|
|
 Previous Article | Table of Contents | Next Article 
Blood, Vol. 91 No. 1 (January 1), 1998:
pp. 154-164
A Novel P-Selectin Glycoprotein Ligand-1 Monoclonal Antibody
Recognizes an Epitope Within the Tyrosine Sulfate Motif of Human PSGL-1
and Blocks Recognition of Both P- and L-Selectin
By
Karen R. Snapp,
Han Ding,
Kristin Atkins,
Roger Warnke,
Francis W. Luscinskas, and
Geoffrey S. Kansas
From the Department of Microbiology/Immunology, Northwestern
University Medical School, Chicago, IL; Vascular Research Division, the
Department of Pathology, Brigham and Women's Hospital and Harvard
Medical School, Boston, MA; and the Department of Pathology, Stanford
University Medical School, Palo Alto, CA.
 |
ABSTRACT |
Interactions between P-selectin and P-selectin glycoprotein ligand-1
(PSGL-1) mediate the earliest "rolling" of leukocytes on the
lumenal surface of endothelial cells at sites of inflammation.
Previously, PSGL-1 has been shown to be the primary mediator of
interactions between neutrophils and P-selectin, but studies on the
ability of PSGL-1 to mediate interactions between P-selectin and other
subsets of leukocytes have yielded variable and conflicting results. A
novel IgG monoclonal antibody (MoAb) to human PSGL-1 was generated, and
the specificity of this MoAb was confirmed by both flow cytometric
analysis and Western blotting of cells transfected with human PSGL-1.
This newly developed MoAb, KPL1, inhibited interactions between
P-selectin expressing COS cells and either HL60 cells, neutrophils, or
lymphocytes. Furthermore, KPL1 completely inhibited interactions
between P-selectin and either purified CD4 T cells or neutrophils in a
flow assay under physiological conditions, but had no effect on
interactions of T cells or neutrophils with E-selectin. In addition,
KPL1 blocked interactions between lymphoid cells transfected with
L-selectin and COS cells expressing PSGL-1. The KPL1 epitope was mapped
to a site within a consensus tyrosine sulfation motif of PSGL-1,
previously shown to be essential for interaction with P-selectin and
now shown to be essential for interaction with L-selectin, and to be
distinct from the epitope identified by the PL1 function blocking
anti-PSGL-1 MoAb. Two-color flow cytometry of normal leukocytes showed
that while natural killer (NK) cells (CD16+),
monocytes, CD4 and CD8 T cells, and  / and  / T cells were
uniformly positive for PSGL-1, B cells expressed low levels of the KPL1
epitope. This low level of KPL1 staining was also observed
immunohistologically in germinal centers, which had no detectable KPL1
staining, whereas T-cell areas (interfollicular region) were positive
for KPL1. Interestingly, plasma cells in situ and
interleukin-6-dependent myeloma cell lines were KPL1+.
Thus, PSGL-1 is expressed on essentially all blood neutrophils, NK
cells, B cells, T cells, and monocytes. Variation in tyrosine sulfation
during B-cell differentiation may affect the ability of B cells to
interact with P- and L-selectin.
| |
INTRODUCTION |
THE SELECTIN FAMILY of adhesion molecules
mediates the initial rolling of leukocytes on lumenal surfaces of
vascular endothelium.1 L-selectin is constitutively
expressed on all blood neutrophils, monocytes, the majority of T and B
cells, eosinophils, and most bone marrow cells. E-selectin in expressed
on endothelium in response to inflammatory mediators such as
interleukin-1 (IL-1), tumor necrosis factor (TNF-), or bacterial
lipopolysaccharide (LPS). P-selectin is stored in granules of
platelets and in Weibel-Palade bodies of endothelial cells and is
rapidly transported to the cell surface after stimulation with
thrombin, histamine, or phorbol esters. All three selectins are
structurally similar, with their C-terminal lectin domain playing the
major role in recognition of and adhesion to their sialylated,
fucosylated, and/or sulfated carbohydrate ligands.
A glycoprotein ligand for P-selectin (PSGL-1) has been identified and
cloned.2,3 PSGL-1 is a mucinlike disulfide-linked homodimer
consisting of two identical 120-kD glycoprotein chains. The sequence of
human PSGL-1 contains a cleavage site for paired basic amino acid
converting enzymes (PACE).3 Three potential tyrosine
sulfation sites are located in a consensus sequence just downstream of
the PACE cleavage site, followed by 15 decamer repeats high in proline,
serine, and threonine. The extracellular portion of the molecule
contains three potential N-linked glycosylation sites. The remaining
C-terminal sequence consists of a single transmembrane spanning domain
followed by a 69-residue cytoplasmic tail. PSGL-1 has numerous
sialylated fucosylated O-linked oligosaccharides branches, many of
which terminate in the sialyl Lewis x (sLex) epitope.4-6 In
addition to fucose and sialic acid, interactions between PSGL-1 and
P-selectin require at least one tyrosine sulfate located in the amino
terminal consensus sequence.7-9
PSGL-1 is expressed by essentially all blood leukocytes including
neutrophils, monocytes, and lymphocytes, and has been shown to mediate
the rolling of human neutrophils on P-selectin.10 In
addition, PSGL-1 can serve as a ligand for L-selectin to mediate
neutrophil-neutrophil interactions.11,12 However, data
describing the function and expression of PSGL-1 on peripheral T and B
cells are conflicting.13-17 Although some of these
disparate observations may be related to expression of nonfunctional
PSGL-1 by the majority of T cells, some of these discrepancies may be
related to the properties of different anti-PSGL-1 reagents or
differences in posttranslational modifications characteristic of
different cell lineages.
To approach these questions, a new monoclonal antibody (MoAb) to human
PSGL-1, KPL1, was generated. This MoAb inhibited interactions between
PSGL-1 and P-selectin and between PSGL-1 and L-selectin, but had no
effect on leukocyte binding to E-selectin. The KPL1 epitope was mapped
to the tyrosine sulfation consensus motif of PSGL-1. Using two-color
flow cytometry, all T cells (CD4+, CD8+,
/ and /), monocytes, and natural killer (NK)
(CD16+) cells were shown to express high levels of
PSGL-1, whereas B cells expressed lower levels of the KPL1 epitope
compared with other leukocyte subsets.
| |
MATERIALS AND METHODS |
Production of MoAbs to PSGL-1.
A cDNA for human PSGL-110 (kindly provided by Henri
Lichenstein, Amgen Inc, Thousand Oaks, CA) was subcloned into a
modified SR vector18 containing a neoR
cassette, and used to transfect the murine 300.19 pre B-cell line by
electroporation. Transfectants were selected in medium containing 2
mg/mL G418 (geneticin; GIBCO-BRL, Grand Island, NY). Transfected cells
expressing PSGL-1 were initially identified by flow cytometry with PL2,
an MoAb which recognizes human PSGL-1 (kindly provided by Kevin Moore,
Department of Medicine, University of Oklahoma Health Sciences Center),
and subcloned by limiting dilution. Balb/c mice were tolerized with
untransfected 300.19 cells followed 4 hours later by an intraperitoneal
(IP) injection of 5 µg cyclophosphamide (Neosar; Pharmacia Inc,
Columbus, OH). Starting 7 days later, mice were repeatedly immunized IP
with 300.19/PSGL-1 cells. Three days after receiving an intravenous
boost, a standard cell fusion procedure was performed, followed by
selection in hypoxanthine, aminopterin, thymidine (HAT)-containing
medium. Culture supernatants were screened via flow cytometry for
staining of 300.19/PSGL-1 transfectants but not untransfected 300.19
cells, using goat anti-mouse IgG-FITC (Biosource, Camarillo, CA) as a
second stage. Hybridomas were subcloned twice at 0.5 cells/well. After
the second round of subcloning, one clone (KPL1) was rescreened by flow
cytometry on 300.19 cells, 300.19/PSGL-1 transfectants, HL60 cells, and
freshly isolated human neutrophils and lymphocytes. Ascites was
generated from the KPL1 clone, and antibodies were purified using the
Affi-Gel Protein A MAPS II kit (Bio-Rad Laboratories, Hercules, CA).
Conjugation of KPL1 to FITC (fluorescein isothiocyanate; Sigma Chemical
Co, St Louis, MO) and biotin (N-hydroxysuccinimido-biotin; Sigma
Chemical Co) were carried out by standard procedures.
Flow cytometry.
For one-color analysis, 0.5 × 106 cells were incubated in
100 µL of phosphate-buffered saline (PBS)/1% fetal calf serum
(FCS)/NaN3 containing pretitered amounts of the indicated
PSGL-1 MoAb, washed, and incubated in goat anti-mouse IgG-FITC, diluted
1:100. For two-color flow cytometry, 0.5 × 106 cells were
incubated in 100 µL of PBS/1% FCS/NaN3 containing
anti-PSGL-1 MoAb conjugated to FITC or biotin plus a leukocyte subset
specific antibody conjugated to either biotin or FITC. After washing,
cells were stained with Streptavidin-PE (Phycoerythrin;
Fischer Scientific Co, Pittsburgh, PA). Antileukocyte
subset antibodies included MoAb directed against CD4, CD8, CD3, CD16,
CD19, and CD14. MoAbs against T-cell receptor (TCR)-/ and
TCR-/ (WT31) were obtained from Becton Dickinson (San Jose, CA).
Samples were analyzed on a Becton Dickinson FACScan and data were
collected on a total of 10,000 to 20,000 light scatter gated events.
Data were analyzed with CELLQuest software
(Becton Dickinson) and both fluorescence histograms and two-color
contour plots were displayed on a 4-decade logarithmic scale.
Western blotting.
HL60, 300.19, 300.19/PSGL-1 cells, or COS cells transiently transfected
with either human PSGL-1 cDNA or a control plasmid were washed twice in
RPMI. HL60, 300.19, and 300.19/PSGL-1 cells were resuspended at 2 ×
107 cells/mL in lysis buffer (1% Triton-X-100 [Sigma; St
Louis, MO]; 150 mmol/L NaCl; 10 mmol/L Tris-HCl, pH 7.6;
1 mmol/L CaCl2; 1 mmol/L MgCl2; 1 mmol/L
aprotinin; 1 mmol/L phenylmethylsulfonyl fluoride (PMSF); 1 mmol/L
leupeptin; and 1 mmol/L pepstatin A) and incubated on ice for 30
minutes. COS cells were resuspended at 2 × 106 cells/mL
in lysis buffer containing 2% Triton-X-100 on ice for 30 minutes.
Samples were clarified by centrifugation at 14,000g for 30
minutes at 4°C, and supernatants were transferred to fresh tubes. For
certain experiments, aliquots of lysates were treated overnight at
37°C with 1 U of Aerobacter aerogenes arylsulfatase (Sigma).
Samples were boiled for 5 minutes in sodium dodecyl
sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) sample buffer
with or without 6.5% 2-mercaptoethanol, electrophoresed on a 6%
polyacrylamide gel, and transferred to nitrocellulose. The
nitrocellulose was blocked with 2% gelatin (Bio-Rad) in Tris-buffered
saline-Tween (TBS-T; 20 mmol/L Tris HCl, pH 7.6; 137 mmol/L NaCl; and
0.1% Tween-20) for 2 hours at room temperature. Blots were probed with
KPL1 or PL2 in TBS-T plus 2% gelatin for 30 minutes, washed five times
in TBS-T, and then incubated with goat anti-mouse IgG conjugated to
horseradish peroxidase (Biosource) for 30 minutes. After five washes in
TBS-T, blots were visualized by chemiluminescence generated after the
addition of enhanced chemiluminescence (Amersham,
Arlington Heights, IL) and exposed to Hyperfilm (Amersham).
Generation and expression of full-length FFFE/PSGL-1 cDNA.
The plasmid pED.FFFE.148.Fc9 was kindly provided by Gray
Shaw (Genetics Institute, Cambridge, MA). This chimeric protein
contains the extracellular 148 amino acids of PSGL-1 fused to the heavy
chain CH2-CH3 region of IgG, and replaces the
three tyrosines at positions 5, 7, and 10 of PSGL-1 with phenylalanine
and the aspartic acid at position 11 with glutamic acid.9 A
423-bp Xma I/Stu I insert containing the FFFE mutation
was ligated into the full-length PSGL-1 cDNA in pBluescript, and the
full-length FFFE/PSGL-1 was inserted into a modified SR vector
containing a neoR cassette. Approximately 15 µg of the
FFFE or wild-type PSGL-1 were used to transiently transfect 1.5 ×
106 300.19 cells by electroporation.
Neutrophil, peripheral blood mononuclear cell (PBMC), and
CD4+ T-cell purification.
Human neutrophils were isolated from heparinized blood by dextran
sedimentation, Ficoll density gradient centrifugation (Histopaque-1077;
Sigma), and hypotonic lysis of the neutrophil-rich pellet. PBMCs were
isolated from heparinized whole blood by dextran sedimentation followed
by Ficoll density gradient centrifugation. CD4 T cells were purified
from single donor human platelet pheresis residues by sequential
density gradient centrifugation and elutriation followed by culture
overnight and positive selection on Dynabeads (model M-450 CD4) and
Dynal DETACHaBEAD (Dynal, Great Neck, NY) as described.16
Low shear force COS cell adhesion assay.
This assay was performed as previously described.19-22
Briefly, COS cells were transfected with either P-selectin or
E-selectin cDNA by the DEAE-dextran method in 100-mm tissue culture
grade petri dishes. For binding of 300.19/L-selectin (300.19/L) or
300.19/P-selectin (300.19/P) cells,19,23 COS cells were
cotransfected with plasmids encoding PSGL-1 or FFFE/PSGL-1, FucT-VII,
and C2GnT. The following day, COS cells were replated on 35-mm dishes
(assay plates), and allowed to readhere overnight. The next day, HL60
cells, freshly purified human neutrophils, or PBMC were washed twice in
RPMI 1640 and resuspended at 2 × 106 cells in a total
volume of 100 µL containing saturating amounts of either KPL1 ascites
or control antibody and placed on ice for 15 minutes. Alternatively,
for 300.19/L or 300.19/P cells binding to transfected COS cells, the
COS cells were incubated with saturating amounts of KPL1 or control
ascites. After cells had been washed and resuspended in 0.6 mL of RPMI,
each petri dish was washed three times with unsupplemented RPMI 1640,
followed by the addition of appropriate cells, and incubated on a
constantly rocking platform for 15 minutes at 4°C. The plates were
washed five times with RPMI 1640 followed by fixation with cold 0.37%
formaldehyde/RPMI 1640. Mean number of cells bound per COS cell was
determined by counting the number of cells bound/COS cell on ~125 COS
cells in multiple 40× fields using a standard inverted light
microscope.
Parallel plate flow chamber adhesion assay.
The flow chamber apparatus used in these studies has been described
previously.15,24,25 Chinese hamster ovary (CHO)/P-selectin
or CHO/E-selectin monolayers were grown to confluence on glass
coverslips and inserted into the flow chamber. A defined flow level of
1.8 dynes/cm2 was obtained by drawing media containing the
desired cell population through the chamber using a syringe pump.
Purified CD4+ T cells or neutrophils, 1 ×
106, were assayed in each experiment. The flow chamber was
mounted on an inverted microscope (Nikon Diaphot; Melville, NY) and
each 5-minute perfusion period was recorded on videotape by a video
camera and video cassette recorder. Leukocyte adhesion was
determined as previously described.15,24,25 In some
experiments, monolayers of CHO/P cells were preincubated with HDPG2/3,
a blocking anti-P-selectin antibody3 at 10 µg/mL for 30
minutes at 37°C. Monolayers of CHO/E were incubated with HEL3/2, a
blocking anti-E-selectin antibody.26 W6/32 was used as an
isotype control for both monolayers. In other experiments, cells
(1 × 106) were incubated with either KPL1 ascites or
control ascites at a 1:200 dilution for 15 minutes at 4°C before use
in assays.
Immunohistology.
Immunohistology was performed using an indirect biotin-streptavidin
method.27 Incubation with primary antibody was followed by
incubation with a biotinylated goat anti-mouse second stage antibody
(Jackson Immunoresearch Laboratories, Inc, West Grove, PA) followed by
peroxidase-conjugated streptavidin (Jackson Immunoresearch
Laboratories). Diaminobenzedene was used as a substrate for the
horseradish peroxidase. Frozen sections were fixed in acetone for 10
minutes at 4°C before staining. Formalin-fixed deparaffinized
sections were stained without pretreatment or microwaved for 15 minutes
in 0.01 mol/L citrate buffer at pH 6.0 before staining.
| |
RESULTS |
Development and characterization of a new MoAb to human PSGL-1.
Balb/c mice were immunized with 300.19/PSGL-1 cells as described in
Materials and Methods. After routine immunization and fusion protocols,
hybridoma supernatants were screened by flow cytometry for negative
staining on untransfected 300.19 cells and positive staining on
300.19/PSGL-1 cells (Fig 1A). One MoAb,
KPL1, was identified. The KPL1 MoAb stained the 300.19/PSGL-1 cells at
high levels but did not stain untransfected 300.19 cells (Fig 1A). To
further confirm the specificity of KPL1, Western blot analysis of cells
known to express PSGL-1 or transfected with PSGL-1 cDNA was performed.
PSGL-1 is a homodimer of ~240 to 250 kD under
nonreducing conditions and ~120 kD under reducing
conditions.2,10 PSGL-1 is difficult to completely reduce,
allowing visualization of both reduced and nonreduced forms of PSGL-1.
KPL1 reacted with bands of ~120 and ~240 kD in lysates from HL60
cells (a myeloid cell line known to express PSGL-1), 300.19/PSGL-1
transfectants, or COS/PSGL-1 transfectants, but not in lysates from
untransfected 300.19 cells or COS cells transfected with a control
plasmid (Fig 1B). The slight differences in molecular weight between
PSGL-1 expressed by HL60 and the two transfectants were most likely
caused by differences in glycosylation in these different cell types.
Finally, KPL1 stained HL60 cells, freshly purified neutrophils, and
peripheral blood lymphocytes by flow cytometry (see
below).
View larger version (19K):
[in this window]
[in a new window]
View larger version (58K):
[in this window]
[in a new window]
| Fig 1.
Specificity of KPL1 for PSGL-1 by flow cytometry and
Western blotting. (A) Untransfected 300.19 cells or 300.19 cells
transfected with human PSGL-1 cDNA were stained with a negative control
(top) or KPL1 (bottom) as described in Materials and Methods. KPL1 does
not stain untransfected 300.19 cells, whereas it stains 300.19/PSGL-1
transfectants. (B) Western blotting of whole cell lysates made from
HL60 cells (lane 1), 300.19 cells (lane 2), 300.19 cells transfected
with human PSGL-1 cDNA (lane 3), COS cells transfected with a control
plasmid (lane 4), or COS cells transfected with human PSGL-1 cDNA (lane
5) with the KPL1 antibody. Bands of the appropriate molecular weight
(~120 and ~240 KD) were seen only in lysates from cells expressing
PSGL-1 endogenously (HL60 cells in lane 1) or transfected with human
PSGL-1 cDNA (lanes 3 and 5).
|
|
KPL1 recognizes a unique epitope within the tyrosine sulfation motif
of PSGL-1.
Because of apparent differences in staining results with different
anti-PSGL-1 antibodies (see Introduction and below), the epitope
defined by KPL1 was explored in some detail. Neuraminidase treatment of
HL60 cells did not affect expression of the KPL1 epitope, but removed
all surface sLex carbohydrates as measured by lack of HECA452 staining
(data not shown). In addition, KPL1 stained cells regardless of the
presence (neutrophils, HL60 cells) or absence (300.19/PSGL-1 cells) of
either FucT-VII or C2GnT (Fig 1 and data not shown; by
RT-PCR,21,22,28 300.19 cells do not detectably express mRNA
for either FucT-VII or C2GnT). Thus, the KPL1 epitope appears to be
independent of carbohydrate modifications, including terminal sialic
acid residues, branched O-linked glycans, or fucosylation.
We have shown elsewhere that KPL1 fails to recognize a PSGL-1 mutant in
which amino acid residues 5-11 (YEYLDYD), comprising a consensus
tyrosine sulfation motif,29 have been
deleted.30 To more precisely evaluate the potential
contribution of residues comprising the tyrosine sulfate motif to the
KPL-1 epitope, we analyzed binding of KPL1 to a PSGL-1 mutant in which
all three tyrosines were simultaneously replaced by phenylalanines
(FFFE9). 300.19 cells were transiently transfected with
cDNA encoding wild-type PSGL-1 or the FFFE mutant, and analyzed by flow
cytometry. Replacement of these three tyrosines at positions 5, 7, and
10 with phenylalanine resulted in complete loss of the KPL1 epitope
(Fig 2A). However, epitopes recognized by
two other anti-PSGL-1 antibodies were still present (Fig 2A),
confirming surface expression of the mutated form of PSGL-1. Thus, KPL1
interacts with the tyrosine sulfation motif of PSGL-1.
View larger version (19K):
[in this window]
[in a new window]
View larger version (64K):
[in this window]
[in a new window]
| Fig 2.
KPL1 recognizes the tyrosine sulfation motif of PSGL-1.
(A) 300.19 cells were transiently transfected with either full length
PSGL-1 (top) or a mutated form of PSGL-1 referred to as FFFE/PSGL-1
(bottom) in which the tyrosines at positions 5, 7, and 10 were replaced
with phenylalanine. Twenty-four hours posttransfection, the 300.19
cells were stained with either a negative control antibody, KPL1, PL2,
or PSL275. PL2 and PSL275 recognize both the full length and mutated
form of PSGL-1, whereas KPL1 was able to interact with full length
PSGL-1 but did not recognize FFFE/PSGL-1. (B) Arylsulfatase treatment
of whole cell lysates abrogates binding of KPL1. Whole cell lysates
were prepared as described in Materials and Methods and treated with 1U
of arylsulfatase, and Western blotting was performed as described for
Fig 1, using KPL1 (left) or PL2 (right).
|
|
To determine whether binding of KPL1 required sulfation of PSGL-1, cell
lysates were treated with a bacterial arylsulfatase, which cleaves
sulfates from tyrosine residues within proteins but does not digest
sulfated carbohydrates, or with buffer alone, and Western blotting was
performed with either KPL1 or PL2. As shown in Fig 2B, treatment of
cell lysates with arylsulfatase nearly abrogated recognition with KPL1,
but did not affect recognition by PL2. Therefore, binding of KPL1 to
PSGL-1 is dependent on sulfates within the PSGL-1 tyrosine sulfate
motif.
Other investigators have suggested that at least some B cells express
non-PACE-processed PSGL-1, and therefore that PSGL-1 on B cells is
expressed in a form that is not recognized by antibodies generated
against or specific for PACE-cleaved PSGL-1.14 After a
24-hour incubation with PACE secreting or control CHO
cells,14 several B-cell lines were analyzed by flow
cytometry with MoAb PSL-275, which recognizes only PACE-cleaved
PSGL-1.14 One cell line showed a significant increase in
PSL-275 staining, but these cells stained identically with KPL1 whether
they were incubated with control or CHO-PACE cells (data not shown).
Therefore, KPL1 binding is independent of PACE cleavage of PSGL-1.
KPL1 MoAb inhibits binding of normal leukocytes and HL60 cells to
P-selectin but not E-selectin.
Adhesion assays were performed to determine if KPL1 could inhibit
interactions between normal leukocytes or HL60 cells and P- or
E-selectin. In the absence of antibody, HL60 cells, PBMCs, and
neutrophils each bound well to COS cells transfected with either P- or
E-selectin, with neutrophils binding slightly better to both P- and
E-selectin compared with PBMC or HL60 cells (Fig
3). Consistent with a requirement for
tyrosine sulfation of PSGL-1 for interaction with P-selectin,
preincubation of leukocytes with KPL1 virtually completely blocked
(>95%) interactions of HL60 cells and neutrophils with P-selectin
and almost completely (>90%) inhibited interactions of PBMCs with
P-selectin (Fig 3A). In contrast, KPL1 had no effect on binding to
E-selectin (Fig 3B).
View larger version (29K):
[in this window]
[in a new window]
| Fig 3.
KPL1 inhibits the binding of HL60 cells, neutrophils, and
PBMCs to P-selectin but not E-selectin expressing COS cells in a low
shear adhesion assay. (A) Adhesion of HL60 cells ( ), neutrophils
( ), and PBMC ( ) to COS cells transiently transfected with
P-selectin was performed as described in Materials and Methods. All
three cell types adhered well to P-selectin. KPL1 almost completely
blocked these interactions, whereas a control antibody had no effect on
adhesion. (B) HL60 cells (), neutrophils () and PBMC () bound
well to E-selectin, but neither KPL1 nor a control antibody inhibited
adhesion to E-selectin expressing COS cells. Values are mean ±SD; one
of at least five experiments.
|
|
KPL1 MoAb inhibits rolling of freshly isolated human neutrophils and
CD4 T cells on P-selectin but not E-selectin.
The results from the low shear stress COS cell adhesion assay were
extended to analysis of rolling under defined shear stress (Figs 4 and
5). Purified human neutrophils
or purified CD4
T cells were incubated with media containing either KPL1 or control
antibody, or the monolayers were incubated with either HDPG2/3, an
anti-P-selectin antibody (CHO/P monolayer); HEL3/2, an
anti-E-selectin antibody (CHO/E monolayer); or an isotype control
antibody (W6/32). Preincubation of the monolayers with MoAb to the
appropriate selectin completed blocked rolling of neutrophils.
Preincubation of neutrophils with KPL1 also completely inhibited
rolling of neutrophils on P-selectin (Fig 4A), but had no effect on
interactions with E-selectin (Fig 4B). Similarly, preincubation of
transfected monolayers with the appropriate anti-selectin antibody
completely inhibited rolling of CD4 T cells, and, as observed with
neutrophils, KPL1 completely inhibited rolling of CD4 T cells on CHO/P
but had no effect on CHO/E (Fig 5A and B).
View larger version (14K):
[in this window]
[in a new window]
| Fig 4.
KPL1 inhibits rolling of neutrophils on CHO cells
transfected with P-selectin but not E-selectin under defined shear
stress. (A) The rolling of neutrophils on CHO cells expressing
P-selectin in the presence of W6/32 (isotype control), HDPG2/3 (a
blocking anti-P-selectin antibody), KPL1, or a control antibody for
KPL1 was performed as described in Materials and Methods. Both KPL1 and
HDPG2/3 completely blocked rolling of neutrophils on P-selectin. No
effect on rolling was observed when the control antibodies were used.
(B) The ability of freshly isolated neutrophils to roll on E-selectin
transfected COS cells in the presence of W6/32, HEL3/2 (a blocking
anti-E-selectin antibody), KPL1, or control antibody was performed as
described in (A). Only the anti-E-selectin antibody inhibited rolling
of neutrophils on E-selectin. No inhibition of rolling was observed
when neutrophils were pre-incubated with KPL1 or control antibody, or
when monolayers were exposed to W6/32. Values are mean ±SD; one of at
least three experiments.
|
|
View larger version (14K):
[in this window]
[in a new window]
| Fig 5.
KPL1 inhibits rolling of all CD4+ T cells
on CHO cells transfected with P-selectin but not E-selectin under
defined shear stress. Freshly isolated CD4 T cells were assayed for
their ability to roll on CHO cells transfected with either P- or
E-selectin. (A) Rolling was assessed as described in Materials and
Methods after exposure to either W6/32 (isotype control antibody),
HDPG2/3 (a blocking anti-P-selectin antibody), KPL1, or a control
antibody. Rolling of freshly isolated CD4 T cells was completely
inhibited by pre-incubation of CD4 cells with KPL1 or monolayers with
HDPG2/3. (B) Rolling of CD4 T cells on CHO cells transfected with
E-selectin after exposure to either W6/32, HEL3/2 (a blocking
anti-E-selectin antibody), KPL1, or a control antibody for KPL1 was
performed as described for (A). Rolling of CD4 T cells was completely
inhibited by preincubation of the E-selectin expressing monolayer with
HEL3/2, whereas incubation of CD4 cells with either KPL1 or control
antibodies had no effect. Values are mean ±SD; one of at least three
experiments.
|
|
KPL1 inhibits interactions between L-selectin and PSGL-1.
PSGL-1 has recently been identified as a leukocyte ligand for
L-selectin, and appears to mediate neutrophil-neutrophil interactions
which may be important in amplifying an inflammatory
response.11,12 Because normal neutrophils express ligands
for L-selectin in addition to PSGL-1,11,31 we analyzed the
effect of KPL1 on L-selectin/PSGL-1 interactions using L-selectin
transfectants binding to PSGL-1 transfectants, which isolates the
molecular interaction of interest. COS cells were cotransfected with
individual plasmids containing cDNA encoding PSGL-1, FucT-VII, and
C2GnT. For the assay, these COS cells were preincubated with either
KPL1 or control antibody, washed, and incubated with 300.19 cells
stably transfected with either L-selectin (300.19/L) or P-selectin
(300.19/P). Preliminary studies (data not shown) demonstrated that
binding of either 300.19/P or 300.19/L was absolutely dependent on
expression of both PSGL-1 and FucT-VII, and that binding of 300.19/L
was enhanced by cotransfection with C2GnT cDNA. Binding of 300.19/P was
not significantly enhanced by transfection with C2GnT cDNA, presumably
because of the endogenous expression of C2GnT by COS cells.
Both 300.19/P and 300.19/L cells bound to COS cells coexpressing
PSGL-1, FucT-VII, and C2GnT (Fig 6A).
Binding of both 300.19/P and 300.19/L was completely blocked in the
presence of KPL1 (Fig 6A). In addition, no binding of either 300.19/L
or 300.19/P to the FFFE/PSGL-1 mutant was detected (Fig 6B). These data
strongly indicate that interaction between PSGL-1 and either P-selectin
or L-selectin involves an identical or overlapping region of PSGL-1,
which includes the KPL1 epitope within the tyrosine sulfation motif.
View larger version (14K):
[in this window]
[in a new window]
| Fig 6.
Interactions between PSGL-1 and L-selectin depend on the
tyrosine sulfate motif. (A) COS cells were cotransfected with cDNA
encoding PSGL-1, FucT-VII, and C2GnT. COS cells were preincubated with
either KPL1 or control antibody for 15 minutes. After extensive
washing, 300.19/P or 300.19/L cells were added to the COS cells and the
assay was performed as described in Materials and Methods. Adhesion of
both cell types was completely inhibited by preincubation of
transfected COS cells with KPL1, but not control MoAb (not shown). (B)
COS cells were cotransfected with cDNA encoding FucT-VII, C2GnT, and
either PSGL-1 or FFFE/PSGL-1. Binding of 300.19/L or 300.19/P cells to
the FFFE mutant was undetectable. Values are mean ±SD; one of three
experiments.
|
|
The KPL1 epitope is expressed at high levels on circulating T cells,
monocytes, and natural killer (NK) cells, but at low levels on B cells.
All circulating leukocyte subsets have been reported to express PSGL-1,
although different researchers have reported variable levels of
expression, or lack of expression, on specific subsets. Some of these
differences may be caused by properties of different PSGL-1 specific
antibodies. Staining of PBMCs with KPL1 in one-color flow cytometry
showed a small but distinct subpopulation expressing lower levels of
PSGL-1 (data not shown; see below), whereas most anti-PSGL-1 MoAb
exhibit a single peak on lymphocytes10,17 (data not shown).
To identify this KPL1lo subpopulation, two-color flow
cytometry was performed with a leukocyte lineage marker in one color
and KPL1 in a second color. Appropriate electronic scatter gates were
set for either monocytes or lymphocytes. All NK cells
(CD16+), monocytes (CD14+ ), and T
cells (CD3+) (Fig 7A),
including all CD4+, CD8+, /, and /
T cells (Fig 7B), were uniformly positive for PSGL-1 (Fig 7A). The
KPL1lo cells were identified as B cells by virtue of their
staining with CD19; all detectable KPL1lo cells were
CD19+, and no CD19 cells were
KPL1lo (Fig 7A). These findings were observed with B cells
analyzed from four different donors. This relatively low level of KPL1
staining was also observed on a panel of Epstein-Barr
virus-transformed B-cell lines (data not shown).
View larger version (33K):
[in this window]
[in a new window]
View larger version (37K):
[in this window]
[in a new window]
| Fig 7.
Expression of PSGL-1 on leukocyte subsets. (A) Human PBMC
were isolated by Ficoll density gradient centrifugation and stained for
two-color flow cytometry as described in Materials and Methods.
Electronic scatter gating was used to select specific subpopulations
for detailed analysis. All NK cells (CD16+), monocytes
(CD14+), and T cells (CD3+) were
positive for PSGL-1 as measured by KPL1, whereas B cells
(CD19+) expressed low levels of the KPL1 epitope. (B)
All CD4, CD8, / (WT31 positive), or / (anti-TCR-/-1
positive) cells express uniform levels PSGL-1 as measured by KPL1.
Horizontal and vertical lines delineating positive and negative
staining were set with appropriate negative control MoAb.
|
|
Expression of PSGL-1 in lymphoid tissues.
Both paraffin-embedded tissues and frozen sections (with or without
heat antigen retrieval) were prepared for immunohistology as described
in Materials and Methods. KPL1 stained numerous T cells in the T-zone
(TZ) of a human tonsil (Fig 8A), but did
not stain B cells in the mantle zone (M) or germinal center (GC) of the
secondary follicles. Macrophages in the germinal center stained but
follicular dendritic cells did not (Fig 8A). Just outside the germinal
center, intense KPL1 staining of overlying plasma cells was observed. A
high-power magnification (Fig 8B) shows this intense staining of plasma
cells which surround the germinal centers. Subepithelial plasma cells
also stained with KPL1 (data not shown), as did plasma cells around
vessels in the skin (Fig 8C); these plasma cells costained with the
plasma cell marker VS38 (Fig 8D). Thus, B cells located in germinal
centers either lack the KPL1 epitope, or express it at levels which are
not detected by these methods, whereas plasma cells in numerous sites
express high levels. Similarly, four IL-6-dependent human myeloma cell
lines expressed high levels of the KPL1 epitope (Diane Jelinek,
personal communication, March 1997). Both cortical and medullary
lymphocytes in the thymus stained with KPL1 (data not
shown). Langerhans cells, bone marrow-derived antigen
presenting cells which reside in the suprabasilar region of the
epidermis, were also positive for KPL1 (Fig 8E). Langerhans cells in
the tonsillar epithelium also stained with KPL1 (data not shown).
Langerhans cells in soft tissue also stain with both KPL1 (Fig 8F) and
CD1a (Fig 8G).
Finally, an extensive tissue survey failed to detect KPL1 staining on
stomach, colon, liver, pancreas, uterus (endometrium and myometrium),
ovary, prostate, testis, brain, lung, heart, thyroid, parathyroid, and
skeletal muscle (data not shown). However, leukocytes, including
dendritic cells and macrophages, reacted in many of the tissues, eg,
the Kupffer cells in the liver. PSGL-1 expression, as assessed by KPL1
staining, therefore appears to be limited to the hematopoietic system,
at least in humans.
| |
DISCUSSION |
We have generated a novel MoAb against PSGL-1, designated KPL1. The
specificity of the KPL1 MoAb for PSGL-1 was confirmed by both flow
cytometry and Western blotting of multiple PSGL-1-expressing
transfectants. The epitope defined by KPL1 is independent of
carbohydrate modifications, including the presence or absence of
sialylation, fucosylation, or branched O-linked structures, because
KPL1 stains cells equivalently regardless of whether the cells express
no or high levels of FucT-VII or C2GnT, and KPL1 staining was not
affected by treatment of cells with neuraminidase. In addition, this
epitope is independent of PACE processing. However, the KPL1 epitope
requires sulfation of at least one tyrosine contained within a
consensus tyrosine sulfation motif (YEYLDYD)29 at the
extreme amino terminus of PSGL-1. Deletion of these seven amino acids
eliminated binding of KPL1,30 as did simultaneous mutation
of all three tyrosines to phenylalanine (Fig 2A) or sulfatase treatment
of PSGL-1 (Fig 2B). Thus, the KPL1 epitope maps to the tyrosine
sulfation motif of PSGL-1.
Consistent with the KPL1 epitope mapping to the tyrosine sulfate motif,
and the requirement for tyrosine sulfation of PSGL-1 in recognition of
P-selectin,7-9 the KPL1 MoAb essentially completely
inhibited adhesion between P-selectin and HL60 cells, neutrophils, or
PBMCs in a low shear adhesion assay, and between P-selectin and CD4 T
cells or neutrophils in a parallel plate flow assay. In contrast, KPL1
had no effect on interactions between these different cell types and
E-selectin. Although PSGL-1 is not required for adhesion to
E-selectin,21 PSGL-1 appears capable of interacting with
both P- and E-selectin,9,32,33 with different (or multiple)
regions of PSGL-1 mediating adhesion to E-selectin.30 It is
currently not known if an identical site or closely overlapping site
within the first 19 amino acids of PSGL-1 mediates interaction with
both P- and E-selectin, or whether this or another site on PSGL-1 is
functional during normal leukocyte interactions with E-selectin.
PSGL-1 also interacts with L-selectin, an interaction likely to be of
importance in neutrophil-neutrophil interactions during
inflammation.11,12,34 Because normal neutrophils express
ligands for L-selectin in addition to PSGL-1,11,31 we
analyzed whether KPL1 could inhibit this molecular interaction in a
"pure" system, consisting of 300.19/L-selectin transfectants
binding to COS cells cotransfected with cDNA encoding PSGL-1, FucT-VII,
and C2GnT. All three of these gene products are required for binding to
P-selectin.3,10,21,26,33,35 Untransfected 300.19 cells do
not bind to COS cells cotransfected with cDNA encoding PSGL-1,
FucT-VII, and C2GnT, and binding of 300.19/L or 300.19/P cells is
completely dependent on transfection of COS cells with cDNA encoding
both PSGL-1 and FucT-VII. We found that KPL1 virtually completely
blocked interactions between L-selectin and PSGL-1 (Fig 6A). Binding of
L-selectin, like binding of KPL1 (Fig 2) or P-selectin,7-9
was also blocked by mutation of all three tyrosines in the PSGL-1
tyrosine sulfation motif to phenylalanine (Fig 6B). P-selectin and
L-selectin (and KPL1) therefore recognize identical or closely
overlapping sites on PSGL-1, and PSGL-1 does not appear to have
multiple L-selectin binding sites.
Both PSGL-1 and at least some L-selectin ligands require
sulfation,7-9,36 and for PSGL-1 this sulfation occurs
exclusively on tyrosine.8,9 In contrast, sulfation of
L-selectin HEV ligands36 occurs on specific carbohydrate
side chains of CD34, GlyCAM-1, and possibly sgp200.37-39
However, recognition of PSGL-1 by L-selectin is dependent on sulfation
of tyrosines within the PSGL-1 tyrosine sulfation motif (Fig 6). Taken
together, these data show that L-selectin recognizes an array of
distinct sulfated ligands whose sulfate moiety can be attached to
either carbohydrate or protein.
Our findings clearly establish that PSGL-1 is the principal or sole
ligand for P-selectin on T cells. Disparate results were previously
found using a polyclonal antiserum against recombinant PSGL-1 (termed
Rb 3026). Alon et al13 found that this reagent inhibited
T-cell interaction with P-selectin, whereas Vachino et al14
failed to detect inhibition. This same reagent previously was found to
inhibit monocyte adhesion to P-selectin.15 Rb 3026 also
failed to detect a subset of , T cells.40 In
contrast, we and others17 have clearly shown that all
subsets of blood T cells express PSGL-1; PSGL-1 T cells
were never detected in blood from any donor. However, that same
study17 failed to detect PSGL-1 on T cells in tissues,
whereas we observed significant staining with KPL1 in T cells areas of
tonsil (Fig 8). PSGL-1 levels do not change significantly with T-cell
activation in vitro,14,16 making it unlikely that
activation within tissues can explain differences in staining between
blood (by FACS) and tissues (by immunohistology), especially by the
same MoAb. The basis for this variation remains unknown.
B cells in peripheral blood stained at lower levels with KPL1 than
other leukocytes, but were clearly positive (Fig 7). We also found
PSGL-1 on 7/7 EBV B-cell lines (data not shown). These results contrast
with those of Vachino et al,14 who detected PSGL-1
expression on only ~30% of blood B cells. Laszik et al17
also found lower levels of PSGL-1 on B cells, but whether B cells
expressed uniformly lower levels or whether a subpopulation of B cells
do not express PSGL-1 was not specified in that study.17 In
contrast, Moore et al10 previously found uniform expression
of PSGL-1 on essentially all lymphocytes. The basis for these
discrepancies is unclear. B cells within mantle zones or germinal
centers of tonsil failed to stain detectably with KPL1. The basis for
the relatively low level of KPL1 staining on B cells is not known, but
may relate to the state of differentiation of the B cell, as both
plasma cells in situ (Fig 8) and cultured, IL-6-dependent myeloma
cells (Diane Jelinek, personal communication, March 1997) stain
brightly with KPL1. These results, in combination with the
localization of the KPL1 epitope to the tyrosine sulfation motif,
suggest that B cells may regulate sulfation of PSGL-1 in a manner
distinct from the apparently constitutive sulfation characteristic of
other types of leukocytes. These putative differences in sulfation of
PSGL-1 may affect the migratory capacity of B cells, and may explain
why B cells are rarely found at most sites of inflammation.
A recent report documented strong PSGL-1 mRNA expression in most
tissues of the mouse,41 implying that PSGL-1 is expressed
on some nonhematopoietic cells. However, extensive immunohistochemical
analysis by ourselves and others17 failed to detect PSGL-1
on any nonhematopoietic cells in any tissues examined. In addition, we
failed to detect PSGL-1 staining on cultured normal endothelium, smooth
muscle cells, fibroblasts, or keratinocytes (data not shown). The
precise cellular source of the strong mRNA signals from multiple murine
tissues is presently unknown, and the apparent species difference
between humans and mice is unexpected.
The previously described function-blocking anti-PSGL-1 MoAb
PL110 and the KPL1 MoAb described in the present report
share a number of functional characteristics, including essentially
complete inhibition of leukocyte binding to P-selectin10
(Figs 3-5) and L-selectin binding to PSGL-111 (Fig
6). However, these two MoAbs map to distinct epitopes near the amino
terminus of PSGL-1. In particular, KPL1 recognizes the tyrosine
sulfation motif comprising residues 5-11 (Fig 2), whereas PL1 sees the
nontyrosine-sulfated sequence centered around residues 13-17
(LPETE).42 Although these data do not exclude the
possibility that each MoAb sterically obscures the neighboring site as
well, the data are most consistent with the idea that these two MoAbs
are recognizing distinct components of physiological ligands for both
L- and P-selectin. Thus, KPL1 prevents recognition of the tyrosine
sulfates, whereas PL1 probably blocks recognition of sialylated and
fucosylated carbohydrates attached to threonine 16, mutation of which
inhibits recognition of P-selectin.8,9 Therefore, these
observations provide support for the idea of a "discontiguous
carbohydrate epitope"5,43 as a physiological recognition
structure for selectins.
Generation of MoAb against human cell surface glycoproteins by
immunizing with 300.19 transfectants stably expressing the molecule of
interest has previously produced large numbers of MoAbs, many of which
recognize evolutionarily conserved epitopes present of the homologues
in other mammalian species.44,45 In contrast, we produced
only a single MoAb to PSGL-1 with this approach, and this MoAb does not
recognize leukocytes from any mammalian species thus far tested,
including cat, dog, horse, cow, or sheep (Mark Jutila, personal
communication, April 1997). Similarly, the PL1 MoAb does
not see leukocytes from any other mammalian species tested (Kevin
Moore, personal communication, March 1997). In part, this may relate to
the relatively low conservation of amino acid sequence at the amino
terminus of PSGL-1: only 7 of 16 residues are identical or similar
between human and mouse PSGL-1, and the tyrosine sulfation motifs of
the two species are at dissimilar relative positions.3,41
In summary, using a novel MoAb directed against the functionally
essential tyrosine sulfation motif of human PSGL-1, we show that PSGL-1
is expressed on all circulating leukocytes, including neutrophils,
monocytes, all subsets of T cells, NK cells, and B cells, and is the
principal or sole ligand for P-selectin on at least T cells and
neutrophils. The expression and function of PSGL-1 on B cells requires
further investigation, as these cells may uniquely regulate tyrosine
sulfation of PSGL-1.
| |
FOOTNOTES |
Submitted April 7, 1997;
accepted August 29, 1997.
Supported by the American Cancer Society Grant No. CB-204 (G.S.K.) and
HL36028 (F.W.L.) and National Institutes of Health Grant No. CA34233
(R.W.). G.S.K. is an Established Investigator of the American Heart
Association.
Address reprint requests to Geoffrey S. Kansas, PhD, Department of
Microbiology-Immunology, Northwestern Medical School, 303 E Chicago
Ave, Chicago, IL 60611.
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.
| |
ACKNOWLEDGMENT |
The authors gratefully acknowledge Kevin Moore (University of Oklahoma
Health Sciences Center, Oklahoma City, OK) for supplying PL2 MoAb;
Henri Lichenstein (Amgen, Inc, Boulder, CO) for PSGL-1 cDNA; and Gray
Shaw, Ray Camphausen, and Gloria Vachino (Genetics Institute, Inc,
Cambridge, MA) for PSL275 MoAb, CHO/PACE cells, and the FFFE PSGL-1
mutant and helpful discussions; and M. Snapp for
inspiration.
| |
REFERENCES |
1.
Kansas GS:
Selectins and their ligands: Current concepts and controversies.
Blood
88:3259,
1996[Free Full Text]
2.
Moore KL,
Stults NL,
Diaz S,
Smith DF,
Cummings RD,
Varki A,
McEver RP:
Identification of a specific glycoprotein ligand for P-selectin (CD62) on myeloid cells.
J Cell Biol
118:445,
1992[Abstract/Free Full Text]
3.
Sako D,
Chang X-J,
Barone KM,
Vachino G,
White HM,
Shaw G,
Veldman GM,
Bean KB,
Ahern TJ,
Furie B,
Cumming DA,
Larsen GA:
Expression cloning of a functional glycoprotein ligand for P-selectin.
Cell
75:1179,
1993[Medline]
[Order article via Infotrieve]
4.
Moore KL,
Eaton SF,
Lyons DE,
Lichenstein HS,
Cummings RD,
McEver RP:
The P-selectin glycoprotein ligand from human neutrophils displays sialylated, fucosylated, O-linked poly-N-acetyllactosamine.
J Biol Chem
269:23318,
1994[Abstract/Free Full Text]
5.
Norgard KE,
Moore KL,
Diaz S,
Stults N,
Ushiyama S,
McEver RP,
Cummings RD,
Varki A:
Characterization of a specific ligand for P-selectin on myeloid cells. A minor glycoprotein with sialylated O-linked oligosaccharides.
J Biol Chem
268:12764,
1993[Abstract/Free Full Text]
6.
Wilkins PP,
McEver RP,
Cummings RD:
Structure of the O glycans on P-selectin glycoprotein ligand-1 from HL60 cells.
J Biol Chem
271:18732,
1996[Abstract/Free Full Text]
7.
Wilkins PP,
Moore KL,
McEver RP,
Cummings RD:
Tyrosine sulfation of P-selectin glycoprotein ligand-1 is required for high affinity binding to P-selectin.
J Biol Chem
270:22677,
1995[Abstract/Free Full Text]
8.
Pouyani T,
Seed B:
PSGL-1 recognition of P-selectin is controlled by a tyrosine sulfation consensus at the PSGL-1 amino terminus.
Cell
83:333,
1995[Medline]
[Order article via Infotrieve]
9.
Sako D,
Comess KM,
Barone KM,
Camphausen RT,
Cumming DA,
Shaw GD:
A sulfated peptide segment at the amino terminus of PSGL-1 is critical for P-selectin binding.
Cell
83:323,
1995[Medline]
[Order article via Infotrieve]
10.
Moore KL,
Patel KD,
Breuhl RE,
Fugang L,
Johnson DL,
Lichenstein HS,
Cummings RD,
Bainton DF,
McEver RP:
P-selectin glycoprotein ligand-1 mediates rolling of human neutrophils on P-selectin.
J Cell Biol
128:661,
1995[Abstract/Free Full Text]
11.
Walchek B,
Moore KL,
McEver RP,
Kishimoto TK:
Neutrophil-neutrophil interactions under hydrodynamic shear stress involve L-selectin and PSGL-1.
J Clin Invest
98:1081,
1996[Medline]
[Order article via Infotrieve]
12.
Guyer GA,
Moore KL,
Lynam EB,
Schammel CMG,
Rogelj S,
McEver RP,
Sklar LA:
P-selectin glycoprotein ligand-1 (PSGL-1) is a ligand for L-selectin in neutrophil aggregation.
Blood
88:2415,
1996[Abstract/Free Full Text]
13.
Alon R,
Rossiter H,
Wang X,
Springer TA,
Kupper TS:
Distinct cell surface ligands mediate T lymphocyte attachment and rolling on P and E-selectin under physiological flow.
J Cell Biol
127:1485,
1994[Abstract/Free Full Text]
14.
Vachino G,
Chang XJ,
Veldman GM,
Kumar R,
Sako D,
Fouser LA,
Berndt MC,
Cumming DA:
P-selectin glycoprotein ligand-1 is the major counter-receptor for P-selectin on stimulated T cells and is widely distributed in non-functional form on many lymphocytic cells.
J Biol Chem
270:21966,
1995[Abstract/Free Full Text]
15.
Luscinskas FW,
Ding H,
Tan P,
Cumming D,
Tedder TF,
Gerritsen ME:
L- and P-selectins but not VLA-4 (41) integrins mediate monocyte initial attachment to TNF- activated endothelium under flow in vitro.
J Immunol
157:326,
1996[Abstract]
16.
Lichtman AH,
Ding H,
Henault L,
Vachino G,
Camphausen R,
Cumming D,
Luscinskas FW:
CD45RA R0+ (memory) but not CD45RA+R0 (naive) T cells roll efficiently on E- and P-selectin and VCAM-1 under flow.
J Immunol
158:3640,
1997[Abstract]
17.
Laszik Z,
Jansen PJ,
Cummings RD,
Tedder TF,
McEver RP,
Moore KL:
P-selectin glycoprotein ligand-1 is broadly expressed in cells of myeloid, lymphoid, and dendritic lineage and in some nonhematopoietic cells.
Blood
88:3010,
1996[Abstract/Free Full Text]
18.
Takebe Y,
Seiki M,
Fujisawa JI,
Hoy P,
Yokota K,
Arai KI,
Yoshida M,
Arai N:
SR promoter: An efficient and versatile mammalian cDNA expression system composed of the Simian virus 40 early promoter and the R-U5 segment of human T-cell leukemia virus type 1 long terminal repeat.
Mol Cell Biol
8:466,
1988[Abstract/Free Full Text]
19.
Kansas GS,
Saunders KB,
Ley K,
Zakrzewicz A,
Gibson RM,
Furie BC,
Furie B,
Tedder TF:
A role for the epidermal growth factor-like domain of P-selectin in ligand recognition and cell adhesion.
J Cell Biol
124:609,
1994[Abstract/Free Full Text]
20.
Kansas GS,
Pavalko FM:
The cytoplasmic domains of E- and P-selectin do not constitutively interact with -actinin and are not essential for leukocyte adhesion.
J Immunol
157:321,
1996[Abstract]
21.
Snapp KR,
Wagers AJ,
Craig R,
Stoolman LM,
Kansas GS:
P-selectin glycoprotein ligand-1 (PSGL-1) is essential for adhesion to P-selectin but not E-selectin in stably transfected hematopoietic cell lines.
Blood
89:896,
1996[Abstract/Free Full Text]
22.
Wagers AJ,
Lowe JB,
Kansas GS:
An important role for the 1,3-fucosyltransferase FucT-VII in leukocyte adhesion to E-selectin.
Blood
88:2125,
1996[Abstract/Free Full Text]
23.
Ley K,
Tedder TF,
Kansas GS:
L-selectin can mediate rolling independent of E- and P-selectin in mesenteric venules in vivo.
Blood
82:1632,
1993[Abstract/Free Full Text]
24.
Luscinskas FW,
Kansas GS,
Ding H,
Pizcueta P,
Schlieffenbaum B,
Tedder TF,
Gimbrone MA:
Monocyte rolling, arrest and spreading on IL-4 activated vascular endothelium under flow is mediated via sequential action of L-selectin, 1 and 2 integrins.
J Cell Biol
125:1417,
1994[Abstract/Free Full Text]
25.
Luscinskas FW,
Ding H,
Lichtman AH:
P-selectin and vascular cell adhesion molecule 1 mediate rolling and arrest, respectively, of CD4+ T lymphocytes on tumor necrosis factor- activated vascular endothelium under flow.
J Exp Med
181:1179,
1995[Abstract/Free Full Text]
26.
Kumar R,
Camphausen RT,
Sullivan FX,
Cumming DA:
Core 2 -1,6-N-Acetylglucosaminyltransferase enzyme activity is critical for P-selectin glycoprotein ligand-1 binding to P-selectin.
Blood
88:3872,
1996[Abstract/Free Full Text]
27.
Wood GS,
Warnke RA:
The immunologic phenotyping of bone marrow biopsies and aspirates: frozen section techniques.
Blood
59:913,
1982[Abstract/Free Full Text]
28.
Wagers AJ,
Stoolman LM,
Kannagi R,
Craig R,
Kansas GS:
Expression of leukocyte fucosyltransferases regulates binding to E-selectin. Relationship to previously implicated carbohydrate epitopes.
J Immunol
159:1917,
1997[Abstract]
29.
Huttner WB:
Tyrosine sulfation and the secretory pathway.
Annu Rev Physiol
50:363,
1988[Medline]
[Order article via Infotrieve]
30.
Goetz DJ,
Greif DM,
Ding H,
Camphausen RT,
Howes S,
Comess KM,
Snapp KR,
Kansas GS,
Luscinskas FW:
Isolated P-selectin glycoprotein ligand-1 dynamic adhesion to P- and E-selectin.
J Cell Biol
137:509,
1997[Abstract/Free Full Text]
31.
Fuhlbrigge RC,
Alon R,
Puri KD,
Lowe JB,
Springer TA:
Sialylated, fucosylated ligands for L-selectin expressed on leukocytes mediate tethering and rolling adhesions in physiologic flow conditions.
J Cell Biol
135:837,
1996[Abstract/Free Full Text]
32.
Asa D,
Raycroft L,
Ma L,
Aeed PA,
Kaytes PS,
Elhammer AP,
Geng J-G:
The P-selectin glycoprotein ligand functions as a common human leukocoyte ligand for P- and E-selectins.
J Biol Chem
270:11662,
1995[Abstract/Free Full Text]
33.
Li F,
Wilkins PP,
Crawley S,
Weinstein J,
Cummings RD,
McEver RP:
Post-translational modifications of recombinant P-selectin glycoprotein ligand-1 required for binding to P- and E-selectin.
J Biol Chem
271:3255,
1996[Abstract/Free Full Text]
34.
Bargatze RF,
Kurk S,
Butcher EC,
Jutila MA:
Neutrophils roll on adherent neutrophils bound to cytokine-induced endothelial cells via L-selectin on the rolling cells.
J Exp Med
180:1785,
1994[Abstract/Free Full Text]
35.
Maly P,
Thall AD,
Petryniak B,
Rogers CE,
Smith PL,
Marks RM,
Kelly RJ,
Gersten KM,
Cheng G,
Saunders TL,
Camper SA,
Camphausen RT,
Sullivan FX,
Isogai Y,
Hindsgaul O,
von Andrian UH,
Lowe JB:
The (1,3) fucosyltransferase FucT-VII controls leukocyte trafficking through an essential role in L-, E-, and P-selectin ligand biosynthesis.
Cell
86:643,
1996[Medline]
[Order article via Infotrieve]
36.
Imai Y,
Lasky LA,
Rosen SD:
Sulphation requirement for GlyCAM-1, an endothelial ligand for L-selectin.
Nature
361:555,
1993[Medline]
[Order article via Infotrieve]
37.
Baumhueter S,
Singer MS,
Henzel W,
Hemmerich S,
Renz M,
Rosen SD,
Lasky LA:
Binding of L-selectin to the vascular sialomucin CD34.
Science
262:436,
1993[Abstract/Free Full Text]
38.
Hemmerich S,
Rosen SD:
6 -Sulfated sialyl Lewis x is a major capping group of GlyCAM-1.
Biochemistry
33:4830,
1994[Medline]
[Order article via Infotrieve]
39.
Hemmerich S,
Bertozzi CR,
Leffler H,
Rosen SD:
Identification of the sulfated monosaccharides of GlyCAM-1, an endothelial-derived ligand for L-selectin.
Biochemistry
33:4820,
1994[Medline]
[Order article via Infotrieve]
40.
Diacovo TG,
Roth SJ,
Morita CT,
Rosat J-P,
Brenner MB,
Springer TA:
Interactions of human / and / T lymphocyte subsets in shear flow with E-selectin and P-selectin.
J Exp Med
183:1193,
1996[Abstract/Free Full Text]
41.
Yang J,
Galipeau J,
Kozak CA,
Furie BC,
Furie B:
Mouse P-selectin glycoprotein ligand-1: Molecular cloning, chromosomal localization and expression of a functional P-selectin receptor.
Blood
87:4176,
1996[Abstract/Free Full Text]
42.
Li F,
Erickson HP,
James JA,
Moore KL,
Cummings RD,
McEver RP:
Visualization of P-selectin glycoprotein-1 as a highly extended molecule and mapping of protein epitopes for monoclonal antibodies.
J Biol Chem
271:6342,
1996[Abstract/Free Full Text]
43.
Varki A:
Selectin ligands.
Proc Natl Acad Sci USA
91:7390,
1994[Abstract/Free Full Text]
44.
Spertini O,
Kansas GS,
Reimann KA,
Mackay CR,
Tedder TF:
Functional and evolutionary conservation of distinct epitopes on the leukocyte adhesion molecule-1 (LAM-1) that regulate leukocyte migration.
J Immunol
147:942,
1991[Abstract]
45.
Engel P,
Nojima Y,
Rothstein D,
Zhou L-J,
Wilson GL,
Kehrl JH,
Tedder TF:
The same epitope on CD22 of B lymphocytes mediates the adhesion of erythrocytes, T and B lymphocytes, neutrophils and monocytes.
J Immunol
150:4719,
1993[Abstract]
 CiteULike  Connotea  Del.icio.us  Digg  Reddit  Technorati What's this?
This article has been cited by other articles:
|
|
|
|
 
K. A. Volcik, D. Catellier, A. R. Folsom, N. Matijevic, B. Wasserman, and E. Boerwinkle
SELP and SELPLG Genetic Variation Is Associated with Cell Surface Measures of SELP and SELPLG: The Atherosclerosis Risk in Communities Carotid MRI Study
Clin. Chem.,
June 1, 2009;
55(6):
1076 - 1082.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. C. Kerr, C. B. Fieger, K. R. Snapp, and S. D. Rosen
Endoglycan, a Member of the CD34 Family of Sialomucins, Is a Ligand for the Vascular Selectins
J. Immunol.,
July 15, 2008;
181(2):
1480 - 1490.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. Nonomura, J. Kikuchi, N. Kiyokawa, H. Ozaki, K. Mitsunaga, H. Ando, A. Kanamori, R. Kannagi, J. Fujimoto, K. Muroi, et al.
CD43, but not P-Selectin Glycoprotein Ligand-1, Functions as an E-Selectin Counter-Receptor in Human Pre-B-Cell Leukemia NALL-1
Cancer Res.,
February 1, 2008;
68(3):
790 - 799.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
M. Sarkar, D. V. Reneer, and J. A. Carlyon
Sialyl-Lewis x-Independent Infection of Human Myeloid Cells by Anaplasma phagocytophilum Strains HZ and HGE1
Infect. Immun.,
December 1, 2007;
75(12):
5720 - 5725.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
M. Shimoda, G. Hashimoto, S. Mochizuki, E. Ikeda, N. Nagai, S. Ishida, and Y. Okada
Binding of ADAM28 to P-selectin Glycoprotein Ligand-1 Enhances P-selectin-mediated Leukocyte Adhesion to Endothelial Cells
J. Biol. Chem.,
August 31, 2007;
282(35):
25864 - 25874.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. Parolini, A. Santoro, E. Marcenaro, W. Luini, L. Massardi, F. Facchetti, D. Communi, M. Parmentier, A. Majorana, M. Sironi, et al.
The role of chemerin in the colocalization of NK and dendritic cell subsets into inflamed tissues
Blood,
May 1, 2007;
109(9):
3625 - 3632.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. Bestebroer, M. J. J. G. Poppelier, L. H. Ulfman, P. J. Lenting, C. V. Denis, K. P. M. van Kessel, J. A. G. van Strijp, and C. J. C. de Haas
Staphylococcal superantigen-like 5 binds PSGL-1 and inhibits P-selectin-mediated neutrophil rolling
Blood,
April 1, 2007;
109(7):
2936 - 2943.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. W. Kehoe, N. Velappan, M. Walbolt, J. Rasmussen, D. King, J. Lou, K. Knopp, P. Pavlik, J. D. Marks, C. R. Bertozzi, et al.
Using Phage Display to Select Antibodies Recognizing Post-translational Modifications Independently of Sequence Context
Mol. Cell. Proteomics,
December 1, 2006;
5(12):
2350 - 2363.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. B. Park and N. Schoene
Clovamide-Type Phenylpropenoic Acid Amides, N-Coumaroyldopamine and N-Caffeoyldopamine, Inhibit Platelet-Leukocyte Interactions via Suppressing P-Selectin Expression
J. Pharmacol. Exp. Ther.,
May 1, 2006;
317(2):
813 - 819.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. Ohmori, F. Fukui, M. Kiso, T. Imai, O. Yoshie, H. Hasegawa, K. Matsushima, and R. Kannagi
Identification of cutaneous lymphocyte-associated antigen as sialyl 6-sulfo Lewis X, a selectin ligand expressed on a subset of skin-homing helper memory T cells
Blood,
April 15, 2006;
107(8):
3197 - 3204.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
I. del Conde, F. Nabi, R. Tonda, P. Thiagarajan, J. A. Lopez, and N. S. Kleiman
Effect of P-Selectin on Phosphatidylserine Exposure and Surface-Dependent Thrombin Generation on Monocytes
Arterioscler. Thromb. Vasc. Biol.,
May 1, 2005;
25(5):
1065 - 1070.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. S. Merzaban, J. Zuccolo, S. Y. Corbel, M. J. Williams, and H. J. Ziltener
An Alternate Core 2 {beta}1,6-N-Acetylglucosaminyltransferase Selectively Contributes to P-Selectin Ligand Formation in Activated CD8 T Cells
J. Immunol.,
April 1, 2005;
174(7):
4051 - 4059.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
T. Inoue, Y. Tsuzuki, K. Matsuzaki, H. Matsunaga, J. Miyazaki, R. Hokari, Y. Okada, A. Kawaguchi, S. Nagao, K. Itoh, et al.
Blockade of PSGL-1 attenuates CD14+ monocytic cell recruitment in intestinal mucosa and ameliorates ileitis in SAMP1/Yit mice
J. Leukoc. Biol.,
March 1, 2005;
77(3):
287 - 295.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. Ivetic, O. Florey, J. Deka, D. O. Haskard, A. Ager, and A. J. Ridley
Mutagenesis of the Ezrin-Radixin-Moesin Binding Domain of L-selectin Tail Affects Shedding, Microvillar Positioning, and Leukocyte Tethering
J. Biol. Chem.,
August 6, 2004;
279(32):
33263 - 33272.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. Yanaba, K. Komura, M. Horikawa, Y. Matsushita, K. Takehara, and S. Sato
P-selectin glycoprotein ligand-1 is required for the development of cutaneous vasculitis induced by immune complex deposition
J. Leukoc. Biol.,
August 1, 2004;
76(2):
374 - 382.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
I. G. Winkler, K. R. Snapp, P. J. Simmons, and J.-P. Levesque
Adhesion to E-selectin promotes growth inhibition and apoptosis of human and murine hematopoietic progenitor cells independent of PSGL-1
Blood,
March 1, 2004;
103(5):
1685 - 1692.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. K. Sarangapani, T. Yago, A. G. Klopocki, M. B. Lawrence, C. B. Fieger, S. D. Rosen, R. P. McEver, and C. Zhu
Low Force Decelerates L-selectin Dissociation from P-selectin Glycoprotein Ligand-1 and Endoglycan
J. Biol. Chem.,
January 16, 2004;
279(3):
2291 - 2298.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
O. Dwir, A. Solomon, S. Mangan, G. S. Kansas, U. S. Schwarz, and R. Alon
Avidity enhancement of L-selectin bonds by flow: shear-promoted rotation of leukocytes turn labile bonds into functional tethers
J. Cell Biol.,
November 10, 2003;
163(3):
649 - 659.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K.-S. Choi, J. Garyu, J. Park, and J. S. Dumler
Diminished Adhesion of Anaplasma phagocytophilum-Infected Neutrophils to Endothelial Cells Is Associated with Reduced Expression of Leukocyte Surface Selectin
Infect. Immun.,
August 1, 2003;
71(8):
4586 - 4594.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. B. Fieger, C. M. Sassetti, and S. D. Rosen
Endoglycan, a Member of the CD34 Family, Functions as an L-selectin Ligand through Modification with Tyrosine Sulfation and Sialyl Lewis x
J. Biol. Chem.,
July 25, 2003;
278(30):
27390 - 27398.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
L. Xia, V. Ramachandran, J. M. McDaniel, K. N. Nguyen, R. D. Cummings, and R. P. McEver
N-terminal residues in murine P-selectin glycoprotein ligand-1 required for binding to murine P-selectin
Blood,
January 15, 2003;
101(2):
552 - 559.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
B. Dang, S. Wiehler, and K. D. Patel
Increased PSGL-1 expression on granulocytes from allergic-asthmatic subjects results in enhanced leukocyte recruitment under flow conditions
J. Leukoc. Biol.,
October 1, 2002;
72(4):
702 - 710.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. Thatte, S. Ficarro, K. R. Snapp, M. K. Wild, D. Vestweber, D. F. Hunt, and K. F. Ley
Binding of function-blocking mAbs to mouse and human P-selectin glycoprotein ligand-1 peptides with and without tyrosine sulfation
J. Leukoc. Biol.,
September 1, 2002;
72(3):
470 - 477.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. Kanamori, N. Kojima, K. Uchimura, T. Muramatsu, T. Tamatani, M. C. Berndt, G. S. Kansas, and R. Kannagi
Distinct Sulfation Requirements of Selectins Disclosed Using Cells That Support Rolling Mediated by All Three Selectins under Shear Flow. L-SELECTIN PREFERS CARBOHYDRATE 6-SULFATION TO TYROSINE SULFATION, WHEREAS P-SELECTIN DOES NOT
J. Biol. Chem.,
August 30, 2002;
277(36):
32578 - 32586.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. Tarte, J. De Vos, T. Thykjaer, F. Zhan, G. Fiol, V. Costes, T. Reme, E. Legouffe, J.-F. Rossi, J. Shaughnessy Jr, et al.
Generation of polyclonal plasmablasts from peripheral blood B cells: a normal counterpart of malignant plasmablasts
Blood,
July 30, 2002;
100(4):
1113 - 1122.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. R. Snapp, C. E. Heitzig, and G. S. Kansas
Attachment of the PSGL-1 cytoplasmic domain to the actin cytoskeleton is essential for leukocyte rolling on P-selectin
Blood,
May 29, 2002;
99(12):
4494 - 4502.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. M. Fong, S. M. Alam, T. Imai, B. Haribabu, and D. D. Patel
CX3CR1 Tyrosine Sulfation Enhances Fractalkine-induced Cell Adhesion
J. Biol. Chem.,
May 24, 2002;
277(22):
19418 - 19423.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. A. Boyce, E. A. Mellor, B. Perkins, Y.-C. Lim, and F. W. Luscinskas
Human mast cell progenitors use alpha 4-integrin, VCAM-1, and PSGL-1 E-selectin for adhesive interactions with human vascular endothelium under flow conditions
Blood,
April 15, 2002;
99(8):
2890 - 2896.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
G. H. Underhill, H. A. Minges-Wols, J. L. Fornek, P. L. Witte, and G. S. Kansas
IgG plasma cells display a unique spectrum of leukocyte adhesion and homing molecules
Blood,
April 15, 2002;
99(8):
2905 - 2912.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. J. Dimitroff, J. Y. Lee, K. S. Schor, B. M. Sandmaier, and R. Sackstein
Differential L-Selectin Binding Activities of Human Hematopoietic Cell L-Selectin Ligands, HCELL and PSGL-1
J. Biol. Chem.,
December 7, 2001;
276(50):
47623 - 47631.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. Janowska-Wieczorek, M. Majka, J. Kijowski, M. Baj-Krzyworzeka, R. Reca, A. R. Turner, J. Ratajczak, S. G. Emerson, M. A. Kowalska, and M. Z. Ratajczak
Platelet-derived microparticles bind to hematopoietic stem/progenitor cells and enhance their engraftment
Blood,
November 15, 2001;
98(10):
3143 - 3149.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
Y.-C. Lim, H. Xie, C. E. Come, S. I. Alexander, M. J. Grusby, A. H. Lichtman, and F. W. Luscinskas
IL-12, STAT4-Dependent Up-Regulation of CD4+ T Cell Core 2 {beta}-1,6-n-Acetylglucosaminyltransferase, an Enzyme Essential for Biosynthesis of P-Selectin Ligands
J. Immunol.,
October 15, 2001;
167(8):
4476 - 4484.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. A. DiVietro, M. J. Smith, B. R. E. Smith, L. Petruzzelli, R. S. Larson, and M. B. Lawrence
Immobilized IL-8 Triggers Progressive Activation of Neutrophils Rolling In Vitro on P-Selectin and Intercellular Adhesion Molecule-1
J. Immunol.,
October 1, 2001;
167(7):
4017 - 4025.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J.-F. Theoret, J.-G. Bienvenu, A. Kumar, and Y. Merhi
P-Selectin Antagonism with Recombinant P-Selectin Glycoprotein Ligand-1 (rPSGL-Ig) Inhibits Circulating Activated Platelet Binding to Neutrophils Induced by Damaged Arterial Surfaces
J. Pharmacol. Exp. Ther.,
August 1, 2001;
298(2):
658 - 664.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. J. White, G. H. Underhill, M. H. Kaplan, and G. S. Kansas
Cutting Edge: Differential Requirements for Stat4 in Expression of Glycosyltransferases Responsible for Selectin Ligand Formation in Th1 Cells
J. Immunol.,
July 15, 2001;
167(2):
628 - 631.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
E. Evans, A. Leung, D. Hammer, and S. Simon
Chemically distinct transition states govern rapid dissociation of single L-selectin bonds under force
PNAS,
March 7, 2001;
(2001)
61324998.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
C. J. Dimitroff, J. Y. Lee, R. C. Fuhlbrigge, and R. Sackstein
A distinct glycoform of CD44 is an L-selectin ligand on human hematopoietic cells
PNAS,
November 22, 2000;
(2000)
250484797.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
R. Sackstein and C. J. Dimitroff
A hematopoietic cell L-selectin ligand that is distinct from PSGL-1 and displays N-glycan-dependent binding activity
Blood,
October 15, 2000;
96(8):
2765 - 2774.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. Nomura, F. Okamae, M. Abe, M. Hosokawa, M. Yamaoka, T. Ohtani, S. Onishi, T. Matsuzaki, A. Teraoka, T. Ishida, et al.
Platelets Expressing P-Selectin and Platelet-Derived Microparticles in Stored Platelet Concentrates Bind to PSGL-1 on Filtrated Leukocytes
Clinical and Applied Thrombosis/Hemostasis,
October 1, 2000;
6(4):
213 - 221.
[Abstract]
[PDF]
|
|
|
|
|
|
|
M. J. Herron, C. M. Nelson, J. Larson, K. R. Snapp, G. S. Kansas, and J. L. Goodman
Intracellular Parasitism by the Human Granulocytic Ehrlichiosis Bacterium Through the P-Selectin Ligand, PSGL-1
Science,
June 2, 2000;
288(5471):
1653 - 1656.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
P. S. Frenette, C. V. Denis, L. Weiss, K. Jurk, S. Subbarao, B. Kehrel, J. H. Hartwig, D. Vestweber, and D. D. Wagner
P-Selectin Glycoprotein Ligand 1 (PSGL-1) Is Expressed on Platelets and Can Mediate Platelet-Endothelial Interactions In Vivo
J. Exp. Med.,
April 18, 2000;
191(8):
1413 - 1422.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. L. Alonso-Lebrero, J. M. Serrador, C. Dominguez-Jimenez, O. Barreiro, A. Luque, M. A. del Pozo, K. Snapp, G. Kansas, R. Schwartz-Albiez, H. Furthmayr, et al.
Polarization and interaction of adhesion molecules P-selectin glycoprotein ligand 1 and intercellular adhesion molecule 3 with moesin and ezrin in myeloid cells
Blood,
April 1, 2000;
95(7):
2413 - 2419.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
E. R. Hentzen, S. Neelamegham, G. S. Kansas, J. A. Benanti, L. V. McIntire, C. W. Smith, and S. I. Simon
Sequential binding of CD11a/CD18 and CD11b/CD18 defines neutrophil capture and stable adhesion to intercellular adhesion molecule-1
Blood,
February 1, 2000;
95(3):
911 - 920.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
L. Tu, P. G. Murphy, X. Li, and T. F. Tedder
L-Selectin Ligands Expressed by Human Leukocytes Are HECA-452 Antibody-Defined Carbohydrate Epitopes Preferentially Displayed by P-Selectin Glycoprotein Ligand-1
J. Immunol.,
November 1, 1999;
163(9):
5070 - 5078.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. Leppanen, P. Mehta, Y.-B. Ouyang, T. Ju, J. Helin, K. L. Moore, I. van Die, W. M. Canfield, R. P. McEver, and R. D. Cummings
A Novel Glycosulfopeptide Binds to P-selectin and Inhibits Leukocyte Adhesion to P-selectin
J. Biol. Chem.,
August 27, 1999;
274(35):
24838 - 24848.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. L. Ramos, Y. Huo, U. Jung, S. Ghosh, D. R. Manka, I. J. Sarembock, and K. Ley
Direct Demonstration of P-Selectin– and VCAM-1–Dependent Mononuclear Cell Rolling in Early Atherosclerotic Lesions of Apolipoprotein E–Deficient Mice
Circ. Res.,
June 11, 1999;
84(11):
1237 - 1244.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. Bistrup, S. Bhakta, J. K. Lee, Y. Y. Belov, M. D. Gunn, F.-R. Zuo, C.-C. Huang, R. Kannagi, S. D. Rosen, and S. Hemmerich
Sulfotransferases of Two Specificities Function in the Reconstitution of High Endothelial Cell Ligands for L-selectin
J. Cell Biol.,
May 17, 1999;
145(4):
899 - 910.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
Y.-C. Lim, L. Henault, A. J. Wagers, G. S. Kansas, F. W. Luscinskas, and A. H. Lichtman
Expression of Functional Selectin Ligands on Th Cells Is Differentially Regulated by IL-12 and IL-4
J. Immunol.,
March 15, 1999;
162(6):
3193 - 3201.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
R. Mukasa, T. Homma, T. Ohtsuki, O. Hosono, A. Souta, T. Kitamura, M. Fukuda, S. Watanabe, and C. Morimoto
Core 2-containing O-glycans on CD43 are preferentially expressed in the memory subset of human CD4 T cells
Int. Immunol.,
February 1, 1999;
11(2):
259 - 268.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
Y.-C. Lim, K. Snapp, G. S. Kansas, R. Camphausen, H. Ding, and F. W. Luscinskas
Important Contributions of P-Selectin Glycoprotein Ligand-1-Mediated Secondary Capture to Human Monocyte Adhesion to P-Selectin, E-Selectin, and TNF-{alpha}-Activated Endothelium Under Flow In Vitro
J. Immunol.,
September 1, 1998;
161(5):
2501 - 2508.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. R. Snapp, R. Craig, M. Herron, R. D. Nelson, L. M. Stoolman, and G. S. Kansas
Dimerization of P-Selectin Glycoprotein Ligand-1 (PSGL-1) Required for Optimal Recognition of P-Selectin
J. Cell Biol.,
July 13, 1998;
142(1):
263 - 270.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. J. Wagers, L. M. Stoolman, R. Craig, R. N. Knibbs, and G. S. Kansas
An sLex-Deficient Variant of HL60 Cells Exhibits High Levels of Adhesion to Vascular Selectins: Further Evidence That HECA-452 and CSLEX1 Monoclonal Antibody Epitopes Are Not Essential for High Avidity Binding to Vascular Selectins
J. Immunol.,
May 15, 1998;
160(10):
5122 - 5129.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
P. Andre, O. Spertini, S. Guia, P. Rihet, F. Dignat-George, H. Brailly, J. Sampol, P. J. Anderson, and E. Vivier
Modification of P-selectin glycoprotein ligand-1 with a natural killer cell-restricted sulfated lactosamine creates an alternate ligand for L-selectin
PNAS,
March 28, 2000;
97(7):
3400 - 3405.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
E. Evans, A. Leung, D. Hammer, and S. Simon
Chemically distinct transition states govern rapid dissociation of single L-selectin bonds under force
PNAS,
March 27, 2001;
98(7):
3784 - 3789.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. J. Dimitroff, J. Y. Lee, R. C. Fuhlbrigge, and R. Sackstein
A distinct glycoform of CD44 is an L-selectin ligand on human hematopoietic cells
PNAS,
December 5, 2000;
97(25):
13841 - 13846.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
Abstract
Introduction
Abstract
