Induction of neutrophilic granulocytosis in mice by administration of purified human native granulocyte colony-stimulating factor (G-CSF). Biochem. BiOpILyS.
Effect of recombinant
human
macrophage-colony
factor on marrow, splenic, progenitor cells in mice Takeshi Tanabe
Yamauchi,
Central
Abstract: colony-stimulating splenic, and
Research
The
Kouji Yada,
Akio
Umemura,
The
Cross
Corporation,
Laboratory,
effects factor
Green
of human (M-CSF) progenitor
peripheral and
macrophage on marrow, cells (CFU-M,
CFU-GM, administered tg/kg).
CFU-G) recombinant Single injection
were investigated human M-CSF of 4,000 igJkg
resulted
in
in
a
decrease
progenitor cells day 2 followed the
original
type
of
the
level
on
day
4
or
progenitors
5.
of
and
In
CFU-G) returning
for
800
j.tgfkg
of M-CSF CFU-G, as
marrow
in well
CFU-G.
The
present
of mice
with
pharmacological
man
M-CSF
affect
the
indicate
to
action
of
blood
and
J.
M-CSF.
cells
lineage
not
also of coincidence between cells and increase of cells suggests that from bone marrow
reseeded
Rio!.
spleen 59:
by the
296-301;
Words:
colony-forming
unit
[16-22].
the progenitor M-CSF affects
macrophage-CSF blood cells
However,
In the present human M-CSF
little
intensive (M-CSF) been
has
is known
progenitor
cells
[17,
of purified
M-CSF
about
its
19, 21]
ef-
because
in sufficient
cells in mice. We show the number of progenitor
progenitor cells from bone marrow lowed by settlement in the spleen
cheon the reported
quan-
study we utilized purified to clarify its hematopoietic
recombieffect on
here cells
mononuclear phagocyte lineage but lineage. In addition, the present results M-CSF may stimulate the mobilization
MATERIALS
that not
human only of
also of granulocyte suggest that human of hematopoietic
to peripheral in mice.
blood
fol-
AND
METHODS
Mice C3H/HeN
7- to 8-week-old
Japan)
were
Recombinant
prolferation
male
mice
(Charles
River
Japan,
Kanagawa,
used.
CSFs bridge,
1996. Key
of human of peripheral
transplantation
after
but
to the
Leukoc.
cell
stem
of hu-
ofprogemtor
only of monocyte/macrophage granulocyte lineage. Also, the decrease of marrow progenitor splenic and peripheral progenitor the progenitor cells are released to peripheral
4- to day 3 also of
blood
of granulocytopenia
unavailability
tities. nant
and Toshizumi
Japan
on hematopoietic
of the
Hanamura,
treatments
concentrations
number
Takuji
of autologous
previously
each
started
that
573,
source
fect
mice caused a decrease in as an increase in splenic
results
Osaka
The effect proliferation
on to
increase markedly on day 2, reaching a level 15-fold higher than that of the basal value on or 4. Peripheral CFU-M, CFU-GM, and CFU-G increased on day 2. In addition, administration
Eiji Asakura,
the
marrow
contrast,
tested
hematopoietic
and for treatment motherapy [9-15].
in mice (8-4,000 of M-CSF
number
(CFU-M, CFU-GM, by a gradual increase,
splenic
and peripheral
stimulating
human
MA.
from
the
cific
activity
It was
culture assay
>99% resis.
purity
with
supplied
by
units/mg mouse
bone
content
cells
cells
pg/1O
product
purified
had
a spe-
in a colony-
[23],
and gel
units/ml
Cam-
and
determined
sulfatelyacrylamide < 10
was
final
as
marrow
Institute,
ovary
The
protein,
dodecyl
Genetics
hamster
to homogeneity.
x 10
on sodium
Endotoxin
was
in Chinese
medium of 4
formation
M-CSF produced
as shown
revealed electrophoby Limlus
INTRODUCTION Colony-stimulating teins that progenitor jects and
(CSFs)
are
a family
stimulate colony formation from cells [1-4]. When administered
or experimental granulocyte-macrophage
proliferation tissues
factors
and
animals,
of the
progenitor
peripheral
blood,
hematopoietic to human sub-
granulocyte-CSF CSF (GM-CSF) cells as well
in
Abbreviations:
of glycopro-
the
(G-CSF) stimulate blood
the
296
59,
1996
Leukocyte
Biology
Volume
February
red
CSF,
colony-stimulating
granulocyte
blood
large-cell fraction white blood cells; FCS,
Reprint
cells
cells in hematopoietic important
of
RBC,
CFU;
in the peripheral blood [5-15]. Progenitor peripheral blood that are mobilized from the tissues by G-CSF or GM-CSF are clinically
Journal
G-CSF,
CSF;
cell;
WBC,
of white CFU-M,
tory, Osaka
573,
as
29,
Green
white
CFU; calf
requests:
The
Received tember
fetal
factor;
GM-CSF,
CFU-G,
M-CSF,
macrophage
granulocyte-macrophage
blood
blood cells; macrophage
granulocyte-macrophage
hematopoietic
as mature
CSF;
cell;
PLT,
CSF;
platelet;
WBC-LCC,
WBC-SCC, colony-forming
small-cell units;
granulocyte
CFU;
fraction CFU-GM,
CFU-S,
of
spleen
serum. Dr.
Cross
Takeshi
Yamauchi,
Corporation
2-25-1,
revised
September
Central
Research
Shodai-Ohtani,
LaborsHirakata,
Japan. July 1995.
17,
1995;
28,
1995;
accepted
Sep-
Table
ts of M-CSF
1. Effec
Treatment
progenitor
on the
Cells
on
Day
ber
Num
CFU-GM,
of Marrow
were
2.
and
added
or CFU-G cells
CFU-G
CFU-M
CFU-GM
x
103/femur
X l03/femur
Saline
5.13
±
6.86
±
1.82
1.29
±
8
5.10
± 0.90
6.86
±
0.48
1.12
± 0.13
80
3.84
±
4.11
± 0.71
0.81
± 0.12
800
3.30
± 0.09
3.98
±
0.33
±
Treatment (M-CGF,
gIkg)
0.85
0.30
0.11
Human
prepared
M-CSF
dilution
saline.
was
of
Control
intravenously
the
mice
original
were
injected
material
into
in
administered
an
0.
mice
2 ml
equal
Recombinant
bridge,
per
of hematological milliliter
(PLT),
WBC
of
total
(0.
2 ml)
of
determined
and
with
blood
blood
cells
small-cell
were
Genzyme
a Sysmex
blood
(WBC),
(Cam-
cells
large-cell
of WBC
Micro-cell-counter
(RBC),
fraction
of
(WBC-SCC)
F-800
day
before
three
mice
were
(day killed
obtained
by
a cardiac
spleens
were
excised
equal
volume
of
temperature (150
mm).
5
saline,
mixed ml
of
dosteum
were
with
McCoy’s
upper
McCoy’s
5A 5A
spleen
22.
on
in
resuspended
the
after
2 x
Splenic and
ice
for
suspensions in
reached
the
levels
counted,
and
McCoy’s
5A
numbers
seen
in control
Macrophage
assayed
Briefly,
bone
tured
in the
presence
medium
for
marrow,
humidified cells
made
splenic,
Petri
CO2
atmosphere
scored
as
granulocyte
pipetted
at
conditions
were
for the
determined blood.
of human
M-CSF,
added 1,
5.
0.0
;(
4.
(D :S
2.0
U.
0
were desired
0.0
0.3%
dishes.
agar The
and
colonies.
All
0.
5,
CFU-M,
10
to each and
inoculated
20%
cells
were
fetal
were
2.0
cul-
calf
serum
incubated
7 days.
cultures cell
Clones
were
ng of mouse
dish, 5
iO
and
cells/dish
for CFU-G
GM-CSF,
respectively.
x
number
except
CFU-GM,
or
In the
assay
were
4.0
semisolid
in 1 ml of McCoy’s
cultures
at 37#{176}Cfor
b
units
in
blood
or G-CSF
experimentally
For
6.0
C’)
colony-forming activities
or peripheral GM-CSF,
with
plastic were
peripheral
cells,
4 or
in
0
in a
performed
10
and the
cell
assays
CSF
.
the means
M-CSF
of the
(open
absolute
frotn
effects
Mice
were
symbols)
of marrow the
±SE
study
progenitors.
number
represent
G-CSF
Kinetic
marrow
of human
in units
5
4
of M-CSF
or saline
progenitors number
3 anitnals
of human
was point.
the
oti
intravenously (closed
counted
of progenitor
at each
M-CSF
injected
cells
days
per
P
