large
and
glycogenolysis,
utilized
for
relatively
glycogen
much
to initiate
of LaVerne
analyses2
it has
glvcogen
In order
cules
contain
as
Furthermore,
assi.slance
biochemical
contain
basis.2
B. Scorr
ROBERT
technical
PLATELETS
onstrated
Platelets
hi1
Institute
work
U.
was
6,
S.
and
Public
time
))resented
health
Blood
in
Serrice
Re,searcim
part
at
time
Researcim
Fund
of
meeting
time
of
Grant
CA08482
Medical
College
time American
of
Society
Virginia.
for
Hematology,
1966.1
submitted
B. Scour,
Jan.
29,
NI.D.:
1967;
accepted
Assistant
for
Professor
publication
of
Marc/i
27,
Medicine,
Medical
VOL.
30, No.
1967.
College
of
Virginia,
Virginia.
321
BLoon,
3
(SEPTEMBER),
1967
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322
ROBERT
and to determine
glycogen
stores.
the
relative
dependence
of coagulation
and
clot
B.
SCOrE
retraction
on
METHODS
glassware
was siliconized.
Normal
donor
blood
was obtained
through
plastic
tubing
into iced centrifuge
tubes
with one-tenth
volume
of 2 per cent disodium
ethylenediaminetetraacetate
( EDTA).
All operations
prior
to incubation
were
performed
at 4 C. Platelet
rich plasma
( PRP)
was removed
after centrifuging
the blood
at 200 g for 20 minutes.
A
few
lymphocytes
were
usually
present
in the plasma.
The PRP was mixed
well,
samples
taken
for platelet
counts,
and the remainder
then
divided
into separate
aliquots
for incubation.
Separate
aliquots
were
incubated
for each
time interval
to be measured.
A pair of 1 or 2 ml. samples
were incubated
for each interval
glycogen
determination
and a 10 to 20 ml. aliquot
for each enzyme
assay.
When
incubations
were to be made
in buffered
saline or Hanks
solution,
the aliquots
of PRP were centrifuged
at 10,000
g for 20 minutes
( to insure
complete
recovery
of the platelets
) and pellets
gently
resuspended
in the salt solution.
Hanks
solution
included
0.2 ml. EDTA
per 10 ml. soiution.
Samples
for glycogen
assay
were
resuspended
in 1 or 2 ml. volumes
of salt solution
and
All
samples
for
enzyme
assay
out in a 37 C. water
were
resuspended
in
2
ml.
volumes.
Incubations
were
carried
terminated
by icing
the samples.
Platelets
were then centrifuged
from suspension
at 10,000
g.
Isotonic
saline
solution
for suspension
was buffered
to p11 7.4 with
0.01 NI Tris-HC1
and
sterilized
by membrane
filtration.
Complete
Hanks
solution
was
obtained
commercially
( Baltimore
Biological
Laboratory
) . Glucose-free
Hanks
solution”
was prepared
in the
laboratory
and sterilized
by membrane
filtration.
Glycogen
bath
and
Determination
Duplicate
samples
treated
with
precipitated
pended
in
glucose
of
distilled
which
had
been
centrifuged
KOH
at 100 C. for 20 minutes
The precipitate
was
centrifuged
platelets
1 ml. of 30
with
1.1 ml.
per cent
ethanol.
water.
Clycogen
was
estimated
l)y
the
from
and
suspension
the
at
of
resus-
Seifter,’2
using
Assay
Platelets
which
had been
centrifuged
from suspending
me(lia
were
containing
0.34
NI sucrose,
0.001 NI EDTA,
0.1 NI NaF,
were
disrupted
by 10 seconds
sonication
at 3 amp. in a Branson
Niodel
were
quickly
reiced.
Previous
experiments
showed
that this period
of
minimum
sufficient
to release
97 per
cent of the glycogen
into
a 1000
ml. of a solution
disruption,
the
enzyme
sonicate
activity
acid-washed
added
was
and
synthesis
sodium
cent.
the
method
added.
Each
performed
most
of
assay
activity
The
after
inorganic
method.’
and a controi
activity
was
Results
-15
were
of protein.
was determined
Enzyme
run
found
expressed
according
found
arid
of
platelet
to
to Lowry
the
0.002
suspensions
mixtures
from
NI
the
blanks
5’-AMP.
the
the
inorganic
et al.’ Because
M
sodium
sonicate
was
Assays
were
enzyme
loses
at 37 C. for
incubated
run
of
0.02
M
of
glucose-i-phosphate
were
direction
contained
since
were
with
without
the
ml.
for
treated
0.05
0.05
After
activity
in
mixture
0.4
assayed
was
reduce
which
without
Reaction
micromoles
g supernate.
supernate
performed
reaction
without
glycogen
in
to be linear
throughout
in
to
in
at pH 7.0. Platelets
5-75 sonifler
and
sonication
was the
(charcoal-treated)
ml.)
with
reagent
the
resuspended
supernate
was
The
glycogen
liberated
and
and
the
assay
0.5
C.
phosphorous
extract
Enzyme
used.
cent
incubation
at
was
of Steiner.14
both
minutes
experiments,
This
volume
performed
overnight
early
10
Phosphorylase
per
(total
Comori
platelet
activity.
milligram
Protein
was
immediately
its
minutes.
the
1
6.5
g for
5’-ANIP.
per
glucose-i-phosphate,
pH
2000
in
remove
15
following
glycerophosphate,
at
Except
to
about
by
spun
protein.
charcoala
5’-AMP
glycogen
tions
was
and
standards.
Phosphorylase
by
g
1000
method
were
glycogen
was
concurrently
with
mixture
the range
showed
of protein
phosphate
(P1)
of turbidity
due
20
estimated
each
insignificant
concenhaper
hour
to glycogen,
per
From www.bloodjournal.org by guest on June 18, 2017. For personal use only.
ACFIVATION
samples
OF
of
the
GLYCOCEN
sonicate
) and
concentration
Topical
PHOSPHORYLASE
were
first
redissolved
thrombin,
precipitated
BLOOD
with
323
PLATELETS
trichloracetic
(5
acid
per
cent
final
in 0.1 N NaOH.
NIH
100
IN
units
per
ml.,
was
dialyzed
against
2
changes
of
distilled
water
overnight
to remove
part of the anthrone-reacting
material
in it. Dialyzed
samples
were treated
with hot alkali and ethanol
precipitation
to obtain
correction
factors
for the
glycogen-like
material
it contained.
Glucose-C14
( uniformly
labeled
with specific
activity
182 mc. per mM ) was dissolved
in
water after removal
of alcohol
in a vacuum
dessicator.
The final concentration
was 500 zc.
per ml. For incubation,
platelets
from 1 ml. aliquots
of plasma
were resuspended
in 0.5 ml.
of
Hanks
solution
made
without
glucose.
After
incubation,
1 mg. carrier
glycogen
was
added,
and glycogen
was
isolated
by KOH
treatment
and
reprecipitated
tilled water.
Aliquots
of the final solution
were dried
on copper
planchets
counted
in a low background
gas flow counter.
Appropriate
corrections
self-absorption.
Statistical
Additional
analyses
were
samples
performed
were incubated
and assayed
according
to Snedecor.’7
once
from
dis-
and radioactivity
were
made
for
glycogen
content.
for
RESULTS
Effect
of
Incubation
Medium
on
Phosphorylase
Phosjmhorylase
Activity.
Enzyme
Activation
and
Glycogen
Depletion
Total
activity
in the
designated
“total”
activity
since
it also includes
present
without
added
5’-AMP.
True
total activity
ured
if every
phosphorylase
interest
in these
experiments,
of activation
of
genolysis
has
When
plasma,
sively
phosphorylase
during
1 hour
the
incubation
platelets
were
salt
solution.
buffered
represents
protein
prior
assays
at 37
In
platelets
saline
made
on
activity
once
group
in the
most
closely
from
experiments
5’-AMP
(presumed
percentage
of the
from
charcoal-treated
which
corrections
after
active
glyco-
in Hanks
to
“total”
the
be
were
the
activity
homogenates
vivo
incubated
active
form
in
made
value,
1. In this
of
or in
activation
plasma,
centri-
contaminating
protein
interfered
from
little
simultaneously
activation
phosphorylase)
took
zero time
in plasma,
is 0.944
case
plasma
with
designated
“plasma”
in plasma,
centri-
0.605-1.389).
Figure
2 represents
in Figure
7.4)
phosphorylase
experiments,
the
The
initial
activity
in
occurred
( pH
in plasma
and significant
when
The lowest
curve
in Figure
1
Although
mg. protein
(range
without
5’-AMP.
shown
activation
saline
that
were
solution.
from
platelet-rich
increased
progres-
) . Similar
preparations.
The
curve
assays
on platelets
incubated
represents
hour
per
Activity
same
1 it is seen
of plasma
incubation
washed
platelets.
moles
P1 per
Phosphorylose
the
Figure
calculations,
washed
1
and not washed
to remove
Since
the contaminating
and unwashed
platelet
platelets)
represents
during
either
was
greater
( Fig.
C.
in Tris-buffered
platelets
washed
(washed
probably
and
resuspended
“total”
activity
were
incubated
or Hanks
solution.
at the end of incubation,
before
enzyme
assay.
and
is
activated.
Of particular
of the relative
state
glycogenolysis
resuspended
specific
fuged,
to
of 5’-AMP
which
would
be
that activity
meas-
begun.
was negligible
when
incubated
in buffered
fuged
were
maximally
is the determination
platelets
were
isolated,
carefully
and incubated
in salt solutions,
the
whether
Hanks
molecule
however,
presence
that activity
would
be
±
0.187
additional
the
activity
is recorded
at intervals
of incubation
at 37 C. Only
are shown.
In untreated
homogenates
place
activity
which
(SE)
data
without
as a
results
the
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324
ROBERT
PLATELET
B.
SCOTE
PHOSPHORYLASE
6.0
Fig.
specific
5.0
1.-Glycogen
activity
in
phosphorylase
platelets
suspended
media.
The activi-
.
1
4.0
E
in
various
tv
the
is
incubation
“total”
of three
30
activity
assay
the
or
bars
more
plasma
taminating
washing
(enzyme
#{149}
in
platelets);
washed
prior
made
for
to
con-
#{149}
in Hanks
proteins);
plasma
means
Q incubation
plasma
iii
(platelets
not
but
correction
assay
are
determinations;
range;
..
after
in
included
points
separate
saline;
assayed
1.0
The
represent
buffered
ao
(ANIP
mixture).
solution.
0
20
40
MINUTES
60
INCUBATION
80
Fig. 2.-AMP-independent
phosphorylase activity
during
incubation
of platelets
in various
media.
All enzyme
PreParations
were
pretreated
to
remove
AMP
before
aSSay.
Each
point
represents
the
mean
except
for a single
determination
60
4O
10
at
20
niinutes
curve;
01
0
20
40
buffered
solution;
60
MINUTES
starting
values
were
slightly
higher
bt
the
curves
were
the
as active
without
Glycogen
at least
and
5’-AMP
as
Degradation.
in saline
in the
salt
cant,
whereas
than
solution.
The
glycogen
and
plasma,
t test
AMP
similar.
increased
to
resulted
in a decrease
in
1) continued
to increase
enzyme
the
60-minute
incubation
is only
with
the
differences
in saline
glycogen
values
correlating
revealed
depletion
Hanks
(1umalitativelv
requiring
or Hanks
the
1 represents
zero
in
saline
range;
media.
about
10 p’
assays
of
cent
it is Present.
when
Table
periments
in Figure
2.
The difference
between
greater
in the
unincubated,
incubation
represent
medium;
plasma.
throughout
60 minutes,
plasma
bars
#{149}
incubation
the proportion
of activity
not
at about
20 minutes.
Further
incubation
even
though
the total
activity
(Fig.
in plasma
the
saline
During
the
a maximum
this fraction
In platelets
on
horizontal
was
in each
greater
the
experiment
enzyme
in plasma
statistically
cxwas
activation
to l)e insignifisignificant
(p
<
0.025).
In
mation,
(not
the
preceding
or
shown)
clot
experiments
retraction;
were
performed
yet
there
glycogenolysis
in which
was
no platelet
agglutination,
occurred.
coagulation
was
fibrin
Additional
experiments
initiated
by recalcifica-
for-
From www.bloodjournal.org by guest on June 18, 2017. For personal use only.
ACHVATION
OF
Table
CLYCOCEN
PHOSPHORYLASE
1.-Degradation
of Platelet
Incubation
Me(lium
4
Plasnia
5
gig. glvcogcii/lO
*
tion
or
As
to
tion
would
these
for
Continued
possibility,
Clycogen
results
and
incubating
from
the
11.9
mg.
retraction
that
minutes.
coagulation
time
was
Thus,
of
solution,
incubated
incul)atiOnS.
at
TIm
10, 30,
and
final
C.
amount
60 minutes
Radioactivity
the
5
37
at
was
specific
in-
activity
) . \%Then the same experiment
ml. ) added
at zero time,
the
2
per
during
and
was
not
a cessation
were
time
of glycogen
Glycogen
and
of incubation
for
results
retraction
constant
the
made
maximum
glycoagula-
shown
in Figure
time
changed
lengthened
glvcogen
although
of
was
content,
coagulation
a decreasing
relatively
cent
the
the
complete
with
stores
( to insure
solution
measured,
taken
measure
A
on glycogen
at intervals.
correlated
to 15 per
Glycogenolysis.
retraction
Hanks
4 hours
remained
reduced
deple-
To investigate
Hanks
was
and
there
aliquots
was
synthesis.
glycogenolysis.
Time
but
This
units
In addi-
glvcogen
isolated.
( Table
glycogen
time
unclotted
Glycogenoltjsis.
parallel
intervals
( 5 NIH
during
phosphorylase
in the
glucose-free
been
all
in glucose-free
and
additional
activated
samples
had
at
recalcifying
he seen
centration
33.0*
5.7
mixture
on
glycogen
of coagulation
2.5 to 3.5 minutes,
13 to 25
12.0*
phosphorylase,
in
the
duplicate
similar.
platelets
time,
Miii.
57.9±
of glycogen
and
glycogen
Retraction
and
3. It can
determined
period
dependence
cogenolysis)
tion
was
per
of
suspended
added,
thrombin
the
Coagulation
by
Media
‘call
I)ifference
89.2±
during
activation
was
qualitatively
during
Synthesis
the
by cessation
the
c.p.m.
with
were
relative
to
into
platelet
X 10
synthesis
in Various
conditions.
were
after
into
performed
already
platelets
determined
4.96
significant
incorporated
corporated
was
no
was
favored
concentration
was
expect,
enzyme
Glycogen
glucose-C14
of radioactivity
was
(SE)*
the
due
be further
ml.
60
11.1
experimental
glycogenolysis
per
/sC.
Incubation
would
since
under
Evidence
tion
during
16.6
±
325
PLATELETS
Time
91.0±
101.1
one
occurred,
suspension
Zero
BLOOD
platelets.
thrombin.
activation
this
Glycogen
No. of
Lxierinents
Saline
IN
from
content.
the
Thus
glvcogen
con-
of normal.
DISCUSSION
In
these
experiments,
unphysiologic
vation
of platelet
results
in
activity
tion
(Fig.
turbed
trauma
it, like
was
of the
may
than some
the mature
in phosphorylase
suspended
vivo.
indicates
It may
The
no
be
that
platelets
to
resulted
that
slight
even
solution
greater
platelets
solutions,
total
resuspension.
somewhat
other
cells.
erythrocvte,
of
salt
in Hanks
during
represent
in
transfer
increases
in Platelets
platelets
that
as buffered
This
samples
also
seen
such
phosphorylase.
detectable
occurring
1)
it
environment,
a
The
or saline
represents
activity
are
than
more
acti-
manipulation
activity.
Accordingly,
relatively
in the
higher
the
present
sensitive
initial
activa-
lower
curve
in
undis-
to
such
platelet
is such a highly
specialized
cell that
longer
has the capacity
to alter its metabolism
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326
ROBERT
Table
2.-Incorporation
of
GIucoseC14
Solution
and
in Hanks
into Platelet
Glycogen
after Thrombin
Addition
Glucose.Free
Solution
Specific/Activity
1nculation
lime
Hanks
during
Ilanks
l’lus
S;ecitic
‘
B.
Incubation
Solution
Thronthin
:tctivity
10 nih.
2.55
X l0
5.98
X l0
30
miii.
2.91
X
10’
7.75
X
60
niii.
4 96 X
101
* Counts
minute
per
ler
lig.
SCOTT
10
14.35 X
101
glvcogen.
Fig. 3.-Coagulation
and clot retraction times correlated
with glycogenolysis
during
incubation
of platelets
in Hanks
solution.
Clotting
was
instituted
by recalcification
of
the
EDTA-containing
00
60
20
medium.
.
)
tion
0
60
120
180
clotting
glycogen
Q
time;
temperature
#{149}
retraction
Incuba-
time;
concentration.
was
37
C.
240
MINUTES
except
in a limited
may
be one
When
measured
activity
zyme,
indicates
prior
to
course
Later
late
decline
ase
phosphatase
the
data.
tion
of the
that
platelets
some
liver
AMP-dependent
kinase
is present
to stress.
per
cent
of
resembles
enzyme
AMP-independent
activity
continued
to
phosphorylase
contributing
both
phosphorylase
fraction
began
increase,
of ATP
the muscle
enis in the h form.5
through
an
If the platelet
dependence
and
in saline,
the
final
The
activation
As pointed
react
90
the enzyme
all of the
a burst
can
increase
in
enzyme
is
on AMP
to the
activa-
activities
in-
to decrease.
more
than
one
Since
activation
must
he implicated.
Activation
mechanisms
other
than
the b to a
are known
to exist since
in certain
mice whose
skeletal
muscle
lacks
h kinase,
near-normal
glycogenolysis
takes
place
during
muscle
contraction.1
6-phosphate,
to provide
platelet
in plasma,
respect
nearly
of activation
AMP-dependent
maximum
the
incubation
a phosphorylase
the
initially.
mechanism
transition
phosphorylase
of glycogen
of ways
platelet
phosphorylase
occurred
both
and the fraction
independent
of AMP.
to muscle
phosphorylase,
then decreasing
that
During
creased
the
Degradation
number
required
AMP.
In this
where
prior
to contraction
Activation
of
“total”
activity
indeed
similar
tion.
way.
of a limited
activity
possible,
(with
since
in AMP-independent
or
some
out
by
Morgan
and
inorganic
activities
type
and
of
not
could
Parmeggiani,11
enzyme
b and
of liver
were
shown
levels
also
present,
in Figure
and
may
a. Another
muscle
this
still
followed
represents
mechanism
orthophosphate
a mixture
activity
at 60 minutes
was
activity
other
of phosphorylase
contain
AMP)
this
might
1. Leukocytes,
may
further.
the
action
not
be
the
the
of phosphoryl-
AMP,
a role
possible
types
be
ascertained
of ATP,
play
not
\Vhether
from
glucose-
in the
regula-
consideration
of phosphorylase.
explain
in which
the
increase
phosphorylase
is
If
in
From www.bloodjournal.org by guest on June 18, 2017. For personal use only.
ACTIVATION
OF
resembles
tion
with
The
glycogen
GLYCOCEN
the
liver
ATP
and
type,
show
turnover,
as has
AMP-dependent
reported
for
been
should
during
of the
taking
glycogen
synthesis
measured
during
part in the coagulation
process
has
Unger4
traction
of
clotting.
Similarly,
depleted,
porcine
the
greater
Since
ATP
levels,
it is not
Under
utilized
of
surprising
may
ruled
of
on
lessened
that
adequate
an
retraction
of
high
relatively
place.
Under
a great
deal
intracellular
than
did
was
glycogen
attempt
also
re-
glycogen
stores
to
maintain
is correlated
with
concentration
additional
ATP
compared
may
be
with
required
for
in vivo conditions
with
vastly
greater
fibrin
more
glycogen
would
be degraded.
Also, the
platelet
in these
platelet
less
fibrinogen23
experiments
and
could
fibrinogen
not
absorbed
fibrinogen
retraction
of muscle
myosin
and
both
to
formation,
contribution
to the
outer
perof
platelet
be ascertained.
The fact that much
glycogen
degradation
is associated
with retraction
sistent
with
the present
concept
of a contractile
protein
in platelets
the retraction.
The contractile
protein,
thrombosthenin,
has many
of the
ties
fully
) that
glycogen
of their
some
not
conditions
much
is probably
that
possibility
be in platelets
out.
amounts
until
retraction
breakdown
no quanthat glycosolution
and
in Hanks
different
larger
C14-glucose
into
were
in a state
of
Although
The
clotting
not been
platelets
dependence
glycogen
conditions
surface
incuba-
levels.2
concentration,
take
haps
after
it is evident
place.
somewhat
preincubating
of
glycogen.21
taking
( under
platelets
by
evident.
ATP
were
showed
was
lowered
retraction
liver
be attempted,
glycogenolysis
coagulation
and
activity
the incorporation
if platelet
glycogen
while
Weber
327
PLATELETS
utilizing
to learn
of the data
active
both
and
BLOOD
ions.2#{176}
reported
primarily
titative
interpretation
gen synthesis
is still
IN
increased
magnesium
experiments
were
done
constant
PHOSPHORYLASE
have
ATPase
activity.24
is concausing
proper-
Phosphorylase
activa-
tion and glycogen
degradation
are concomitants
of the action
of both
these
ATP requiring
contractile
systems.
Both platelets
and muscle
are of mesodermal
origin.
It is possible
that the platelet
and muscle
enzymes
will be shown
to be
similar;
this suggestion
is supported
by the properties
described
in these
experiments.
The muscle
enzyme
is believed
to differ
from the liver
and Hela
cell
enzyme,
both
Although
functions
may
of which
clotting
which
result
in
phagocytose
this
since
platelets
would
If
result
and,
energy
and
from
ectodermal
be
assuming
may
of the
lysosomal
structures.
most
important
glycogen
to leukocyte
which
to
phagocy-
breakdown.
of these
granules
breakdown
and
platelet
be other
circumstances
have
been
demonstrated
in glycogen
some
rupture
the
analogy
result
granules,29
part
then
from
may
there
may
Platelets
matter,27
lysosomal
so,
derived
retraction
glycogenolysis,
glycogenolysis.
require
contain
amylase.
clotting
the
require
platelet
would
tissues
clot
particulate
tosis,
contain
are
and
Also,
may
occurring
disintegration
of
well
during
platelets
in
clot.
These
studies
changing
standing
which
have
glycogen
of the
are
described
requirements
mechanisms
intimately
associated
ways
in which
a single
enzyme
behaves
during
in a specialized
cell.
A more
complete
under-
of the
enzymes
involved
with
will
require
particulate
data
glycogen
on
all
in the
platelet.
For
From www.bloodjournal.org by guest on June 18, 2017. For personal use only.
328
ROBERT
instance,
Evidence
even
than
what
happens
to glycogen
synthetase
presented
suggests
that some glycogen
during
clotting.
that of glycogen
Obviously
synthetase
the effective
under
these
activity
synthesis
activity
during
takes
B.
SCOTT
glycogenolysis?
place,
perhaps
of phosphorylase
is greater
conditions.
SUMMARY
In platelets
1.
the
degree
the
incubated
in vitro,
of activation
roughly
platelets
and is greatest
2. Activation
proceeds
active
activating
3.
Total
tinues
enzyme
suggesting
more
During
4.
induced
retraction
was
while
the
than
one
glvcogen
by
suggesting
kinase,
( activity
activity
to increase
of manipulation
degree
with
activating
)
AMP
of
medium.
enzyme
the
likewise
fraction
activated,
of
which
is
phosphorylase-
increases
begins
and
and
con-
to decrease
again,
mechanism.
and
during
glycogen
synthesis
to be more
dependent
coagulation
was
and
clot
shown
to continue
on adequate
glycogen
retraction
in vitro.
stores
Clot
than
coagulation.
SUMMARIO
1 . Phosphorvlase
(IC
was
to the
the action
in platelets.
AMP-independent
breakdown,
thrombin,
was found
phosphorylase
in the least physiologic
incubation
with
an increasing
proportion
of
the absence
of AMP,
enzyme,
phosphorylase
in
glycogen
corresponds
corresponde
activation
1)1t15
(IC glycogeno
marcate
in
Ic
de
procecle
(IC
activatori,
Le
accresce
monophosphat()
de
incul)ation
Ic
minus
plachettas
in vitro.
incubate
manipulation
dcl
de activation.
in vitro
que
position
de
inciucite
per
glycogeno
e
thrombina.
dependeva
plus
le
Esseva
que
le
le
durante
trovate,
coagulation
lo
synthese
de
processos
de
plus
de
un
grados
Illos
es
Ic
physiologic.
declinar,
recomencia
Le
plachettas.
con
(1#{128}’
ll(IellOSlflll
(lii
4.
in
al grados
in
le
adenosina,
sol mechanismo
Esseva
monstrate
(itIC
INTERLINGUA
activate
urn crescente
proportion
ic enzyma
active
Jo que
suggestiona
que
que
Ic cnzyma
kinase
de phosphorylase
es active
in Ic plachettas.
activitate
enzymatic
total
( i.e., activitate
con
monophosphato
similemente
.
Illo
continua
accrescer
quando
le fraction
non
monopliosphato
3.
grossierrnente
medio
2. Le activation
IN
esseva
que
suggestiona
glycogeno
e
specificamente,
adequate
de
de
adenosina)
(IC
de
durante
plus
decom-
IC
(IC
retraction
del
coagulo
le
retraction
del
coagulo
que
reserva
(iC
(iepen(ientc
k’ existentia
continua
coagulation
absentia
phosphorylaso-
glycogeno.
ACKNOWLEDGMENTS
\Ve
of
wish
this
thank
to
Dr.
manuscript,
analyses,
and
Dr.
Dr.
C.
J.
Aaron
K.
Marcus
Mullen
\Vatson
James,
for
and
Mr.
S.
III,
for
his
many
E.
helpful
Ketner
continued
suggestions
for
in
assistance
interest
the
with
and
preparation
thC
statistical
support.
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1967 30: 321-330
Activation of Glycogen Phosphorylase in Blood Platelets
ROBERT B. SCOTT and LaVerne W. Cooper
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