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Blood, Vol. 91 No. 11 (June 1), 1998:
pp. 4099-4105
Anti-LFA-1 Blocking Antibodies Prevent Mobilization of
Hematopoietic Progenitor Cells Induced by Interleukin-8
By
Johannes F.M. Pruijt,
Yvette van Kooyk,
Carl G. Figdor,
Ivan J.D. Lindley,
Roel Willemze, and
Willem E. Fibbe
From the Laboratory of Experimental Hematology, the Department of
Hematology, Leiden University Medical Center, Leiden; the Department of
Tumor Immunology, University Hospital Nijmegen, The Netherlands; and
Novartis Forschungsinstitut, Vienna, Austria.
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ABSTRACT |
Previously, we have shown that interleukin (IL)-8 induces the rapid
(15 to 30 minutes) mobilization of hematopoietic progenitor cells (HPC)
in mice. Because integrins are essential for adhesion and
transendothelial migration of HPC, we studied the involvement of the
 2-integrin leukocyte function-associated antigen-1
(LFA-1) in IL-8-induced mobilization. After a single injection of
blocking anti-LFA-1 antibodies, no mobilization of colony-forming
cells was observed. In addition, when mice were pretreated with
anti-LFA-1 or saline and subsequently injected with 30 µg of IL-8,
mobilization of HPC was completely blocked. We showed that this was not
due to anti-LFA-1 antibodies affecting colony formation, as addition of anti-LFA-1 antibodies to colony cultures in semisolid medium had no
inhibitory activity. Also, anti-intercellular adhesion molecule
(ICAM)-1 antibodies, directed to the main ligand of LFA-1 significantly inhibited the IL-8-induced mobilization. Furthermore, IL-1-induced mobilization was significantly inhibited by anti-LFA-1 antibodies. Because LFA-1 is reported to be expressed on more differentiated HPC, it was considered that the IL-8-induced
mobilization of more primitive HPC would not be blocked by anti-LFA-1
antibodies. Transplantation of blood-derived mononuclear cells (MNC)
from IL-8-mobilized animals pretreated with anti-LFA-1 antibodies
protected only 25% of lethally irradiated recipient mice, whereas the
radioprotection rate of control mice transplanted with MNC derived from
IL-8-mobilized animals was 86% (P < .01). Anti-LFA-1
antibodies did not interfere with stem cell homing, as transplantation
of IL-8-mobilized blood MNC, incubated in vitro with these antibodies
resulted in 100% radioprotection. We conclude that anti-LFA-1
antibodies completely prevent the rapid mobilization of colony-forming
cells and of cells with radioprotective capacity induced by IL-8. These
results indicate a major role for the 2-integrin LFA-1
in the IL-8-induced mobilization of hematopoietic stem cells.
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INTRODUCTION |
TRANSPLANTATION WITH mobilized progenitor
cells is increasingly being applied in the treatment of a variety of
hematologic disorders. The use of blood-derived stem cells is
associated with a more rapid restoration of the marrow function in
comparison with autologous bone marrow transplantation.1-3
This advantage is obtained only when the numbers of circulating stem
cells are greatly expanded using mobilization regimens. Although stem
cell mobilization is a property of most hematopoietic growth factors (granulocyte colony-stimulating factor [G-CSF],4-6
granulocyte-macrophage colony-stimulating factor
[GM-CSF],1 interleukin [IL]-3,7,8 stem cell
factor [SCF],9 flt-3 ligand10), relatively
prolonged treatment is required to induce mobilization. Only a few
cytokines, including IL-1,11 IL-8,12,13 and
macrophage inflammatory protein (MIP-1 ),14
induce rapid mobilization after a single injection. We have recently
shown that IL-8 induces rapid mobilization of cells with lymphomyeloid
repopulating ability in mice12 and of progenitor cells in
monkeys.13
Adhesion molecules have been implicated to play a role in retaining
progenitor cells in the bone marrow microenvironment.15-21 Among these, integrins are involved in the cellular interactions between hematopoietic progenitor cells, stromal cells, and components of the extracellular matrix. From the two major families, the 1-integrins, very late antigen (VLA)-4 (CD49d/CD29) and
VLA-5 (CD49e/CD29), as well as the 2-integrins leukocyte
function-associated antigen-1 (LFA-1) (CD11a/CD18) and Mac-1
(CD11b/CD18) have been reported to be expressed on progenitor
cells.16,20,22-27 In addition, ligands for these integrins
are abundantly expressed on cells of the bone marrow
microenvironment.17,19,28 For instance, fibronectin, a
component of the extracellular matrix, is a ligand for VLA-4, and
vascular cell adhesion molecule-1 (VCAM-1), as well as
intercellular adhesion molecule (ICAM)-1, which are ligands for VLA-4
and LFA-1, respectively, are both expressed on activated endothelial
and bone marrow stromal cells.16,17,19,28,29 Adhesion
blocking experiments have indicated that the VLA-4/VCAM-1, VLA-5/fibronectin, and 2-integrins/ICAM-1 pathways play a role in
the attachment of CD34+ cells to stromal
cells.16 The prominent role of VLA-4 in retaining progenitor cells in the bone marrow is shown by Papayannopoulou and
Nakamoto,30 who demonstrated that anti-VLA-4 antibodies, but not anti-2 antibodies, induce mobilization of HPC in primates. Likewise, adhesion molecules have been implicated to play a role in the
induction of stem cell mobilization.
In the present study, we have used antibodies against 1- or
2-integrins to study the role of these molecules in IL-8-induced stem cell mobilization. We found that anti-LFA-1 antibodies, but not
anti-VLA-4 antibodies, completely prevented the rapid mobilization of
colony-forming cells, as well as cells with radioprotective capacity,
without interfering with stem cell homing. These results indicate a
major role for the 2-integrin LFA-1 in the IL-8-induced mobilization of stem cells.
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MATERIALS AND METHODS |
Mice.
Balb/c mice, with an age ranging between 8 and 12 weeks and weight
between 20 and 25 g, were purchased from Broekman BV, Someren, The
Netherlands. Donor animals were fed commercial rodent chow and
acidified water ad libitum. Recipient animals were maintained in a
pathogen-free environment and fed water containing ciprofloxacin 1 mg/mL (Bayer Nederland BV, Mijdrecht, The Netherlands), polymyxin-B 70 µg/mL and saccharose 2 g/100 mL.
Experimental design.
Mobilization of HPC was induced by a single intraperitoneal (IP)
injection of 30 µg of IL-8. After 20 minutes, the mice were killed by
CO2 asphyxiation and peripheral blood was obtained by intracardiac puncture. In blocking experiments, mice were pretreated with an IP injection of anti-LFA-1 blocking antibodies or saline. The
following day, 30 µg of IL-8 was administered IP, and blood was
obtained after 20 minutes. In transplantation experiments, recipient
mice were placed in a polymethylmetaacetate (PMMA) box and given total
body irradiation (8.5 Gy, Philips SL 75-5/6 mV linear accelerator,
Philips Medical Systems, Best, The Netherlands), divided in two parts
in posterior-anterior and anterior-posterior position, at a dose rate
of 4 Gy/min. In survival experiments, a fixed number of 5 × 105 blood-derived mononuclear cells (MNC) obtained from
IL-8-mobilized donor animals pretreated with anti-LFA-1 antibodies or
saline, was injected in the tail vein of lethally irradiated
recipients. The experimental protocol was approved by the institutional
ethical committee on animal experiments.
Cytokines.
Recombinant human IL-8 was purified from Escherichia coli
(E coli) expressing a synthetic gene31 and provided
by the Novartis Forschungsinstitut, Vienna, Austria. IL-8 has no
colony-stimulating activity as reported previously.32 The
concentration of endotoxin was less than 0.05 EU/mL as determined by
the Limulus amoebocyte lysate assay. Human rIL-1 was kindly provided
by Hoffmann-La Roche (Nutley, NJ) and does not contain detectable
levels of endotoxin (<60 pg/mL). For in vivo experiments, all agents
were diluted to the desired concentration in endotoxin-free
phosphate-buffered saline (PBS) containing 0.1% bovine serum albumin
(BSA) and administered as an IP injection.
Monoclonal antibodies and detection of free circulating antibody.
Rat antimurine monoclonal antibodies directed against different
adhesion molecules were used: H154.163 (anti-CD11a, LFA-1, IgG2a,
kindly provided by Dr M. Pierres, Centre d'Immunologie, Marseille,
France),33 R1-2 (anti-CD49d, VLA-4, IgG2b,
kindly provided by Dr B. Holzmann, Department of Pathology, Stanford University, Stanford, CA),35 and YN1/1.7
(anti- CD54, ICAM-1, IgG2b).34 To detect circulating free
antibody, plasma was obtained from mice at various time intervals after
a single IP injection of 100 µg anti-LFA-1. A volume of 50 µL
plasma was incubated for 30 minutes at 4°C with 2 × 105 peripheral blood leukocytes of untreated mice. After
washing and labeling with phycoerythrin-conjugated goat antirat-IgG
(GaRa-PE) (Caltag, San Francisco, CA), fluorescence intensity was
analyzed by fluorescence-activated cell sorting (FACS) (Becton
Dickinson, Mountain View, CA).
Preparation of cell suspensions.
Mice were killed by CO2 asphyxiation. Blood was obtained by
intracardiac puncture and cell counts were performed on a Sysmex F800
(TOA Medical Electronics Co, LTD, Kobe, Japan). Manual neutrophil counts were performed after May Grünwald-Giemsa staining.
Blood-derived MNC suspensions were obtained by Ficoll separation as
described.11
Progenitor cell assays.
Colony-forming unit-granulocyte macrophage (CFU-GM) were cultured as
described previously.11 Briefly, peripheral blood MNC were
cultured in 3.5-cm dishes containing 5 × 105 cells
per mL in semisolid medium in the presence of murine GM-CSF (1.25 ng/mL). After 6 days of culture in a fully humidified atmosphere of
37°C containing 5% CO2, the number of colonies
(defined as aggregates of > 20 cells) were scored using an inverted
microscope.
Statistical analysis.
Differences were evaluated using the Student's t-test. In
survival analysis, differences were evaluated using the Mantel-Haenszel test for linear association. P values of < .05 were
considered statistically significant.
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RESULTS |
Effect of pretreatment with anti-LFA-1 antibodies on the mobilization
of progenitor cells by IL-8.
Balb/c mice were injected IP with a single dose of 100 µg of
anti-LFA-1 antibodies or saline. At various time intervals ranging from 30 minutes to 24 hours, the numbers of circulating progenitor cells were assessed. Anti-LFA-1 antibodies alone did not induce the
mobilization of progenitor cells. Free circulating antibodies could be
detected up to 72 hours after a single injection (data not shown).
Subsequently, mice pretreated with a single dose of anti-LFA-1
antibodies (100 µg) were treated after 24 hours with a single dose of
30 µg IL-8 IP. After 20 minutes, the mice were killed and peripheral
blood was obtained by cardiac puncture for progenitor cell assays.
Pretreatment with anti-LFA-1 antibodies completely prevented the
IL-8-induced mobilization of circulating progenitor cells (anti-LFA-1 + IL-8 46 ± 34, n = 21 v saline + IL-8 570 ± 724 CFU-GM/mL blood, n = 18; mean ± standard deviation [SD], P < .01, Fig
1). This effect appeared to be dose dependent, complete blocking being
obtained after injecting a dose of 100 µg
(Fig 2). Addition of anti-LFA-1 antibodies
to colony cultures in semisolid medium of IL-8-mobilized blood or
steady-state bone marrow had no inhibitory activity, indicating that
the lack of mobilization after pretreatment with anti-LFA-1 was not
due to interference of the antibody with colony growth in vitro (data not shown). Also, blocking antibodies to ICAM-1, the main ligand of
LFA-1, significantly inhibited the IL-8-induced mobilization of
colony-forming cells, although inhibition was not complete (108 ± 107 CFU-GM/mL blood, n = 8; mean ± SD, P < .01, Fig 1). In contrast, anti-VLA-4 antibodies did not block the IL-8-induced mobilization of HPC (728 ± 462 CFU-GM/mL blood, n = 10; mean ± SD, Fig 1), nor did they induce themselves the peripheralization of
progenitor cells. After a single injection of 300 µg anti-VLA-4, no
significant increase in the number of circulating progenitor cells was
observed (anti-VLA-4 19 ± 13 v saline 20 ± 17 CFU-GM/mL blood, n = 3). In addition, administration of anti-VLA-4
(300 µg) for 3 consecutive days did not induce mobilization of HPC (data not shown).
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| Fig 1.
Effect of pretreatment with anti-LFA-1 on the
mobilization of progenitor cells by IL-8. Mice were pretreated with a
single IP injection of 100 µg of anti-LFA-1 (H154.163, n = 20),
300 µg of anti-VLA-4 (R1/2, n = 10), 300 µg of anti-ICAM-11
(YN1/1.7, n = 8) or saline (n = 20). The following day 30 µg of
IL-8 was administered as a single IP injection 20 minutes before
harvesting peripheral blood. Results are expressed as mean ± SD. *,
P < .01 as compared with saline pretreated controls.
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| Fig 2.
Prevention of IL-8-induced mobilization of progenitor
cells by anti-LFA-1 is dose dependent. Mice were pretreated with a
single injection of increasing doses of anti-LFA-1 (10, 30, and 100 µg) or saline IP. The following day 30 µg of IL-8 was administered IP 20 minutes before harvesting peripheral blood. Results are expressed
as mean (n = 20 for saline and 100 µg anti-LFA-1, and n = 2 for
10 and 30 µg anti-LFA-1).
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Effect of anti-LFA-1 antibodies on blood cell counts.
Mice pretreated with anti-LFA-1 antibodies showed a consistent
thrombocytopenia 24 hours after injection, whereas mice treated with
saline or control antibodies did not (312 ± 60, n = 21 v 739 ± 109, n = 18; mean ± SD, P < .001, Table 1). The decrease in platelets started
2 hours after injection of the antibody, was maximal at 8 hours, and
lasted several days. Concomitant with the decline of free circulating
antibodies, the platelet count returned to baseline values. No
significant effect on the numbers of white blood cells, red blood
cells, or total number of granulocytes was observed after 24 hours. The
instant neutropenia and subsequent neutrophilia seen after IL-8
injection was unaltered in mice pretreated with blocking anti-LFA-1
antibodies before IL-8 injection (data not shown).
Effect of anti-LFA-1 antibodies on IL-1-induced mobilization of
progenitor cells.
Because IL-1 is a potent inducer of IL-836 and because it
also induces the mobilization of hematopoietic stem cells after a
single injection,11 we studied the involvement of LFA-1 in IL-1-induced progenitor cell mobilization. In these experiments, mice
were pretreated with anti-LFA-1 antibodies (100 µg) at 24 hours
before a single IP injection of IL-1 (1 µg). Circulating HPC numbers
were assessed at 5 hours after injection of IL-1. In comparison with
IL-1-mobilized control animals, mice treated with IL-1 after
pretreatment with anti-LFA-1 had significantly lower numbers of
circulating HPC (anti-LAF-1 + IL-1 423 ± 191 v saline + IL-1 628 ± 146 CFU-GM/mL blood; n = 6 per treatment group, mean ± SD, P < .05, Fig 3),
showing the involvement of LFA-1 in IL-1-induced progenitor cell
mobilization.
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| Fig 3.
Effect of pretreatment with anti-LFA-1 on IL-1-induced
mobilization of progenitor cells. Mice were pretreated with 100 µg of
anti-LFA-1 or saline. The following day, 1 µg of IL-1 (n = 6 per
treatment group) or 30 µg of IL-8 (n = 20 per treatment group) was
administered as a single IP injection, 4 to 6 hours or 20 minutes
before harvesting peripheral blood, respectively. Results are expressed
as mean ± SD. **, P < .01 and *, P < .05 as
compared with saline pretreated controls.
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Effect of anti-LFA-1 antibodies on the mobilization of hematopoietic
progenitor cells with radioprotective capacity.
The possibility was considered that pretreatment with anti-LFA-1
antibodies, while blocking mobilization of HPC, did not preclude the
IL-8-induced mobilization of more primitive HPC. To test this hypothesis, we studied the radioprotective capacity of the mobilized MNC fraction. Recipient mice were lethally irradiated (8.5 Gy) and
subsequently transplanted with 5 ×105 MNC, derived
from the peripheral blood of pretreated donor mice. The survival of
recipients transplanted with (5 × 105) MNC derived
from IL-8-mobilized animals pretreated with 100 µg anti-LFA-1 was
only 25%, not different from control recipients injected with saline
only (19%). In contrast, transplantation of (5 × 105) MNC from IL-8-mobilized animals pretreated with
saline protected 86% of the recipient mice (n = 20 per group in two
experiments, P < .01, Fig 4).
These results show that anti-LFA-1 blocking antibodies prevent the
IL-8-induced mobilization of progenitor cells with radioprotective
capacity. In control experiments, animals transplanted with 5 × 105 IL-8-mobilized MNC incubated in vitro with anti-LFA-1
blocking antibodies before injection into lethally irradiated recipient animals (100 µg per 5 × 106 cells for 30 minutes at
4°C) showed a 100% survival. Moreover, pretreatment of recipient
mice with 100 µg anti-LFA-1 antibodies, followed by transplantation
with 5 × 105 IL-8-mobilized MNC resulted in a
survival rate of 90%, indicating that these antibodies did not
interfere with homing (n = 10 per group, Fig 4).
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| Fig 4.
Survival of lethally irradiated recipients at 33 days
after transplantation. Recipient mice were transplanted with 5 × 105 blood-derived MNC from IL-8-mobilized animals (30 µg
IP) pretreated with anti-LFA-1 (100 µg IP) (anti-LFA-1 + IL-8) or
saline (saline + IL-8) at day -1. To exclude the possibility that
anti-LFA-1 antibodies would interfere with homing of HPC,
IL-8-mobilized blood was incubated with anti-LFA-1 (IL-8/anti-LFA-1)
in one control experiment. Furthermore, recipient mice were pretreated
with anti-LFA-1 (IL-8 + anti-LFA-1) as another control. Survival
data are expressed as absolute percentages of two experiments with 10 mice transplanted per group in each experiment.
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DISCUSSION |
Since the observation that cytokines are capable of inducing stem cell
mobilization, adhesion molecules have been suggested to play a role in
the process of mobilization.15,21,22,25,37 However, direct
evidence that these molecules are essential for mobilization has been
lacking. In this report, evidence is presented that the functional
expression of the 2-integrin LFA-1 is required for the IL-8-induced
mobilization of hematopoietic progenitor cells and shows the direct
involvement of the 2-integrin LFA-1 in cytokine-induced
mobilization. A single injection of anti-LFA-1 antibodies completely
prevented IL-8-induced stem cell mobilization. This was not due to the
effect of anti-LFA-1 antibodies on colony formation, as addition of
these antibodies to colony cultures in semisolid medium had no
inhibitory activity. As LFA-1 is reported to be expressed on more
differentiated HPC,22,23 it was considered that the
IL-8-induced mobilization of more primitive HPC would not be blocked
by anti-LFA-1 antibodies. This appeared not to be the case because the
radioprotective capacity of IL-8-induced MNC derived from anti-LFA-1
pretreated donor mice was not different from control recipients
injected with saline only. Moreover, transplantation of IL-8-mobilized
MNC in recipients pretreated with anti-LFA-1 antibodies or
transplantation with in vitro-incubated IL-8-mobilized MNC engrafted
normally, indicating that these antibodies did not interfere with stem
cell homing. These results are in accordance with the reported improved
engraftment in immunodeficient children after treatment with
anti-LAF-1antibodies as part of their conditioning regimen for
allogeneic bone marrow transplantation.38,39
Anti-LAF-1 antibodies have been reported to inhibit the migration of
lymphocytes through IL-1-stimulated human umbilical vein endothelial
cells.40 Möhle et al41 have used blocking
anti-LFA-1 antibodies to inhibit migration of CD34+ cells
in a transmigration assay using a bone marrow endothelial cell-derived
cell line. In the transmigrated cell population, primitive
CD34+ CD38 progenitor cells were virtually
absent, while in comparison with the starting cell population, the
transmigrated cells had a significantly higher plating efficiency.
These data indicate differences in the ability to migrate between
primitive and lineage committed progenitor cells. Indeed, LFA-1 was
found to be preferentially expressed on more mature HPC, while immature
progenitor cells seemed to lack expression.22,23 It is
therefore possible that the effect of anti-LFA-1 antibodies on
IL-8-induced mobilization is mediated by the inhibition of the
transmigration of progenitor cells through the bone marrow endothelium.
In accordance with this hypothesis, also anti-ICAM-1 antibodies,
directed to the main ligand of LFA-1 and expressed on endothelial
cells,41-44 significantly inhibited IL-8-induced
mobilization of HPC. The lack of expression of LFA-1 on primitive HPC,
however, would argue against such a mechanism and suggests the
involvement of an accessory cell expressing both LFA-1 and receptors
for IL-8.
The inhibitory effect of anti-LFA-1 on the mobilization of progenitor
cells does not only affect IL-8-induced mobilization, but also
mobilization induced by IL-1. In previous studies in our laboratory,
IL-1 has been found to induce HPC mobilization after an interval of 4 to 6 hours.11 Because IL-1 is a potent inducer of other
cytokines,36,45 the possibility was considered that an
intermediate cytokine, induced by IL-1, was responsible for this
effect. Because the production of IL-8 by endothelial cells is induced
by IL-1,36 the observations in this study are compatible
with the hypothesis that IL-1-induced mobilization is at least in part
mediated by the induction of IL-8.
The interaction between progenitor cells and bone marrow stromal cells
with 1-integrins appears to be essential for adhesion of
hematopoietic progenitor cells. Leavesley et
al46 reported that VLA-4, but not LFA-1, contributes to the
adhesive phenotype of steady-state bone marrow-derived
CD34+ cells. In vitro experiments have shown that blocking
anti-VLA-4 antibodies completely abrogate lymphopoiesis and severely
reduce myelopoiesis in long-term bone marrow cultures.47
Furthermore, in vivo, incubation of bone marrow cells with
anti-1-integrin antibodies before transplantation into lethally
irradiated mice inhibited the formation of CFU-S derived spleen
colonies, as well as the reconstitution of medullary
hematopoiesis.18 These data indicate the prominent role of
1-integrins in the retention and homing of progenitor cells in the
bone marrow microenvironment. In our experiments, we have used
anti-VLA- 4 antibodies to study the effect on IL-8-induced
mobilization. The antibodies did not influence mobilization or induce
mobilization when administered alone. These data are in contrast with
those reported by Papayannopoulou who found that VLA-4 antibodies were
able to induce the peripheralization of colony-forming progenitor cells
in primates and mice.30,48 These discrepancies may be due
to the use of other antibodies, recognizing different epitopes of the
VLA-4 molecule.
Mice treated with anti-LFA-1 antibodies exhibited distinct
thrombocytopenia. Studies from Tavassoli and Makoto49,50
indicated that megakaryocyte adhesion to endothelial cells is an
important physiologic process for platelet formation. Subsequently,
Avraham et al51 showed that LFA-1 is expressed on
megakaryocytes and that anti-CD18 antibodies inhibit the binding of
megakaryocytes to cytokine-stimulated endothelial cells. These
observations suggest a direct interaction between the binding of the
anti-LFA-1 antibody and platelet production. Because the
thrombocytopenia was already observed at 2 hours after injection, a
direct interaction with platelets is also likely. At present, the
sequence of events underlying this thrombocytopenia remains unclear.
In conclusion, we have found that anti-LFA-1 antibodies completely
prevent the IL-8-induced mobilization of progenitor cells with
colony-forming or radioprotective capacity. We showed that these
antibodies did not interfere with homing of IL-8-induced mobilized
progenitor cells after transplantation. Our data clearly show the
important role of the 2-integrin, LFA-1, in IL-8- and IL-1-induced
mobilization of HPC. Whether LFA-1 expression on hematopoietic
progenitor cells or on accessory cells is responsible for this activity
remains to be determined. Recent experiments from our group indicate
that the majority of the colony-forming cells in murine steady-state
bone marrow are LFA-1 negative.52 These observations do not
imply a direct inhibition of the anti-LFA-1 antibodies on the
egression of HPC through the bone marrow endothelium and indicate the
involvement of an intermediate cell population expressing receptors for
IL-8, as well as LFA-1. We are currently studying the hypothesis that
neutrophils may fulfill such a role. In accordance with this hypothesis
Liu et al53 have found severely impaired IL-8-induced
mobilization in G-CSF-receptor-deficient mice. Furthermore, activation
of neutrophils by IL-8 induces the release of the metalloproteinase
gelatinase-B (MMP-9), involved in the degradation of extracellular
matrix molecules.54 Preliminary experiments in monkeys have
indicated the rapid induction of MMP-9 by IL-8 and the blocking of
IL-8-induced mobilization of HPC by antibodies against
MMP-9.55 Taken together, these data are in line with a
central role for neutrophils and metalloproteinases in the induction of
IL-8-induced stem cell mobilization.
| |
FOOTNOTES |
Submitted November 11, 1997;
accepted January 26, 1998.
Supported by the Dutch Cancer Society (NKB) (Amsterdam, The
Netherlands) Grant No. RUL 95-1091.
Address reprint requests to Johannes F.M. Pruijt, MD, Department of
Hematology, Leiden University Medical Center, Bldg 1, C2-R, PO Box
9600, 2300 RC Leiden, The Netherlands.
The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" is accordance with 18 U.S.C. section 1734 solely to indicate this fact.
| |
ACKNOWLEDGMENT |
The authors thank Arie Boon of the Department of Radiotherapy for his
technical assistance in irradiating the animals and Peter de Jong of
the facilities for laboratory animals for his excellent animal care.
| |
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Abstract
Introduction
Abstract