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 Previous Article | Table of Contents | Next Article 
Blood, Vol. 93 No. 10 (May 15), 1999:
pp. 3317-3326
Modulation of Integrin Function in Hematopoietic Progenitor Cells by
CD43 Engagement: Possible Involvement of Protein Tyrosine Kinase
and Phospholipase C-
By
Naoyuki Anzai,
Akihiko Gotoh,
Hirohiko Shibayama, and
Hal E. Broxmeyer
From the Departments of Microbiology/Immunology, Medicine, and the
Walther Oncology Center, Indiana University School of Medicine,
Indianapolis, IN.
 |
ABSTRACT |
Attachment of cells to extracellular matrix components is critical
for the regulation of hematopoiesis. CD43 is a mucin-like transmembrane
sialoglycoprotein expressed on the surface of almost all hematopoietic
cells. A highly extended structure of extracellular mucin with negative
charge may function as a repulsive barrier to hematopoietic cells.
However, some investigators have shown that CD43 has proadhesive
properties, and engagement of CD43 has been reported to upregulate
integrin-mediated cell adhesion in T cells. We found that cross-linking
of CD43 with monoclonal antibodies (MoAbs) enhanced integrin  4 1
(very late antigen [VLA]-4) and 5 1 (VLA-5)-dependent adhesion
of human cord blood CD34+ cells to fibronectin.
CD34+ CD38hi, but not
CD34+CD38 /low cells responded
significantly to the stimulus, suggesting that committed, but not stem
and more immature progenitors are sensitive to CD43-mediated activation
of integrin. To elucidate the molecular mechanism leading to integrin
activation, we used the growth factor-dependent cell line MO7e.
Cross-linking of CD43 induced tyrosine phosphorylation of several
intracellular molecules including the protein tyrosine kinase Syk, the
proto-oncogene product Cbl, and phospholipase C (PLC)- 2 in MO7e
cells. Moreover, protein tyrosine kinase inhibitor herbimycin A and PLC
inhibitor U73122 both blocked CD43-induced enhancement of adhesion to
fibronectin. These results indicate that signals mediated through CD43
may increase integrin affinity to fibronectin via a pathway dependent
on protein tyrosine kinase and PLC- activation in hematopoietic progenitors.
© 1999 by The American Society of Hematology.
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INTRODUCTION |
CD43 IS A MAJOR transmembrane
sialoglycoprotein expressed on almost all hematopoietic cells except
for erythrocytes and a population of B cells.1-3
Pluripotent hematopoietic stem cells have been reported to express CD43
molecule.4 Multiple glycoforms are generated by
modifications in the O-glycans attached to the protein core, giving
rise to activation- or development-dependent isoforms.5-7
Altered expression of CD43 has been reported in Wiskott-Aldrich
syndrome8 and human immunodeficiency virus (HIV)
infection.9
CD43 has a highly conserved cytoplasmic domain across
species10 and cross-linking of CD43 with specific
monoclonal antibodies (MoAbs) has various effects depending on cell
types. These include activation of monocytes,11
proliferation and activation of T cells,12-14 enhancement
of natural killer (NK) cell activity,15 and induction of
apoptosis in hematopoietic progenitors.16 An isoform-specific response has been also reported.7
Intracellular signaling events reported are the generation of
diacylglycerol and inositol phosphate, with calcium mobilization and
protein kinase C (PKC) activation, and the activation of tyrosine
kinase-dependent pathways in T cells.17-19
CD43 is a prototypic member of a family of cell surface-associated
mucins, which are characterized by the presence of extensive O-linked
glycan substitutions.20 A highly extended structure of
extracellular mucin with negative charge may function as a repulsive
barrier. The antiadhesive role of CD43, which prevents nonspecific
binding, has been stressed in a number of studies.21-23 On
the other hand, many mucins are long enough to gain optimal exposure of
their terminal sugars, thus being accessible to interact with specific
carbohydrate receptors. Some cell surface-associated mucins appear to
be physiological ligands of selectins, and interaction of selectins
with mucins initiate the adhesion cascade by which leukocytes move from
the blood into tissue.20 In this context, a recent study
suggested the significance of CD43-endotherial cell interaction in
T-cell homing.24 Moreover, engagement of CD43 with specific
MoAbs has been reported to enhance 1 and 2 integrin affinity in T
cells.25
Integrins are heterodimeric transmembrane glycoproteins consisting of
 and subunits. The ligand-binding ability of integrin is
regulated by cytoplasmic signals triggered on various external stimuli
(inside-out signaling).26 Integrin-mediated adhesion is
believed to be crucial in the homing and anchoring of hematopoietic cells to the bone marrow environment. In particular, the importance of
very late antigen (VLA)-4 and VLA-5, which interact with fibronectin and vascular adhesion molecule-1, has been implicated in
hematopoiesis.27,28 In this study, we show that
cross-linking of CD43 with MoAbs enhances adhesion of human primary
cord blood hematopoietic progenitors to immobilized fibronectin.
Intracellular signaling events leading to integrin activation after
engagement of CD43 were assessed using the human factor-dependent cell
line, MO7e.
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MATERIAL AND METHODS |
Cytokine, antibodies, and reagents.
Recombinant human granulocyte-macrophage colony-stimulating factor
(rhuGM-CSF) and steel factor were kindly provided by Immunex Corp
(Seattle, WA). Anti-integrin 1 MoAb (clone Lia1/2), anti-integrin 4 MoAb (clone HP2/1), anti-integrin 5 MoAb (clone SAM1),
anti-CD43 MoAb (clone DF-T1), anti-CD31 MoAb (clone 5.6E), anti-CD50
antibody (clone HP2/19), and F(ab')2 fragment goat
antimouse IgG were purchased from Immunotech (Westbrook, ME). Anti-Syk
polyclonal Ab, raised against an epitope corresponding to amino acids
257-352 mapping within the linker region of Syk, anti-Zap-70 polyclonal
Ab and anti-Cbl polyclonal Ab were from Santa Cruz Biotechnology (Santa Cruz, CA). Anti-CD43 (clone MEM-59), fluorescein isothiocyanate (FITC)-conjugated anti-CD38, phycoerythrin (PE)-conjugated anti-CD34 and PE-Cy5-conjugated F(ab')2 fragment goat antimouse IgG
were from Caltag Laboratories (Burlingame, CA). Antiphosphotyrosine (anti-pTyr) MoAb (4G10) and antiphospholipase C (PLC)-1 mixed MoAbs
were from Upstate Biothechnology (Lake Placid, NY). Anti-PLC-2 polyclonal antibody was from Pharmingen (San Diego, CA). Wortmannin, H7, HA1004, herbimycin A, U73122, and U73343 were obtained from
Calbiochem (San Diego, CA). Human fibronectin was purchased from
Collaborative Biomedical (Bedford, MA).
Cell lines.
The human growth factor-dependent myeloid cell lines, MO7e and TF-1,
were cultured in RPMI 1640 supplemented with 20% fetal bovine serum
and 100 U/mL rhuGM-CSF. The biological characteristics of these cell
lines have been described previously.29-31 Before cross-linking or cytokine stimulation, cells were washed twice, resuspended in RPMI 1640 supplemented with 1% bovine serum albumin (BSA), and then incubated for 18 hours at 37°C.
Cell sorting.
Mononuclear cells were isolated from cord blood by Ficoll-Hypaque
(Pharmacia, Piscataway, NJ) gradient centrifugation. Next, mononuclear
cells were preenriched for CD34+ cells using a Magnetic
Cell Sorting (MACS) system (Miltenyi Biotec, Auburn, CA). Cells were
then incubated with PE-conjugated anti-CD34 with or without
FITC-conjugated anti-CD38 for 30 minutes at 4°C. Subsequently,
CD34+ cells were sorted on a FACStar Plus (Becton
Dickinson, Franklin Lakes, NJ). Sorted CD34+ cells were
starved overnight in serum-free medium (QBSF58, Sigma, St Louis, MO)
before adhesion assay.
Flow cytometory analysis.
Cord blood CD34+ cells isolated by MACS were first stained
with anti-CD43 (DF-T1) followed by PE-Cy5-conjugated
F(ab')2 fragment goat antimouse IgG. After blocking the
free binding sites of secondary antibody with mouse IgG, cells were
stained with PE-conjugated anti-CD34 and FITC-conjugated anti-CD38.
CD43 expression was analyzed on gated CD34+ cells by
FACScan (Becton Dickinson). Cells stained with isotype-matched irrelevant MoAbs were used as a negative control.
Adhesion assay.
Human fibronectin was diluted in phosphate-buffered saline (PBS) at a
concentration of 20 µg/mL and distributed in 96-well tissue culture
flat-bottomed plates (Corning, Cambridge, NY). After overnight
incubation at 4°C, the coated wells were washed twice with PBS and
blocked by adding PBS with 1% BSA at 37°C for 1 hour. The wells
were then washed twice with PBS. Cells were labeled with
51Cr (Amersham, Arlington Heights, IL) (100 µCi/5 × 106 cells) at 37°C for 1 hour, washed twice, and
resuspended in RPMI with 0.5% BSA. A total of 100 µL of the cell
suspension was added to the protein-coated wells, centrifuged at 600 rpm for 1 minute, and incubated at 37°C for 20 minutes, except
where otherwise indicated. Unattached cells were removed by two washes
with prewarmed RPMI with 0.5% BSA. Adherent cells were solubilized
with 1% sodium dodecyl sulfate (SDS), and then radioactivity was
quantified by a scintillation counter. Percent adhesion was calculated
as the ratio of the radioactivity in adherent cells to that of input, after correction of nonspecific binding to BSA-coated wells.
Engagement of CD43.
Cells were treated with 20 µg/mL anti-CD43 MoAb for 20 minutes at
4°C. Next, 10 µg/mL F(ab')2 fragment goat
antimouse IgG was added for 10 minutes at 4°C. Then, cells were
incubated for indicated time periods at 37°C. Although anti-CD43
antibody alone had some effect on cell adhesion, we observed that
secondary antibody enhanced the effect in preliminarily experiments
using MO7e and cord blood CD34+ cells. Thus, we
used secondary antibody in all experiments.
Immunoprecipitation and immunoblotting.
Cells were lysed in lysis buffer (20 mmol/L Tris-HCl [pH 7.4], 150 mmol/L NaCl, 10% glycerol, 1% Nonidet P-40, 1 mmol/L phenylmethyl sulfonyl fluoride [PMSF], 10 µg/mL aprotinin, 10 µg/mL leupeptin, 100 mmol/L NaF, and 1 mmol/L sodium orthovanadate) on ice for 20 minutes, and insoluble fractions were removed by centrifugation at
14,000g for 20 minutes. Equal amounts of protein were used for
immunoprecipitation. Cell extracts were mixed with appropriate Ab at
4°C for 2 hours. Antigen-Ab complexes were collected with protein
A-Sepharose beads (Pharmacia). Immunoprecipitates were washed with
lysis buffer four times and separated by SDS-polyacrylamide gel
electrophoresis (PAGE) and transferred to polyvinylidene difluoride (PVDF) membrane (Millipore, Bedford, MA). Membranes
were blocked in Tris-buffered saline containing 0.5% Tween 20 and 1%
BSA for 1 hour at room temperature and incubated with appropriate
primary Abs for 2 hours. Blots were visualized using horseradish
peroxidase-conjugated secondary Ab and an enhanced chemiluminescence
system (ECL, Amersham). To reprobe with another first Ab, membranes
were incubated in striping buffer (62.5 mmol/L Tris-HCl [pH 6.7], 100 mmol/L 2 mercaptoethanol [2ME], and 2% SDS) for 30 minutes at 50°C, washed, and then used for further study.
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RESULTS |
Engagement of CD43 enhances adhesion of hematopoietic progenitors to
fibronectin.
We have used MO7e and TF-1 cells, CD34+ factor-dependent
cell lines having primitive myeloid lineage characteristics, as a model
system of integrin-mediated adhesion.32,33 MO7e and TF-1 cells have been reported to express a set of integrins similar to those
of primary CD34+ cells.34 A line of studies
suggests that integrin function in CD34+ hematopoietic
progenitors can be activated by a number of cytokines.34-36 Figure 1 shows that cross-linking of CD43
with MoAb DF-T1 also stimulates adhesion of MO7e and TF-1 cells to
immobilized fibronectin. Similar effects were observed with another
CD43 MoAb MEM-59 (data not shown). CD31 was expressed on MO7e and TF-1
cells (data not shown), however cross-linking of CD31 with MoAb 5.6E
had no effect on attachment to fibronectin. Although MO7e cells are
positive for a receptor for laminin (VLA-6), cross-linking of CD43 did not enhance attachment of MO7e cells to laminin (data not shown). The
effect of CD43 engagement was prolonged
(Fig 2) in comparison with those of
cytokines, which induce transient cell attachment to
fibronectin.32-34 Next, we investigated the effect of CD43
engagement on the adhesion of CD34+ cord blood progenitors
to immobilized fibronectin. Because subpopulations of CD34+
cells may display different sensitivity to the anti-CD43 MoAb treatment,16 we separated cord blood CD34+
cells into three fractions on the basis of CD38 antigen expression (Fig 3A). Adhesion of
CD34+CD38hi cells to fibronectin was enhanced
in response to cross-linking of CD43. In contrast, cross-linking of
CD43 did not have a significant effect on
CD34+CD38/low and
CD34+CD38low cells (Fig 3B). We further
examined the expression of CD43 in cord blood CD34+ cells.
Most of the CD34+ cells were positive for CD43 and the
expression of CD43 tended to correlate with that of CD38 (Fig
3C).
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| Fig 1.
CD43-mediated enhancement of (A) MO7e and (B) TF-1 cell
adhesion to immobilized fibronectin. Radiolabeled cells were pretreated
with medium (RPMI 1640 medium containing 1% BSA) alone, 20 µg/mL
control IgG, anti-CD43 MoAb (clone DF-T1), or anti-CD31 MoAb (clone
5.6E) for 20 minutes at 4°C. Next, 10 µg/mL goat antimouse IgG
was added for 10 minutes at 4°C. Cells were then distributed in
flat-bottom 96-well plate coated with fibronectin. The plate was
centrifuged at 600 rpm for 1 minute and incubated for 20 minutes at
37°C. Cell adhesion assay was performed as described in Materials
and Methods. Data represent the mean ± standard error (SE) of
triplicate assays for one of three reproducible experiments each.
*P < .0001 versus control **P < .001 versus
control.
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| Fig 2.
Kinetics of MO7e cell adhesion to immobilized
fibronectin. MO7e cells stimulated with or without anti-CD43 MoAb
(clone DF-T1) and goat antimouse IgG were distributed in
fibronectin-coated 96-well plate. The plate was incubated for the
indicated time periods at 37°C and subjected to adhesion assay.
Data represent the mean ± SE of triplicate assays for one of three
reproducible experiments. *P < .01, **P < .001, ***P < .0001 versus control at each time point. ( ),
control; ( ), CD43-stimulated.
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| Fig 3.
Influence of CD43 on cord blood CD34+ cell
subsets. (A) Gates used to select cord blood CD34+
subpopulations based on CD38 expression. Cells were sorted by FACSstar
Plus after MACS preenrichment. R1:
CD34+CD38/low cells; R2:
CD34+CD38low cells; R3:
CD34+CD38hi cells. (B) The effect of CD43
engagement on adhesion of CD34+ subpopulations to
immobilized fibronectin. Cells were stimulated with isotype-matched
control IgG or anti-CD43 MoAb (clone DF-T1) followed by goat antimouse
IgG, then subjected to adhesion assay. Data represent the mean ± SE
from three separate experiments with triplicate samples for each group
in every experiment. *P < .05 versus control. ( ), control;
( ) CD43-stimulated. (C) Expression of CD38 and CD43 on the
CD34+ cell population. Three-color analysis was performed
on the cells isolated from cord blood by MACS. Expression of CD38 and
CD43 was analyzed on the gated CD34+ cell population by
FACScan.
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CD43 enhanced-adhesion to fibronectin is integrin-dependent in MO7e
and cord blood CD34+ cells.
To test if CD43 enhanced-adhesion to fibronectin was mediated by
integrin, function-blocking anti-integrin antibodies were included in
adhesion assay. Although anti-4 MoAb (HP2/1) only partially blocked
CD43 enhanced-adhesion to fibronectin in MO7e cells, the attachment was
completely abrogated by anti-5 MoAb (SAM1) or anti-1 MoAb
(Lia1/2) (Fig 4A). As for cord blood
CD34+ cells, the enhancement of adhesion to fibronectin was
partially inhibited by HP2/1 or SAM1. Complete inhibition was observed
by the combined use of HP2/1 and SAM1 or by Lia1/2 (Fig 4B). These results indicate that CD43-enhanced adhesion is mediated through VLA-4
and VLA-5 in hematopoietic progenitors.
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| Fig 4.
Effect of anti-integrin antibody on CD43-enhanced MO7e
and cord blood CD34+ cell adhesion to fibronectin. (A)
MO7e cells were treated with or without anti-CD43 MoAb (clone DF-T1)
and goat antimouse IgG. CD43-stimulated cells were incubated with 10 µg/mL control IgG or anti-integrin antibody for an additional 20 minutes at 4°C and subjected to adhesion assay. Data represent the
mean ± SE of triplicate assays for one of two reproducible
experiments. *P < .0001, **P < .001 versus
CD43-stimulated adhesion without additional antibodies. (B) Sorted
CD34+ cells were treated with 20 µg/mL control IgG or
CD43 (clone DF-T1) followed by goat antimouse IgG. CD43-stimulated
cells were incubated with 10 µg/mL anti-integrin antibody for an
additional 20 minutes at 4°C and subjected to adhesion assay. Data
represent the mean ± SE from three separate experiments with
triplicate samples for each group in every experiment. *P < .01, **P < .05 versus CD43-stimulated adhesion without
additional antibodies.
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CD43 enhanced-adhesion is differentially inhibited by second
messenger inhibitors in MO7e cells.
To clarify the pathway leading to integrin activation after engagement
of CD43, we treated MO7e cells with second messenger inhibitors before
adhesion assay (Fig 5A). Cholera toxin, an
activator of adenyl cyclase, and wortmannin, a phosphatidylinositol 3 (PI3)-kinase inhibitor failed to suppress CD43 enhanced-adhesion to
fibronectin. The same concentration of wortmannin greatly reduced
enhancement of adhesion induced by steel factor (Fig 5B). Herbimycin A,
a protein tyrosine kinase inhibitor, substantially suppressed adhesion augmented with CD43 cross-linking. H7, which inhibits both PKC and
protein kinase A (PKA), but not HA1007, which inhibits
PKA, but not PKC, almost completely blocked CD43-enhanced adhesion. Herbimycin A and H7 also suppressed background adhesion significantly. This may suggest the possibility that PKC and tyrosine kinase were
necessary for constitutive adhesion to fibronectin in MO7e cells.
Because PKC is considered to be one of the downstream signals of PLC
and in some systems a PLC-PKC pathway is suggested to be involved in
integrin inside-out signaling,36,37 we used the putative
PLC inhibitor, U73122. This reagent suppressed CD43-enhanced adhesion
in a dose-dependent manner (Fig 5C). U73343, a close analog to U73122,
had no significant inhibitory effect on MO7e cell adhesion to
fibronectin.
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| Fig 5.
Effect of inhibitors on CD43-stimulated MO7e
cell adhesion to fibronectin. (A) Cells were pretreated with 1 µmol/L
wortmannin, 0.2 µg/mL chorea toxin, 50 µmol/L HA1004, or 50 µmol/L H7 for 30 minutes or 1 µg/mL herbimycin A for 18 hours at
37°C. Then, cells were cross-linked with anti-CD43 MoAb (clone
DF-T1) and subjected to adhesion assay. *P < .01 versus
control, **P < .01 versus CD43-stimulated adhesion without
inhibitors. (), control; ( ), CD43-stimulated; (B) Cells
pretreated with 1 µmol/L wortmannin for 30 minutes at 37°C were
cross-linked with anti-CD43 MoAb (clone DF-T1) or treated with 10 ng/mL
steel factor and subjected to adhesion assay. *P < .01 versus
steel factor-stimulated adhesion without wortmannin. (), control;
(), CD43-stimulated; (), steel factor-stimulated. (C) Cells were
pretreated with U73122 at the indicated concentrations for 30 minutes
at 37°C. Then, cells were cross-linked with anti-CD43 MoAb (clone
DF-T1) and subjected to adhesion assay. *P < .05, **P < .001 versus CD43-stimulated adhesion without U73122. (),
control; (), CD43-stimulated; ( ) pretreated with U733343 (2 µmol/L) before CD43 cross-linking; Data represent the mean ± SE of
triplicate assays for one representative experiment of three
reproducible experiments.
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Engagement of CD43 induced tyrosine phosphorylation of
PLC-2 p72Syk and p120Cbl in
MO7e cells.
Signals via CD43 in T cells are suggested to be mediated through
protein tyrosine kinases, PLC and PKC.17-19 The result of second messenger inhibitors suggested involvement of these pathways in
CD43-mediated integrin inside-out signaling in MO7e cells. PLC-
isozymes are known to be activated through phosphorylation. We
investigated whether PLC- became phosphorylated on cross-linking of
CD43. PLC-1 was only marginally affected (data not shown), but
PLC-2 was significantly tyrosine phosphorylated by CD43 engagement (Fig 6). The implication of Syk family
tyrosine kinase in the phosphorylation of PLC- has been
suggested.38,39 We found that engagement of CD43 induced
tyrosine phosphorylation of Syk protein tyrosine kinase
(Fig 7). Tyrosine phosphorylation of
ZAP-70, a family member of Syk, was not detectable (data not shown).
The proto-oncogene product, c-Cbl, may play a critical role in signal transduction pathways mediated by cytokines in hematopoietic
cells40,41 and an interaction with Syk has been
reported.42 These observations led us to examine the
possibility of involvement of c-Cbl in CD43-mediated signaling. c-Cbl
was tyrosine phosphorylated in a time-dependent manner and the peak
phosphorylation was observed 10 minutes after CD43 engagement. The
degree of tyrosine phosphorylation of c-Cbl was almost equal to that
observed after stimulation with 100 U/mL GM-CSF
(Fig 8). We could not detect association
of Syk or PLC- with Cbl by a combination of immunoprecipitation and
immunoblotting (data not shown). We further examined whether engagement
of other adhesion molecules expressed on MO7e cells could modify
tyrosine phosphorylation of these proteins. Cross-linking of CD31 or
CD50 induced no significant tyrosine phosphorylation of Syk and Cbl in
MO7e cells (Fig 9). Similar results were
observed in the case of PLC-2 (data not shown). These results
suggest that the tyrosine phosphorylation events we observed were
specific to CD43-mediated signaling and a potential involvement of
Fc receptor was unlikely.
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| Fig 6.
Tyrosine phosphorylation of PLC-2 by engagement of
CD43. MO7e cells cross-linked with anti-CD43 MoAb (clone DF-T1) were
incubated for the indicated time periods at 37°C. Cells treated
with isotype-matched control IgG and goat antimouse IgG were incubated
for 10 minutes at 37°C (Lane C). Cell lysates were
immunoprecipitated with anti-PLC-2 Ab. Upper panel: these
immunoprecipitates were separated by 7.5% SDS-PAGE and immunoblotted
with antiphosphotyrosin MoAb; lower panel: the same membrane was
reprobed with anti-PLC-2 Ab. These are the representative results
from three separate experiments.
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| Fig 7.
Tyrosine phosphorylation of p72Syk by
engagement of CD43. MO7e cells cross-linked with anti-CD43 MoAb (clone
DF-T1) were incubated for the indicated time periods at 37°C. Cells
treated with isotype-matched control IgG and goat antimouse IgG were
incubated for 10 minutes at 37°C (Lane C). Cell lysates were
immunoprecipitated with anti-Syk Ab. Upper panel: these
immunoprecipitates were separated by 7.5% SDS-PAGE and immunoblotted
with antiphosphotyrosine MoAb; lower panel: the same membrane was
reprobed with anti-Syk. These are the representative results from three
separate experiments.
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| Fig 8.
Tyrosine phosphorylation of p120Cbl by
engagement of CD43. (A) MO7e cells treated with isotype-matched control
IgG or anti-CD43 MoAb (clone DF-T1) followed by goat antimouse IgG or
100 U/mL GM-CSF were incubated for 10 minutes at 37°C. Cell lysates
were immunoprecipitated with anti-Cbl Ab or preimmune rabbit serum
(lane C). Upper panel: these immunoprecipitates were separated by 7.5%
SDS-PAGE and immunoblotted with antiphosphotyrosine MoAb; lower panel:
the same membrane was reprobed with anti-Cbl. (B) MO7e cells
cross-linked with anti-CD43 MoAb (clone DF-T1) were incubated for the
indicated time periods at 37°C. Cell lysates were
immunoprecipitated with anti-Cbl Ab and followed by immunoblotting with
antiphosphotyrosine MoAb (upper panel) and reprobing with anti-Cbl Ab
(lower panel). These are the representative results from three separate
experiments.
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| Fig 9.
Effect of engagement of cell surface molecules on
tyrosine phosphorylation of p72Syk and p120Cbl.
MO7e cells were cross-linked with the indicated antibodies for 10 minutes at 37°C. (A) Cell lysates were immunoprecipitated with
anti-Syk Ab and followed by immunoblotting with antiphosphotyrosine
MoAb (upper panel) and reprobing with anti-Syk Ab (lower panel). (B)
Cell lysates were immunoprecipitated with anti-Cbl Ab and followed by
immunoblotting with antiphosphotyrosin MoAb (upper panel) and reprobing
with anti-Cbl Ab (lower panel). The tyrosine-phosphorylated band around
130 kD in the lanes from cell lysates stimulated with
anti-CD31 MoAb is supposed to be CD31 molecule bound to protein A
through the antibody used for cross-linking. These are the
representative results from two separate experiments.
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| |
DISCUSSION |
Integrin affinity of CD34+ hematopoietic progenitors is
reported to be modulated by a number of cytokines34-36 or
engagement of adhesion molecules such as CD31.43 Here, we
presented data showing that the cell surface mucin CD43 could mediate
signals, which upregulate integrin functional activity in hematopoietic progenitors.
Evidence for CD43 as a barrier molecule, which negatively regulates
cell-cell and cell-ligand interaction, has been presented. Transfection
of CD43 into HeLa cell interfered with interaction of T cells to these
cells21 and targeted disruption of CD43 gene enhanced
T-cell adhesiveness.22 Cross-linking of CD43 may provide a
mechanism such as capping of surface mucin to one side of cells and
unmasking integrins.44 However, failure of enhancement of
adhesion to laminin made such an unspecific mechanism unlikely. In this
regard, our results are similar to upregulation of integrin affinity by
CD43 engagement in T cells.25
Recent studies have shown the importance of PI3-kinase in integrin
inside-out signaling.45-47 Adhesion induced by
hematopoietic growth factors was sensitive to the PI3-kinase inhibitor
wortmannin.31,48 However, wortmannin failed to modulate
CD43-mediated attachment to fibronectin, suggesting a negligible role
for PI3-kinase in this system. Platelet-derived growth factor
(PDGF) receptor expressed in mast cells could initiate
two independent pathways leading to integrin activation, one dependent
on PI3-kinase and the other dependent on PLC- and PKC.37
The tyrosine phosphorylation of PLC- after engagement of CD43 and
the blocking effect of the PKC inhibitor, H7, suggest the possibility
that the PLC- and PKC-dependent pathway might be also activated in
this system. In further support of the role of PLC in integrin
inside-out signaling, the putative PLC inhibitor, U73122, which has
been reported to inhibit integrin-mediated polymorphonuclear neutrophil
(PMN) adhesion,49 blocked CD43-enhanced
adhesion of MO7e cells to fibronectin. Considering the difference in
sensitivity to inhibitors and kinetics of adhesion, CD43 may elicit
signals leading to integrin activation that are distinct from those
mediated by hematopoietic growth factors.
CD43 is reported to be associated with Fyn and Lck tyrosine
kinases19,50 and may modulate tyrosine phosphorylation of
specific cellular proteins in T cells.51 Recently, tyrosine
phosphorylation of the proto-oncogene product Vav and activaton of
mitogen-activated protein kinase have been recognized as downstream
signals.52 Our results suggest that signaling via CD43
could affect the tyrosine phosphorylation status of some cellular
proteins also in primitive myeloid cells. We identified some of these
proteins as PLC-2, the protein tyrosine kinase p72syk,
and the proto-oncogene product p120cbl. The fact that the
protein tyrosine kinase inhibitor, herbimycin A, reduced CD43-enhanced
MO7e cell adhesion to fibronectin may indicate the possible
participation of some of these proteins in integrin inside-out signaling.
The role of Syk kinase family in primitive myeloid cells is largely
unknown. We recently reported that integrin cross-linking, but not
cytokines, induced Syk tyrosine phosphorylation.33 This study presents another possible pathway leading to Syk activation, although the involvement of Syk in PLC- phosphorylation or integrin inside-out signaling is uncertain at present.
The proto-oncogene product, c-Cbl, is inducibly phosphorylated after
engagement of many types of receptors. c-Cbl contains a proline-rich
domain and several potential tyrosine phosphorylation sites, which can
function as ligands for SH2 and SH3 domains of multiple signaling
proteins. c-Cbl is suggested to function as a complex adapter molecule
modulating signaling pathways such as RAS, JAK/STAT, and
PI3-kinase.53-55 In mast cells, overexpression of c-Cbl
resulted in inhibition of kinase activity of Syk, suggesting that c-Cbl
may have a direct effect on enzyme activity with which it
associates.42 Our study suggests that c-Cbl also has a
potential role in CD43-mediated signaling. Although the mechanism of
c-Cbl involvement remains to be elucidated, we suspect c-Cbl may be an
important signal transducer because of its major phosphorylation compatible with that induced by GM-CSF.
CD43 has been reported to mediate signals leading to apoptosis in
cytokine-activated dividing population of CD34+ bone marrow
cells, but not in quiescent stem cells.16 The
CD34+CD38 immunophenotype defined a
quiescent subpopulation in both cord blood and bone marrow and the
percentage of CD34+ cells in cycle directly correlated with
increasing CD38 expression.56 Our results showed that the
CD34+CD38hi subpopulation responded most to
integrin activation through CD43. Thus, it is highly possible that at
least a part of CD34+ cells are susceptible to both
apoptosis and integrin inside-out signals. CD43-mediated apoptosis
might be induced in an isoform-specific manner.7 However,
the following observations suggest that apoptosis and integrin
inside-out signals may be mediated through the same isoform of CD43.
MEM-59, which has been reported to induce apoptosis in
CD34+ bone marrow cells, also enhanced attachment to
fibronectin in MO7e cells. DF-T1 and MEM-59 has been reported to bind
the same or closely related sialic acid-dependent
epitope,57 and both MoAbs recognized the identical
molecular weight protein in Western blotting of MO7e and TF-1 cell
lysate (data not shown). Taking the correlation of CD38 and CD43
expression in CD34+ cells into account, CD43hi
cell might react to cross-linking of CD43. CD34+ cells in a
certain stage of commitment may be effected preferentially by signaling
mediated through CD43. The execution of apoptosis by CD43 required
immobilized anti-CD43 MoAb and the presence of cytokine.16
Compared with integrin inside-out signaling, more profound engagement
of CD43 and other stimuli may be necessary for apoptosis.
Supposing an unidentified ligand for CD43 is expressed on the bone
marrow stromal cells, CD43 may mediate signaling regulating cell
adhesion or survival in the bone marrow microenvironment. Our results
suggest that signaling via CD43 may be distinct from cytokine signaling
and further studies on the cooperative effect of cytokine receptors and
CD43 could provide novel insight into understanding mechanisms
regulating the development of hematopoietic progenitors in the microenvironment.
| |
ACKNOWLEDGMENT |
We thank Charlie Mantel for helpful discussion.
| |
FOOTNOTES |
Submitted June 8, 1998; accepted January 8, 1999.
Supported by U.S. Public Health Service Grants No. R01 DK53674, R01
HL56416, R01 HL54037, and a project in P01 HL53586 from the National
Institutes of Health (NIH) (to H.E.B.).
The publication costs of this
article were defrayed in part by
page charge payment. This article
must therefore be hereby marked
"advertisement"
in accordance with 18 U.S.C. section
1734 solely to indicate this fact.
Address reprint requests to Hal E. Broxmeyer, PhD, Walther Oncology
Center, Indiana University School of Medicine, 1044 W. Walnut St, Rm
302, Indianapolis, IN 46202; e-mail:hbroxmey{at}iupui.edu.
| |
REFERENCES |
1.
Remold-O'Donnell E, Zimmerman C, Kenney D, Rosen F:
Expression on blood cells of sialophorin, the surface glycoprotein that is defective in Wiskott-Aldrich Syndrome.
Blood
70:104, 1987[Abstract/Free Full Text]
2.
Bettaieb A, Farace F, Mijavila MT, Mishal Z, Dokhelar MC, Tursz T, Breton-Gorius J, Vainchenker W, Kieffer N:
Use of a monoclonal antibody (GA3) to demonstrate lineage restricted o-glycosylation on leukosialin during terminal erythroid differentiation.
Blood
71:1226, 1988[Abstract/Free Full Text]
3.
Wiken M, Bjorck P, Axelsson B, Perlmann P:
Induction of CD43 expression during activation and terminal differentiation of human B cells.
Scand J Immunol
28:457, 1988[Medline]
[Order article via Infotrieve]
4.
Moore T, Huang S, Terstappen LWMM, Bennett M, Kumar V:
Expression of CD43 on murine and human pluripotent hematopoietic stem cells.
J Immunol
153:4978, 1994[Abstract]
5.
Piller F, Piller V, Fox RI, Fukuda M:
Human T-lymphocyte activation is associated with changes in O-glycan biosynththesis.
J Biol Chem
263:15146, 1988[Abstract/Free Full Text]
6.
Ellies LG, Tao W, Fellinger W, Teh H-S, Ziltener HJ:
The CD43 130-kD peripheral T-cell activation antigen is downregulated in thymic positive selection.
Blood
88:1725, 1996[Abstract/Free Full Text]
7.
Joseph Brown T, Shuford WW, Wang W-C, Nadler SG, Bailey TS, Marquardt H, Mittler RS:
Characterization of a CD43/leukosialin-mediated pathways for inducing apoptosis in human T-lymphoblastoid cells.
J Biol Chem
271:27686, 1996[Abstract/Free Full Text]
8.
Remold-O'Donnell E, Kennyey DM, Perrine S, Rosen FS:
Characterization of human lymphocyte sialoglycoprotein that is defective in Wiskott-Aldrich syndrome.
J Exp Med
159:1705, 1984[Abstract/Free Full Text]
9.
Giordanengo V, Limouse M, Du Roure LD, Cottalorda J, Doglio A, Passeron A, Fuzibet JG, Lefebvre JC:
Autoantibodies directed against CD43 molecules with an altered glycosylation status on human immunodifficiency virus type 1 (HIV-1)-infected CEM cells are found in all HIV-1+ individuals.
Blood
86:2302, 1995[Abstract/Free Full Text]
10.
Shelley CS, Remold-O'Donnell E, Davis AE III, Bruns GAP, Rosen FS, Carroll MC, Whitehead AS:
Molecular caracterization of sialophorin (CD43), the lymphocyte surface sialoglycoprotein defective in Wiskott-Aldrich syndrome.
Proc Natl Acad Sci USA
86:2819, 1989[Abstract/Free Full Text]
11.
Nong y-H, Remold-O'Donnell E, LeBien TW, Remold HG:
A monoclonal antibody to sialophorin (CD43) induces homotypic adhesion and activation of human monocytes.
J Exp Med
170:259, 1989[Abstract/Free Full Text]
12.
Mentzer SJ, Remold-O'Donnell E, Crimmins MA, Bierer BE, Rosen FS, Burakoff SJ:
Sialophorin, a surface sialoglycoprotein defective in the Wiskott-Aldrich syndrome, is involved in human T lymphocyte proliferation.
J Exp Med
165:1383, 1987[Abstract/Free Full Text]
13.
Axelsson B, Youseffi-Etemad R, Hammarström S, Perlmann P:
Induction of aggregation and enhancement of proliferation and IL-2 secretion in human T cells by antibodies to CD43.
J Immunol
141:2912, 1988[Abstract]
14.
Park JK, Rosenstein YJ, Remold-O'Donnell E, Bierer BE, Rosen FS, Burakoff SJ:
Enhancement of T-cell activation by the CD43 molecule whose expression is defective in Wiskott-Aldrich syndrome.
Nature
350:706, 1991[Medline]
[Order article via Infotrieve]
15.
Vagas-Cortes M, Axelsson B, Larsson A, Berzin T, Permann P:
Enhancement of human spontaneous cell-mediated cytotoxicity by a monoclonal antibody against the large sialoglycoprotein (CD43) on peripheral blood lymphocytes.
Scan J Immunol
27:661, 1988[Medline]
[Order article via Infotrieve]
16.
Bazil V, Brandt J, Chen S, Roeding M, Luens K, Tsukamoto A, Hoffman R:
A monoclonal antibody recoganizing CD43 (leukosialin) initiates apoptosis of human hematopoietic progenitor cells but not stem cells.
Blood
87:1272, 1996[Abstract/Free Full Text]
17.
Silverman LB, Wong RCK, Remold-O'Donnell E, Vercelli D, Sancho J, TerHorst C, Rosen F, Ceha R, Chatila T:
Mechanism of mononuclear cell activation by an anti-CD43 antibody.
J Immunol
141:4194, 1989
18.
Wong RCK, Remold-O'Donnell E, Vercelli D, Sancho J, Terhost C, Rosen F, Geha R, Chatila T:
Signal transduction via leukocyte antigen CD43. Feedback regulation by protein kinase C.
J Immunol
144:1455, 1990[Abstract]
19.
Pedraza-Alva G, Mérida LB, Burakoff SJ, Rosenstein Y:
CD43-specific activation of T cells induces association of CD43 to fyn kinase.
J Biol Chem
271:27654, 1996
20.
Shimizu Y, Shaw S:
Mucins in the mainstream.
Nature
366:630, 1993[Medline]
[Order article via Infotrieve]
21.
Ardman B, Sikorski MA, Staunton DE:
CD43 interferes with T-lymphocyte adhesion.
Proc Natl Acad Sci USA
89:5001, 1992[Abstract/Free Full Text]
22.
Manjunath N, Johnson RS, Staunton DE, Pasqualini R, Ardman B:
Targeted disruption of CD43 gene enhances T lymphocyte adhesion.
J Immunol
151:1528, 1993[Abstract]
23.
Manjunath N, Correa M, Ardman M, Ardman B:
Negative regulation of T-cell adhesion and activation by CD43.
Nature
377:535, 1995[Medline]
[Order article via Infotrieve]
24.
McEvoy LM, Sun H, Frelinger JG, Butcher EC:
Anti-CD43 inhibition of T cell homing.
J Exp Med
185:1493, 1997[Abstract/Free Full Text]
25.
Sánches-Mateos P, Campanero MR, del Pozo MA, Sánchez-Madrid F:
Regulatory role of CD43 leukosialin on integrin-mediated T-cell adhesion to endotherial and extracellular matrix ligands and its polar redistribution to a cellular uropod.
Blood
86:2228, 1995[Abstract/Free Full Text]
26.
Hynes RO:
Integrins: Versatility, modulation and signaling in cell adhesion.
Cell
69:11, 1992[Medline]
[Order article via Infotrieve]
27.
Verfaillie CM, McCarthy JB, McGlave PB:
Differentiation of primitive human multipotent hematopoietic progenitors into single lineage colonogenic progenitors is accompanied by alteration in their interaction with fibronectin.
J Exp Med
174:693, 1991[Abstract/Free Full Text]
28.
Williams DA, Rios M, Stephens C, Patel VP:
Fibronectin and VLA-4 in haematopoietic stem cell-microenviroment interactions.
Nature
352:438, 1991[Medline]
[Order article via Infotrieve]
29.
Avanzi GC, Lista P, Giovinazzo B, Miniero R, Saglio G, Benetton G, Coda R, Cattoretti G, Pegoraro L:
Selective growth response to IL-3 of a human leukemic cell line with megakaryoblastic features.
Br J Haematol
69:359, 1988[Medline]
[Order article via Infotrieve]
30.
Hendrie PC, Miyazawa K, Yang Y-C, Langefeld CD, Broxmeyer HE:
Mast cell growth factor (c-kit ligand) enhances cytokine stimulation of proliferation of human factor dependent cell line, MO7e.
Exp Hematol
19:1031, 1991[Medline]
[Order article via Infotrieve]
31.
Kitamura T, Tange T, Terasawa T, Chiba S, Kuwaki T, Miyagawa K, Piao YF, Miyazono K, Urabe A, Takaku F:
Establishment and characterization of unique human cell line that proliferates dependently on GM-CSF, IL-3 or erythropoietin.
J Cell Physiol
140:323, 1989[Medline]
[Order article via Infotrieve]
32.
Takahira H, Gotoh A, Ritchie A, Broxmeyer HE:
Steel factor enhances integrin-mediated tyrosine phosphorylation of focal adhesion kinase (pp125FAK) and paxillin.
Blood
89:1574, 1997[Abstract/Free Full Text]
33.
Gotoh A, Takahira H, Geahlen RL, Broxmeyer HE:
Cross-linking of integrins induces tyrosine phosphorylation of the proto-oncogene product vav and the protein kinase syk in human factor-dependent myeloid cells.
Cell Growth Differ
8:721, 1997[Abstract]
34.
Lévesque JP, Leavesley DI, Niutta S, Vadas M, Simmons PJ:
Cytokines increase human hematopoietic cell adhesiveness by very late antigen (VLA)-4 and VLA-5 integrins.
J Exp Med
181:1805, 1995[Abstract/Free Full Text]
35.
Lévesque JP, Haylock DN, Simmons PJ:
Cytokine regulation of proliferation and cell adhesion are correlated events in human CD34+ hematopoietic progenitors.
Blood
88:1168, 1996[Abstract/Free Full Text]
36.
Cui Li, Ramsfjell V, Borge OJ, Veiby OP, Lok Si, Jacobsen SEW:
Thrombopoietin promotes adhesion of primitive human hematopoietic cells to fibronectin and vascular cell adhesion molecule-1. Role of activation of very late antigen (VLA)-4 and VLA-5.
J Immunol
159:1961, 1997[Abstract]
37.
Kinashi T, Escaobedo JA, Williams LT, Takatsu K, Springer TA:
Receptor tyrosine kinase stimulates cell-matrix adhesion by phosphatidylinositol 3 kinase and phospholipase C-1 pathways.
Blood
86:2086, 1995[Abstract/Free Full Text]
38.
Tanaka M, Sabe H, Hata A, Inazu T, Homma Y, Nukada T, Yamamura H, Kurosaki T:
Tyrosine kinases Lyn and Syk regulate B cell receptor-coupled Ca2+ mobilization through distinct pathways.
EMBO J
6:1341, 1994
39.
Law C-L, Chandran KA, Sidrenko SP, Clark EA:
Phospholipase C-1 interacts with conserved phosphotyrosyl residues in the linker region of Syk and is a a substrate for Syk.
Mol Cell Biol
16:1305, 1996[Abstract]
40.
Odai H, Sasaki K, Iwamatsu A, Hanazono Y, Tanaka T, Mitani K, Yazaki Y, Hirai H:
The proto-oncogene product c-Cbl becomes tyrosine phosphorylated by stimulation with GM-CSF or Epo and constitutively binds to the SH3 domein of Grb2/Ash in human hematopoietic cells.
J Biol Chem
270:10800, 1995[Abstract/Free Full Text]
41.
Brizzi MF, Dentelli P, Lanfrancone L, Rosso A, Pelicci PG, Peoraro L:
Discrete protein interactions with Grb2/c-Cbl complex in SCF- and TPO-mediated myeloid cell proliferation.
Oncogene
13:2067, 1996[Medline]
[Order article via Infotrieve]
42.
Ota Y, Samelson LE:
The product of the proto-oncogene c-cbl: A negative regulator of the syk tyrosine kinase.
Science
276:416, 1997
43.
Leavesley DI, Oliver JM, Swart BW, Berndt MC, Haylock DN, Simmons PJ:
Signals from platelet/endothelial cell adhesion molecule enhance the adhesive activity of very late antigen-4 integrin of human CD34+ hematopoietic progenitor cells.
J Immunol
153:4673, 1994[Abstract]
44.
Wesseling J, van der Valk SW, Vos HL, Sonnenberg A, Hilkens J:
Episialin (MUC1) overexpression inhibits integrin-mediated cell adhesion to extracellular matrix components.
J Cell Biol
129:225, 1995
45.
Zell T, Hunt SW III, Mobley JL, Finkelstein LD, Shimizu Y:
CD28-mediated up-regulation of 1-integrin adhesion involves phosphatidylinositol 3-kinase.
J Immunol
156:883, 1996[Abstract]
46.
Chan ASH, Mobley JL, Fields GB, Shimizu Y:
CD7-mediated regulation of integrin adhesiveness on human T cells involves tyrosine phospholiration-dependent activation of phosphatidylinositol 3-kinase.
J Immunol
159:934, 1997[Abstract]
47.
Zauli G, Bassini A, Vitale M, Gibellini D, Celeghini C, Caramelli E, Pierpaoli S, Guidotti L, Capitani S:
Thrombopoietin enhances the II3-dependent adhesion of megakaryocytic cells to fibrinogen or fibronectin through PI 3 kinase.
Blood
89:883, 1997[Abstract/Free Full Text]
48.
Gotoh A, Ritchie A, Takahira H, Broxmeyer HE:
Thrombopoietin and erythropoietin activate inside-out signaling of integrin and enhance adhesion to immobilized fibronectin in human growth-factor-dependent hematopoietic cells.
Ann Hematol
75:207, 1997[Medline]
[Order article via Infotrieve]
49.
Smith RJ, Justen JM, McNab AR, Rosenbloom CL, Steele AN, Detmers PA, Anderson DC, Manning AM:
U-73122: A potent inhibitor of human polymorphonuclear neutrophil adhesion on biological surfaces and adhesion-related effector functions.
J Pharmacol Exp Ther
278:320, 1996[Abstract/Free Full Text]
50.
Alvarado M, Klassen C, Cerny J, Horejsí V, Schmidt RE:
MEM-59 monoclonal antibody detects a CD43 epitope involved in lymphocyte activation.
Eur J Immunol
25:1051, 1995[Medline]
[Order article via Infotrieve]
51.
Manjunath N, Ardman B:
CD43 regulates tyrosine phosphorylation of a 93-kD protein in T lymphocytes.
Blood
86:4194, 1995[Abstract/Free Full Text]
52.
Pedraza-Alva G, Mérida LB, Burakoff SJ, Rosenstein Y:
T cell activation through the CD43 molecule leads to vav tyrosine phosphorylation and mitogen-activatated protein kinase pathway activation.
J Biol Chem
273:14218, 1998[Abstract/Free Full Text]
53.
Rellahan BL, Graham LJ, Stoica B, DeBell KE, Bonvini E:
Cbl-mediated regulation of T cell receptor-induced AP1 activation. Implication for activation via the ras signaling pathway.
J Biol Chem
272:30806, 1997[Abstract/Free Full Text]
54.
Ueno H, Sasaki K, Miyagawa K, Honda H, Mitani K, Yazaki Y, Hirai H:
Antisense repression of proto-oncogene c-Cbl enhances activation of the JAK-STAT pathway but not the ras pathway in epidermal growth factor receptor signaling.
J Biol Chem
272:8739, 1997[Abstract/Free Full Text]
55.
Ueno H, Sasaki K, Honda H, Nakamoto T, Yamagata T, Miyagawa K, Mitani K, Yazaki Y, Hirai H:
c-Cbl is tyrosine-phosphorylated by interleukin-4 and enhances mitogenic and survival signals of interleukin-4 receptor by linking with the phosphatidlinositol 3'-kinase pathway.
Blood
91:465, 1998
56.
Hao Q-L, Shah AJ, Thiemann FT, Smogorzewska EM, Crooks GM:
A functional comparison of CD34+ CD38 cells in cord blood and bone marrow.
Blood
86:3745, 1995[Abstract/Free Full Text]
57.
De Smet W, Walter H, Van Hove L:
A new monoclonal antibody induces homotypic aggregation of human leukocytes through a CD11a/CD18-dependent and -independent mechanism.
Immunology
79:46, 1993[Medline]
[Order article via Infotrieve]
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ABSTRACT
INTRODUCTION