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HEMOSTASIS, THROMBOSIS, AND VASCULAR BIOLOGY
From the Department of Chemical Engineering, Department
of Medicine, Johns Hopkins University, Baltimore, MD; DuPont
Pharmaceuticals Co, Wilmington, DE.
Accumulating evidence suggests that successful metastatic spread
may depend on the ability of tumor cells to undergo extensive interactions with platelets. However, the mechanisms mediating tumor
cell adhesion to platelets under conditions of flow remain largely
unknown. Therefore, this study was designed to analyze the ability of 3 human colon carcinoma cell lines (LS174T, COLO205, and HCT-8) to bind
to surface-anchored platelets under flow and to identify the receptors
involved in these processes. Immobilized platelets support LS174T cell
adhesion at wall shear stresses up to 1.4 dyn/cm2. Our data
suggest that platelets primarily recruit LS174T cells through a 2-step,
sequential process of adhesive interactions that shares common features
but is distinct from that elaborated for neutrophils. Platelet
P-selectin mediates LS174T cell tethering and rolling in a PSGL-1- and
CD24-independent manner. Moreover, platelet
Blood-borne metastasis is a highly regulated and
dynamic process in which cancerous cells separate from a primary tumor,
migrate across blood vessel walls into the bloodstream, and disperse
throughout the body to generate new colonies. During their transit into
the circulatory system, tumor cells are exposed to fluid mechanical forces, plasma proteins, and vascular cells such as platelets and
neutrophils, all of which may affect their survival and extravasation from the vasculature.
Several lines of evidence suggest that platelets facilitate
hematogenous dissemination of tumor cells. The most convincing evidence
is the inhibition of metastasis by experimental thrombocytopenia and
the restoration of metastatic potential by platelet
infusion.1,2 Morphologic observations of tumor cells
arrested in capillaries have documented the close association of tumor
cells with activated platelets.3 However, the mechanisms
by which platelets assist tumor metastasis are not clearly understood.
Some studies suggest that platelets, by adhering to tumor cells,
provide a protective shield that masks them from the cytotoxic activity
of natural killer cells.4,5 Alternatively, platelets may
facilitate tumor extravasation by potentiating tumor cell adhesive
interactions with extracellular matrix proteins of the vessel
wall,4,6,7 thereby assisting tumor cells to escape from
the circulation into the interstitium and form a distinct secondary
colony. Moreover, activated platelets may release a number of growth
factors such as platelet-derived growth factor that has been reported
to stimulate the growth of highly metastatic colon carcinoma
cells.4 Along with these lines, it has recently been
hypothesized that platelets may contribute to tumor-induced
angiogenesis.8 Therefore, elucidating the molecular
mechanisms of tumor cell-platelet adhesion may eventually provide a
rational basis for the development of novel therapeutic strategies to
combat metastasis.
Platelets have been reported to bind to certain melanoma, breast,
and colon cancer cells through the integrin receptor
Most of the prior work on tumor cell adhesion to platelets was carried
out in vitro mainly under static conditions.9,14-16 However, these interactions typically occur in the bloodstream, in the
presence of shear flow. As has been argued in the literature, data
obtained in vitro using static assays may not be relevant to the
fluid dynamic environment of the vasculature.17 For
instance, melanoma cells fail to adhere directly to collagen I
extracellular matrix under flow, despite their ability to interact
extensively with this matrix under static conditions.6
Furthermore, previous studies on leukocyte-endothelial cell adhesion as
well as on platelet binding to extracellular matrix proteins have
clearly established that the shear stress generated by blood flow
critically affects cell adhesive interactions.12,18,19
Consequently, the present study was undertaken to investigate the
molecular mechanisms of tumor cell attachment to surface-bound platelet
layers under flow conditions. Prior work has shown that platelets
adherent to thrombogenic surfaces support neutrophil recruitment
through a multistep, sequential process of adhesive interactions.
P-selectin expressed on the surface of immobilized, activated platelets
mediates the initial tethering and rolling of neutrophils in shear
flow.20,21 Activation-dependent attachments of
The present study demonstrates that surface-adherent platelets support
extensive adhesive interactions of sLex-bearing colon
cancer cells, L5174T and COLD205, under flow conditions simulated in
vitro using a parallel-plate flow chamber. Our results indicate that
tumor cell tethering and rolling involve binding of a sialylated
molecule on the tumor cell surface to platelet P-selectin that is
distinct from P-selection glycoprotein ligand-1 (PSGL-1) and
CD24. Furthermore, platelet Monoclonal antibodies
Cell lines and culture
Enzyme treatment To remove terminal cell surface sialic acid residues, tumor cells (107/mL) were incubated with 0.1 U/mL Vibrio cholerae neuraminidase (Boehringer Mannheim, Indianapolis, IN) for 30 minutes at room temperature.31,32 In independent experiments, tumor cells were treated with 1 U/mL PI-PLC (Sigma, Glyko, Novato, CA) for 1 hour at 37°C to cleave GPI-linked molecules such as CD24.33 Following enzyme treatment, tumor cells were washed once and analyzed by flow cytometry or infused into the flow chamber for adhesion assays.Generation of P-selectin-coated surfaces Recombinant P-selectin (R&D Systems, Minneapolis, MN) was diluted with D-phosphate-buffered saline (D-PBS) containing Ca++/Mg++ to a final concentration of 4 µg/mL, layered on a 35-mm tissue culture dish (Corning, Corning, NY) and allowed to coat overnight at 4°C. Subsequently the dishes were washed 3 times with D-PBS before incubation for 2 hours with D-PBS/1% BSA at 4°C to eliminate nonspecific interactions.Immobilization of platelet layers on glass slides To provide a substrate that readily binds platelets, glass slides (24 × 50 mm) were coated with 3-aminopropyltriethoxysilane (APES; Sigma).20 Human blood was drawn by venipuncture from healthy volunteers and a patient with Glanzmann thrombasthenia (GT)34 into sodium citrate (0.38% wt/vol) anticoagulant. Platelet-rich plasma (PRP) was prepared by centrifugation of whole blood at 160g for 15 minutes. The PRP count was adjusted to 2 × 108/mL before being bound to APES-treated glass slides for 60 minutes. Nonspecific binding was blocked with 0.1% BSA for 10 minutes at 37°C. Under these conditions, a confluent layer of platelets was formed as evaluated by light microscopy for each experiment (Figure 1A). The density (17 580 ± 971/mm2) and confluency of platelet layers were not affected during the flow experiment.
Flow adhesion assays Tumor cell adhesion to immobilized platelets (and purified P-selectin) was quantitated under flow conditions using a parallel-plate flow chamber. A platelet-coated glass slide was assembled to a flow chamber (150-µm channel depth, 1.26-cm channel width) and mounted on the stage of an inverted microscope (Nikon TE300) equipped with × 10 and × 20 phase objectives (Nikon, Melville, NY), a × 0.55 projection lens (Nikon) and a CCD100 camera (Dage-MTI, Michigan City, IN) connected to a VCR and a TV monitor. Surface-adherent platelets were then incubated with either 1 U/mL thrombin (only in experiments involving LS174T cells) or D-PBS/0.1% BSA for 10 minutes at 37°C. After washing the platelet layer with D-PBS/0.1% BSA for approximately 2 minutes, tumor cells were perfused through the chamber for 10 minutes at the appropriate flow rates to obtain wall shear stresses of 0.6 to 1.4 dyn/cm2, thereby mimicking the fluid mechanical environment of the microcirculation and postcapillary venules. The flow system was maintained at 37°C in an air curtain incubator. Interactions between tumor cells and surface-adherent platelets were visualized in real-time by phase-contrast videomicroscopy. A single field of view (× 20; 0.14 mm2) was monitored during the 10 minutes of the experiment, and at the end 5 fields of view (× 10; 0.55 mm2) were monitored for 15 seconds each. Three parameters were quantified in the analysis: (1) the number of total interacting cells during the entire 10-minute experiment; (2) the number of firmly adherent cells after 10 minutes of shear flow; and (3) the average rolling velocity. Interacting cells were defined as those that interacted with the platelet layer for at least 2 seconds, and included both immediately arrested and fast rolling or skipping cells that interacted intermittently with the monolayer. Their number was determined manually by reviewing the videotapes. Firmly adherent cells were considered as those that remained stationary for at least 10 seconds at the end of the 10-minute run. To quantify their number, images were digitized from the videotape recorder using a Scion frame grabber and a personal computer and processed with OPTIMAS 6.1 software package (Argis-Schoen Vision Systems Inc, Alexandria, VA). Rolling velocity was computed as the distance traveled by the centroid of the translating cell divided by the time interval, using OPTIMAS 6.1 software.For some inhibition studies, tumor cells were pretreated for 10 minutes with mAbs (20 µg/mL, unless otherwise stated), which were kept present during the flow assays. For others, surface-adherent platelets were preincubated with mAbs (20 µg/mL), XV454 (150 nmol/L), GRGDSP or GRGESP (500 µmol/L), or anti-vWf (1:250 dilution) for 10 minutes during the thrombin or buffer incubation. Saturating concentrations were also maintained in the flow buffer only for the GRGDSP and GRGESP peptides, anti-vWf, antifibronectin, and antifibrinogen antibodies. The extent of LS174T cell attachment to surface-anchored platelets was unaffected by platelet exposure to thrombin before tumor cell perfusion (data not shown), indicating a high degree of platelet activation had occurred as previously shown.35 Furthermore, LS174T cell adhesion to immobilized platelets was unaltered by the presence or absence of an IgG control mAb (untreated samples: 249 ± 54 firmly adherent cells/mm2 versus nonspecific IgG-treated samples: 247 ± 39 firmly adherent cells/mm2; n = 3). Similar results were also observed for COLO205 and HCT-8 cells (data not shown). Flow cytometry Indirect single-color immunofluorescence assays were performed to determine tumor cell surface glycoprotein expression.31 Tumor cells were incubated with saturating concentrations of each mAb for 30 minutes on ice, and then washed with D-PBS/0.1% BSA. After an additional 30 minutes of incubation with 15 µg/mL phycoerythrin (PE)-labeled IgG or IgM antibody (Vector Laboratories, Burlingame, CA), the specimens were washed again, fixed with 1% formaldehyde, and analyzed in a FACScan flow cytometer (Becton Dickinson). Tumor cells were distinguished from debris on the basis of their characteristic forward- and side-scatter profiles, and the geometric mean PE fluorescence of each specimen was recorded. Appropriate isotype-matched IgG or IgM mAbs were also included for background fluorescence determination.Statistics Data are expressed as the mean ± SEM. Statistical significance of differences between means was determined by ANOVA. If means were shown to be significantly different, multiple comparisons by pairs were performed by the Tukey test. Probability values of P < .05 were selected to be statistically significant.
Surface-adherent platelets support LS174T cell adhesion under flow Previous studies have shown that platelets interact with a number of cell lines derived from colon carcinomas, including LS174T, but cell adhesion was studied exclusively under static conditions.14,16 To determine whether LS174T cells also adhere to platelets under flow, tumor cells were perfused through a parallel-plate flow chamber whose lower plate was coated with a layer of platelets (Figure 1). Figure 2 shows that immobilized platelets supported extensive LS174T cell adhesion under flow in a shear stress-dependent manner. A progressive decrease in the extent of adhesion was detected between 0.6 dyn/cm2 and 1.4 dyn/cm2. No interaction was observed at higher stress levels. In distinct contrast, freshly isolated neutrophils tethered to platelets even at shear stress levels in the excess of 3.0 dyn/cm2 (data not shown), an observation that is in agreement with previously published work.21 LS174T cells that tethered from the fluid stream to the platelet layer became firmly adherent instantaneously (Figure 1B). Under these control settings, rolling or irregular translocation of LS174T cells along the platelet surface were rather rare events (firmly adherent cells: 184 ± 18 cells/mm2 versus total interacting cells: 208 ± 11 cells/mm2 at 0.8 dyn/cm2).
Roles of platelet P-selectin and
 IIb 3.14 Hence, as a first
step, we examined the potential role of platelet P-selectin in these
adhesive interactions under dynamic flow conditions. The results
indicate that incubation of the platelet layer with a function-blocking
anti-P-selectin antibody alone resulted in an approximately 70% to
75% reduction of adhesion (Figure 3).
Subsequent experiments aimed to identify the platelet receptor(s)
responsible for the residual LS174T cell attachment to immobilized platelets in the presence of an anti-P-selectin antibody. The integrin
To further assess the role of platelet
To further validate these findings, we used blood from a patient with
GT whose platelets are devoid of
Dependence of LS174T cell adhesion on platelet
IIb3-integrin that supports LS174T cell
firm adhesion using function-blocking mAbs directed against
IIb and 3 subunits. Figure
4 illustrates that an
anti-3 antibody, SZ21, did not significantly affect the
extent and pattern of these adhesive interactions. Similar results were
also obtained using another anti-3-blocking mAb, B3A
(control samples: 327 ± 35 firmly adherent cells/mm2
versus anti-3-treated samples: 307 ± 51 firmly
adherent cells/mm2; n = 3). In contrast, an
anti-IIb mAb, P2,39 dramatically inhibited
the number of firmly adherent LS174T cells to the platelet layers, an
effect that was accompanied by a marked increase in the number of
tethered cells (Figure 4). Similar results were also obtained using the
RGD-containing peptide GRGDSP (Figure 4). As expected, a GRGESP peptide
did not alter the extent of LS174T cell attachment to immobilized
platelets (control samples: 327 ± 35 firmly adherent
cells/mm2 versus GRGESP-treated samples: 285 ± 33 firmly
adherent cells/mm2; n = 3). Taken together, these data
suggest that the platelet IIb3-integrin
is involved in the formation of stable adhesive interactions between
LS174T cells and surface-adherent platelets via an RGD-dependent
mechanism. Although an intact
IIb3-complex is required in this process,
it is likely that the ligand binding site resides in the
IIb subunit.
We next explored the potential involvement in this process of
adhesive proteins such as vWf, fibrinogen, and fibronectin, which are
anchored on the surface of activated platelets predominantly via
Experiments were performed in the presence of 5 mmol/L EDTA to assess
the divalent cation requirements. These ions are necessary for
selectin- and integrin-mediated adhesion.12 When EDTA was added to the perfusion buffer, LS174T (Figure 4), cell interactions with immobilized platelets were totally abrogated, a finding that is
consistent with the platelet P-selectin and
Characterization of LS174T counter-receptors involved in adhesive interactions with platelets We next aimed to identify the counter-receptor for platelet P-selectin on the LS174T cell surface. Prior studies have demonstrated that the major ligand on leukocytes involved in the recruitment to P-selectin is PSGL-1.40,41 Therefore, its contribution to LS174T binding to surface-adherent platelets was examined. Flow cytometric analysis of LS174T adhesion receptor expression suggests that PSGL-1 may be marginally present on the tumor cell surface (Table 1). Blocking PSGL-1 function with a mAb failed to substantially reduce the extent of LS174T cell tethering toIIb3-blocked platelet surfaces (Figure
5A). In contrast, this anti-PSGL-1
antibody was effective in inhibiting neutrophil binding to purified
P-selectin (data not shown). These results are in agreement with recent
reports that suggest that a variety of cell lines from breast and colon carcinomas interact with P-selectin substrates in a PSGL-1-independent manner.30,42
Treatment of LS174T cells with neuraminidase, an enzyme that
cleaves sialic acid residues from cell surfaces (Table
1),31 abolished their tethering to immobilized
In an effort to demonstrate that P-selectin is sufficient to support
LS174T cell tethering, in the absence of any other adhesion receptor(s)
present on the surface of immobilized
We finally aimed to identify the LS174T cell counter-receptor for
platelet COLO205 and HCT-8 cell adhesion to immobilized platelets under flow To validate that immobilized platelets are capable of supporting tethering, rolling, and firm adhesion of colon cancer cells other than LS174T, we chose to look at sLex-bearing (COLO205) and sLex-negative (HCT-8) cell lines (Table 1).9,10,14 Our results indicate that COLO205 tethered and rolled extensively on immobilized platelets at a wall shear stress of 0.8 dyn/cm2 (Figure 6A). A significant number of interacting COLO205 cells became firmly adherent within a 1- to 5-second interval after tethering to the platelet substrate. Blockade of platelet P-selectin or treatment of COLO205 cells with neuraminidase but not PI-PLC nearly eliminated any cell tethering/adhesive interactions. Furthermore, blockade of either plateletIIb3 or platelet-bound vWf
abrogated COLO205 firm adhesion while dramatically increasing the
number of tethered cells (Figure 6A). Taken together, these data
suggest that platelet P-selectin initiates COLO205 tethering and
rolling to platelet layers in shear flow. In accordance with the LS174T data, the P-selectin ligand on COLO205 cells appears to be a sialylated molecule that is distinct from PSGL-1 (Table 1) and
CD24 (Figure 6A). COLO205 tethering and rolling events are occasionally
followed by stable adhesion that is totally dependent on platelet
IIb3 and platelet-bound vWf. It is worth
noting that HCT-8 cells, which are devoid of sLex residues
(Table 1), tether minimally to surface-anchored platelets in shear flow
(Figure 6B), despite their extensive adhesive interactions under static
conditions (data not shown). The very few HCT-8 cells that tethered
from the fluid stream to the platelet layer became firmly adherent
instantaneously with no obvious period of rolling in a
IIb3- and vWf-dependent manner
(Figure 6B).
Pattern of interactions/rolling velocities We observed substantial heterogeneity in the qualitative binding patterns of colon cancer cells to platelet P-selectin. Some LS174T or COLO205 cells that tethered from the fluid stream rolled along the P-selectin surface for some distance, which was variable but sometimes quite extended (the entire path of a field of view) at velocities significantly lower than the critical velocity (Table 2). The latter was experimentally calculated to be approximately 700 µm/s at a wall shear stress of 0.8 dyn/cm2 for noninteracting tumor cells traveling adjacent to the lower wall of the flow chamber.42 Other LS174T or COLO205 cells formed only brief adhesive interactions after which they skipped to velocities near the critical value before they tethered again to the substrate downstream to the site of initial contact. In general, COLO205 cells exhibited a rolling velocity that was significantly lower than that of LS174T cells (Table 2), a finding that correlates inversely with the sLex content of these cell lines.
Both the pattern of interaction and magnitude of rolling velocities of LS174T and COLO205 cells are remarkably different from those of neutrophils isolated with standard procedures.43 The latter rolled stably along the P-selectin substrates with an average velocity of approximately 4.0 µm/s, which is substantially lower than that of tumor cells (approximately 50-100 µm/s; Table 2).30 These data provide evidence that the P-selectin ligand activity on the colon cancer cells appears to be less efficient and more sensitive to shear stress than the PSGL-1 on neutrophils at mediating rolling interactions with P-selectin substrates.
To the best of our knowledge, this is the first study to
demonstrate that surface-bound platelets are capable of mediating colon
cancer cell tethering, rolling, and firm adhesion in shear flow. These
observations suggest that tumor cell recruitment to sites of platelet
deposition may follow a multistep sequential process of adhesive
interactions similar to that outlined for neutrophils.20-23 However, a detailed analysis of LS174T
cell binding to immobilized platelets has revealed several similarities
and disparities to the "neutrophil model." In concert with this
model, platelet P-selectin is required for the efficient capture of
free-flowing LS174T cells (Figure 7). Its
role was established by the ability of function-blocking
anti-P-selectin mAbs to dramatically reduce tumor cell binding to
surface-anchored platelets under flow conditions.
Previous work has demonstrated that P-selectin, expressed either on the
surface of activated endothelial cells or platelets, mediates tethering
and rolling of leukocytes.12,18,19 However, LS174T cells
that tethered to immobilized platelets were instantly arrested with no
obvious period of rolling. We therefore hypothesized that platelet
P-selectin may cooperate with another receptor to stabilize LS174T
cell-platelet interactions. The
The We next explored whether LS174T cell adhesion to immobilized platelets
involves a direct interaction between the 2 cell types or whether
bridging ligands are required. We hypothesized that the platelet
integrin We also aimed to identify the P-selectin ligand on the LS174T
cell surface. Prior work indicates that PSGL-1, expressed on the
neutrophil surface, is the primary ligand for platelet P-selectin under
hydrodynamic flow conditions.43 Using flow cytometry, we
found that PSGL-1 may be marginally expressed on the LS174T cell
surface. Blocking its function with a mAb failed to inhibit the extent
of LS174T cell tethering on the
Recent studies have identified CD24, a mucin-type GPI-linked cell surface glycoprotein, as a ligand for P-selectin.33 Its physiologic relevance became evident by flow experiments showing that CD24 mediates rolling of PSGL-1-negative breast tumor cells to purified P-selectin under flow.42 Using flow cytometry, we documented the presence of CD24 on the LS174T cell surface. To assess its involvement in tumor cell binding to platelet P-selectin, and in the absence of any blocking anti-CD24 mAbs, LS174T cells were treated with PI-PLC, an enzyme that specifically cleaves GPI-anchored molecules from the cell surface. Although this enzyme treatment abolished CD24 expression from the LS174T cell surface, it did not affect tumor cell tethering/rolling on platelet P-selectin. In contrast, treatment of LS174T cells with neuraminidase abrogated the divalent cation-dependent tethering/rolling events. These data collectively suggest that the P-selectin ligand on the LS174T cell surface is a sialylated molecule that is distinct from PSGL-1 and CD24. We next sought to identify a potential counter-receptor on LS174T cells
for the platelet integrin subunit As summarized in Figure 7, our data provide evidence that LS174T
cell recruitment to sites of platelet deposition follows a cascade of
events that shares common features but is distinct from that outlined
for neutrophils.20-23 Platelet P-selectin supports LS174T
cell tethering/rolling by binding to a sialylated molecule that is
distinct from PSGL-1 and CD24. Subsequent platelet
The authors wish to thank Dr William R. Bell (Johns Hopkins University, Baltimore) and his patient for providing us with a blood specimen; Dr John Hemperly (Becton Dickinson, Research Triangle, NC) for supplying us with an anti-L1 polyclonal antibody; Dr Ellen L. Berg (Protein Design Labs, Mountain View, CA) for her generous donation of EP5C7; Dr Bruce S. Bochner (Johns Hopkins University, Baltimore) for insightful comments and reagent donation; and Dr Ronald L. Schnaar (Johns Hopkins University, Baltimore) for helpful discussions.
Submitted October 19, 1999; accepted April 26, 2000.
Supported by a Whitaker Foundation grant (K.K.), a DuPont Young Professor grant (K.K.), and a National Institutes of Health grant HL58564 (P.F.B.).
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734.
Reprints: Konstantinos Konstantopoulos, Department of Chemical Engineering, Johns Hopkins University, 3400 North Charles St, Baltimore, MD 21218-2694; e-mail: konst_k{at}jhu.edu.
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  J. Du, M A. Meledeo, Z. Wang, H. S Khanna, V. D P Paruchuri, and K. J Yarema Metabolic glycoengineering: Sialic acid and beyond Glycobiology, December 1, 2009; 19(12): 1382 - 1401. [Abstract] [Full Text] [PDF]
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 S. N. Thomas, R. L. Schnaar, and K. Konstantopoulos Podocalyxin-like protein is an E-/L-selectin ligand on colon carcinoma cells: comparative biochemical properties of selectin ligands in host and tumor cells Am J Physiol Cell Physiol, March 1, 2009; 296(3): C505 - C513. [Abstract] [Full Text] [PDF]
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 L. Nimrichter, M. M. Burdick, K. Aoki, W. Laroy, M. A. Fierro, S. A. Hudson, C. E. Von Seggern, R. J. Cotter, B. S. Bochner, M. Tiemeyer, et al. E-selectin receptors on human leukocytes Blood, November 1, 2008; 112(9): 3744 - 3752. [Abstract] [Full Text] [PDF]
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S. N. Thomas, F. Zhu, R. L. Schnaar, C. S. Alves, and K. Konstantopoulos Carcinoembryonic Antigen and CD44 Variant Isoforms Cooperate to Mediate Colon Carcinoma Cell Adhesion to E- and L-selectin in Shear Flow J. Biol. Chem., June 6, 2008; 283(23): 15647 - 15655. [Abstract] [Full Text] [PDF]
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C. S. Alves, M. M. Burdick, S. N. Thomas, P. Pawar, and K. Konstantopoulos The dual role of CD44 as a functional P-selectin ligand and fibrin receptor in colon carcinoma cell adhesion Am J Physiol Cell Physiol, April 1, 2008; 294(4): C907 - C916. [Abstract] [Full Text] [PDF]
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S. L. Napier, Z. R. Healy, R. L. Schnaar, and K. Konstantopoulos Selectin Ligand Expression Regulates the Initial Vascular Interactions of Colon Carcinoma Cells: THE ROLES OF CD44V AND ALTERNATIVE SIALOFUCOSYLATED SELECTIN LIGANDS J. Biol. Chem., February 9, 2007; 282(6): 3433 - 3441. [Abstract] [Full Text] [PDF]
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 C. Medina, P. Jurasz, M. J. Santos-Martinez, S. S. Jeong, T. Mitsky, R. Chen, and M. W. Radomski Platelet Aggregation-Induced by Caco-2 Cells: Regulation by Matrix Metalloproteinase-2 and Adenosine Diphosphate J. Pharmacol. Exp. Ther., May 1, 2006; 317(2): 739 - 745. [Abstract] [Full Text] [PDF]
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 L. H. K. Lim, M. M. Burdick, S. A. Hudson, F. B. Mustafa, K. Konstantopoulos, and B. S. Bochner Stimulation of Human Endothelium with IL-3 Induces Selective Basophil Accumulation In Vitro J. Immunol., May 1, 2006; 176(9): 5346 - 5353. [Abstract] [Full Text] [PDF]
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M. M. Burdick and K. Konstantopoulos Platelet-induced enhancement of LS174T colon carcinoma and THP-1 monocytoid cell adhesion to vascular endothelium under flow Am J Physiol Cell Physiol, August 1, 2004; 287(2): C539 - C547. [Abstract] [Full Text] [PDF]
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M. Wei, G. Tai, Y. Gao, N. Li, B. Huang, Y. Zhou, S. Hao, and X. Zeng Modified Heparin Inhibits P-selectin-mediated Cell Adhesion of Human Colon Carcinoma Cells to Immobilized Platelets under Dynamic Flow Conditions J. Biol. Chem., July 9, 2004; 279(28): 29202 - 29210. [Abstract] [Full Text] [PDF]
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O. J. T. McCarty, N. Tien, B. S. Bochner, and K. Konstantopoulos Exogenous eosinophil activation converts PSGL-1-dependent binding to CD18-dependent stable adhesion to platelets in shear flow Am J Physiol Cell Physiol, May 1, 2003; 284(5): C1223 - C1234. [Abstract] [Full Text] [PDF]
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M. M. Burdick, J. M. McCaffery, Y. S. Kim, B. S. Bochner, and K. Konstantopoulos Colon carcinoma cell glycolipids, integrins, and other glycoproteins mediate adhesion to HUVECs under flow Am J Physiol Cell Physiol, April 1, 2003; 284(4): C977 - C987. [Abstract] [Full Text] [PDF]
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W. Hanley, O. McCarty, S. Jadhav, Y. Tseng, D. Wirtz, and K. Konstantopoulos Single Molecule Characterization of P-selectin/Ligand Binding J. Biol. Chem., March 14, 2003; 278(12): 10556 - 10561. [Abstract] [Full Text] [PDF]
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S. K. Khaldoyanidi, V. V. Glinsky, L. Sikora, A. B. Glinskii, V. V. Mossine, T. P. Quinn, G. V. Glinsky, and P. Sriramarao MDA-MB-435 Human Breast Carcinoma Cell Homo- and Heterotypic Adhesion under Flow Conditions Is Mediated in Part by Thomsen-Friedenreich Antigen-Galectin-3 Interactions J. Biol. Chem., January 31, 2003; 278(6): 4127 - 4134. [Abstract] [Full Text] [PDF]
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S. Jadhav and K. Konstantopoulos Fluid shear- and time-dependent modulation of molecular interactions between PMNs and colon carcinomas Am J Physiol Cell Physiol, October 1, 2002; 283(4): C1133 - C1143. [Abstract] [Full Text] [PDF]
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J. Pilch, R. Habermann, and B. Felding-Habermann Unique Ability of Integrin alpha vbeta 3 to Support Tumor Cell Arrest under Dynamic Flow Conditions J. Biol. Chem., June 7, 2002; 277(24): 21930 - 21938. [Abstract] [Full Text] [PDF]
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S. Jadhav, B. S. Bochner, and K. Konstantopoulos Hydrodynamic Shear Regulates the Kinetics and Receptor Specificity of Polymorphonuclear Leukocyte-Colon Carcinoma Cell Adhesive Interactions J. Immunol., November 15, 2001; 167(10): 5986 - 5993. [Abstract] [Full Text] [PDF]
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 J. P. Abulencia, N. Tien, O. J. T. McCarty, D. Plymire, S. A. Mousa, and K. Konstantopoulos Comparative Antiplatelet Efficacy of a Novel, Nonpeptide GPIIb/IIIa Antagonist (XV454) and Abciximab (c7E3) in Flow Models of Thrombosis Arterioscler Thromb Vasc Biol, January 1, 2001; 21(1): 149 - 156. [Abstract] [Full Text] [PDF]
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Top
Abstract
Introduction
effects of antibodies and
peptides.
Closed bars represent the stable adherent cells (stationary for > 10 seconds) counted at the end of the 10-minute experiment, whereas
open bars represent the total interacting cells counted throughout the
entire 10-minute experiment. Immobilized platelets were treated with
antiplatelet agents during the 10-minute thrombin incubation.
Saturating concentrations were also maintained in the flow buffer only
for the GRGDSP peptide, anti-vWf polyclonal antibody, and
antifibrinogen (NYB-4.2) mAb. HIP8 (blocking
