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 Previous Article | Table of Contents | Next Article 
Blood, Vol. 94 No. 9 (November 1), 1999:
pp. 2990-2998
SDF-1 Responsiveness Does Not Correlate With CXCR4 Expression
Levels of Developing Human Bone Marrow B Cells
By
Marek Honczarenko,
Raymond S. Douglas,
Clarissa Mathias,
Benhur Lee,
Mariusz Z. Ratajczak, and
Leslie E. Silberstein
From the Division of Hematology-Oncology, the Departments of
Pathology and Laboratory Medicine and Medicine, University of
Pennsylvania School of Medicine, Philadelphia, PA.
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ABSTRACT |
Chemokines and their receptors are broadly expressed in different
tissues and are involved in diverse biologic processes. Gene
inactivation studies have shown that both stromal cell derived factor-1
(SDF-1) and chemokine receptor 4 (CXCR4) are essential for B
lymphopoiesis. However, it is not yet clear by which mechanisms B
lymphopoiesis is affected. In the present study, we have examined CXCR4
expression and function on primary B cells representing sequential
stages of development (eg, pro-B, pre-B, immature, and
mature B cells) in fetal and adult bone marrow. The expression of CXCR4
was observed to be sinusoidal. Expression was highest on pre-B cells,
decreased as cells developed into immature B cells, and then increased
again upon transition to the mature B-cell stage. The corresponding
ligand SDF-1 was shown to trigger vigorous cell signaling and migration
responses, which are restricted to early lineage B cells. The
responsiveness to SDF-1 was markedly decreased for immature and mature
B cells despite relatively high levels of CXCR4 expression. Thus, the
diminished responsiveness to SDF-1 by more mature B cells was
determined to be disproportionate to the level of CXCR4 expression.
These findings raise the possibility that CXCR4 function is
differentially controlled during B lymphopoiesis and may be relevant to
the compartmentalization of B-cell precursors in the bone marrow.
© 1999 by The American Society of Hematology.
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INTRODUCTION |
CHEMOKINES ARE RESPONSIBLE for numerous
activities in normal and pathologic states.1,2 They
influence migration of lymphocytes through a gradient of chemokine
concentrations and play a role in leukocyte adhesion and infiltration
during inflammation.3-6 Some chemokines affect
hematopoiesis and angiogenesis.7-11 Of these, the chemokine
stromal cell derived factor-1 (SDF-1) and corresponding receptor
chemokine receptor 4 (CXCR4) are essential for B lymphopoiesis and bone
marrow myelopoiesis. Mice, in which either the gene for SDF-1 or CXCR4
has been inactivated, have reduced numbers of both pro-B (progenitor B
cells) cells and (precursor B cells) pre-B cells in fetal liver and
bone marrow.12-14
The mechanism(s) by which loss of SDF-1 or CXCR4 expression affects B
lymphopoiesis is not clear. SDF-1 can induce chemotaxis of murine
B-cell progenitors, thus suggesting that the SDF-1/CXCR4 axis may be
important in directing the migration of B-cell progenitors to the
appropriate bone marrow microenvironment.13,15,16
Alternatively, SDF-1, which can stimulate proliferation of murine
B-cell progenitors in vitro, also may trigger signaling pathways
necessary for the retention and development of early lineage B cells in
the bone marrow.17,18 In this regard, SDF-1 has been shown
to induce calcium mobilization of murine progenitor B cells but,
interestingly, not mature bone marrow B cells.15 In the
current study, we wished to further examine for differential SDF-1
responsiveness relative to CXCR4 expression during B lymphopoiesis. In
addition, the studies were performed on human bone marrow as
differences in biologic responses might exist between human and mouse B
lymphopoiesis.19
We examined primary human B cells representing sequential stages of
development (eg, pro-B, pre-B, immature, and mature B cells) in bone
marrow of the same individual. First, we found distinct CXCR4
expression patterns on bone marrow B-cell populations. Second, we
showed that SDF-1 triggered marked biologic responses of early B
lineage cells (eg, pro-B and pre-B), whereas the same responses of more
mature B cells were diminished or absent. Interestingly, the diminished
or lack of SDF-1 induced responsiveness by more mature B cells was
disproportionate to the level of CXCR4 expression. These findings
suggest that CXCR4 function may be regulated during B-cell development
and, thus, influence the distinct anatomic location of B-cell
progenitors in the bone marrow.
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MATERIALS AND METHODS |
Cells.
The following cell lines were obtained from ATCC: RS4:11 (CRL 1873) , REH (CRL 8286), HS-Sultan (CRL 1484). The JIM-1 cell line20
was provided by Dr Jack W. Singer (Cell Therapeutics Inc, Seattle, WA),
the 697 cell line21 was provided by Dr Max D. Cooper
(Howard Hughes Medical Institute, Birmingham, AL), the RS11846 cell
line22 was provided by Dr John Reed (La Jolla Cancer Institute, La Jolla, CA), the Blin-123 Nalm-16, the Nalm-6
cell lines24 were provided by Dr Tucker LeBien (University
of Minnesota, Minneapolis, MN), the SU-DHL-6 cell line25
was provided by Dr Alan Epstein (University of Southern California, Los
Angeles, CA), and the 20A and 15A26 cell lines were
established in our laboratory from patients with cold agglutinin disease.
Fetal bone marrow was obtained from 18- to 23-week-old fetuses. Adult
heparinized bone marrow was obtained by iliac crest aspiration from
healthy adult individuals. Umbilical cord blood cells were obtained
from full-term deliveries without evidence of maternal or fetal
infection. The isolated bone marrow cells were passed through a 70 µm
filter to remove cellular debris and aggregates. Mononuclear cells were
isolated by Ficoll-Hypaque (Pharmacia, Piscataway, NJ) gradient
centrifugation (density 1.077 g/mL). Pelleted cells were collected and
Ficoll (Pharmacia) was removed by washing the cells 3 times in 1X
phosphate-buffered saline (PBS), pH 7.4. After the third wash, cells
were resuspended in ice-cold staining buffer (1X PBS with 2% fetal
bovine serum [FBS]). Fetal bone marrow cells were stored at 37°C
in Iscove (GIBCO, Grand Island, NY), serum free medium for 24 hours
before staining.
Antibodies.
Surface staining of bone marrow cells was performed with the following
antibodies: fluorescein isothiocyanate (FITC)-labeled anti- Light
chain (Sigma, St Louis, MO), FITC-labeled anti- Light chain
(Pharmingen, San Diego, CA), phycoerythrin (PE)-labeled anti-CD19
(Becton Dickinson, San Jose, CA), Cy-Chrome-labeled anti-CD19
(Pharmingen, San Diego, CA), PECy5-labeled anti-CD34 (Immunotech,
Westbrook, ME), PE-labeled anti-IgD (Pharmingen), PE-labeled anti-CXCR4
(clone 12G5) (Pharmingen). The secondary antibody was goat anti-mouse
IgG-APC labeled (Accurate Chemical & Scientific Corporation, Westbury,
NY). Nonconjugated mouse monoclonal antibody (MoAb) anti-CXCR4 (clone
12G5, kindly provided by Dr James Hoxie, University of
Pennsylvania, Philadelphia, PA).
Cell sorting.
The Ficoll-Hypaque-purified bone marrow cells (Pharmacia) were
suspended in ice-cold staining buffer (1X PBS, 2% FBS) and incubated
for 30 min with the first step reagent. Cells were washed three times
in staining buffer and incubated for 30 minutes with the second step
reagents. Four stages of B-cell differentiation were isolated from bone
marrow cells as previously described. The earliest B-cell population,
designated here as pro-B cells, was
 /, CD19+,
CD34+. The next developmental B-lineage populations were
designated as pre-B cells, identified as
/, CD19+,
CD34; followed by immature B cells identified as
+/+, CD19+,
IgD; and mature B cells identified as
+/+, CD19+, IgD+.
CellQuest software (Becton Dickinson Immunocytometry Systems) was used
to define gates. The gating strategy for the different B-cell subsets
is shown on Fig 1A.
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| Fig 1.
(A) Gating strategy for identifying different subsets of
B cells from fetal, adult bone marrow, and umbilical cord blood. Cells
were first gated based on forward and side scatter identifying the
lymphocyte population. Staining for kappa and lambda light chains
identified early (pro-B/pre-B), eg,
/, and later (immature and mature B
cells), eg, +/+, lineage B cells.
Cells of the lymphocyte phenotype and negative for kappa/lambda
immunoglobulin chains were further separated into CD19+,
CD34+ pro-B cells and CD19+,
CD34 pre-B cells. Cells positive for kappa/lambda
immunoglobulin chains were further identified as immature B cells
(CD19+, IgD) and mature B cells
(CD19+, IgD+). (B) CXCR4 expression during
B-cell development in bone marrow. Expression of CXCR4 on B-cells from
adult (A), fetal (B) bone marrow, and umbilical cord blood (C) is
shown. Cells were incubated with anti-CXCR4 specific antibody and
analyzed by flow cytometry. Triple gates were set on different B-cell
subsets and the percentage of CXCR4 positive cells was determined. The
threshold line for positive cells was based on the maximum staining of
a matched isotypic antibody with irrelevant specificity used in the
same concentration as the anti-CXCR4 antibody. The isotype control
antibody binding was analyzed on the same triple gated B-cell subsets
as for CXCR4 expression. Anti-CXCR4 labeled cells that were brighter
than those stained with the isotype control antibody were defined as
positive for CXCR4. The shaded part of the histogram represents cells
stained with anti-CXCR4 antibody, and the black line represents
staining with the isotype control. The numbers represent the percentage
of CXCR4 positive cells. The results are representative of 5 experiments using adult bone marrow B cells and 3 experiments each of
fetal bone marrow and umbilical cord blood B cells.
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The sorting into highly purified populations was performed on a
fluorescence-activated cell sorter (FACS) Vantage with Turbo Sort
option (Becton Dickinson Immunocytometry Systems), with the purity over
98.5%. The sorter was aligned with glutaraldehyde fixed chicken red
blood cells.
Cells used in chemotaxis and calcium flux assays were stored overnight
at 37°C in Iscove Dulbecco's modified Eagle's medium (DMEM)
(GIBCO) (105 cells/mL), supplemented with 25% of
artificial serum containing 1% delipidated, deionized, and
charcoal-treated bovine serum albumin (BSA), 270 µg/mL iron saturated
transferrin, insulin (20 µg/mL), and 2 mmol/l L-glutamine (all from
Sigma, St Louis, MO).27
Cell surface staining.
Four color immunofluorescence analysis was used to examine the
expression of chemokine receptors on the surfaces of different B-cell
populations. Different subsets of B cells were identified as described
above (Fig 1A). The fourth channel was used to define cell surface
expression of CXCR4. Ten thousand events were acquired in the fourth
channel after triple gating for B-cell subsets, as described above. The
threshold line was based on the maximum staining of a matched isotypic
antibody with irrelevant specificity (mouse IgG2a) used in
the same concentration as the anti-CXCR4 antibody (1 µg/50 µL of
staining buffer). The isotypic antibody binding was analyzed on the
same triple gated B-cell subsets as for CXCR4 expression. The threshold
line for defining CXCR4 positive and negative cells was set such that
less than 1% of isotype positive cells was present to the right of the
threshold line. Anti-CXCR4 labeled cells that were brighter than those
stained with the isotypic antibody (present to the right of isotype
control histogram) were defined as positive for CXCR4.
To detect internalization of CXCR4, cells were incubated for 30 minutes
at 37°C with 125 nmol/L of SDF-1. After incubation, cells were put
on ice to prevent resurfacing of the receptor, washed in ice-cold acid
buffer (50 mmol/L glycine, 100 mmol/L NaCl, pH 3.0) to remove cell
bound SDF-1, which could interfere with 12G5 binding to CXCR4, and then
stained with anti-CXCR4 antibody.
The flow cytometer was calibrated with CaliBRITE-3 beads (Becton
Dickinson Immunocytometry Systems) and FACSComp Software (Becton
Dickinson Immunocytometry Systems). The data were analyzed with the
WinMDI 2.7 software (kindly provided by Dr Joseph Trotter, Scripps
Research Institute, La Jolla, San Diego, CA).
Quantitative FACS.
Quantitative FACS was performed by converting the mean fluorescence
intensity of the cell subset into antibody-binding sites (ABS) by using
a standardized microbeads kit (Sigma, St Louis, MO). This is a mixture
of 5 microbead populations of uniform size, coated with goat anti-mouse
antibodies that have different properties to bind mouse antibodies.
Beads (105 per sample) were incubated with the same,
saturating concentration of 12G5-PE labeled (Pharmingen) anti-CXCR4
antibody as the samples and were processed identically as
the samples being quantified. The beads were then analyzed by using the
same instrument settings as those for the sample cells. The binding
capacities of the stained beads were then regressed against the
corresponding geometric mean of each bead population and the mean
fluorescence intensity of the CXCR4 on the cells was converted to ABS
per cell by comparison with the regression curve generated. The mean
fluorescence intensity of the isotype control (68-70 antibody binding
sites) for each B-cell subset was also converted to ABS and subtracted
from ABS values obtained with the experimental sample. The threshold of detection was 110 ABS per cell. Additional details on the relationship between ABS and mean fluorescence intensity values can be obtained from
the manufacturer (Sigma).
Calcium signaling.
Sorted cells were stored for 12 to 18 hours before the experiment at
37°C in Iscove DMEM (GIBCO) (105 cells/mL),
supplemented with 25% of artificial serum.27 Cells were
suspended in 1 mL of 30°C Hanks' balanced salt solution containing 1.3 mmol/L CaCl2, 0.5 mmol/L MgCl2, 1 µmol/L
Indo-1AM (Molecular Probes, Eugene, OR), 0.01% F-127 Pluronic
detergent (Molecular Probes), and incubated for 45 minutes at 30°C.
Calcium mobilization was analyzed with Becton Dickinson FACStar Plus
flow cytometer (Becton Dickinson Immunocytometry Systems), equipped
with Time Zero Injection Module (Cytek, Fremont, CA) after stimulation
with 100 ng/mL of SDF-1 (R&D Systems, Minneapolis, MN) or 2 µg/mL of anti-IgM MoAb (Sigma). Ultraviolet (UV) excitation
wavelengths were 350 to 364 nm, emission 405 nm, and 510 nm. UV laser
was aligned by using FluoresBRITE carboxyBB 6 µm microspheres (Poly Science, Niles, IL). Cells were incubated at 37°C for 5 minutes before analysis. The data were acquired with Lysis II software (Becton
Dickinson Immunocytometry Systems) and analyzed with WinMDI 2.7 software.
Chemotaxis assay.
In the initial experiments, B-cell subsets were first sorted from total
bone marrow and then placed into the upper compartment of the Transwell
and allowed to migrate as described below. Sorted cells used in these
experiments were stored for 12 to 18 hours before the experiment at
37°C in Iscove DMEM (GIBCO) (105 cells/mL) and
supplemented with 25% of artificial serum.27
The ratio between the number of cells migrated in the presence of
chemokine and the number of cells migrated in the presence of media
alone was defined as the migration index.
In subsequent experiments, total bone marrow Ficoll-purified
lymphocytes were put into the upper compartment (106 cells
per well) of Transwell inserts (6.5 mm diameter, 5 µm filter pore
size) (Costar, Cambridge, MA). The lower compartment contained 600 µL
of serially diluted SDF-1 (R&D Systems) in 0.25% BSA, RPMI 1640. Cells were allowed to migrate for 2 hours at 37°C. Cells that
passed through the membrane to the lower chamber were collected and
stained with B-cell markers. Cells were counted by timed acquisition
(600 seconds each sample) on FACSCalibur flow cytometer (Becton
Dickinson Immunocytometry Systems).
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RESULTS |
Expression of CXCR4 on different B-cell lines.
As a first measure for changes in CXCR4 expression during B-cell
development, we analyzed different B-cell lines representing early and
mature stages of B-cell development (Table
1). Overall, the percentage of CXCR4 expressing cells was highest on
B-cell lines representing early lineage (ie, pro-B/pre-B) B cells,
although a much lower percentage was noted for mature B-cell lines
derived from peripheral lymphomas.
We also examined the function of CXCR4 on different cell lines. We
found that as previously reported,15 CXCR4 mediated
chemotaxis and calcium mobilization was generally high on pro-B and
pre-B cell lines but low or absent on lymphoma derived B cell. However, some exceptions were noted. Certain pro-B cell lines were nonresponsive to SDF-1 (RS 4:11, JIM-1; see Table 1), as measured by calcium mobilization; some mature lymphoma derived cell lines did migrate to
the SDF-1 ligand (RS11846, HS Sultan). Overall, the results of these
studies were intriguing and suggested a correlation between CXCR4
function and developmental B-cell stage that warranted further investigation by using untransformed cells.
Expression of CXCR4 on subsets of primary B cells from adult bone
marrow.
The expression of CXCR4 during B-cell development was examined on
subsets of B lymphocytes from adult bone marrow (Fig 1A). As shown in
Fig 1B, 30% of pro-B cells expressed CXCR4 (Panel A). Among pre-B
cells, 89% of cells stained positive for CXCR4. At the next stage of
B-cell development, the immature B-cell stage, the frequency of CXCR4
positive cells is decreased as compared with the pre-B cell population.
Among mature B cells from bone marrow, the frequency of CXCR4 positive
cells increased to 79%.
Expression of CXCR4 on subsets of B cells from fetal bone marrow and
umbilical cord blood.
We were curious if CXCR4 expression differed during ontogeny.
Therefore, we examined CXCR4 expression on different subsets of B cells
from fetal bone marrow (Fig 1B, Panel B) and umbilical cord blood (Fig
1B, Panel C). Similar to adult bone marrow, the percentage of cells
expressing CXCR4 increases during the pro-B to pre-B cell transition
(Fig 1B). Because of the small number of cells available for analysis,
we were unable to determine CXCR4 expression on fetal immature B cells.
However, the majority of mature B cells expressed CXCR4, which was also
noted for mature B cells from adult bone marrow (Fig 1B). We noted that
CXCR4 expression on pro-B cells from fetal bone marrow and umbilical
cord blood B cells was higher than on pro-B cells from adult bone
marrow. The reason for this difference in expression is currently not clear. However, CXCR4 expression on pre-B and mature B cells was similar during ontogeny.
CXCR4 mediated signal transduction on different B-cell subsets.
One of the events that characterize activation of the G-protein coupled
receptors is mobilization of calcium from intracellular stores, leading
to a transient rise in intracellular calcium
concentration.1,28-30 We tested calcium mobilization after
stimulation of B lineage cell subsets sorted from bone marrow with 100 nmol/L SDF-1 (Fig 2). Eighty-seven
percent of pro-B and 81% of pre-B cells stimulated with SDF-1
showed a significant agonist-dependent calcium response, which appeared
15 to 20 seconds after stimulation and returned to base level after 2 to 3 minutes. The response by pro-B cells was considered particularly
strong considering that on average the percentage of CXCR4 positive
cells was lower than for pre-B and much lower than for mature B cells
(see Fig 1B). In contrast, only 3% of mature B cells showed a calcium
mobilization response to SDF-1, despite their ability to respond to
other stimuli (anti-IgM MoAb) (Fig 2).
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| Fig 2.
SDF-1 induced calcium mobilization of B-cell subsets
sorted from adult bone marrow. Sorted B cells were loaded with Indo-1AM
and stimulated at the indicated time points (arrows) with the SDF-1
(100 nmol/L) or anti-IgM (2 µg/mL). The calcium mobilization is
expressed as the violet/green ratio during the indicated time course.
As a positive control, cells were stimulated with anti-IgM MoAb. The
results are representative of 4 experiments using primary B cells from
adult bone marrow. Two experiments were done on pro-B and pre-B cells
from fetal bone marrow. In addition, data were confirmed by analysis of
calcium mobilization of developmental stage specific B-cell lines: REH
(pro-B), 697 (pre-B), Nalm-6 (pre-B) and 15A (mature).
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Internalization of CXCR4 by exposure to SDF-1.
We also examined whether binding of SDF-1 would cause a similar degree
of CXCR4 internalization on both early and late lineage B cells. CXCR4
was internalized on early and late B cells with a comparable decrease
in expression, ie, an 86% decrease for early and an 87% decrease for
late B-cell subsets (Fig 3).
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| Fig 3.
SDF-1 induced internalization of CXCR4 from surface of
early and mature B cells. Total bone marrow lymphocytes were incubated
for 30 minutes at 37°C with 125 nmol/L of SDF-1 and placed on ice
immediately after incubation to prevent resurfacing of the receptor.
Cells were subsequently washed in ice-cold glycine buffer to remove
cell bound SDF-1. Next, the cells were stained for CXCR4 expression in
parallel with surface markers defining (A) early and (B) late lineage B
cells (see Fig 1A). The early lineage B-cell population contained both
pro-B and pre-B cells, whereas late lineage B cells contained immature
and mature B cells. The shaded part of the histogram represents
untreated cells; the gray line represents cells incubated with SDF-1;
the dashed black line represents antibody isotype control. The results
are representative of 3 experiments with primary B cells.
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Chemotactic responses of different B-cell subsets toward
SDF-1.
Chemotaxis was tested across a wide range of SDF-1 concentrations
(0.125 nmol/L to 125 nmol/L), previously shown to mediate responses of
T cells and macrophages.31,32 In the initial experiments (Fig 4, Panel A), bone marrow B cells were
first sorted into purified populations (see Fig 1A) and then examined
for migration to SDF-1, as previously reported for murine bone marrow B
cells.15 Both pro-B and pre-B cells showed weak chemotactic
responses to SDF-1. The maximal observed migration index (±1 SD)
was 7.55 ± 4.17 for pro-B cells and 4.3 ± 3.7 for pre-B cells.
Immature and mature B cells did not respond to SDF-1; the highest
migration indices (±1 SD) of 1.3 ± 0.42 and 1.4 ± 0.42 for
immature and mature B cells, respectively, were not significantly
different from controls (see Fig 4A).
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| Fig 4.
Migration of different B-cell subsets from adult bone
marrow to human SDF-1. Chemotaxis of FACS-sorted B cells (A) or B
cells from total bone marrow (B) in response to serial dilutions of
SDF-1 (1 to 1,000 ng/mL) or medium alone is shown. In (B), early
lineage B cells define a cell population that includes both pro-B
(CD19+, /,
CD34+) and pre-B (CD19+,
/, CD34) cells.
Results are expressed as the migration index, which is defined as the
number of cells migrated to the lower compartment in the presence of
the chemokine divided by the number of cells that have migrated to the
lower compartment in the presence of medium alone. A migration index 1 indicated the migration of 1.1% of total input cells and a migration
index of 20 equaled 35% migration of total input cells. The results
are representative of 4 experiments with primary B cells from total
bone marrow and 2 with sorted bone marrow B cells.
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Because of concern of cell stress caused by cell sorting, total bone
marrow cells were loaded into Transwell and the migrated cells were
then stained with appropriate markers to define B-cell subsets. Due to
limitation in cell number, we defined the migrated B-cell subsets as
either early lineage, eg, a cell population that included both pro-B
and pre-B (CD19+,
/, CD34+and
CD19+, /,
CD34) (Fig 1A) cells, or late lineage, eg, immature
(CD19+, +/+,
IgD) and mature B cells (CD19+,
+/+, IgD+) (Fig 1A). In
comparison to responses by other primary hematopoietic cells, the early
lineage (pro-B and pre-B) cells showed a strong chemotactic response to
SDF-1.31-33 The maximal migration index (±1 SD) for
early lineage B cells was 21.3 ± 3 at a SDF-1 concentration of
100 ng/mL (12.5 nmol/L) (see Fig 4B). Immature and mature B cells
showed a much weaker chemotactic response toward SDF-1 with highest
migration indices (±1 SD) of 5.4 ± 2 and 7.3 ± 2.2 for
immature and mature B cells, respectively, at a comparable SDF-1
concentration of 100 to 1,000 ng/mL (see Fig 4B).
The chemotaxis data on sorted cells (panel A) are shown for comparison
to previously reported data on sorted murine bone marrow B cells.
Moreover, the data show the influence of mechanical stress conferred by
high-speed cell sorting on the chemotaxis assay.
Comparison of CXCR4 expression and function between early and late
lineage B cells.
For this comparison, we also calculated the number of anti-CXCR4
antibody binding sites per B cell to complement the CXCR4 expression
analyses in Fig 1, which shows the percentage of anti-CXCR4 positive
cells compared with the binding of an isotype control antibody. Due to
a limitation in cell number described above, the migration responses to
SDF-1 by early lineage B cells were calculated on a cell population
that included both pro-B and pre-B cells. Therefore, as shown in Table
2, we compared CXCR4 expression and function of early and late lineage
B cells. The studies on early lineage B cells included both pro-B
(CD19+, /,
CD34+) and pre-B (CD19+,
/, CD34)
cell populations and those on late lineage B cells included immature
(CD19+, +/+,
IgD) and mature (CD19+,
+/+, IgD+) B-cell populations
(see Fig 1A). The number of CXCR4 molecules varied considerably between
3 different donors, with a range of 7,352 to 12,977 for early lineage
(pro/pre) B cells and a range of 4,046 to 9,112 for late lineage
(immature/mature) B cells (see Table 2).
The difference between the number of CXCR4 positive cells and
anti-CXCR4 (12G5) antibody binding sites between early and late stages
of B-cell development was not statistically significant (P > 2) (T-test to compare 2 independent sample means). The decrease in function between early and late lineage B cells was statistically significant as measured by calcium mobilization (P < .01) and chemotaxis (P < .05) (T-test to compare 2 independent
sample means). Taken together, these data suggested that the decrease
in SDF-1 responsiveness was disproportionate to changes in CXCR4
expression on developing human B-cell populations from bone marrow.
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DISCUSSION |
The importance of SDF-1 and CXCR4 in B lymphopoiesis has been shown in
murine studies involving gene inactivation.12-14 However, the mechanisms by which these molecules affect B lymphopoiesis are not
yet clear. In the current analysis of human B cells, we assessed
primary bone marrow B cell and correlated CXCR4 expression and function
at each developmental stage. We observed that unlike early lineage B
cells that responded markedly to SDF-1, the late lineage B cells (eg,
immature and mature) were poorly responsive to SDF-1 stimulation. For
example, SDF-1 stimulation caused a calcium mobilization response of
only 3% of mature B cells as compared with over 84% of early lineage
B cells (Figs 2 and 5). In addition, the
migration responses of immature/mature B cells were greatly diminished
(more than 70%) compared with early lineage B cells (see
Table 2 and Figs 4 and 5). Of note, a
similar decrease in SDF-1 responsiveness has been observed in thymocyte
subsets of different maturation stages.34
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| Fig 5.
The reduction of SDF-1 responsiveness during B
lymphopoiesis is disproportionate to the decrease in CXCR4 expression.
The terms "CXCR4 positive cells" and "anti-CXCR4 (12G5)
antibody binding sites per cell" are defined in Materials and
Methods. Presence of calcium mobilization after stimulation describes
the percentage of cells responding with calcium mobilization after
SDF-1 stimulation. Migration of cells toward SDF-1 describes the ratio
between the number of cells that migrated in the presence of chemokine
and the number of cells that migrated in the presence of media alone.
CXCR4 expression and function data of late lineage (immature and
mature) B cells are normalized to the data of early (pro-B and pre-B)
lineage B cells (defined as equal to 1). The data represent the mean of
experiments shown in Figs 2 and 4 and Table 2.
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The magnitude of the response to SDF-1 could be related to either the
frequency of CXCR4 positive cells or the number of CXCR4 receptor
molecules expressed on B cells. In Table 2, early lineage B cells
(pro-B and pre-B) are compared with late lineage B cells (immature/mature) from bone marrow. The percentage of CXCR4 positive cells among late lineage B cells decreased by ~17%, and the number of 12G5 antibody binding sites on late B cells decreased by ~35% in
comparison with early lineage cells (see Figs 1B and 5 and Table 2).
Nevertheless, mature B cells express similar or higher levels of CXCR4
compared with monocytes or T cells35 and these latter cell
types show vigorous responses to SDF-1 as measured by calcium
mobilization and chemotaxis.33,36-38 Moreover, when receptor signaling is measured as an average increase of calcium mobilization, a strong response is detectable on a cell population with
as few as 11% of cells being CXCR4 positive.39 Therefore, the level of CXCR4 expression on mature B cells appears sufficient to
generate an SDF-1 response as measured by chemotaxis or calcium mobilization.
We also assessed to what extent bound SDF-1 would lead to
internalization of CXCR4 on early and late lineage B cells. As
illustrated in Fig 3, CXCR4 was internalized by comparable levels on
both early and late lineage B cells. At first, we considered the degree of CXCR4 internalized on mature cells to be unexpected in view of the
poor/absent SDF-1 responsiveness measured by chemotaxis and calcium
mobilization. However, internalization of CXCR4 has been shown to be
uncoupled from G-protein-dependent intracellular signaling
pathways.40-42 Rather internalization of CXCR4 by SDF-1 stimulation appears to be regulated by pathways, which do not lead to
an increase in intracellular calcium.43 Thus, these data
suggest that CXCR4 on both early and late lineage B cells can bind to
SDF-1, which subsequently leads to internalization of the receptor.
However, as discussed below there is a lack of correlation between
CXCR4 expression and SDF-1 responsiveness as measured by chemotaxis and
calcium mobilization.
When CXCR4 expression and function of mature B cells are normalized to
early lineage B cells (Fig 5), the reduction in SDF-1 responsiveness
during B lymphopoiesis is disproportionate to the decrease in CXCR4
expression. Based on these analyses, we speculate that CXCR4 function
may be controlled during development by processes involving either RGS
proteins, posttranslational modification and/or cross-desensitization
of the chemokine receptor.38,44-50 With respect to
posttranslational modification, for example, it may be possible that
the tyrosine residues of CXCR4 on mature B cells are less sulfated
compared with early lineage B cells, thus accounting for their poor
response to SDF-1. As shown in vitro for T cells, cellular responses to
chemokines may be influenced by their activation state.51
However, we were unable to elicit a calcium mobilization response to
SDF-1 on mature B cells from bone marrow, which had been activated in
vitro with anti-IgM (data not shown). Nevertheless, it remains possible
that the biologic cellular responses mediated by other chemokine
receptors may be linked to B-cell activation state.
Clearly additional studies are needed to investigate the mechanisms
underlying the lack of correlation between CXCR4 expression and SDF-1
responsiveness on developing bone marrow B cells. It is attractive to
speculate, however, that the observed decrease in SDF-1 responsiveness
during B-cell maturation may be related to the distinct anatomic
location in the bone marrow of early and late lineage B lymphocytes.
The earliest B-cell progenitors are located within or near the
endosteum of the bone marrow. These cells, remaining in close contact
with stromal reticular cells in the peripheral region of bone marrow,
begin to differentiate into more mature B cells. During and after µ chain gene rearrangement, the B cells tend to move toward sinusoids
along the processes of adventitial reticular cells, and after beginning
to express sIgM, the immature B cells move into the sinusoid segment
and are released into blood circulation.52,53
SDF-1-mediated signaling triggers rapid adhesion of peripheral
lymphocytes to ICAM-1 through activation of integrins.54
Therefore, the SDF-1/CXCR4 axis may be important in retaining the early
lineage B cells in the appropriate bone marrow compartment through
activation of their adhesion molecules, eg, VLA-4, which binds to
VCAM-1.55-59
| |
ACKNOWLEDGMENT |
We are grateful to Dr Robert Doms and members of his laboratory for
helpful discussion during the course of the experiments described in
this article. We thank Dr John G. Monroe for critical review of the
manuscript and Dr Jonni Moore and Hank Pletcher for the assistance with
flow cytometry.
| |
FOOTNOTES |
Submitted May 10, 1999; accepted July 6, 1999.
Supported by Grant No. NIH P50 HL54516.
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 Leslie E. Silberstein, MD, Professor,
Departments of Pathology and Laboratory Medicine and Medicine,
University of Pennsylvania School of Medicine, 284 John Morgan
Building, 36th and Hamilton Walk, Philadelphia, PA 19104-6082; e-mail:
silbersl{at}mail.med.upenn.edu.
| |
REFERENCES |
1.
Murphy PM:
The molecular biology of leukocyte chemoattractant receptors.
Annu Rev Immunol
12:593, 1994[Medline]
[Order article via Infotrieve]
2.
Rollins BJ:
Chemokines.
Blood
90:909, 1997[Free Full Text]
3.
Adams DH, Lloyd AR:
Chemokines: Leukocyte recruitment and activation cytokines.
Lancet
349:490, 1997[Medline]
[Order article via Infotrieve]
4.
Conti P, Reale M, Barbacane RC, Frydas S, Felaco M, Grilli A, Placido FC, Cataldo I, Feliciani C, Di Gioacchino M, Anogianakis G, Dimitriadou D, Vacalis D, Trakatellis A:
Massive infiltration of basophilic cells in inflamed tissue after injection of RANTES.
Immunol Lett
58:101, 1997[Medline]
[Order article via Infotrieve]
5.
Lukacs NW, Standiford TJ, Chensue SW, Kunkel RG, Strieter RM, Kunkel SL:
C-C chemokine-induced eosinophil chemotaxis during allergic airway inflammation.
J Leukoc Biol
60:573, 1996[Abstract]
6.
Legler DF, Loetscher M, Roos RS, Clark-Lewis I, Baggiolini M, Moser B:
B cell-attracting chemokine 1, a human CXC chemokine expressed in lymphoid tissues, selectively attracts B lymphocytes via BLR1/CXCR5.
J Exp Med
187:655, 1998[Abstract/Free Full Text]
7.
Arenberg DA, Polverini PJ, Kunkel SL, Shanafelt A, Hesselgesser J, Horuk R, Strieter RM:
The role of CXC chemokines in the regulation of angiogenesis in non-small cell lung cancer.
J Leukoc Biol
62:554, 1997[Abstract]
8.
Strieter RM, Polverini PJ, Arenberg DA, Kunkel SL:
The role of CXC chemokines as regulators of angiogenesis.
Shock
4:155, 1995[Medline]
[Order article via Infotrieve]
9.
Strieter RM, Polverini PJ, Kunkel SL, Arenberg DA, Burdick MD, Kasper J, Dzuiba J, Van Damme J, Walz A, Marriott D, Chan SY, Roczniak S, Shanafelt AB:
The functional role of the ELR motif in CXC chemokine-mediated angiogenesis.
J Biol Chem
270:27348, 1995[Abstract/Free Full Text]
10.
Gao JL, Wynn TA, Chang Y, Lee EJ, Broxmeyer HE, Cooper S, Tiffany HL, Westphal H, Kwon-Chung J, Murphy PM:
Impaired host defense, hematopoiesis, granulomatous inflammation and type 1-type 2 cytokine balance in mice lacking CC chemokine receptor 1.
J Exp Med
185:1959, 1997[Abstract/Free Full Text]
11.
Kim CH, Broxmeyer HE:
In vitro behavior of hematopoietic progenitor cells under the influence of chemoattractants: Stromal cell-derived factor-1, steel factor, and the bone marrow environment.
Blood
91:100, 1998[Abstract/Free Full Text]
12.
Nagasawa T, Hirota S, Tachibana K, Takakura N, Nishikawa S, Kitamura Y, Yoshida N, Kikutani H, Kishimoto T:
Defects of B-cell lymphopoiesis and bone-marrow myelopoiesis in mice lacking the CXC chemokine PBSF/SDF-1.
Nature
382:635, 1996[Medline]
[Order article via Infotrieve]
13.
Zou YR, Kottman AH, Kuroda M, Tanichi I, Littman DR:
Function of the chemokine receptor CXCR4 in haematopoiesis and in cerebellar development.
Nature
393:595, 1998[Medline]
[Order article via Infotrieve]
14.
Ma Q, Jones D, Borghesani PR, Segal RA, Nagasawa T, Kishimoto T, Bronson RT, Springer TA:
Impaired B-lymphopoiesis, myelopoiesis, and derailed cerebellar neuron migration in CXCR4- and SDF-1-deficient mice.
Proc Natl Acad Sci USA
95:9448, 1998[Abstract/Free Full Text]
15.
D'Apuzzo M, Rolink A, Loetscher M, Hoxie JA, Clark-Lewis I, Melchers F, Baggiolini M, Moser B:
The chemokine SDF-1, stromal cell-derived factor 1, attracts early stage B cell precursors via the chemokine receptor CXCR4.
Eur J Immunol
27:1788, 1997[Medline]
[Order article via Infotrieve]
16.
Peled A, Petit I, Kollet O, Magid M, Ponomaryov T, Byk T, Nagler A, Ben-Hur H, Many A, Shultz L, Lider O, Alon R, Zipori D, Lapidot T:
Dependence of human stem cell engraftment and repopulation of NOD/SCID mice on CXCR4.
Science
283:845, 1999[Abstract/Free Full Text]
17.
Nagasawa T, Nakajima T, Tachibana K, Iizasa H, Bleul CC, Yoshie O, Matsushima K, Yoshida N, Springer TA, Kishimoto T:
Molecular cloning and characterization of a murine pre-B-cell growth-stimulating factor/stromal cell-derived factor 1 receptor, a murine homolog of the human immunodeficiency virus 1 entry coreceptor fusin.
Proc Natl Acad Sci USA
382:14726, 1996
18.
Nagasawa T, Kikutani H, Kishimoto T:
Molecular cloning and structure of a pre-B-cell growth-stimulating factor.
Proc Natl Acad Sci USA
91:2305, 1994[Abstract/Free Full Text]
19.
Ghia P, ten Boekel E, Rolink AG, Melchers F:
B-cell development: A comparison between mouse and man.
Immunol Today
19:480, 1998[Medline]
[Order article via Infotrieve]
20.
Lassoued K, Nunez CA, Billips L, Kubagawa H, Monteiro RC, LeBien TW, Cooper MD:
Expression of surrogate light chain receptors is restricted to a late stage in pre-B cell differentiation.
Cell
73:73, 1993[Medline]
[Order article via Infotrieve]
21.
Findley HW, Cooper MD, Kim TH, Alvarado C, Ragab AH:
Two new acute lymphoblastic leukemia cell lines with early B-cell phenotypes.
Blood
60:1305, 1982[Abstract/Free Full Text]
22.
Reed JC, Tsujimoto Y, Epstein SF, Cuddy M, Slabiak T, Nowell PC, Croce CM:
Regulation of bcl-2 gene expression in lymphoid cell lines containing normal:18 or t(14;18) chromosomes.
Oncogene Res
4:271, 1989[Medline]
[Order article via Infotrieve]
23.
Wormann B, Anderson JM, Liberty JA, Gajl-Peczalska K, Brunning R, Silberman TL, Arthur DC, LeBien TW:
Establishment of a leukemic cell model for studying human pre-B to B cell differentiation.
J Immunol
142:110, 1989[Abstract]
24.
Korsmeyer SJ, Arnold AA, Bakashi A, Ravetech JV, Siebenlist U, Hieter PA, Sharrow SO, LeBien TW, Kersey JH, Poplack DG, Leder P, Waldmann TA:
Immunoglobulin gene rearrangement and cell surface antigen expression in acute lymphocytic leukemias of T cell and B cell precursor origins.
J Clin Invest
71:301, 1983
25.
Siminovitch KA, Jensen JP, Epstein AL, Korsmeyer SJ:
Immunoglobulin gene rearrangements and expression in diffuse histiocytic lymphomas reveal cellular lineage, molecular defects, and sites of chromosomal translocation.
Blood
67:391, 1986[Abstract/Free Full Text]
26.
Silberstein LE, Jefferies LC, Goldman J, Friedman D, Moore JS, Nowell PC, Roelcke D, Pruzanski W, Roudier J, Silverman GJ:
Variable region gene analysis of pathologic human autoantibodies to the related i and I red blood cell antigenes.
Blood
78:2372, 1991[Abstract/Free Full Text]
27.
Ratajczak J, Zhang Q, Wojczyk S, Pertusini E, Wasik M, Ratajczak MZ:
The role of insulin, and insulin like growth factor-I in regulating human erythropoiesis. Studies in vitro under serum free conditions Comparison to other cytokines and growth factors.
Leukemia
12:371, 1998[Medline]
[Order article via Infotrieve]
28.
Gutkind JS:
The pathways connecting G protein-coupled receptors to the nucleus through divergent mitogen-activated protein kinase cascades.
J Biol Chem
273:1839, 1998[Free Full Text]
29.
Gether U, Kobilka BK:
G protein-coupled receptors. II. Mechanism of agonist activation.
J Biol Chem
273:17979, 1998[Free Full Text]
30.
Ji TH, Grossmann M, Ji I:
G protein-coupled receptors. I. Diversity of receptor-ligand interactions.
J Biol Chem
273:17299, 1998[Free Full Text]
31.
Sawada S, Gowrishankar K, Kitamura R, Suzuki M, Suzuki G, Tahara S, Koito A:
Disturbed CD4+ T cell homeostasis and in vitro HIV-1 susceptibility in transgenic mice expressing T cell line-tropic HIV-1 receptors.
J Exp Med
187:1439, 1998[Abstract/Free Full Text]
32.
Verani A, Pesenti E, Polo S, Tresoldi E, Scarlatti G, Lusso P, Siccardi AG, Vercelli D:
CXCR4 is a functional coreceptor for infection of human macrophages by CXCR4-dependent primary HIV-1 isolates.
J Immunol
161:2084, 1998[Abstract/Free Full Text]
33.
Oberlin E, Amara A, Bachelerie F, Bessia C, Virelizier JL, Arenzana-Seisdedos F, Schwartz O, Heard JM, Clark-Lewis I, Legler DF, Loetscher M, Baggiolini M, Moser B:
The CXC chemokine SDF-1 is the ligand for LESTR/fusin and prevents infection by T-cell-line-adapted HIV-1.
Nature
382:833, 1996[Medline]
[Order article via Infotrieve]
34.
Zaitseva MB, Lee S, Rabin RL, Tiffany HL, Farber JM, Peden KW, Murphy PM, Golding H:
CXCR4 and CCR5 on human thymocytes: Biological function and role in HIV-1 infection.
J Immunol
161:3103, 1998[Abstract/Free Full Text]
35.
Lee B, Sharron M, Montaner LJ, Weissman D, Doms RW:
Quantification of CD4, CCR5, and CXCR4 levels on lymphocyte subsets, dendritic cells, and differentially conditioned monocyte-derived macrophages.
Proc Natl Acad Sci USA
96:5215, 1999[Abstract/Free Full Text]
36.
Bleul CC, Wu L, Hoxie JA, Springer TA, Mackay CR:
The HIV coreceptors CXCR4 and CCR5 are differentially expressed and regulated on human T lymphocytes.
Proc Natl Acad Sci USA
94:1925, 1997[Abstract/Free Full Text]
37.
Sallusto F, Lenig D, Mackay CR, Lanzavecchia A:
Flexible programs of chemokine receptor expression on human polarized T helper 1 and 2 lymphocytes.
J Exp Med
187:875, 1998[Abstract/Free Full Text]
38.
Lapham CK, Zaitseva MB, Lee S, Romanstseva T, Golding H:
Fusion of monocytes and macrophages with HIV-1 correlates with biochemical properties of CXCR4 and CCR5.
Nat Med
5:303, 1999[Medline]
[Order article via Infotrieve]
39.
Doranz BJ, Orsini MJ, Turner JD, Hoffman TL, Berson JF, Hoxie JA, Peiper SC, Brass LF, Doms RW:
Identification of CXCR4 domains that support coreceptor and chemokine receptor functions.
J Virol
73:2752, 1999[Abstract/Free Full Text]
40.
Forster R, Kremmer E, Schubel A, Breitfeld D, Kleinschmidt A, Nerl C, Bernhardt G, Lipp M:
Intracellular and surface expression of the HIV-1 coreceptor CXCR4/fusin on various leukocyte subsets: Rapid internalization and recycling upon activation.
J Immunol
160:1522, 1998[Abstract/Free Full Text]
41.
Amara A, Gall SL, Schwarzt O, Salamero J, Montres M, Loetscher P, Baggiolini M, Virelizier JL, Arenzana-Seisdedos F:
HIV coreceptor downregulation as antiviral principle: SDF-1alpha dependent internalization of the chemokine receptor CXCR-4 contributes to inhibition of HIV replication.
J Exp Med
186:139, 1997[Abstract/Free Full Text]
42.
Zimmermann N, Conkright JJ, Rothenberg ME:
CC chemokine receptor-3 undergoes prolonged ligand-induced internalization.
J Biol Chem
274:12611, 1999[Abstract/Free Full Text]
43.
Haribabu B, Richardson RM, Fisher I, Sozzani S, Peiper SC, Horuk R, Ali H, Snyderman R:
Regulation of human chemokine receptors CXCR4. Role of phosphorylation in desensitization and internalization.
J Biol Chem
272:28726, 1997[Abstract/Free Full Text]
44.
Farzan M, Mirzabekov T, Kolchinksy P, Wyatt R, Cayabyab M, Gerard NP, Gerard C, Sodroski J, Choe H:
Tyrosine sulfation of the amino terminus of CCR5 facilitates HIV-1 entry.
Cell
96:667, 1999[Medline]
[Order article via Infotrieve]
45.
Kehrl JH:
Heterotrimeric G protein signaling: Roles of immune function and fine-tuning by RGS proteins.
Immunity
8:1, 1998[Medline]
[Order article via Infotrieve]
46.
Bowman EP, Campbell JJ, Druey KM, Scheschonka A, Kehrl JH, Butcher EC:
Regulation of chemotactic and proadhesive responses to chemoattractant receptors by RGS (regulator of G-protein signaling) family members.
J Biol Chem
273:28040, 1998[Abstract/Free Full Text]
47.
Dohlman HG, Thorner J:
RGS proteins and signaling by heterotrimeric G proteins.
J Biol Chem
272:3871, 1997[Free Full Text]
48.
Lefkowitz RJ:
G protein-coupled receptors.
J Biol Chem
273:18677, 1998[Free Full Text]
49.
Ali H, Richardson RM, Haribabu B, Snyderman R:
Chemoattractant receptor cross-desensitization.
J Biol Chem
274:6027, 1999[Free Full Text]
50.
Richardson RM, Pridgen BC, Haribabu B, Ali H, Snyderman R:
Differential cross-regulation of the human chemokine receptors CXCR1 and CXCR2. Evidence for time-dependent signal generation.
J Biol Chem
273:23830, 1998[Abstract/Free Full Text]
51.
Rabin RL, Park MK, Liao F, Swafford R, Stephany D, Farber JM:
Chemokine receptor responses on T cells are achived through regulation of both receptor expression and signaling.
J Immunol
162:3840, 1999[Abstract/Free Full Text]
52.
Jacobsen K, Osmond DG:
Microenvironmental organization and stromal cell associations of B lymphocyte precursor cells in mouse bone marrow.
Eur J Immunol
20:2395, 1990[Medline]
[Order article via Infotrieve]
53.
Osmond DG, Kim N, Manoukian R, Phillips RA, Rico-Vargas SA, Jacobsen K:
Dynamics and localization of early B-lymphocyte precursor cells (pro-B cells) in the bone marrow of scid mice.
Blood
79:1695, 1992[Abstract/Free Full Text]
54.
Campbell JJ, Hedrick J, Zlotnik A, Siani MA, Thompson DA, Butcher EC:
Chemokines and the arrest of lymphocytes rolling under flow conditions.
Science
279:381, 1998[Abstract/Free Full Text]
55.
Oostendorp RA, Dormer P:
VLA-4-mediated interactions between normal human hematopoietic progenitors and stromal cells.
Leuk Lymphoma
24:423, 1997[Medline]
[Order article via Infotrieve]
56.
Miyake K, Medina KL, Hayashi S, Ono S, Hamaoka T, Kincade PW:
Monoclonal antibodies to Pgp-1/CD44 block lympho-hemopoiesis in long-term bone marrow cultures.
J Exp Med
171:477, 1990[Abstract/Free Full Text]
57.
Miyake K, Weissman IL, Greenberger JS, Kincade PW:
Evidence for a role of the integrin VLA-4 in lympho-hemopoiesis.
J Exp Med
173:599, 1991[Abstract/Free Full Text]
58.
Gordon MY, Clarke D, Atkinson J, Greaves MF:
Hemopoietic progenitor cell binding to the stromal microenvironment in vitro.
Exp Hematol
18:837, 1990[Medline]
[Order article via Infotrieve]
59.
Kinashi T, Springer TA:
Adhesion molecules in hematopoietic cells.
Blood Cells
20:25, 1994[Medline]
[Order article via Infotrieve]
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|
|
T. Papayannopoulou, G. V. Priestley, B. Nakamoto, V. Zafiropoulos, and L. M. Scott
Molecular pathways in bone marrow homing: dominant role of {alpha}4{beta}1 over {beta}2-integrins and selectins
Blood,
October 15, 2001;
98(8):
2403 - 2411.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
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|
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F. Baribaud, T. G. Edwards, M. Sharron, A. Brelot, N. Heveker, K. Price, F. Mortari, M. Alizon, M. Tsang, and R. W. Doms
Antigenically Distinct Conformations of CXCR4
J. Virol.,
October 1, 2001;
75(19):
8957 - 8967.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
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S. K. Gupta, K. Pillarisetti, and N. Aiyar
CXCR4 undergoes complex lineage and inducing agent-dependent dissociation of expression and functional responsiveness to SDF-1{alpha} during myeloid differentiation
J. Leukoc. Biol.,
September 1, 2001;
70(3):
431 - 438.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
D. C. Hargreaves, P. L. Hyman, T. T. Lu, V. N. Ngo, A. Bidgol, G. Suzuki, Y.-R. Zou, D. R. Littman, and J. G. Cyster
A Coordinated Change in Chemokine Responsiveness Guides Plasma Cell Movements
J. Exp. Med.,
July 2, 2001;
194(1):
45 - 56.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
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H. Shen, T. Cheng, I. Olszak, E. Garcia-Zepeda, Z. Lu, S. Herrmann, R. Fallon, A. D. Luster, and D. T. Scadden
CXCR-4 Desensitization Is Associated with Tissue Localization of Hemopoietic Progenitor Cells
J. Immunol.,
April 15, 2001;
166(8):
5027 - 5033.
[Abstract]
[Full Text]
[PDF]
|
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|
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C. Voermans, M. L. K. Kooi, S. Rodenhuis, H. van der Lelie, C. E. van der Schoot, and W. R. Gerritsen
In vitro migratory capacity of CD34+ cells is related to hematopoietic recovery after autologous stem cell transplantation
Blood,
February 1, 2001;
97(3):
799 - 804.
[Abstract]
[Full Text]
[PDF]
|
|
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|
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F. Sanz-Rodriguez, A. Hidalgo, and J. Teixido
Chemokine stromal cell-derived factor-1{alpha} modulates VLA-4 integrin-mediated multiple myeloma cell adhesion to CS-1/fibronectin and VCAM-1
Blood,
January 15, 2001;
97(2):
346 - 351.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
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M. Rosu-Myles, L. Gallacher, B. Murdoch, D. A. Hess, M. Keeney, D. Kelvin, L. Dale, S. S. G. Ferguson, D. Wu, F. Fellows, et al.
The human hematopoietic stem cell compartment is heterogeneous for CXCR4 expression
PNAS,
December 19, 2000;
97(26):
14626 - 14631.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. D. Buckley, N. Amft, P. F. Bradfield, D. Pilling, E. Ross, F. Arenzana-Seisdedos, A. Amara, S. J. Curnow, J. M. Lord, D. Scheel-Toellner, et al.
Persistent Induction of the Chemokine Receptor CXCR4 by TGF-{beta}1 on Synovial T Cells Contributes to Their Accumulation Within the Rheumatoid Synovium
J. Immunol.,
September 15, 2000;
165(6):
3423 - 3429.
[Abstract]
[Full Text]
[PDF]
|
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TOP
ABSTRACT
INTRODUCTION