doi: 10.1161/hc4801.099737
(Circulation. 2001;104:2826.)
© 2001 American Heart Association, Inc.
Basic Science Reports |
Sex Steroids Used in Hormonal Treatment Increase Vascular Procoagulant Activity by Inducing Thrombin Receptor (PAR-1) Expression
Role of the Glucocorticoid Receptor
From the Institut für Kardiovaskuläre Physiologie (O.H., R.B., V.B.S.-K.), Zentrum der Frauenheilkunde und Geburtshilfe (O.H., H.K.), and Zentrum der Pharmakologie (J.S.), Klinikum der Johann Wolfgang Goethe-Universität, Frankfurt am Main, Germany.
Correspondence to V.B. Schini-Kerth, PhD, Pharmacologie et Physicochimie des Interactions Cellulaires et Moléculaires, UMR CNRS 7034, Faculté de Pharmacie, 74, route du Rhin BP 24, F-67401 Illkirch, France. E-mail schini{at}aspirine.u-strasbg.fr
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Abstract |
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Background— The use of sex steroids in oral contraception or hormonal replacement therapy is associated with an increased risk of cardiovascular thromboembolic complications. Although both the estrogen and the progestin components have been involved, the underlying mechanisms responsible are unclear.
Methods and Results— This study examined whether sex steroids promote hemostasis indirectly by increasing the procoagulant activity of blood vessels. Treatment of vascular smooth muscle cells with several progestins (progesterone, 3-keto-desogestel, gestodene, and medroxyprogesterone acetate) upregulated proteolytically activatable thrombin receptor (PAR-1) expression, resulting in a potentiated thrombin-induced tissue factor expression and surface procoagulant activity. In contrast, neither the progestins levonorgestrel, norethisterone, and norgestimate nor the synthetic estrogen 17
-ethinylestradiol had such effects. The effect of the stimulatory progestins, which induce glucocorticoid-like effects in several cell systems, was mimicked by dexamethasone and inhibited by the progesterone and glucocorticoid receptor antagonist RU-38486. In addition, long-term administration of progesterone, 3-keto-desogestrel, or medroxyprogesterone acetate to ovariectomized rats increased PAR-1 protein level in the arterial wall, resulting in an increased responsiveness of isolated aortic rings to thrombin.
Conclusions— These data demonstrate that several progestins markedly potentiate the vascular procoagulant effects of thrombin by increasing the availability of membrane thrombin receptors in the smooth muscle, an effect that is most likely due to their glucocorticoid-like activity.
Key Words: thrombin • tissue factor • thrombosis • muscle, smooth • progestin
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Introduction |
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Numerous clinical trials indicate that estrogen-progestin combinations used for oral contraception or postmenopausal replacement therapy increase the relative risk of venous thromboembolic events (VTEs) 2- to 4-fold.1–3 These complications can be observed particularly in first-time users and during the first year of treatment, and they increase further in women with risk factors for venous thromboembolism, such as obesity.1,2 The higher incidence of VTEs was attributed initially to the estrogen component of oral contraceptives (OCs), because the reduction from 50 to 30 µg of estrogen (low-dose estrogen in newer preparations) per tablet had a favorable outcome.4 Several recent independent studies, however, have shown a 2-fold increased relative risk of VTEs with combined OCs containing either gestodene or desogestrel in combination with low-dose estrogen compared with those containing levonorgestrel and similar low-dose estrogen.2,3,5 Although a certain selection bias cannot be entirely excluded,6 these findings highlight a potential determinant role of the progestin component of OCs for the development of VTEs.
Thrombotic events are the result of complex local processes involving not only humoral factors of the hemostatic and fibrinolytic systems but also blood constituents (eg, platelets) and cells from the vascular wall. Progestins appear to have negligible influence on the moderate increase in coagulation and fibrinolytic activity in plasma induced by estrogens in women.7,8 The recent observation of a reversible acquired resistance to activated protein C during intake of OCs, which was more pronounced with formulations containing gestodene and desogestrel than levonorgestrel,9 might help to explain the increased risk of VTEs.
In the present study, we examined whether sex steroids induce vascular tissue factor (TF)–dependent procoagulant activity by either a direct or an indirect effect by potentiating the response to thrombin. The investigations were performed with vascular smooth muscle cells (VSMCs), because they are a major cellular source of TF, the physiological trigger of the blood clotting system, in blood vessels.
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Methods |
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Cell Culture
VSMCs were isolated from Wistar rat aorta and cultured as described.10 Human aortic SMCs were obtained from Clonetics and cultured as recommended. Confluent cells were serum-deprived for 24 hours before experiment.
Northern and Western Blot Analyses
PAR-1 mRNA and TF mRNA levels were assessed by Northern blot analysis as described.10 The PAR-1 cDNA probe was a 1400-bp-long PstI restriction fragment from a Chinese hamster PAR-1 cDNA clone (provided by Dr Van Obberghen-Schilling, Nice, France). The TF cDNA probe was an 1100-bp-long EcoRI restriction fragment from a mouse TF cDNA clone (provided by Dr Nawroth, Tübingen, Germany). Phenol-soluble proteins from VSMCs or homogenized rat aortic rings were separated by SDS-PAGE (8%) and transferred to nitrocellulose membranes. The immunoreactive band was detected by enhanced chemiluminescence (Amersham).
Animal Treatment and Organ Chamber Studies
Wistar rats (16 weeks old) were ovariectomized, followed by a 2-week rest period. They were treated subcutaneously with either vehicle (sesame oil) or a progestin (progesterone 16.7 mg/kg, 3-keto-desogestrel [KDG] 0.5 mg/kg, or medroxyprogesterone acetate [MPA] 16.7 mg/kg) every day for 7 days. The aorta was excised and cut into rings, which were either suspended in organ chambers for measurement of changes in isometric tension in the presence of diclofenac (10 µmol/L) as described11 or processed for determination of PAR-1 protein level by Western blot analysis.
Procoagulant Activity Study
The surface procoagulant activity of human VSMCs was determined essentially as described.12 After treatment, VSMCs were washed with a HEPES-Tyrode’s solution and then incubated with human platelet-poor plasma. The formation of thrombin was initiated by addition of CaCl2 (16.7 mmol/L). Aliquots were removed at different time points, and the formation of thrombin was determined by use of the chromogenic substrate S-2238 (Hemochrom Diagnostica). Optical densities were measured in a spectrophotometer at 405 nm.
Statistical Analysis
All values are reported as mean±SEM. Statistical analysis was performed with Student’s t test or ANOVA followed by Student-Newman-Keuls t test where appropriate. A value of P<0.05 was considered statistically significant.
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Results |
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Sex Steroids Induce Thrombin Receptor Expression
The progestin KDG, the active metabolite of desogestrel, significantly increased PAR-1 mRNA levels in VSMCs in a time-dependent manner (Figure 1A). PAR-1 mRNA levels were also significantly increased by the progestins progesterone, gestodene, and MPA, whereas the progestins levonorgestrel, norgestimate, and norethisterone and the synthetic estrogen 17
-ethinylestradiol had only minor effects (Figure 1B). The stimulatory effect of KDG and MPA on PAR-1 at the mRNA level was observed at concentrations of 
10 nmol/L (Figure 1, C and D). To determine PAR-1 expression at the protein level, Western blot analysis was performed with a monoclonal antibody directed against the peptide sequence GRAVYLNKSRFPPMPPPPFISEDASG in the N-terminus below the thrombin cleavage site of the rat PAR-1 (COR7-6H9, provided by Dr Ramakrishnan, COR Therapeutics). In VSMCs, a faint band of 
70 kDa was detected, which appears to be specific for PAR-1, because it could be eliminated by competition with a 10-fold molar excess of the synthetic immunizing peptide (data not shown). Exposure of VSMCs to MPA for 24 hours increased the intensity of this 70-kDa band, indicating an upregulation of membrane thrombin receptors (Figure 1E). Importantly, the stimulatory effect of progestins is observed at concentrations within the therapeutic range. Peak serum concentrations of KDG in women taking the combination of 30 µg ethinylestradiol and 150 µg desogestrel were 10 nmol/L,13 those of gestodene were 30 nmol/L after intake of the combination of 30 µg ethinylestradiol and 75 µg gestodene,14 and those of MPA were 5 nmol/L after intake of 2 mg 17ß-estradiol and 5 mg MPA.15
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Role of the Glucocorticoid Receptor
The stimulatory progestins (KDG, progesterone, gestodene, and MPA) have been shown to bind not only to the progesterone but also to the glucocorticoid receptor with relatively high affinity and to induce glucocorticoid-like effects in several experimental cell systems. In contrast, the inactive progestins (levonorgestrel, norethisterone, norgestimate) bind almost exclusively to the progesterone receptor and do not bind to the glucocorticoid receptor.16,17 Therefore, we analyzed the role of the glucocorticoid receptor in regulating PAR-1 expression. The glucocorticoid dexamethasone upregulated PAR-1 mRNA and protein levels with a threshold concentration of 1 nmol/L (Figure 2, A and B). The stimulatory effect of progestins and dexamethasone was abolished by RU-38486, a potent progesterone and glucocorticoid receptor antagonist (Figure 2C). Thus, activation of the glucocorticoid rather than the progesterone receptor appears to mediate the stimulatory effect of progestins on PAR-1 expression. The finding that RU-38486 alone slightly induced PAR-1 mRNA expression (Figure 2C) can be explained by the fact that this drug at concentrations >0.1 µmol/L can act as a partial glucocorticoid agonist.16,18
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Progestins Potentiate Vascular Procoagulant Activity
Thrombin induces the expression of TF, the physiological trigger of the blood clotting system, in VSMCs.19 Therefore, we examined the possibility that progestins potentiate the thrombin-induced TF-dependent procoagulant activity. A low concentration of thrombin (0.1 U/mL, 1 hour) induced a modest signal for TF mRNA, which was markedly increased by preincubation of VSMCs with either MPA, KDG, or dexamethasone at concentrations as low as 1 nmol/L for 24 hours (Figure 3, A, B, and C). In contrast, steroids alone did not induce TF mRNA expression (Figure 3A and data not shown). To determine whether increased TF mRNA levels are associated with cell surface procoagulant activity, human VSMCs were exposed to thrombin for 6 hours, a time period that induces maximal TF activity on the surface of VSMCs.20 As expected, thrombin slightly increased the low basal procoagulant activity (significant when the data shown in Figure 4, A and B, were combined), and this response was markedly potentiated by pretreatment of VSMCs with MPA or dexamethasone for 24 hours before the addition of thrombin (Figure 4, A and B). Exposure of VSMCs to MPA or dexamethasone alone did not induce procoagulant activity (Figure 4, A and B). The thrombin-induced surface procoagulant activity of VSMCs is dependent on the availability of membrane-bound TF, because this effect was abolished by a neutralizing antibody directed against TF (Figure 4, A and B). It also involves PAR-1 activation, because an increased procoagulant activity was obtained with the combination of MPA and the direct activator of PAR-1 SFLLRNP, whereas the increased procoagulant activity was abolished by a neutralizing antibody directed against human PAR-1 (Figure 5, A and B). To examine PAR-1 expression in human VSMCs, Western blot analysis was performed with a monoclonal antibody directed against human PAR-1. A faint band of 70 kDa was observed in control VSMCs, and the intensity of this band was markedly increased after exposure of VSMCs to either MPA or dexamethasone for 24 hours (Figure 4, A and B). Altogether, the findings indicate that several sex steroids potentiate the thrombin-induced TF-dependent surface procoagulant activity of VSMCs by increasing the number of membrane-associated PAR-1s.
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In Vivo Treatment With Progestins and Vascular Reactivity
To determine whether long-term administration of progestins potentiates the vascular responsiveness to thrombin, ovariectomized rats were treated with either vehicle (control group) or a progestin (progesterone, KDG, MPA), and the vascular reactivity of the isolated aorta was assessed in an organ chamber. Thrombin (10 U/mL) induced modest but consistent contractions of aortic rings without endothelium, which were significantly increased in the group of rats treated with either KDG or MPA but not progesterone (Figure 6A). In contrast to thrombin, concentration-contraction curves to phenylephrine were similar in the different groups of rats (EC50 values were 30.9±6.2, 44.9±5.5, 18.0±3.0, and 27.5±2.9 nmol/L in control and progesterone-, KDG-, and MPA-treated rats). In addition, the endothelium-dependent and nitric oxide (NO)–mediated relaxations to either thrombin or acetylcholine were similar in the control and sex steroid–treated groups of rats (data not shown). To examine PAR-1 protein levels in the aorta of each group of rats, Western blot analysis with MabCOR7-6H9 was performed. In the control group, a band of 70 kDa was detected, which was not observed in the presence of a 10-fold molar excess of immunizing peptide (Figure 6, B and C). The intensity of this 70-kDa protein band was increased after treatment of rats with either progesterone, KDG, or MPA (Figure 6B). These findings indicate that long-term administration of several progestins to rats potentiates the responsiveness of the arterial smooth muscle to thrombin, presumably by increasing the availability of membrane thrombin receptors.
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Discussion |
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Thrombin is a key enzyme of the coagulation cascade and also a potent inducer of platelet activation and of procoagulant and proinflammatory responses in vascular cells. All of these events play a central role in the initiation and development of cardiovascular pathologies associated with thrombosis, such as stroke and myocardial infarction. Thrombin exerts most of its action on target cells via an irreversible proteolytic activation of PAR-1.21 Cellular responses to thrombin rapidly desensitize despite the irreversible mechanism of activation of PAR-1.22 A major consequence of this unusual mechanism of activation and desensitization of PAR-1 is that the responsiveness of cells to thrombin is critically dependent on the constant supply of new receptor molecules to the plasma membrane.
The present findings indicate that certain progestins substantially upregulate PAR-1 expression in VSMCs, resulting in the subsequent increased availability of membrane receptors. As an important consequence of these progestin-induced changes, the vascular smooth muscle becomes more responsive to thrombin in its TF-dependent procoagulant activity.
Progestins are a heterogeneous family of steroids that, in addition to their interaction with the cytosolic progesterone receptor, can also bind strongly to other steroid receptors, such as those for androgens, mineralocorticoids, and glucocorticoids, to induce biological responses in target cells.23 Surprisingly, upregulation of vascular PAR-1 expression occurred selectively with certain progestins, such as progesterone, gestodene, KDG, and MPA, whereas levonorgestrel, norgestimate, and norethisterone, despite their similarly strong progestin potency, and also the estrogen 17-ethinylestradiol did not. The effect of the stimulatory steroid hormones was inhibited by RU-38486, suggesting the involvement of the progesterone and/or glucocorticoid receptor. Because all stimulatory progestins have distinct intrinsic glucocorticoid activity as opposed to the inactive ones,17 and the glucocorticoid dexamethasone is a potent inducer of vascular smooth muscle PAR-1 expression, activation of the glucocorticoid receptor rather than of the progesterone receptor appears to be a key event in the steroid-induced upregulation of PAR-1 expression. Moreover, the kinetics of PAR-1 expression suggests that progestins control PAR-1 expression by an indirect mechanism possibly involving the generation of secondary regulatory molecules. Among potential candidates are growth factors, such as platelet-derived growth factor, transforming growth factor, and basic fibroblast growth factor, all of which are generated by the vascular smooth muscle and upregulate PAR-1 expression in VSMCs.10,24
The present results also indicate that long-term administration of progestins with partial glucocorticoid activity (progesterone, MPA, and KDG) to ovariectomized rats causes a marked increase of the PAR-1 level in the aorta. The increased availability of membrane PAR-1 subsequently enhances the contraction of the vascular smooth muscle to thrombin. Surprisingly, endothelium-dependent and NO–mediated relaxations to thrombin are unaffected by the progestin treatment. Similarly, increased PAR-1 levels in the aorta of rats after long-term administration of angiotensin II correlate with an increased vasoconstriction to thrombin, whereas the endothelial formation of NO is unaltered.25 Thus, PAR-1 function is likely to be controlled by different mechanisms in VSMCs and in endothelial cells. Consistent with such an idea, the responsiveness of endothelial cells to thrombin appears to be controlled largely by an endogenous pool of preformed thrombin receptors, whereas in VSMCs it is controlled predominantly by de novo synthesis of the receptor.10,26 Alternatively, PAR-1 expression in endothelial cells may be controlled by endogenous regulatory factors, such as NO and prostacyclin, which have been shown to regulate the expression of several proatherosclerotic and prothrombotic genes, including those encoding adhesion molecules and TF.27,28
The stimulatory effect of progestins on vascular PAR-1 expression both in vitro and in vivo is observed at concentrations similar to those found in plasma of women during hormonal treatment.13–15 Therefore, the present findings are likely to be of clinical significance. They suggest that an increased sensitivity of the vascular smooth muscle for expression of procoagulant activity to thrombin by functional upregulation of thrombin receptors (PAR-1) may be a key event in promoting initiation and progression of VTEs in women using OCs. In particular, such a sequence of events might provide an explanation for the 2-fold increased relative risk of VTEs and fatal pulmonary embolism in women taking combined low-dose OCs containing gestodene or desogestrel compared with those taking levonorgestrel and estrogen at a similar low dose.5 Recently, in vitro studies have shown that TF expression in endothelial cells is depressed by the endogenous formation of NO.28 Therefore, it is likely that the potentiating effect of progestins on the TF-dependent vascular procoagulant activity to thrombin may be greater in vascular pathologies characterized by a reduced bioavailability of endothelial NO, such as obesity, hypertension, and hypercholesterolemia, and possibly also in blood vessels exposed to low levels of shear stress. Shear stress is the most important physiological stimulus for the continuous formation of NO in endothelial cells. Consistent with such a concept, the relative risk of VTEs in women during hormonal treatment increases with the presence of such risk factors as high body mass index,2,5 and thrombotic events in OC users occur predominantly in the venous circulation.
An additional important finding of the present study is that, like progestins with partial glucocorticoid activity, the pure glucocorticoid dexamethasone also markedly potentiated TF-dependent procoagulant activity to thrombin by increasing the availability of membrane PAR-1 in human VSMCs. The stimulatory effect of the glucocorticoid, which does not bind to the progesterone receptor,29 is obtained at low concentrations (in the nanomolar range), suggesting that it may be of clinical relevance. Such a sequence of events may contribute to explain the increased risk of thromboembolism in patients under long-term glucocorticoid therapy or with an endogenous hypercortisolism (Cushing’s syndrome).30
In conclusion, the present findings demonstrate that certain progestins currently used in oral contraception and in hormone replacement therapy can remarkably sensitize the vascular smooth muscle for expression of procoagulant activity to thrombin by upregulating the availability of membrane thrombin receptors (PAR-1). They further suggest that the partial glucocorticoid activity of progestins is likely to be responsible for this action and thus might be a key determinant in the induction of a thrombotic state.
Received August 1, 2001; revision received September 25, 2001; accepted September 25, 2001.
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References |
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1. Daly E, Vessey MP, Hawkins MM, et al. Risk of venous thromboembolism in users of hormone replacement therapy. Lancet. 1996; 348: 977–980.[Medline] [Order article via Infotrieve]
2. World Health Organization Collaborative Study of Cardiovascular Disease and Steroid Hormone Contraception. Venous thromboembolic disease and combined oral contraceptives: results of international multicentre case-control. Lancet. 1995; 346: 1575–1582.[Medline] [Order article via Infotrieve]
3. World Health Organization Collaborative Study of Cardiovascular Disease and Steroid Hormone Contraception. Effect of different progestagens in low oestrogen oral contraceptives on venous thromboembolic disease. Lancet. 1995; 346: 1582–1588.[Medline] [Order article via Infotrieve]
4.
Gerstman BB, Piper JM, Tomita DK, et al. Oral contraceptives estrogen dose and the risk of deep venous thromboembolic disease. Am J Epidemiol. 1991; 133: 32–37.
5.
Jick H, Kaye JA, Vasilakis-Scaramozza V, et al. Risk of venous thromboembolism among users of third generation oral contraceptives compared with users of oral contraceptives with levonorgestrel before and after 1995: cohort and case-control analysis. BMJ. 2000; 321: 1190–1195.
6. Farmer RDT, Lawrenson RA, Thompson CR, et al. Population-based study of risk of venous thromboembolism associated with various oral contraceptives. Lancet. 1997; 349: 83–88.[Medline] [Order article via Infotrieve]
7. Kroon U-B, Silferstolpe G, Tengborn L. The effects of transdermal estradiol and oral conjugated estrogens on haemostasis variables. Thromb Haemost. 1994; 71: 420–423.[Medline] [Order article via Infotrieve]
8. Gilabert J, Estelles A, Cano A, et al. The effect of estrogen replacement therapy with or without progestogen on the fibrinolytic system and coagulation inhibitors in postmenopausal status. Am J Obstet Gynecol. 1995; 173: 1849–1854.[Medline] [Order article via Infotrieve]
9. Rosing J, Tans G, Nicolaes GAF, et al. Oral contraceptives and venous thrombosis: different sensitivities to activated protein C in women using second- and third-generation oral contraceptives. Br J Haematol. 1997; 97: 233–238.[Medline] [Order article via Infotrieve]
10.
Schini-Kerth VB, Bassus S, Fisslthaler B, et al. Aggregating platelets stimulate the expression of thrombin receptors in cultured vascular smooth muscle cells via the release of transforming growth factor-ß1 and platelet-derived growth factorAB. Circulation. 1997; 96: 3888–3896.
11.
Schini VB, Vanhoutte PM. L-Arginine evokes both endothelium-dependent and -independent relaxations in L-arginine–depleted aortas of the rat. Circ Res. 1991; 68: 209–216.
12. Beguin S, Lindhout T, Hemker HC. The effect of trace amounts of tissue factor on thrombin generation in platelet rich plasma: its inhibition by heparin. Thromb Haemost. 1989; 61: 25–29.[Medline] [Order article via Infotrieve]
13. Kuhl H, Jung-Hoffmann C, Heidt F. Serum levels of 3-keto-desogestrel and SHBG during 12 cycles of treatment with 30 µg ethinylestradiol and 150 µg desogestrel. Contraception. 1988; 38: 381–390.[Medline] [Order article via Infotrieve]
14. Kuhl H, Jung-Hoffmann C, Heidt F. Alterations in the serum levels of gestodene and SHBG during 12 cycles of treatment with 30 µg ethinylestradiol and 75 µg gestodene. Contraception. 1988; 38: 477–486.[Medline] [Order article via Infotrieve]
15. Svensson L-O, Hedman Johnson S, Olsson S-E. Plasma concentrations of medroxyprogesterone acetate, estradiol and estrone following oral administration of Klimaxil®, Trisequence®/Provera® and Divina®: a randomized, single-blind, triple cross-over bioavailability study in menopausal women. Maturitas. 1994; 18: 229–238.[Medline] [Order article via Infotrieve]
16. Rondinone CM, Baker ME, Rodbard D. Progestins stimulate the differentiation of 3T3-L1 preadipocytes. J Steroid Biochem Mol Biol. 1992; 42: 795–802.[Medline] [Order article via Infotrieve]
17. Kontula K, Paavonen T, Luukkainen T, et al. Binding of progestins to the glucocorticoid receptor: correlation to their glucocorticoid-like effects on in vitro functions of human mononuclear leukocytes. Biochem Pharmacol. 1983; 32: 1511–1518.[Medline] [Order article via Infotrieve]
18. Schweizer-Groyer G, Cadepond F, Groyer A, et al. Stimulation of specific transcription and DNA binding studies suggest that in vitro transformed RU 486-glucocorticosteroid receptor complexes display agonist activity. J Steroid Biochem. 1988; 30: 291–294.[Medline] [Order article via Infotrieve]
19. Taubman MB, Marmur JD, Rosenfield C-L, et al. Agonist-mediated tissue factor expression in cultured vascular smooth muscle cells: role of Ca2+ mobilization and protein kinase C activation. J Clin Invest. 1993; 91: 547–552.
20. Schecter AD, Giesen PLA, Taby O, et al. Tissue factor expression in human arterial smooth muscle cells: TF is present in three cellular pools after growth factor stimulation. J Clin Invest. 1997; 100: 2276–2285.[Medline] [Order article via Infotrieve]
21. Vu T-KH, Hung DT, Wheaton VI, et al. Molecular cloning of a functional thrombin receptor reveals a novel proteolytic mechanism of receptor activation. Cell. 1991; 64: 1057–1068.[Medline] [Order article via Infotrieve]
22.
Brass LF. Homologous desensitization of HEL cell thrombin receptors: distinguishable roles for proteolysis and phosphorylation. J Biol Chem. 1992; 267: 6044–6050.
23. Ojasoo T. Multivariate preclinical evaluation of progestins. Menopause. 1995; 2: 97–107.
24.
Zhong C, Hayzer DJ, Corson MA, et al. Molecular cloning of the rat vascular smooth muscle thrombin receptor: evidence for in vitro regulation by basic fibroblast growth factor. J Biol Chem. 1992; 267: 16975–16979.
25.
Capers QIV, Laursen JB, Fukui T, et al. Vascular thrombin receptor regulation in hypertensive rats. Circ Res. 1997; 80: 838–844.
26. Storck J, Zimmermann ER. Regulation of the thrombin receptor response in human endothelial cells. Thromb Res. 1996; 81: 121–131.[Medline] [Order article via Infotrieve]
27.
Zeiher AM, Fisslthaler B, Schray-Utz B, et al. Nitric oxide modulates the expression of monocyte chemoattractant protein 1 in cultured human endothelial cells. Circ Res. 1995; 76: 980–986.
28.
Yang Y, Loscalzo J. Regulation of tissue factor expression in human microvascular endothelial cells by nitric oxide. Circulation. 2000; 101: 2144–2148.
29.
Spelsberg TC, Graham ML, Berg NJ, et al. A nuclear binding assay to assess the biological activity of steroid receptors in isolated animal and human tissues. Endocrinology. 1987; 121: 631–644.
30. Sjöberg HE, Blombäck M, Granberg PO. Thromboembolic complications, heparin treatment and increase in coagulation factors in Cushing’s syndrome. Acta Med Scand. 1976; 199: 95.
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