|
|||||||||
|
|
|
|
|
|
|
|
|
|
|
HEMOSTASIS, THROMBOSIS, AND VASCULAR BIOLOGY
From the Second Department of Medicine, Kyoto
Prefectural University of Medicine, Kyoto, Japan.
Serotonin (5-hydroxytryptamine, or 5-HT), released from
activated platelets, not only accelerates aggregation of platelets but
also is known to promote mitosis, migration, and contraction of
vascular smooth muscle cells (VSMCs). These effects are considered to
contribute to thrombus formation and atherosclerosis. The aim of this
study was to investigate the effects of 5-HT on the expressions of
coagulative and fibrinolytic factors in rat aortic endothelial cells.
Endothelial cells were stimulated with various concentrations of 5-HT
(0.1~10 µM), and the expressions of tissue factor (TF), tissue
factor pathway inhibitor (TFPI), plasminogen activator inhibitor-1
(PAI-1), and tissue-type plasminogen activator (TPA) messenger RNAs
(mRNAs) were evaluated by Northern blot analysis. The activities of TF
and PAI-1 were also measured. TF and PAI-1 mRNA were increased
significantly in a concentration- and time-dependent manner. However,
TFPI and TPA mRNA expression did not change. The inductions of TF and
PAI-1 mRNAs were inhibited by a
5-HT1/5-HT2 receptor antagonist
(methiothepin) and a selective 5-HT2A receptor antagonist
(MCI-9042). These results indicate that 5-HT increases procoagulant
activity and reduces fibrinolytic activities of endothelial cells
through the 5-HT2A receptor. It was concluded that the
modulation of procoagulant and hypofibrinolytic activities of
endothelial cells by 5-HT synergistically promotes thrombus formation
at the site of vessel injury with the platelet aggregation, VSMC
contraction, and VSMC proliferation.
(Blood. 2001;97:1697-1702) Serotonin (5-hydroxytryptamine, or 5-HT) is
synthesized and secreted into the blood stream by enterochromaffin
cells in the gastrointestinal tract and is rapidly taken up and stored
in small dense granules in platelets.1 When platelets
adhere and aggregate at a site of vessel injury, 5-HT is secreted and
directly accelerates platelet aggregation and potentiates the response
of platelets to other agonists such as adenosine diphosphate, collagen,
and thromboxane A2.2 5-HT also promotes the
proliferation,3-5 migration,6 and
contraction7-9 of vascular smooth muscle cells (VSMCs). In
addition to physiologic hemostasis, these vascular responses play a
pivotal role in the development and progression of thrombotic
diseases.10,11 Although the effects of 5-HT on platelets
and smooth muscle cells (SMCs) are well documented, little information
on its effect in vascular endothelial cells (ECs) is available,
especially concerning the regulation of coagulation and fibrinolysis.
Vascular ECs produce many modulatory proteins that regulate coagulation
and fibrinolysis, such as tissue factor (TF) and plasminogen activator
inhibitor-1 (PAI-1), which accelerate the development of thrombotic
states.12,13 TF is a 47-kd membrane-bound glycoprotein that plays an important role in the extrinsic pathway of blood coagulation.14 TF present on cell membrane surfaces forms
complexes with coagulation factor VII/VIIa. The TF-VIIa complex
activates circulating factors X and IX by limited proteolysis, leading
to thrombin formation and deposition of insoluble fibrin. The
inhibitory regulator of this pathway is tissue factor pathway inhibitor
(TFPI), a 42-kd plasma glycoprotein previously referred to as
lipoprotein-associated coagulation inhibitor, or extrinsic pathway
inhibitor.15 PAI-1 is a 50-kd glycoprotein and the primary
physiologic inhibitor of plasminogen activation. PAI-1 plays a critical
role in the regulation of fibrinolysis, serving as the primary
inhibitor of tissue-type plasminogen activator (TPA).16
These 4 proteins are involved in the regulation of both coagulation and
fibrinolysis by ECs.17,18
In this study we examined the effects of 5-HT on the expression of TF,
TFPI, PAI-1, and TPA in cultured rat aortic endothelial cells (RAECs).
Cell cultures
Measurements of TF, TFPI, PAI-1, and TPA messenger RNA
expression
Total RNA samples (10 µg) were size fractionated on 1%
agarose/formaldehyde gels and capillary transferred onto nitrocellulose membranes, Hybond-N (Amersham International plc, Buckinghamshire, England). After baking at 80°C for 1 hour, the membranes were prehybridized at 42°C for 2 hours in a mixture of 50% formamide, 4 × sodium chloride and sodium citrate (SSC), 5 × Denhardt
solution, 0.2% SDS, and 100 µg/mL of heat-denatured sonicated salmon
sperm DNA. Membranes were hybridized overnight with specific cDNA
probes at 42°C in a shaker, washed with 2 × SSC-0.1% sodium
dodecyl sulfate (SDS) at room temperature for 10 minutes, air dried,
and exposed to X-OMAT AR film (Eastman Kodak Co, Rochester, NY) at
Measurements of TF and PAI-1 activities TF activity of RAEC surfaces was measured by using a chromogenic assay because TF is a membrane-associated protein.20 After stimulation, RAECs were washed 3 times in Tris-buffered saline (50 mM Tris HCl,120 mM NaCl, 2.7 mM KCl, 3 mg/mL BSA, pH 7.4) and incubated with 1.0 mL prothrombin complex (PPSB-HT: factor II, VII, IX, and X concentrates; Nippon Pharmaceutical Co, Tokyo, Japan) diluted 2-fold with Tris-buffered saline containing 10 mM CaCl2. Samples were incubated for 30 minutes at 37°C, and 200 µL supernatant was added to 100 µL of 2 mM S-2765 (Chromogenix AB, Möndal, Sweden), a chromogenic substrate of factor Xa. After a 15-minute incubation at 25°C, the reaction was stopped by the addition of 200 µL of 50% (vol/vol) acetic acid. The amidolytic activity of generated factor Xa was determined by the absorbance at 405 nm. TF activity is expressed as arbitrary units (AU), using reference curves generated with a standard human placenta-derived TF, Thromborel S (Behringwerke AG, Marburg, Germany) containing 106 AU/mL. The curve was linear from 50 to 1000 AU/mL.For the measurement of PAI-1 concentration in the medium, conditioned
medium was immediately collected and stored at Reagents EC growth supplement was obtained from Becton Dickinson Labware (Bedford, MA). DMEM and heat-inactivated FBS were from Gibco BRL Life Technologies Inc (Grand Island, NY). BSA and antibiotics were purchased from Sigma Chemical (St Louis, MO). 5-HT was obtained from Sigma Chemical Co. MCI-9042, 5-HT2A receptor antagonist, was kindly provided by Tokyo Tanabe Inc (Tokyo, Japan). 5-carboxamidotryptamine maleate (a 5-HT1 receptor agonist), 2-methyl-5-hydroxytryptamine hydrochloride (a 5-HT3 receptor agonist), methiothepin maleate (a 5-HT1/5-HT2 receptor antagonist), and MDL-72222 (a 5-HT3 receptor antagonist) were purchased from Tocris Cookson Inc (Ballwin, MO).Data analysis The intensity of bands of interest from Northern blots was quantified by reflectance densitometry. The TF, TFPI, PAI-1, and TPA mRNA expression was normalized GAPDH mRNA expression. The relative TF, TFPI, PAI-1, and TPA mRNA expression levels are expressed as the fold increase in normalized TF, TFPI, PAI-1, and TPA mRNA expression relative to the expression in vehicle-treated or quiescent RAECs. All values are expressed as the mean ± SEM, for n = 3. Statistical analysis was performed by analysis of variance, and Fisher protected least significant difference was used for individual comparisons. Values of P < .05 were considered significant.
Effects of 5-HT on TF, TFPI, PAI-1, and TPA mRNA expression in RAECs After quiescent RAECs were incubated with vehicle or various concentrations of 5-HT for 2 hours, total RNA was isolated from the RAECs to examine TF and TFPI mRNA expression by Northern blot analysis (Figure 1A). In the RAECs cultured with 5-HT at concentrations 1 µM, the expression of TF mRNA
increased significantly in a concentration-dependent manner, whereas
TFPI mRNA expression did not change relative to
vehicle-treated RAECs.
To investigate the time-dependent effects of 5-HT on TF and TFPI mRNA expression, total RNA was isolated from RAECs stimulated with 10 µM 5-HT for various periods of time (Figure 1B). Northern blot analysis of the isolated RNA shows that TF mRNA induction by 5-HT stimulation began within 1 hour and reached a peak expression after 2 hours. The peak TF mRNA expression was 9.23 ± 0.71-fold higher than the basal level. To characterize PAI-1 and TPA mRNA expression, the dose-dependent and
time-dependent effects of 5-HT were studied as described above for TF
and TFPI mRNA. PAI-1 mRNA expression increased significantly in a
concentration-dependent manner, at concentrations of 5-HT
Effects of 5-HT on the activities of TF on the surface of RAECs and PAI-1 in the culture medium To confirm a relationship between mRNA expression and the activity/amount of protein, the activity of TF and the concentration of PAI-1 were examined (Figure 3). After a 48-hour incubation in conditioned medium, RAECs were stimulated with 5-HT (0.01, 0.1, 1, and 10 µM) for 12 hours. 5-HT stimulation resulted in a significant increase in TF activity on the surface of RAECs in a dose-dependent manner (Figure 3A). The time course for changes in TF activity induced by 10 µM 5-HT was examined. A significant increase in TF activity, compared with vehicle-treated controls, was seen at 4 hours of stimulation with 5-HT, and the maximum TF activity on the cell surface was noted 12 hours after the addition of 5-HT (Figure 3B). To quantify functionally active PAI-1 produced by RAECs, they were stimulated with 5-HT (0.1, 0.3, 1, 3, and 10 µM) for 8 hours after a 48-hour incubation in conditioned medium. A significant increase in the PAI-1 concentration in the culture medium was attained at a 5-HT concentration 3 µM (Figure
4A). In the time course study, a
significant increase in the PAI-1 concentration was observed 8 hours
after stimulation (Figure 4B).
Effects of 5-HT receptor antagonists on 5-HT-induced TF and PAI-1 mRNA expression RAECs were incubated with 5-HT receptor antagonists for 30 minutes before stimulation with 1 µM 5-HT. Total RNA was extracted from the RAECs after 2- or 3-hour incubations with 5-HT. Northern blot analysis revealed that preincubation with methiothepin, a 5-HT1/5-HT2 receptor antagonist, blocked the increase in TF and PAI-1 mRNA expression, at a concentration of 10 2 µM and 104 ~ 102
µM, respectively (Figure 5). MDL-72222,
a 5-HT3 receptor antagonist, had no effect on
5-HT-stimulated TF and PAI-1 mRNA expression in RAECs (data not
shown). Pretreatment with MCI-9042, a 5-HT2A receptor
antagonist, decreased TF and PAI-1 mRNA induction by 5-HT, at
concentrations 1 µM and completely blocked induction of both
at a concentration of 10 µM (Figure 6).
The 3 major factors responsible for thrombus formation are "abnormality of the vascular wall," "decrease in blood flow," and "imbalance of coagulative and fibrinolytic factors," which are known as the Virchow triad.21,22 Injury of the endothelium begins in the first place, which would cause platelets to aggregate and release 5-HT. 5-HT directly accelerates platelet aggregation and also potentiates the response of platelets to other agonists such as adenosine diphosphate, collagen, and thromboxane A2.2 5-HT is known to promote the contraction of VSMCs.7-9 Vikenes et al23 recently reported that elevated concentrations of 5-HT in platelet- rich plasma promote platelet aggregation and VSMC contraction and is associated with the development of coronary artery disease and cardiac events. Vasoconstriction, together with platelet aggregation, would reduce arterial blood flow and would increase the risk of thrombus formation.24 5-HT also promotes the proliferation3-5 and migration of VSMCs6 and may contribute to the progression of atherosclerosis, which is considered to be the major risk factor of arterial thrombotic disease. 5-HT not only interacts synergistically with platelet-derived growth factor4 but also has a direct mitogenic effect on cultured aortic VSMCs.3-5,25,26 It has also been reported that 5-HT stimulates the expression of thrombin receptors in VSMCs probably by 5-HT2 receptors, which would further potentiate the contraction and proliferation of VSMCs by thrombin.27 These effects could cause narrowing of the vessel lumen and contribute to both the development of atherosclerosis and the restenosis observed after angioplasty.11,28,29 In respect to the imbalance of coagulative and fibrinolytic factors, vascular ECs play an important role by producing modulatory proteins such as TF, TFPI, PAI-1, and TPA. Many factors, including vasoactive substances affect this regulation.30-32 Expression and balance of these proteins is known to control the development of thrombosis. Following platelet aggregation, the activation of coagulation cascade would result in thrombin activation and fibrin formation, which is known as secondary hemostasis. When platelets are activated, the expressions of phosphatidyl-serine (PS), one of the major negatively charged phospholipids on the surface of platelets, are enhanced and bind activated factors V and X, which is supposed to augment the activation of following coagulation reactions.33 However, little information is available about the effects of 5-HT released from activated platelets on this regulation. In the present study, 5-HT was shown to increase the TF and PAI-1 mRNA and protein expression in a concentration- and time-dependent manner in RAECs. In contrast, 5-HT had no significant effect on the expressions of TFPI and TPA. On the basis of these results, 5-HT causes ECs to be procoagulative and hypofibrinolytic by increasing TF relative to TFPI and increasing PAI-1 relative to TPA. These findings suggest that 5-HT is involved in physiologic hemostasis by modifying the properties of vascular ECs, in addition to causing platelet aggregation and SMC contraction. In applying these findings to in vivo status, however, several limitations must be considered. Recent studies demonstrate that the function of cultured ECs is different, depending on the source of endothelial cells. For example, the differences of adhesion molecule expressions or permeability to various vasoactive substances between cultured human arterial and venous ECs were reported.34,35 RAECs were used in this study, because platelets participate more potently in the arterial thrombus formation than in the venous thrombus formation. Whether the same reactions are seen in venous ECs or not remain uncertain. Second, the concentration of 5-HT used in this investigation must be compared with those in vivo. In healthy humans and animals, the normal concentration of 5-HT in plasma is reported to be low, because 5-HT is rapidly taken up by platelets or metabolized by monoamine oxidase in ECs and SMCs. Rat plasma concentration of 5-HT is reported to be approximately 0.1 µM,36 which is equivalent to the minimum concentration of 5-HT used in our experiments. This concentration is regarded as the basal concentration that ECs are usually exposed in vivo. With platelet aggregation, the local 5-HT concentration is considered to increase from basal level, which can trigger the local actions involving ECs. In the 5-HT concentration-dependent study, the significant induction of TF and PAI-1 mRNA was obtained at the concentrations of 5-HT 1 µM and higher. These results from concentration-dependent study seem to be relevant. 5-HT is known to be rapidly taken by platelets; therefore, the local concentration of an injured vessel might be rapidly decreased at local sites. As Ashton et al37 demonstrated, however, the persistent mobilization of platelets from circulating blood and the adherence of the activated platelets to the initial layer contribute the increased 5-HT concentration at the site. This means that physiologic primary hemostasis is maintained by activated platelets, which can exist until secondary hemostasis is completed. In a rabbit model of angioplasty involving the femoral artery, Pakala et al38 found that there was platelet deposition at the site of vascular injury with progressive accumulation of 5-HT. The concentration of 5-HT measured at the local vascular tissue in their model exceeded the concentrations required in vitro to stimulate ECs. Moreover, under these circumstances, the dysfunction of the ECs, which accompanies a decrease in monoamine oxidase activity, may result in the local accumulation of 5-HT by the decrease in its breakdown.39 In the present study, the stimulation of RAECs to produce TF and PAI-1 required high concentration and prolonged incubation of 5-HT; however, these are in agreement with the earlier reports in other cell types and in other responses related to 5-HT.7,27,40,41 On the basis of these observations, it is considered to be relevant that high 5-HT concentration levels persist at local sites until TF and PAI-1 proteins express. The 5-HT receptors, 5HT1-7 receptors, have been characterized pharmacologically.1 The 5-HT2 receptor is responsible for the platelet aggregation induced by 5-HT, because the effect of 5-HT is blocked by antagonists such as ketanserin.2 The 5-HT2 receptor is also involved in the proliferation of VSMCs induced by 5-HT.4,42 Moreover, Pakala et al43 suggested that the mitogenic effect of 5-HT to canine aortic ECs is mediated by the 5-HT2 receptor. Because MCI-9042, a selective 5-HT2A receptor antagonist,44 inhibited the induction of TF and PAI-1 mRNA by 5-HT in the present study, it was suggested that the responsible receptor for these reactions is the 5-HT2A receptor. The effect of 5-carboxamidotryptamine maleate, a 5-HT1 receptor agonist, and 2-methyl-5-hydroxytryptamine, a 5-HT3 receptor agonist, were also investigated. Both agonists induced TF and PAI-1 mRNA expression in a dose-dependent manner, respectively, which was contrary to the results we had expected. However, because MCI-9042 completely inhibited these effects, we consider that these results were due to cross-reaction of these agonists with the 5-HT2A receptor (data not shown). Taken together, we concluded that the effects of 5-HT were mediated by the activation of 5-HT2A receptors on the endothelial surface. The existence of endothelial 5-HT2A receptors was confirmed by RT-PCR (data not shown). In conclusion, 5-HT causes ECs to become thrombogenic via the 5-HT2A receptor, which is a novel and interesting property of 5-HT, in light of the interactions between platelets and ECs. These results demonstrate the contribution of 5-HT to the development of thrombotic diseases in combination with other properties, such as platelet aggregation, VSMC contraction, and VSMC proliferation.
Submitted June 23, 2000; accepted November 8, 2000.
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: Hidehiko Kawano, Second Department of Medicine, Kyoto Prefectural University of Medicine, Kawaramachi-Hirokoji, Kamigyo-ku, Kyoto, Japan; e-mail: kawano{at}gemini.kpu-m.ac.jp.
1.
Fanburg BL, Lee SL.
A new role for an old molecule: serotonin as a mitogen.
Am J Physiol.
1997;272:L795-806
2.
Clerck FFPD.
Serotonin and amplification mechanisms in platelet reactions.
NIPS.
1989;4:130-133
3.
Crowley ST, Dempsey EC, Horwitz KB, Horwitz LD.
Platelet-induced vascular smooth muscle cell proliferation is modulated by the growth amplification factors serotonin and adenosine diphosphate.
Circulation.
1994;90:1908-1918
4.
Nemeck G, Coughlin S, Handley D, Moskowitz M.
Stimulation of aortic smooth muscle cell mitogenesis by serotonin.
Proc Natl Acad Sci U S A.
1986;83:674-678
5.
Pakala R, Willerson JT, Benedict CR.
Effect of serotonin, thromboxane A2, and specific receptor antagonists on vascular smooth muscle cell proliferation.
Circulation.
1997;96:2280-2286 6. Tamura K, Kanzaki T, Saito Y, Otabe M, Saito Y, Morisaki N. Serotonin (5-hydroxytryptamine, 5-HT) enhances migration of rat aortic smooth muscle cells through 5-HT2 receptors. Atherosclerosis. 1997;132:139-143[CrossRef][Medline] [Order article via Infotrieve].
7.
Forstermann U, Mugge A, Bode SM, Frolich JC.
Response of human coronary arteries to aggregating platelets: importance of endothelium-derived relaxing factor and prostanoids.
Circ Res.
1988;63:306-312
8.
Fukai T, Egashira K.
Serotonin-induced coronary spasm in a swine model.
Circulation.
1993;88:1922-1930
9.
Shimokawa H, Aarhus LL, Vanhoutte PM.
Porcine coronary arteries with regenerated endothelium have a reduced endothelium-dependent responsiveness to aggregating platelets and serotonin.
Circ Res.
1987;61:256-270 10. Farstad M. The role of blood platelets in coronary atherosclerosis and thrombosis. Scand J Clin Lab Invest. 1998;58:1-10[CrossRef][Medline] [Order article via Infotrieve]. 11. Ross R. The pathogenesis of atherosclerosis: a perspective for the 1990s. Nature. 1993;362:801-809[CrossRef][Medline] [Order article via Infotrieve].
12.
Schneiderman J, Sawdey MS, Keeton MR, et al.
Increased type 1 plasminogen activator inhibitor gene expression in atherosclerotic human arteries.
Proc Natl Acad Sci U S A.
1992;89:6998-7002 13. Tremoli E, Camera M, Toschi V, Colli S. Tissue factor in atherosclerosis [In Process Citation]. Atherosclerosis. 1999;144:273-283[CrossRef][Medline] [Order article via Infotrieve].
14.
Nemerson Y.
Tissue factor and hemostasis [published erratum appears in Blood 1988 Apr;71(4):1178].
Blood.
1988;71:1-8 15. Novotny WF. Tissue factor pathway inhibitor. Semin Thromb Hemost. 1994;20:101-108[Medline] [Order article via Infotrieve].
16.
Sprengers E, Kluft C.
Plasminogen activator inhibitors.
Blood.
1987;69:381-387
17.
Dieval J, Nguyen G, Gross S, Delobel J, Kruithof EK.
A lifelong bleeding disorder associated with a deficiency of plasminogen activator inhibitor type 1.
Blood.
1991;77:528-532
18.
Sandset PM, Warn-Cramer BJ, Maki SL, Rapaport SI.
Immunodepletion of extrinsic pathway inhibitor sensitizes rabbits to endotoxin-induced intravascular coagulation and the generalized Shwartzman reaction.
Blood.
1991;78:1496-1502 19. Mcguire P, Orkin R. Isolation of rat aortic endothelial cells by primary esplant technique and phenotypic modulation by defined substrata. Lab Invest. 1987;57:94-105[Medline] [Order article via Infotrieve]. 20. Drake T, Morrisey J, Edginton T. Selective cellular expression of tissue factor in human tissues. Am Pathol. 1989;134:1087-1097[Abstract].
21.
Lip GY, Gibbs CR.
Does heart failure confer a hypercoagulable state? Virchow's triad revisited.
J Am Coll Cardiol.
1999;33:1424-1426 22. Janssens S, Van de Werf F. Cardiology. Acute coronary syndromes: Virchow's triad revisited. Lancet. 1996;342(suppl 2):sII2.
23.
Vikenes K, Farstad M, Nordrehaug JE.
Serotonin is associated with coronary artery disease and cardiac events.
Circulation.
1999;100:483-489 24. Willerson JT, Campbell WB, Winniford MD, et al. Conversion from chronic to acute coronary artery disease: speculation regarding mechanisms. Am J Cardiol. 1984;54:1349-1354[CrossRef][Medline] [Order article via Infotrieve].
25.
Ross R, Glomset J, Kariya B, Harker L.
A platelet-dependent serum factor that stimulates the proliferation of arterial smooth muscle cells in vitro.
Proc Natl Acad Sci U S A.
1974;71:1207-1210
26.
Williams LT, Tremble P, Antoniades HN.
Platelet-derived growth factor binds specifically to receptors on vascular smooth muscle cells and the binding becomes nondissociable.
Proc Natl Acad Sci U S A.
1982;79:5867-5870
27.
Schini-Kerth VB, Fisslthaler B, Van Obberghen-Schilling E, Busse R.
Serotonin stimulates the expression of thrombin receptors in cultured vascular smooth muscle cells. Role of protein kinase C and protein tyrosine kinases.
Circulation.
1996;93:2170-2177 28. Adams PC, Badimon JJ, Badimon L, Chesebro JH, Fuster V. Role of platelets in atherogenesis: relevance to coronary arterial restenosis after angioplasty. Cardiovasc Clin. 1987;18:49-71[Medline] [Order article via Infotrieve]. 29. Epstein SE, Speir E, Unger EF, Guzman RJ, Finkel T. The basis of molecular strategies for treating coronary restenosis after angioplasty. J Am Coll Cardiol. 1994;23:1278-1288[Abstract]. 30. Gallicchio M, Hufnagl P, Wojta J, Tipping P. IFN-gamma inhibits thrombin- and endotoxin-induced plasminogen activator inhibitor type 1 in human endothelial cells. J Immunol. 1996;157:2610-2617[Abstract]. 31. Lopez-Aguirre Y, Paramo JA. Endothelial cell and hemostatic activation in relation to cytokines in patients with sepsis. Thromb Res. 1999;94:95-101[CrossRef][Medline] [Order article via Infotrieve]. 32. Nishimura H, Tsuji H, Masuda H, et al. Angiotensin II increases plasminogen activator inhibitor-1 and tissue factor mRNA expression without changing that of tissue type plasminogen activator or tissue factor pathway inhibitor in cultured rat endothelial cells. Thromb Haemost. 1997;77:1189-1195[Medline] [Order article via Infotrieve]. 33. Larson PJ, Camire RM, Wong D, et al. Structure/function analyses of recombinant variants of human factor Xa: factor Xa incorporation into prothrombinase on the thrombin-activated platelet surface is not mimicked by synthetic phospholipid vesicles. Biochemistry. 1998;37:5029-5038[CrossRef][Medline] [Order article via Infotrieve].
34.
Kalogeris TJ, Kevil CG, Laroux FS, Coe LL, Phifer TJ, Alexander JS.
Differential monocyte adhesion and adhesion molecule expression in venous and arterial endothelial cells.
Am J Physiol.
1999;276:L9-L19 35. Ikeda K, Utoguchi N, Makimoto H, Mizuguchi H, Nakagawa S, Mayumi T. Different reactions of aortic and venular endothelial cell monolayers to histamine on macromolecular permeability: role of cAMP, cytosolic Ca2+ and F-actin. Inflammation. 1999;23:87-97[CrossRef][Medline] [Order article via Infotrieve]. 36. Teramoto Y, Urano T, Nagai N, Takada Y, Ikeda K, Takada A. Plasma levels of 5-HT and 5-HIAA increased after intestinal ischemia/reperfusion in rats. Jpn J Physiol. 1998;48:333-339[CrossRef][Medline] [Order article via Infotrieve].
37.
Ashton JH, Benedict CR, Fitzgerald C, et al.
Serotonin as a mediator of cyclic flow variations in stenosed canine coronary arteries.
Circulation.
1986;73:572-578 38. Pakala R, Benedict CR. Effect of serotonin and thromboxane A2 on endothelial cell proliferation: effect of specific receptor antagonists. J Lab Clin Med. 1998;131:527-537[CrossRef][Medline] [Order article via Infotrieve].
39.
Vanhoutte PM, Houston DS.
Platelets, endothelium, and vasospasm.
Circulation.
1985;72:728-734 40. McDuffie JE, Motley ED, Limbird LE, Maleque MA. 5-hydroxytryptamine stimulates phosphorylation of p44/p42 mitogen-activated protein kinase activation in bovine aortic endothelial cell cultures. J Cardiovasc Pharmacol. 2000;35:398-402[CrossRef][Medline] [Order article via Infotrieve]. 41. McDuffie JE, Coaxum SD, Maleque MA. 5-hydroxytryptamine evokes endothelial nitric oxide synthase activation in bovine aortic endothelial cell cultures. Proc Soc Exp Biol Med. 1999;221:386-390[CrossRef][Medline] [Order article via Infotrieve].
42.
Corson MA, Alexander RW, Berk BC.
5-HT2 receptor mRNA is overexpressed in cultured rat aortic smooth muscle cells relative to normal aorta.
Am J Physiol.
1992;262:C309-C315
43.
Pakala R, Willerson JT, Benedict CR.
Mitogenic effect of serotonin on vascular endothelial cells.
Circulation.
1994;90:1919-1926 44. Nishio H, Inoue A, Nakata Y. Binding affinity of sarpogrelate, a new antiplatelet agent, and its metabolite for serotonin receptor subtypes. Arch Int Pharmacodyn Ther. 1996;331:189-202[Medline] [Order article via Infotrieve].
© 2001 by The American Society of Hematology.
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
 |
|
 |
|
  A. Fabre, J. Marchal-Somme, S. Marchand-Adam, C. Quesnel, R. Borie, M. Dehoux, C. Ruffie, J. Callebert, J-M. Launay, D. Henin, et al. Modulation of bleomycin-induced lung fibrosis by serotonin receptor antagonists in mice Eur. Respir. J., August 1, 2008; 32(2): 426 - 436. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. E. Linder, W. Ni, J. L. Diaz, T. Szasz, R. Burnett, and S. W. Watts Serotonin (5-HT) in Veins: Not All in Vain J. Pharmacol. Exp. Ther., November 1, 2007; 323(2): 415 - 421. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. Kotzailias, T. Andonovski, A. Dukic, V. L. Serebruany, and B. Jilma Antiplatelet Activity During Coadministration of the Selective Serotonin Reuptake Inhibitor Paroxetine and Aspirin in Male Smokers: A Randomized, Placebo-Controlled, Double-blind Trial. J. Clin. Pharmacol., April 1, 2006; 46(4): 468 - 475. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Steffel, T. F. Luscher, and F. C. Tanner Tissue Factor in Cardiovascular Diseases: Molecular Mechanisms and Clinical Implications Circulation, February 7, 2006; 113(5): 722 - 731. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Steffel, A. Akhmedov, H. Greutert, T. F. Luscher, and F. C. Tanner Histamine Induces Tissue Factor Expression: Implications for Acute Coronary Syndromes Circulation, July 19, 2005; 112(3): 341 - 349. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Slominski, J. Wortsman, and D. J. Tobin The cutaneous serotoninergic/melatoninergic system: securing a place under the sun FASEB J, February 1, 2005; 19(2): 176 - 194. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Hironaka, M. Hongo, A. Sakai, E. Mawatari, F. Terasawa, N. Okumura, A. Yamazaki, Y. Ushiyama, Y. Yazaki, and O. Kinoshita Serotonin receptor antagonist inhibits monocrotaline-induced pulmonary hypertension and prolongs survival in rats Cardiovasc Res, December 1, 2003; 60(3): 692 - 699. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
 |
 |
 |
|||||||
 |
 |
 |
 |
 |
 |
 |
 |
||
| Copyright © 2001 by American Society of Hematology Online ISSN: 1528-0020 | |||||||||
Top
Abstract
Introduction
80°C.
; TFPI, 
. Each bar represents the
mean ± SEM of 3 independent experiments (*P < .05
versus vehicle-treated controls). Relative TF and TFPI mRNA expressions
represent the fold increase in normalized TF and TFPI mRNA expression
compared to vehicle-treated controls, respectively.
