© 1999 American Heart Association, Inc.
Original Contributions |
Adhesive Interaction Between P-Selectin and Sialyl Lewisx Plays an Important Role in Recurrent Coronary Arterial Thrombosis in Dogs
From the Department of Internal Medicine III (H.I., T.U., T.M., H.Y., N.H., H.E., A.K., Y.T., I.O., T.U., T.I.) Kurume University School of Medicine, Kurume, Japan; and Sumitomo Pharmaceuticals Research Center (S.J.T.), Osaka, Japan.
Correspondence to Hisao Ikeda, MD, PhD, Department of Internal Medicine III, Kurume University School of Medicine, 67 Asahi-machi, Kurume 830, Japan. E-mail ikeikeda{at}med.kurume-u.ac.jp
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Abstract |
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Abstract—Cell adhesion molecules may play an important role in the disease process of acute coronary syndromes. We have shown a neutralizing anti-P-selectin monoclonal antibody and a sialyl Lewisx-containing oligosaccharide (SLex-OS), an analogue of selectin ligand on leukocytes, reduce cyclic flow variations (CFVs) in a canine model of recurrent coronary arterial thrombosis, suggesting the important interaction between P-selectin and SLex for the pathophysiology of these syndromes. However, the functional role of these adhesion molecules in the thrombotic process remains unclear. Therefore, we investigated effects of SLex-OS on CFVs, platelet P-selectin expression, and morphology of the stenotic site in the same model. Anesthetized open-chest dogs (n=34) were randomly divided into 4 groups after developing CFVs. Dogs intravenously received saline or graded doses of SLex-OS (5, 20, or 40 mg/kg bolus) infusion followed by a continuous infusion (5 mg · kg-1 · h-1) for 60 minutes. By flow cytometric analysis, P-selectin expression on platelets after CFVs was significantly upregulated during CFVs. Immunohistochemical analysis revealed the incorporation of platelets with upregulated P-selectin within thrombi at the stenotic site. Microscopic observations revealed the presence of numerous platelets adhered to leukocytes at the stenotic site on the damaged endothelium. SLex-OS significantly reduced CFVs, inhibited the P-selectin expression on platelets, and prevented the adherence of platelets and leukocytes. These findings further support the notion that the adhesive interaction between P-selectin on platelets and SLex on leukocytes plays an important role in platelet-mediated thrombus formation in this model.
Key Words: thrombosis • platelets • leukocytes • P-selectin • sialyl Lewisx
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Introduction |
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P-selectin, a member of the selectin family adhesion molecules, is an integral membrane glycoprotein located in both
-granules of platelets1 and the Weibel-Palade
bodies of endothelial cells.2 3 On
stimulation of these cells by agonists such as thrombin,4
histamine,5 or oxygen free radicals,6
P-selectin is rapidly translocated onto the cell surface within minutes
and adheres to a sialylated carbohydrate structure, sialyl
Lewisx (SLex), on
leukocytes through a calcium-dependent mechanism.7 8 A
previous study has shown that activated platelets enhance
extracellular oxygen free radical generation by leukocytes through
P-selectin,9 and that P-selectin mediates "rolling" of
leukocytes on the endothelium10 and
activated platelets.11 Furthermore,
analysis of complementary DNA demonstrated the presence of a
circulating form of P-selectin that possesses a deleted transmembrane
segment by alternative splicing of mRNA.12 13 This soluble
form of P-selectin is shown to inhibit leukocyte ß2-integrin
adhesion, and thus may protect against thrombosis and inflammatory
reactions.14 Therefore, P-selectin and
SLex are important adhesion molecules that
initially mediate the cellular interaction of platelets or
endothelial cells with leukocytes. Platelet-mediated thrombus formation secondary to plaque disruption at the site of atherosclerotic lesion plays a fundamental role in the development of acute coronary syndromes including unstable angina and acute myocardial infarction.15 16 The process of these syndromes may be mediated by a complex cascade of cellular interplay among platelets, endothelial cells, and leukocytes through various adhesion molecules.17 18 Using an experimental canine model with cyclic flow variations (CFVs), which is a well established experimental canine model of recurrent coronary arterial thrombosis,16 19 20 21 we have recently shown that the combination of neutralizing anti-P-selectin monoclonal antibody (PB1.3) and carbohydrate analogue of SLex (SLex-OS) significantly reduces CFVs,22 suggesting that the adhesive interaction between P-selectin and SLex may be involved in mediation of thrombus formation in vivo. However, we had no evidence as to (1) whether P-selectin expression is upregulated on the surface of platelets during CFVs, (2) P-selectin expression is indeed observed within the thrombi at the stenotic site, (3) whether platelets and leukocytes adhere to the injured endothelium at the stenotic site, and (4) whether treatment with SLex-OS dose-dependently reduces CFVs and inhibits P-selectin expression on platelets during the episode of CFVs. In the present study, to further investigate the pathophysiology of the acute coronary syndromes, we tested the above issues by examining the expression of P-selectin on platelets by flow cytometry and by immunohistochemical staining in control dogs with CFVs and SLex-OS treated dogs. SLex-OS was used to investigate the functional role of the adhesive interaction between platelet P-selectin and leukocyte SLex. By electron microscopy, we also examined whether both platelets and leukocytes adhere to the coronary lumen at the stenotic site.
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Methods |
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Surgical Preparation
All experiments were conducted in accordance with the guidelines for animal experimentation of the Animal Research Committee of the Kurume University School of Medicine. Healthy mongrel dogs (15 to 20 kg), anesthetized with pentobarbital sodium (30 mg/kg), were mechanically ventilated. A micromanometer was placed in the femoral artery to monitor the arterial pressure. A thoracotomy was performed in the fifth left intercostal space and the heart was suspended in the pericardial cradle. Polyethylene catheters were placed in the right atrium for drug infusion and blood sampling. A segment of the left anterior descending (LAD) coronary artery was gently dissected free from the surrounding tissue and a pulse Doppler flow probe (Hartley Instruments) was placed proximal to the constricting cylinder. Coronary blood flow (CBF) velocity was measured with a pulsed Doppler flow system (Model VF-1, Crystal Biotech). Arterial blood gasses and body temperature were maintained within normal physiological ranges. Before the coronary arterial constriction, dogs were allowed to stabilize for 30 minutes. Heart rate, systolic and diastolic blood pressures, and phasic and mean CBF velocities were continuously recorded throughout the study. After baseline measurements were obtained, the endothelium of the exposed LAD coronary artery was injured by gently squeezing the artery with cushioned forceps. A cylindrical constrictor was then placed around the injured coronary artery distal to the flow probe to reduce the phasic CBF velocity to approximately 40% of the baseline level in order to eliminate reactive hyperemia after 15 seconds of temporary coronary occlusion. Subsequently, CFVs developed in 40 out of 56 dogs.
Experimental Protocol
The CBF velocity was monitored for 60 minutes to obtain the
baseline value, and drugs were administered intravenously
according to the following protocols. Dogs (n=34) were divided into 4
treatment groups. The control group (n=8) received a bolus of saline
followed by a continuous infusion of saline (1 mL/h). The
SLex-OS treated groups received a bolus of 5
mg/kg (n=9), 20 mg/kg (n=9), or 40 mg/kg (n=9) of
SLex-OS followed by a continuous infusion (5
mg · kg-1 ·
h-1) for 60 minutes (the generous supply of
SLex-OS was from Sumitomo Pharmaceutical,
Osaka). To assess the effect of treatments, the severity of CFVs
was evaluated by monitoring mean coronary blood flow (mL/min),
phasic and mean nadir CBF velocities (% control), and the frequency
(cycles/h) for 60 minutes before and after the treatments. CBF was
determined as described previously.21 22 Briefly, CBF
velocity near the center of the vessel was recorded by utilizing
the pulsed Doppler principle; CBF velocity was calculated by a
digital planimeter. The cross-sectional area of the vessel was
approximated to an inside diameter of the Doppler flow probe,
ranging in size from 2.0 to 2.5 mm. Mean CBF was then derived by
multiplying mean CBF velocity by the cross-sectional area. The peak and
nadir flow velocities in both phasic and mean CBF were expressed as a
percentage of unconstricted CBF velocity (control). The nadir flow
velocity was calculated by averaging the 3 lowest flow velocities
before and after treatments.22 In dogs that exhibited only
2 flow restorations after the treatment, nadir flow velocity was
calculated by averaging the 2.
Flow Cytometric Analysis
We first evaluated cross-reactivity of CRC81 (Biodesign
International), a mouse monoclonal antibody directed against human
P-selectin, to freshly isolated dog platelets. Platelet
immunostaining was performed as previously
described.23 24 25 Briefly, blood samples (2 mL) were drawn
from the right atrium of 6 pentobarbital-anesthetized dogs into
collecting tubes containing 7.5% EDTA. One mL of blood was stimulated
with thrombin (0.5 U/mL) for 5 minutes at room temperature. A 100-µL
aliquot of activated blood was immediately fixed with 1%
paraformaldehyde (1 mL) for 3 hours (4°C) and
platelet pellets were obtained by centrifuge at
1200g for 5 minutes at room temperature. The platelet
pellets were washed with PBS containing 0.1% sodium azide
(PBS/NaN3), then resuspended in PBS containing
0.1% sodium azide and 2% calf serum (PBS/NaN3/CS). An additional
100-µL aliquot of blood was treated in a similar manner, without the
thrombin treatment, as an inactivated control. The
platelets were incubated with the primary antibody (CRC81; 5
µg/mL) or a nonspecific mouse IgG1 (0.1 mL; 5
µg/mL) for 20 minutes. The platelets were then washed in
PBS/NaN3 to remove any excess of the unbound
antibodies. A secondary antibody (0.1 mL; final concentration 5
µg/mL), FITC-conjugated goat anti-mouse IgG (TAGO Co; Cat #4349), was
added to the pellet for 20 minutes. After incubation with the secondary
antibody, the pellet was washed and suspended with
PBS/NaN3 and then analyzed by flow
cytometry (FACScan, Becton Dickinson). At least 10 000 platelets
were counted, gated using the same procedure except using an anti-CD61
(glycoprotein IIb/IIIa) monoclonal antibody. The results
were expressed as the percentage of specific FITC-positive
platelets. Aliquots of dog platelets (n=7) incubated with the
control IgG1 were 2.6±0.8% positive. In
contrast, addition of thrombin (0.5 U/mL) to dog platelets
increased the percent of positive cells to CRC81 (42.4±4.5%). Binding
of CRC81 to nonthrombin activated platelets was minimal
(2.2±0.7%) and not significantly different from mouse IgG1 controls.
Thus, CRC81 does react with thrombin-activated dog
platelets. A representative histogram of the CRC81
binding to thrombin-stimulated dog platelets is shown in Figure 1
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To further examine P-selectin expression on platelets before and after CFVs, and the effect of saline (n=5) or SLex-OS (n=6) on P-selectin expression on platelets, flow cytometric analysis was performed as described above.
Morphological and Immunohistochemical Studies
To assess the morphological changes of the coronary
arteries before and after treatment with SLex-OS,
an additional 3 dogs were rendered to develop CFVs. After rapid removal
of the heart, a catheter was placed into the left coronary
ostium. Then, 2% glutaraldehyde in PBS was perfused
through the catheter at 100 mm Hg pressure for 10 minutes. The
LAD coronary arteries were carefully dissected and the
constrictor was removed. The segments were longitudinally dissected and
visualized. The specimens were incubated in the same fixation buffer
for 2 hours, and rinsed with 0.1 mol/L cacodylated buffer containing
0.1 mol/L sucrose for 12 hours. The specimens were immersed in 1%
osmium tetroxide for 1 hour, dehydrated in a series of graded
concentrations of cold ethanol, dried by the critical-point drying
method, mounted on silver blocks, coated with about 10 nm of gold, and
observed under a scanning electron microscope (S-800, Hitachi) at 20
KV.
Immunohistochemical study of the coronary stenotic lesion was performed using CRC81. An additional 3 dogs were rendered to develop CFVs for this purpose. After the hearts were rapidly removed, the LAD coronary arteries were carefully dissected. The isolated coronary arteries were embedded in optimal cutting temperature compound and snap frozen by liquid nitrogen. Serial 4-µm-thick sections were adhered to poly-L-lysine-coated slides and fixed in cold acetone for 10 minutes. The labeled streptavidin-biotin method was used for immunohistochemical staining (DAKO LSAB kit). Briefly, specimens were treated with 3% hydrogen peroxide for 5 minutes to inhibit endogenous peroxidase activity, then incubated with 1% BSA. Next, they were incubated with 10 µg/mL of CRC81 or a similar amount of nonimmune mouse IgG (control) for 1 hour at room temperature. After washing 3 times in PBS (pH 7.4), biotinylated anti-mouse IgG secondary antibody was applied, followed by peroxidase-labeled streptavidin. Bound peroxidase activity was visualized with 3-amino-9-ethylcarbazole, and the sections were faintly counterstained with Mayer's hematoxylin.
Statistical Analysis
Values are presented as means±SEM. Repeated measures of
ANOVA with a post hoc Scheffé's test were applied for multiple
comparisons. Differences were considered statistically significant at
P<0.05.
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Results |
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The hemodynamic data from the 4 treatment groups of dogs are shown in the Table
, and
Figures 2 and 3.
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Before Development of CFVs (Stenosis in Table
)
Endothelial injury and coronary
constriction decreased the averaged peak phasic CBF velocity to 39% to
43% of baseline (control) and mean CBF velocity to 48% to 51% of
baseline (control). Heart rate, aortic pressure, and peak phasic and
mean flow velocities were comparable among the 4 groups. These data are
consistent with those reported by others26
and us.22
After Developing CFVs and Before Treatment (60-minute CFVs in
Table)
No significant changes were observed in heart rates or
systolic and diastolic aortic blood pressures after
the development of CFVs. The peak phasic and mean CBF velocities were
similarly decreased among the 4 groups. The phasic coronary
nadir flow velocity was decreased to 7% to 8% of control and mean
coronary nadir flow velocity decreased to 11% to 12% of
control; these values were not significantly different among the
groups. The frequency and the mean CBF of CFVs was 8.3 to 8.5
cycles/h and 6.6 to 6.9 mL/min, respectively (Figure 3). These
values were also comparable among the groups. Thus, the severity of
CFVs was comparable among the groups. These data are in agreement with
those reported by others26 and us.22
Effects of Treatments on CFVs (After Treatment in Table)
The effects of saline and SLex-OS on
hemodynamics and severity of CFVs are shown in Table,
and Figures 2 and 3. There were no significant effects of
treatments on heart rate and aortic pressure in the 4 groups. Treatment
with saline or SLex-OS (a bolus of 5 mg/kg) did
not cause a significant change in the nadir CBF velocity, nor the
frequency or the mean CBF of CFVs. Treatment with
SLex-OS (a bolus of 20 mg/kg) did not increase
the nadir CBF velocity (Table), but significantly decreased the
frequency of CFVs (P<0.05) and significantly increased the
mean CBF (P<0.05). Treatment with
SLex-OS (a bolus of 40 mg/kg) significantly
increased the nadir CBF velocity (P<0.05) and the mean CBF
(P<0.05), and significantly decreased the frequency of CFVs
(P<0.05). Thus, there were dose-dependent effects of
SLex-OS on CFVs.
Expression of P-Selectin on Platelets and Immunohistochemical
Localization of P-selectin in Thrombi
Flow cytometric detection of bound CRC81, a monoclonal antibody
against P-selectin during the episode of CFVs is shown in Figure 4. The upper panel demonstrates the
expression of P-selectin on platelets in a control dog with CFVs,
and the lower panel in a SLex-OS treated dog.
Summarized data are shown in Figure 5.
The P-selectin expressions on platelets before treatments (baseline
and stenosis) were similar between the saline and
SLex-OS treated groups. After development of
CFVs, the extent of P-selectin expression similarly and significantly
increased in both groups (P<0.05). Although treatment with
saline did not affect P-selectin expression, treatment with
SLex-OS significantly decreased P-selectin
expression (P<0.05). The immunohistochemical staining of
P-selectin within thrombi at the stenotic site was intense, but
the expression of P-selectin on the damaged endothelium
was undetectable (Figure 6).
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P<0.05 compared with
stenosis.
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Morphology of Coronary Arteries
Scanning electron photomicrograms are illustrated in Figure 7. Figure 7A demonstrates the left
circumflex coronary artery with the intact
endothelium. Numerous platelets and leukocytes were
observed at the stenotic site with the mechanically damaged
endothelium (Figure 7B). After treatment with
SLex-OS, leukocytes no longer attached and a few
platelets were observed at the stenotic site (Figure 7C).
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Discussion |
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In the present study, the important findings were as follows: (1) The surface expression of P-selectin on platelets was significantly upregulated after developing CFVs. (2) Immunohistochemical analysis showed the upregulated P-selectin expression within thrombi at the coronary stenotic site with the damaged endothelium. (3) Microscopic observations revealed the presence of numerous platelets with leukocytes at the stenotic site. (4) Treatments with SLex-OS, a unique soluble carbohydrate analogue of SLex, significantly reduced CFVs in a dose-dependent manner, suppressed P-selectin expression of platelets, and inhibited the attachment of platelets and leukocytes on the damaged endothelium. Thus, these immunohistochemical and morphological findings suggest that the adhesive interaction between P-selectin on platelets and SLex on leukocytes plays an important role in mediating platelet-mediated thrombus formation in the stenosed and endothelium-injured canine coronary arteries.
We have previously reported that conscious dogs with CFVs manifest a pathophysiology similar to acute coronary syndromes including unstable angina, acute myocardial infarction, and ischemic sudden death in humans.21 Episodes of CFVs have been observed during coronary angioplasty in some patients with unstable angina.27 As originally described by Folts et al,19 CFVs are caused by brief and repeated episodes of coronary occlusion secondary to focal platelet aggregation and subsequent dislodgment at the stenotic site of the coronary artery.16 In the present study, we used this canine model of coronary arterial thrombosis to examine the role of the adhesion molecules in the thrombotic process in vivo. We have reported that soluble P-selectin is increased in blood in patients with acute coronary syndrome,28 29 and that the combination of neutralizing anti-P-selectin monoclonal antibody (PB1.3) and carbohydrate analogue of SLex (SLex-OS) significantly reduces CFVs.22 These results suggest that the adhesive interaction between P-selectin and SLex may be involved in thrombus formation in vivo. In the present study, we demonstrated not only the upregulation of platelet P-selectin expression after developing CFVs by flow cytometric analysis, but also the incorporation of platelets with upregulated P-selectin expression into thrombi at the coronary stenotic site. Moreover, in the present study, scanning electron micrographic observations revealed the presence of numerous platelets with leukocytes on the damaged endothelium at the stenotic site. Our findings are additive to the results of previous clinical studies demonstrating the upregulation of platelet P-selectin expression in patients with acute coronary syndromes,30 subacute occlusive coronary stent thrombosis,31 and primary antiphospholipid syndrome,32 and are consistent with the results of a previous experimental study showing positive immunostaining for P-selectin within the thrombus in a primate model of femoral arterial thrombosis.33 These findings further support the functional role of platelet P-selectin interaction with leukocyte SLex during thrombus formation of CFVs in this model.
To further investigate the role of adhesive interaction between P-selectin and SLex in the thrombotic process of CFVs, effects of SLex-OS an unique soluble carbohydrate analogue of SLex, were examined on CFVs and platelet P-selectin expression. SLex functions as a ligand for selectins.7 8 Soluble SLex-OS is thus assumed to compete with native SLex expressed on leukocytes and inhibit the interaction between platelets and leukocytes.34 Recently, we have shown that a high dose of SLex-OS (40 mg/kg) significantly reduced CFVs in dogs.22 However, the dose-dependent effects of SLex-OS on CFVs were not examined in our previous study. In the present study, SLex-OS reduced CFVs in a dose-dependent manner and decreased the expression of P-selectin on platelets. Moreover, the scanning electronmicrogram demonstrated that SLex-OS prevented attachments of leukocytes and platelets onto the damaged endothelium. Thus, these findings suggest that blocking the function of leukocyte SLex by SLex-OS downregulates the expression of P-selectin on platelets, inhibits the interaction among platelets, leukocytes and the endothelium, and thus prevents the formation of thrombi at the stenotic site.
Coronary thrombus formation at the site of culprit lesion plays an important role in the development of acute coronary syndromes including unstable angina and acute myocardial infarction.15 16 The process of these syndromes may be mediated by a complex cascade of cellular interplay among platelets, endothelial cells, and leukocytes through various adhesion molecules such as integrins, selectins, and immunoglobulin superfamilies.17 18 Among these adhesion molecules, P-selectin is rapidly translocated to cell surfaces on platelets or endothelial cells and adheres to a sialylated fucosylated carbohydrate structure such as SLex on leukocytes,7 8 when these cells are activated by thrombin4 or oxygen free radicals.6 Previous studies have indicated that activated platelets enhance extracellular oxygen free radical generation by leukocytes through P-selectin,9 and that P-selectin mediates "rolling" of leukocytes on the endothelium10 and activated platelets.11 In animal models of myocardial reperfusion injury, either monoclonal antibody to P-selectin35 36 or SLex-OS34 37 protected against reperfusion-induced endothelial and myocardial injuries. Thus, both in vitro and in vivo experimental studies have shown that the adhesive interaction between P-selectin and SLex may have an active role in modulating vascular and tissue injuries. However, the functional role of P-selectin in the coronary thrombus formation has been fully unknown in vivo. In the previous study, Toombs et al33 demonstrated that occlusive thrombi formed in the presence of P-selectin antagonism lyse more rapidly in the presence of pharmacological thrombolysis in a primate model of electrically induced femoral arterial thrombosis, which is characterized by persistent fibrin-rich, platelet-poor thrombi. These results are suggestive of the role of P-selectin in stabilizing fibrin-rich, platelet-poor thrombi. The present model of cyclic flow variations is characterized by recurrent platelet-rich thrombi.16 19 20 Thus, the present study focused on the role of P-selectin in platelet adhesion and aggregation in vivo. Based on the results from this and the previous study, the adhesive interaction between P-selectin on platelets and SLex on leukocytes is shown to be an important regulator of not only platelet adhesion and aggregation but also the cascade of coagulation and fibrinolysis in vivo.
The present study may provide important clinical implications. Increases in the surface expression of P-selectin on platelets have been shown in patients with acute coronary syndromes.30 Recently, we have shown that leukocyte adherence to the endothelium via P-selectin plays an important role in injuries of the endothelium of the coronary artery distal to the thrombotic site in vivo in dogs,38 and that treatment with SLex-OS prevented endothelial injuries distal to the thrombotic site.38 Taken together with the findings of the present study, it is likely that SLex-OS not only prevents thrombus formation at the stenotic site but also preserves endothelial function of the artery distal to the thrombotic site. Because the small sugar moiety of SLex-OS has low antigenicity when used in vivo and has a potential efficacy with oral administration,39 SLex-OS could become an attractive therapeutic modality of acute coronary syndromes in humans.
The present study may have some limitations. We did not examine the expression of P-selectin on platelets after small doses of SLex-OS, and thus the exact relationship between the expression of P-selectin on platelets and the severity of CFVs was undetermined. We did not examine which mechanisms induce the P-selectin expression on platelets in the current model. P-selectin is rapidly translocated onto the cell surface after stimulation with thrombin4 and/or oxygen free radicals.6 Thrombin40 and oxygen free radicals41 42 43 are important mediators of CFVs. Accordingly, it is possible that these agonists are generated by mechanical manipulations (endothelial damage and constriction) and induce P-selectin expression on platelets in vivo.
In conclusion, the present study is the first demonstration that the adhesive interaction between P-selectin on platelets and SLex on leukocytes plays an important role in platelet-mediated thrombus formation in stenosed and endothelium-injured canine coronary arteries in vivo. SLex-OS as used in this study may provide the salutary effects against acute thrombotic events in the coronary artery in vivo.
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Acknowledgments |
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The authors are grateful to Kimiko Kimura and Aya Shimizu for their excellent technical assistance. This study was supported in part by a grant-in-aid for scientific research (08670836) from the Ministry of Education, Science and Culture, Tokyo and by a research grant from the Foundation for the Advancement of Clinical Medicine, Fukuoka and by a research grant from the Kimura Memorial Heart Foundation, Kurume, Japan.
Received July 7, 1998; accepted October 22, 1998.
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References |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
1. Stenberg PE, McEver RP, Shuman MA, Jacques YV, Bainton DF. A platelet alpha-granule membrane protein (GMP-140) is expressed on the plasma membrane after activation. J Cell Biol. 1985;101:880–886.
2. McEver RP, Beckstead JH, Moore KL, Marshall-Carlson L, Bainton DF. GMP-140, a platelet a-granule membrane protein, is also synthesized by vascular endothelial cells and is localized in Weibel-Palade bodies. J Clin Invest. 1989;84:92–99.
3. Johnston GI, Cook RG, McEver RP. Cloning of GMP-140, a granule membrane protein of platelets and endothelium: sequence similarity to proteins involved in cell adhesion and inflammation. Cell. 1989;56:1033–1044.[Medline] [Order article via Infotrieve]
4.
Johnston GI, Pickett EB, McEver RP, George JN.
Heterogeneity of platelet secretion in response to
thrombin demonstrated by fluorescence flow cytometry.
Blood. 1987;69:1401–1403.
5.
Hattori R, Hamilton KK, Fugate RD, McEver RD, Sims PJ.
Stimulated secretion of endothelial von
Willebrand factor is accompanied by rapid redistribution to the
cell surface of the intracellular granule membrane protein GMP-140.
J Biol Chem. 1989;264:7768–7771.
6.
Patel KD, Zimmerman GA, Prescott SM, McEver RP,
McIntyre TM. Oxygen radicals induce human endothelial
cells to express GMP-140 and bind neutrophils. J Cell
Biol. 1991;112:749–759.
7. Geng J-G, Bevilacqua MP, Moore KL, McIntyre TM, Prescott SM, Kim JM, Bliss GA, Zimmerman GA, McEver RP. Rapid neutrophil adhesion to activated endothelium mediated by GMP-140. Nature. 1990;343:757–760.[Medline] [Order article via Infotrieve]
8.
Polley MJ, Phillips ML, Wayner E, Nudelman E, Singhal
AK, Hakomori S, Paulson JC. CD62 and endothelial
cell-leukocyte adhesion molecule 1 (ELAM-1) recognize the same
carbohydrate ligand, sialyl-Lewis X. Proc Natl Acad Sci
U S A. 1991;88:6224–6228.
9. Nagata K, Tsuji T, Todoroki N, Katagiri Y, Tanoue K, Yamazaki H, Hanai N, Irimura T. Activated platelets induce superoxide anion release by monocytes and neutrophils through P-selectin (CD62). J Immunol. 1993;151:3267–3273.[Abstract]
10. Lawrence MB, Springer TA. Leukocytes roll on a selectin at physiologic flow rates: distinction from and prerequisite for adhesion through integrins. Cell. 1991;65:859–873.[Medline] [Order article via Infotrieve]
11.
Yeo EL, Sheppard JAI, Feuerstein IA. Role of P-selectin
and leukocyte activation in polymorphonuclear cell adhesion to
surface adherent activated platelets under physiologic
shear conditions (an injury vessel wall model). Blood. 1994;83:2498–2507.
12.
Johnston GI, Bliss GA, Newman PJ, McEver RP. Structure
of the human gene encoding granule membrane protein-140, a member of
the selectin family of adhesion receptors for leukocytes. J
Biol Chem. 1990;265:21381–21385.
13.
Ishiwata N, Takio K, Katayama M, Watanabe K, Titani K,
Ikeda Y, Handa M. Alternatively spliced isoform of P-selectin is
present in vivo as soluble molecule. J Biol Chem. 1994;269:23708–23715.
14.
Gamble JR, Skinner MP, Berndt MC, Vadas MA. Prevention
of activated neutrophil adhesion to endothelium
by soluble adhesion protein GMP140. Science. 1990;249:414–417.
15.
Fuster V, Badimon L, Cohen M, Ambrose JA, Badimon JJ,
Chesebro J. Insights into the pathogenesis of acute
ischemic syndromes. Circulation. 1988;77:1213–1220.
16.
Willerson JT, Golino P, Eidt J, Campbel WB, Buja LM.
Specific platelet mediators and unstable coronary artery
lesions: experimental evidence and potential clinical implications.
Circulation. 1989;80:198–205.
17.
Entman ML, Ballantyne CM. Inflammation in acute
coronary syndromes. Circulation. 1993;88:800–803.
18.
Entman ML, Ballantyne CM. Association of neutrophils
with platelet aggregates in unstable angina. should we alter
therapy? Circulation. 1996;94:1206–1208.
19.
Folts JD, Crowell EB, Rowe GG. Platelet aggregation
in partially obstructed vessels and its elimination with aspirin.
Circulation. 1976;54:365–370.
20. Bush LR, Shebuski RJ. In vivo models of arterial thrombosis and thrombolysis. FASEB J. 1990;4:3087–3098.[Abstract]
21. Ikeda H, Koga Y, Kuwano K, Nakayama H, Ueno T, Yoshida N, Adachi K, Park IS, Toshima H. Cyclic flow variations in a conscious dog model of coronary artery stenoses and endothelial injury correlate with acute ischemic heart disease syndromes in humans. J Am Coll Cardiol. 1993;21:1008–1017.[Abstract]
22.
Ueyama T, Ikeda H, Haramaki N, Kuwano K, Imaizumi T.
Effects of monoclonal antibody to P-selectin and analogue of sialyl
Lewis X on cyclic flow variations in stenosed and
endothelium-injured canine coronary arteries.
Circulation. 1997;95:1554–1559.
23.
Ginsberg M, Frelinger A, Lam S, Forsyt J, McMillan R,
Plow E, Shattil S. Analysis of platelet aggregation
disorders based on flow cytometric analysis of platelet
membrane glycoprotein GPIIb-IIIa with conformation specific
monoclonal antibodies. Blood. 1990;76:2017–2023.
24. Gawaz M, Ward R. Effects of hemodialysis on platelet derived thrombospondin. Kidney Int. 1991;40:257–265.[Medline] [Order article via Infotrieve]
25.
Gawaz M, Ott I, Reininger AJ, Heinzmann U, Neumann FJ.
Agglutination of isolated platelet membranes. Arteroscler
Thromb Vasc Biol. 1996;16:621–627.
26.
Ashton JH, Ogletree ML, Michel IM, Golino P, McNatt JM,
Taylor AL, Raheja S, Schmitz J, Buja LM, Campbell WB, Willerson JT.
Cooperative mediation by serotonin S2
and thromboxane
A2/prostaglandin
H2 receptor activation of cyclic flow variations
in dogs with severe coronary artery stenoses.
Circulation. 1987;76:952–959.
27. Eichhorn EJ, Grayburn PA, Willard JE, Anderson HV, Bedotto JB, Carry M, Kahn JK, Willerson JT. Spontaneous alternations in coronary blood flow velocity before and after coronary angioplasty in patients with severe angina. J Am Coll Cardiol. 1991;17:43–52.[Abstract]
28. Ikeda H, Nakayama H, Oda T, Kuwano K, Muraishi A, Sugi K, Koga Y, Toshima H. Soluble form of P-selectin in patients with acute myocardial infarction. Coron Artery Dis. 1994;5:515–518.[Medline] [Order article via Infotrieve]
29.
Ikeda H, Takajo Y, Ichiki K, Ueno T, Maki S, Noda T,
Sugi K, Imaizumi T. Increased soluble form of P-selectin in patients
with unstable angina. Circulation. 1995;92:1693–1696.
30. Ott I, Neumann FJ, Gawaz M, Schmitt M, Schömig A. Increased neutrophil-platelet adhesion in patients with unstable angina. Circulation. 1996;92:1239–1246.
31. Gawaz M, Neumann FJ, Ott I, May A, Rüdiger S, Schömig A. Role of activation-dependent platelet membrane glycoproteins in development of subacute occlusive coronary stent thrombosis. Coron Artery Dis. 1997;8:121–128.[Medline] [Order article via Infotrieve]
32.
Fanelli A, Bergamini C, Rapi S, Caldini A, Spinelli A,
Buggiani A, Emmi L. Flow cytometric detection of circulating
activated platelets in primary antiphospholipid syndrome.
correction with thrombocytopenia and anticardiolipin antibodies.
Lupus. 1997;6:261–267.
33.
Toombs CF, Degraaf GL, Martin JP, Geng JG, Anderson DC,
Shebuski RJ. Pretreatment with a blocking monoclonal antibody to
P-selectin accelerates pharmacological thrombolysis in
a primate model of atrial thrombosis. J Pharmacol Exp
Ther. 1995;275:941–949.
34.
Lefer DJ, Flynn DM, Phillips ML, Ratcliffe M,
Buda AJ. A novel sialyl Lewis X analog attenuates neutrophil
accumulation and myocardial necrosis after ischemia and
reperfusion. Circulation. 1994;90:2390–2401.
35. Weyrich AS, Ma X-L, Lefer DJ, Albertine KH, Lefer AM. In vivo neutralization of P-selectin protects feline heart and endothelium in myocardial ischemia and reperfusion injury. J Clin Invest. 1993;91:2620–2629.
36.
Chen LY, Nichols WW, Hendricks JB, Yang BC, Mehta JL.
Monoclonal antibody to P-selectin (PB1.3) protects against myocardial
reperfusion injury in the dog. Cardiovasc Res. 1994;28:1414–1422.
37. Buerke M, Weyrich AS, Zheng Z, Gaeta FCA, Forrest MJ, Lefer AM. Sialyl Lewis X - containing oligosaccharide attenuates myocardial reperfusion injury in cats. J Clin Invest. 1994;93:1140–1148.
38.
Eguchi H, Ikeda H, Murohara T, Yasukawa H,
Haramaki N, Sakisaka S, Imaizumi T. Endothelial injuries of coronary
arteries distal to thrombotic sites: role of adhesive interaction
between enodthelial P-selectin and leukocyte sialyl LewisX.
Circ Res. 1999;84:525–535.
39. Williams TJ, Hellewell PG. Adhesion molecules involved in the microvascular inflammatory response. Am Rev Respir Dis. 1992;146:S45–S50.[Medline] [Order article via Infotrieve]
40. Eidt JF, Allison P, Noble S, Ashton J, Golino P, McNatt J, Buja LM, Willerson JT. Thrombin is an important mediator of platelet aggregation in stenosed canine coronary arteries with endothelial injury. J Clin Invest. 1989;84:18–27.
41. Ikeda H, Koga Y, Oda T, Kuwano K, Nakayama H, Ueno T, Toshima H, Michael LH, Entman ML. Free oxygen radicals contribute to platelet aggregation and cyclic flow variations in stenosed and endothelium-injured canine coronary arteries. J Am Coll Cardiol. 1994;24:1749–1756.[Abstract]
42.
Kuwano K, Ikeda H, Oda T, Nakayama H, Koga Y,
Toshima H, Imaizumi T. Xanthine oxidase mediates cyclic flow variations
in a canine model of coronary arterial thrombosis.
Am J Physiol. 1996;270:H1993–H1999.
43.
Yao S-K, Ober JC, Gonenne A, Clubb FJJ, Krishnaswami A,
Ferguson JJ, Anderson HV, Gorecki M, Buja LM, Willerson JT. Active
oxygen species play a role in mediating platelet aggregation and
cyclic flow variations in severely stenosed and
endothelium-injured coronary arteries.
Circ Res. 1993;73:952–967.
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