(Hypertension. 2001;37:640.)
© 2001 American Heart Association, Inc.
Scientific Contributions |
Estradiol Metabolites Inhibit Endothelin Synthesis by an Estrogen Receptor-Independent Mechanism
From the Department of Obstetrics and Gynecology (R.K.D., P.J.K., B.I., M.R.), Clinic for Endocrinology, University Hospital, Zurich, Switzerland, and Departments of Medicine (R.K.D., E.K.J.), and Pharmacology (E.K.J.) Center for Clinical Pharmacology, University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania.
Correspondence to Raghvendra K. Dubey, PhD, Department of Obstetrics and Gynecology, Clinic for Endocrinology, D215, NORD-1; Frauenklinik, University Hospital Zurich, 8091 Zurich, Switzerland. E-mail raghvendra.Dubey{at}fhk.usz.ch
 |
Abstract |
|---|
 Top Abstract IntroductionMethodsResultsDiscussionReferences |
|---|
Estradiol inhibits endothelin-1 synthesis, an effect that may contribute to the cardiovascular protective effects of estradiol. Recent findings that estradiol inhibits neointima formation in mice lacking estrogen receptors suggests that the cardiovascular protective effects of estradiol may be mediated by means of an estrogen receptor-independent mechanism. Because 2-hydroxyestradiol and 2-methoxyestradiol, metabolites of estradiol with little/no affinity for estrogen receptors, are more potent than estradiol in inhibiting vascular smooth muscle cell growth, we investigated whether these metabolites also inhibit endothelin-1 synthesis by means of an receptor-independent mechanism. Treatment of porcine coronary artery endothelial cells for 4 to 24 hours with 0.001 to 1 µmol/L of estradiol, 2-hydroxyestradiol, or 2-methoxyestradiol concentration-dependently inhibited basal as well as serum-induced (2.5%), TNF
-induced (10 ng/mL),
angiotensin II–induced (100 nmol/L), and thrombin-induced
(4 U/mL) endothelin-1 synthesis. Estradiol, 2-hydroxyestradiol, and
2-methoxyestradiol also inhibited serum-induced
mitogen-activated protein kinase activity. As compared with
estradiol, its metabolites were more potent in inhibiting endothelin-1
secretion and mitogen activated protein kinase activity. The
inhibitory effects of 2-hydroxyestradiol and
2-methoxyestradiol on endothelin-1 release and
mitogen-activated protein kinase activity were not blocked by
ICI182780 (50 µmol/L), an estrogen receptor antagonist.
Our findings indicate that the estradiol metabolites 2-hydroxyestradiol
and 2-methoxyestradiol potently inhibit endothelin-1 synthesis by means
of an estrogen receptor-independent mechanism. This effect of estradiol
metabolites may be mediated by inhibition of mitogen activated
protein kinase activity and may contribute to the cardioprotective
effects of estradiol.
Key Words: postmenopause • endothelial cells • vascular remodeling • cardiovascular disease • estradiol metabolites • coronary artery
|
Introduction |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
Increased endothelin-1 (ET-1) synthesis is associated with vaso-occlusive disorders,1 2 and estradiol inhibits ET-1 synthesis, perhaps by means of estrogen receptor (ER) -dependent pathways.3 4 Thus, inhibition of ET-1 production by estradiol may contribute to its cardioprotective effects. However, the mechanisms by which estradiol inhibits ET-1 synthesis remain unclear.
In this regard, although the biological effects of estradiol
are thought to be ER
mediated,1 recent findings
indicate that estradiol inhibits neointima formation in
mice lacking either ER or ERß, suggesting that the
cardiovascular protective effects of estradiol may be
mediated in part by means of ER-independent
mechansims.5 6 This
notion is further supported by the observation that estradiol is unable
to inhibit neointima formation in nongonadectomized male
rats7 even though these rats
express high levels of ER and
ERß.8
2-Hydroxyestradiol and 2-methoxyestradiol are major endogenous metabolites of estradiol that have low or no affinity for ERs.1 Even so, these estradiol metabolites are more potent than estradiol in inhibiting vascular smooth muscle cell growth.9 10 Thus, we hypothesize that these metabolites play a major role in mediating the cardiovascular protective effects of estradiol by means of an ER-independent mechanism.
The main goal of the present study was to investigate whether estradiol metabolites inhibit ET-1 synthesis in coronary artery endothelial cells by means of an ER-independent mechanism. Another objective was to compare the efficacy of various clinically used estrogens in inhibiting ET-1 synthesis. Finally, we investigated whether inhibition of mitogen activated protein kinase (MAPK) activity could participate in the effects of estradiol metabolites on endothelial biology.
|
Methods |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
Isolation and Culture of Porcine Coronary Artery Endothelial Cells
Hearts from female pigs were obtained from a local slaughter house and placed in ice-cold oxygenated DMEM containing antibiotics. The epicardial coronary arteries were isolated, and the fat and connective tissue were removed. Endothelial cells were isolated by collagenase/dispase digestion.11 The isolated cells were washed in DMEM/F12 medium supplemented with 10% fetal calf serum (FCS) and endothelial cell growth supplement (Clonetics), and cells were grown to confluence under standard tissue culture conditions. The purity of the cultures, which was characterized by immunostaining with factor VIII antigen and by assaying the preferential uptake of the fluorescent probe Dil-acetylated low density lipoprotein,11 was 98%. Cells were passaged by trypsinization, and cells in second and third passage were used for all experiments.
Treatment Protocols for Endothelin-1
Synthesis Studies
To study the effects of estradiol and its metabolites
on ET-1 synthesis in porcine coronary artery
endothelial cells (PCAECs), confluent monolayers of
PCAECs were washed twice with HBSS and treated with DMEM/Ham’s F12
supplemented with 0.4% bovine serum albumin (BSA) containing
or lacking 0.001 to 1 µmol/L of estradiol, 2-hydroxyestradiol, or
2-methoxyestradiol. After 24 hours of treatment, the monolayers were
treated for an additional 4 hours with fresh treatments in the presence
of vehicle, angiotensin II (100 nmol/L), TNF (10 ng/mL),
thrombin (4 U/mL), or FCS (2.5%). The supernatants were collected for
ET-1 analysis, and samples were frozen at -70°C until
analysis. ET-1 levels were measured with the Biotrak,
high-sensitivity endothelin-1 human ELISA system.
To evaluate whether the effects of estradiol and its
metabolites were ER mediated, the cells were treated as above and in
the presence and absence of the ER antagonist, ICI182780
(50 µmol/L; Tocris Cookson Ltd). To analyze whether the
inhibitory effects of estradiol were mediated by means of
ERß, we investigated the effects of genistein (0.001 to 10 µmol/L).
To evaluate whether the effects of estradiol were mimicked by various
clinically used estrogens, the cells were treated as above, in the
presence and absence of ICI182780, with 10 nmol/L of estradiol
valerate, estradiol cypionate, estradiol benzoate, 17-estradiol,
estrone, estriol or estrone sulfate, and the ET-1 synthesis in response
to FCS (2.5%) was analyzed.
To analyze the effects of estradiol and its metabolites on the basal synthesis of ET-1, cells were pretreated with or without the various agents and with and without ICI182780 for 24 hours and were fed fresh DMEM/F12 supplemented with 0.4% BSA and the respective treatments. The supernatants were collected after 4, 8, 12, and 24 hours and the ET-1 levels analyzed.
Protocols for MAPK Activity Measurement
PCAECs were grown to confluence in
35-mm2 culture dishes and were made
quiescent by feeding DMEM containing 0.4% BSA for 48 hours. Growth
arrested PCAECs washed with PBS and pretreated for 24 hours with or
without various test agents were stimulated with FCS (2.5%) for 10
minutes. Some cells were pretreated for 1 hour with ICI182780 before
treatment with the test agents. After stimulation, the cells were
washed with ice-cold PBS and extraction buffer (50 mmol/L
ß-glycerophosphate, 1.5 mmol/L EGTA, 1 mmol/L
dithiothreitol, 100 µmol/L
Na3VO4, 10 µg/mL
aprotinin, 5 µg/mL pepstatin, 20 g/mL leupeptin, and 1 mmol/L
benzamidine), scraped off the plates, and sonicated for 20 seconds in
0.5 mL of extraction buffer. The extracts were collected, the cytosolic
fraction were separated by centrifuging the extracts at 100
000g for 20 minutes at 4°C,
and the supernatants were diluted to a concentration of 1 mg protein/mL
and stored at -70°C for MAPK activity assays. The MAPK activity in
the cytosolic extracts was quantified as previously described by
us.12
|
Results |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
ET-1 levels were increased in a time-dependent manner in the medium collected from PCAECs incubated for 4 to 24 hours under basal conditions (Figure 1). Preliminary studies showed that treatment of PCAECs with FCS stimulated ET-1 secretion maximally at 4 hours, and this time point was selected for all subsequent studies. As compared with vehicle-treated controls, treatment of PCAECs for 4 hours with FCS, angiotensin II, thrombin, and TNF
induced ET-1
secretion by 138±14%, 116±7%, 98±5%, and 120±8% (percentage
increase), respectively
(Figure 2). Treatment of PCAECs for 8 hours with
physiological concentrations (2 nmol/L) of
estradiol inhibited basal synthesis of ET-1 by 19±2%
(P<0.05). Significant
reductions in ET-1 levels were also evident in PCAECs treated under
basal conditions for 12 and 24 hours
(Figure 1).
|
|
Estradiol inhibited FCS-, angiotensin II-,
thrombin-, and TNF-induced ET-1 secretion in a
concentration-dependent manner
(Figure 2). Physiological concentrations
(1 nmol/L) of estradiol significantly
(P<0.05) decreased ET-1
secretion induced in response to FCS (from 138±16% to 82±8%),
angiotensin II (from 116±7% to 73±6%), thrombin (from
98±6% to 53±4%), and TNF (from 120±8% to 68±5%).
The concentration-dependent inhibitory
effects of estradiol on basal and stimulated ET-1 secretion were
mimicked by its metabolites 2-hydroxyestradiol and 2-methoxyestradiol
(Figures 1 and 2). Compared with estradiol, its metabolites
were more effective in inhibiting basal and stimulated ET-1 secretion,
the order of potency being 2-methoxyestradiol > 2-hydroxyestradiol >
estradiol
(Figures 1, and 2). At physiological
concentrations (2 nmol/L), estradiol, 2-hydroxyestradiol, and
2-methoxyestradiol inhibited basal ET secretion in PCAECs incubated for
4, 8, 12, and 24 hours
(Figure 1). Moreover, FCS-, angiotensin II-,
thrombin-, and TNF-induced ET-1 secretion were inhibited by 31%,
33%, 40% and 35% by 2-hydroxyestradiol (1 nmol/L;
Figure 2); by 44%, 39%, 46%, and 56% by
2-methoxyestradiol (1 nmol/L;
Figure 2); and by 22%, 20%, 24%, and 26% by estradiol (1
nmol/L;
Figure 2).
Similar to estradiol, FCS-induced ET-1 synthesis was
inhibited by estradiol valerate, estradiol cypionate, and estradiol
benzoate
(Figure 3). In contrast, estrone, estrone sulfate, estriol,
and 17-estradiol were unable to lower ET-1 secretion. For all the
tested estrogens, the potency order for inhibition of ET-1 synthesis
was as follows: estradiol > estradiol valerate 
estradiol cypionate
> estradiol benzoate > estrone estriol estrone sulfate
17-estradiol.
|
Treatment of PCAECs with 50 µmol/L ICI182780 did not influence basal or FCS-stimulated ET-1 synthesis. ICI182789 completely blocked the inhibitory effects of estradiol, but not 2-hydroxyestradiol and 2-methoxyestradiol, on FCS-stimulated ET-1 secretion (Figure 4), as well as on basal ET-1 release (data not shown). Similar to estradiol the inhibitory effects of estradiol valerate, estradiol cypionate, and estradiol benzoate were reversed by ICI182780 (Figure 3).
|
Genistein, an ERß ligand at nanomolar conentrations,13 inhibited basal (data not shown) as well as FCS-stimulated (Figure 5) ET-1 secretion by PCAECs in a concentration-dependent manner. The inhibitory effects of genistein were observed at concentrations of 1 µmol/L and higher, moreover the inhibitory effects of genistein were not reversed by ICI182780 (Figure 5).
|
Treatment of growth-arrested cells with FCS (2.5%) increased MAPK activity from 0.187 to 7.25 pmol/min/mg protein, and the stimulatory effects of FCS were inhibited by the MEK inhibitor PD98059 (10 µmol/L) to 0.7 pmol/min/mg protein. In cells pretreated for 24 hours with estradiol, 2-hydroxyestradiol, or 2-methoxyestradiol, the stimulatory effects of FCS on MAPK activity were inhibited in a concentration-dependent manner (Figure 6). Compared with estradiol, 2-hydroxyestradiol and 2-methoxyestradiol were more potent in inhibiting FCS-induced MAPK activity. In cells pretreated with ICI182780 (50 µmol/L), the inhibitory effects of estradiol, but not 2-hydroxyestradiol or 2-methoxyestradiol, on MAPK activity were completely reversed (Figure 6). In PCAECs pretreated for 24 hours with physiological concentrations (1 nmol/L) of estradiol, 2-hydroxyestradiol, and 2-methoxyestradiol, FCS-induced MAPK activity was inhibited by 19±2.6%, 34±4%, and 46±4.7%, respectively.
|
|
Discussion |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
ET-1 is a potent vasoconstrictor peptide and enhances the mitogenic effects of mitogens on smooth muscle cell growth.1 Moreover, increases in ET-1 synthesis/release are associated with vaso-occlusive disorders.2 Because estradiol inhibits ET-1 synthesis, it is hypothesized that the protective effects of estradiol on the vessel wall are in part mediated by means of this mechanism. Many of the biologic effects of estradiol are mediated by means of ERs,1 and some studies suggest that the inhibitory effect of estradiol on ET-1 synthesis by endothelial cells is ER mediated.4 Consistent with the previous report,4 we also observe that estradiol inhibits the release of ET-1 and these effects are inhibited by ICI182780, an ER antagonist. However, recent data from our laboratory indicate that ICI182780 is not only an ER antagonist, but also inhibits the metabolism of estradiol to 2-hydroxyestradiol.14 Therefore, it is possible that blockade of the effects of estradiol on ET-1 production by ICI182780 is due in part to inhibition of metabolism of estradiol to 2-hydroxyestradiol, as well as antagonism of ERs.
Although the inhibitory effects of
estradiol on ET-1 release/secretion are inhibited by ICI182780, the
findings of two recent studies that estradiol prevents balloon
injury–induced neointima formation in mice lacking
functional ER and
ERß5 6 suggest that
ER-independent mechanisms may also participate in mediating the
cardiovascular protective effects of estradiol. Under
in vivo conditions, estradiol is efficiently metabolized by means of
multiple CYP450 enzymes.1 Our
previous studies indicate that 2-hydroxyestradiol and
2-methoxyestradiol, major endogenous metabolites of
estradiol, possess biological activity and are more potent than
estradiol in inhibiting smooth muscle cell and cardiac fibroblast
proliferation.10 14 15
Because these metabolites possess minimal binding affinity for the
ERs,1 we hypothesize that the
in vivo cardiovascular protective effects of estradiol
may be mediated in part by means of estradiol metabolites and by means
of ER-independent mechanisms. This notion is supported by our
present findings that basal as well as stimulated release of ET-1
by PCAECs are inhibited by 2-hydroxyestradiol and 2-methoxyestradiol,
and this effect is not blocked by ICI182780.
Genistein is a phytoestrogen with a high affinity for the
ERß.13 In the present
study, we used this agent to study the role of ERß in regulating ET-1
synthesis. Our observation that genistein inhibits ET-1 synthesis in
micromolar, but not nanomolar, concentrations suggests that the
inhibitory effects of estradiol are not mediated by ERß.
This contention is supported by the fact that genistein acts as an
ERß ligand at nanomolar concentrations and inhibits tyrosine kinase
activity at higher concentrations. Because inhibition of tyrosine
kinase inhibits ET-1
synthesis,2 the observed
inhibitory effects of genistein on ET-1 synthesis could be
attributed to its inhibitory effects on tyrosine kinase.
This conclusion is further supported by the fact that the
inhibitory effect of genistein on ET-1 synthesis is not
blocked by ICI182780. Taken together, these findings provide evidence
that the inhibitory effects of estradiol on ET-1 synthesis
may be mediated in part by means of ER. This notion is supported by
the recent observation that overexpression of ER in
endothelial cells results in a dramatic decrease in
ET-1 secretion.16
Although the inhibitory effects of estradiol on stimulated ET-1 release are well documented,3 its inhibitory effects on basal synthesis of endothelin-1 remain controversial. Our finding that estradiol metabolites inhibit basal ET-1 synthesis suggest that, under in vivo conditions, estradiol would downregulate ET-1 release. This contention is indirectly supported by the findings that the expression of preproendothelin-1 is significantly upregulated in ovariectomized pigs.17
Our finding that estradiol is effective in inhibiting ET-1
synthesis, whereas 17-estradiol, estrone, estrone sulfate, and
estriol are much less active in this regard, indicates that the effects
of estrogens on ET-1 production vary considerably. This
conclusion is also supported by our finding that estradiol benzoate is
significantly less potent than estradiol in inhibiting ET-1 synthesis.
Our previous studies reveal similar differential
antimitogenic effects of these clinically used estrogens on
human vascular smooth muscle cells and cardiac
fibroblasts.12 15
Thus, the inconsistent and confusing effects of hormone
replacement therapy on cardiovascular disease
risk18 may be due in part to
the differential effects of clinically used estrogens on ET-1 synthesis
by endothelial cells.
ET-1 synthesis is known to be regulated by the MAPK
pathway.3 Inhibition of MEK by
PD98059 in endothelial cells is associated with
inhibition of ET-1 synthesis; whereas, stimulation of MAPK activity by
mitogens such as angiotensin II and FCS is known to
stimulate ET-1 synthesis.3
Because estradiol and its metabolites inhibit mitogen stimulated MAPK
activity in vascular smooth muscle
cells,12 14 we
hypothesize that inhibition of MAPK activity in
endothelial cells may provide a common pathway by which
estradiol and its metabolites inhibit ET-1 synthesis. Indeed, the
findings of the present study indicate that FCS-stimulated MAPK
activity is inhibited by estradiol and its metabolites. That TNF,
thrombin, angiotensin II, and FCS induce MAPK
activity,2 3 together
with our finding that estradiol and its metabolites inhibit MAPK
activity, suggests that the inhibitory effects of estradiol
and its metabolites on ET-1 synthesis are due to their
inhibitory effects on MAPK activity.
Could our findings be of physiological and clinical significance? Metabolism of estradiol to catecholestrogens is one of the most prominent pathways for estradiol metabolism, and catecholestrogens are precursors for methoxyestradiols. Thus, substantial amounts of estradiol metabolites would be available in vivo. Although the physiological levels of methoxyestradiol in humans are not known with accuracy, the serum levels of methoxyestrogens in pregnant women are 30 nmol/L.19 Moreover, rough estimates suggest that 2-methoxyestradiol levels may be several-fold higher than the levels of estradiol.20 With regard to catecholestrogens, the levels range between 0.12 to 0.3 µmol/L in peripheral blood and the rate of urinary excretion of 2-hydroxyestradiol is 20 to 180 µg/24 hours in urine.21 Both VSMCs and endothelial cells are well endowed with COMT,1 the enzyme responsible for metabolizing catecholestrogens to methoxyestrogens, thus likely ensuring pharmacologically active steady state levels of methoxyestradiol in the blood vessel wall.
That 2-hydroxyestradiol and 2-methoxyestradiol are more potent than estradiol in inhibiting ET-1 synthesis suggests that metabolism of estradiol may play an important role in mediating the overall cardiovascular protective effects of estradiol. This suggests that the cardioprotective effects of estradiol may vary and be dependent on the metabolic capability of the individual. Notably, estrogen replacement therapy is not beneficial in all postmenopausal women,1 and previous results indicate that estrogen replacement therapy in postmenopausal women differentially increases nitric oxide synthesis22 23 and decreases LDL levels.22 Moreover, recent studies show that estradiol must be metabolized to prevent LDL oxidation.24 On the basis of these findings, it is possible that the variable cardioprotective effects of estrogen that are observed in postmenopausal women may be due to lack of metabolism of estradiol to 2-hydroxyestradiol and 2-methoxyestradiol. This contention is further strengthened by our recent findings that methoxyestradiols and catecholestradiols mediate the antimitogenic effects of estradiol on vascular smooth muscle cell growth by means of an ER-independent mechanism.14
With regard to clinical relevance of the present findings, the overall protective effects of hormone replacement therapy may depend on the type of estrogen used. That estradiol and its metabolites inhibit ET-1 synthesis, whereas some clinically used estrogens are ineffective, suggests that estrogens may have differential protective effects on vascular biology.
In conclusion, we provide the first evidence that 2-hydroxyestradiol and 2-methoxyestradiol, the major endogenous metabolites of estradiol with no affinity for ERs, inhibit ET-1 secretion by PCAECs by an ER-independent mechanism. Moreover, we demonstrate that the inhibitory effects of estradiol on ET-1 synthesis are not mediated by ERß. Finally, estradiol and its metabolites may attenuate ET-1 synthesis by inhibiting MAPK activity.
|
Acknowledgments |
|---|
This study was supported by the Swiss National Science Foundation (grants 32-54172.98; 32-64040.00 and 32-55738.98), by the National Institutes of Health (grants HL35909 and HL5314), and by an unconditional educational grant from Schering (Schweiz AG).
Received October 25, 2000; first decision December 7, 2000; accepted December 19, 2000.
|
References |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
1. Dubey RK, Jackson EK. Estrogen-induced cardiorenal protection: potential cellular, biochemical and molecular mechanisms. Am J Physiol Renal Physiol. 2001;280:F365–F388.
2. Miyauchi T, Masaki T. Pathophysiology of endothelin in the cardiovascular system. Annu Rev Physiol. 1999;61:391–415.[Medline] [Order article via Infotrieve]
3. Morey AK, Razandi M, Pedram A, Hu R-M, Prins BA, Levin ER. Oestrogen and progesterone inhibit the stimulated production of endothelin-1. Biochem J. 1998;330:1097–1105.
4. Akishita M, Kozaki K, Eto M, Yoshizumi M, Ishikawa M, Toba K, Orimo H, Ouchi Y. Estrogen attenuates ednothelin-1 production by bovine endothelial cells via estrogen receptor. Biochem Biophys Res Commun.1998;251:17–21.
5. Iafrati MD, Karas RH, Aronovitz M, Kim S, Sullivan TR Jr, Lubahn DB, O’Donell TF Jr, Korach KS, Mendelsohn ME. Estrogen inhibits the vascular injury response in estrogen receptor alpha-deficient mice. Nat Med. 1997;3:545–548.[Medline] [Order article via Infotrieve]
6.
Karas RH, Hodgin JB,
Kwoun M, Krege JH, Aronovitz M, Mackey W, Gustafsson JA, Korach KS,
Smithies O, Mendelsohn ME. Estrogen inhibits the vascular injury
response in estrogen receptor beta-deficient female mice.
Proc Natl Acad Sci
U S A. 1999;96:15133–15136.
7.
Oparil S, Levine R,
Chen S-J, Durand J, Chen YF. Sexually dimorphic response of the
balloon-injured rat carotid artery to hormone treatment.
Circulation. 1997;95:1301–1307.
8.
Linder V, Kim
SK, Karas RH, Kuiper GGJM, Gustafsson J-A, Mendelsohn ME. Increased
expression of estrogen receptor ß mRNA in male blood vessels after
vascular injury. Circ Res. 1998;83:224–229.
9. Nishigaki I, Sasaguri Y, Yagi K. Anti-proliferative effect of 2-methoxyestradiol on cultured smooth muscle cells from rabbit aorta. Atherosclerosis. 1995;113:167–170.[Medline] [Order article via Infotrieve]
10.
Dubey RK, Tyurina
YY, Tyurin VA, Gillespie DG, Branch RA, Jackson EK, Kagan VE. Estrogen
and tamoxifen metabolites protect smooth muscle cell membrane
phospholipids against peroxidation and inhibit cell growth.
Circ Res. 1999;84:229–239.
11. Ohbayashi A, Hiraga T, Okubo M, Mutase T, Matsushita H, Hara M. Characteristics of porcine coronary artery endothelial cells in culture: comparison with aortic endothelium. Biochem Biophys Res Commun. 1994;202:504–511.[Medline] [Order article via Infotrieve]
12.
Dubey RK, Jackson
EK, Gillespie DG, Zacharia LC, Imthurn B, Keller P. Clinically used
estrogens differentially inhibit human aortic smooth muscle cell growth
and mitogen-activated protein kinase activity.
Arterioscler Thromb Vasc Biol. 2000;20:964–972.
13.
Kuiper GGJM,
Lemmen JG, Carlsson B, Corton JC, Safe SH, van der Saag PT, van der
Burg B, Gustafsson J-A. Interaction of estrogenic chemicals and
phytoestrogens with estrogen receptor ß.
Endocrinology. 1998;139:4252.4263.
14. Dubey RK, Gillespie DG, Zacharia LC, Rosselli M, Korzekwa KR, Fingerle J, Jackson EK. Methoxyestradiols mediate the antimitogenic effects of estradiol on vascular smooth muscle cells via estrogen receptor-independent mechanisms. Biochem Biophys Res Commun. 2000;278:27–33.[Medline] [Order article via Infotrieve]
15. Dubey RK, Gillespie DG, Jackson EK, Keller PJ. 17ß-estradiol, its metabolites, and progesterone inhibit cardiac fibroblast growth. Hypertension. 1998;31(pt 2):522–528.
16.
Ali SH, O’Donnell
AL, Mohamed S, Mousa S, Dandona P. Stable over-expression of estrogen
receptor- in ECV304 cells inhibits proliferation and levels of
secreted endothelin-1 and vascular endothelial growth
factor. Mol Cell Endocrinol. 1999;152:1–9.[Medline]
[Order article via Infotrieve]
17. Wang X, Barber DA, Lewis DA, McGregor CGA, Sieck GC, Fitzpatrick LA, Miller VM. Gender and transcriptional regulation of NO synthase and ET-1 in porcine aortic endothelial cells. Am J Physiol. 1997;273:H1962–H1967.
18. Oparil S. Hormones and Vasoprotection. Hypertension 1999;33(pt 2):170–176.
19.
Berg D, Sonsalla
R, Kuss E. Concentrations of 2-methoxyestradiol in human serum measured
by a heterologous immunoassay with an 125I-labelled ligand.
Acta Endocrinol (Copenh). 1983;103:282–288.
20.
Zhu BT, Conney AH.
Is 2-methoxyestradiol an endogenous estrogen metabolite
that inhibits mammary carcinogenesis.
Cancer Res. 1998;58:2269–2277.
21. Gelbke HP, Ball P, Knuppen R. 2-Hydroxyoestrogens: chemistry, biogenesis, metabolism and physiological significance. In: Briggs MH, Christie GA, eds. Advances in Steroid Biochemistry and Pharmacology. Vol 6. London, New York: Academic Press; 1977:81–154.
22.
Imthurn B,
Rosselli M, Jaeger AW, Keller PJ, Dubey RK. Differential effects of
hormone-replacement therapy on endogenous nitric oxide
(nitrate/nitrite) levels in postmenopausal women substituted with
17ß-estradiol valerate and cyproterone acetate or
medroxyprogesterone acetate.
J Clin Endocrinol Metab. 1997;82:388–394.
23.
Rosselli M,
Imthurn B, Keller PJ, Jackson EK, Dubey RK. Circulating nitric oxide
(nitrite/nitrate) levels in postmenopausal women substituted with
17ß-estradiol and norethisterone acetate: a 2 year follow-up-study.
Hypertension. 1995;25:848–853.
24.
Shwaery GT, Vita
JA, Keaney Jr JF. Antioxidant protection of LDL by
physiological concentrations of 17ß-estradiol:
requirement for estradiol modification.
Circulation. 1997;95:1378–1385.
This article has been cited by other articles:
 |
|
 |
|
  K. Athirakul, J. A. Bradbury, J. P. Graves, L. M. DeGraff, J. Ma, Y. Zhao, J. F. Couse, R. Quigley, D. R. Harder, X. Zhao, et al. Increased blood pressure in mice lacking cytochrome P450 2J5 FASEB J, December 1, 2008; 22(12): 4096 - 4108. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. L Jeanes, C. Tabor, D. Black, A. Ederveen, and G. A Gray Oestrogen-mediated cardioprotection following ischaemia and reperfusion is mimicked by an oestrogen receptor (ER){alpha} agonist and unaffected by an ER{beta} antagonist J. Endocrinol., June 1, 2008; 197(3): 493 - 501. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 V. M. Miller and S. P. Duckles Vascular Actions of Estrogens: Functional Implications Pharmacol. Rev., June 1, 2008; 60(2): 210 - 241. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. V. Casarez, M. E. Dunlap-Brown, M. R. Conaway, and G. P. Amorino Radiosensitization and Modulation of p44/42 Mitogen-Activated Protein Kinase by 2-Methoxyestradiol in Prostate Cancer Models Cancer Res., September 1, 2007; 67(17): 8316 - 8324. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Bourghardt, G. Bergstrom, A. Krettek, S. Sjoberg, J. Boren, and A. Tivesten The Endogenous Estradiol Metabolite 2-Methoxyestradiol Reduces Atherosclerotic Lesion Formation in Female Apolipoprotein E-Deficient Mice Endocrinology, September 1, 2007; 148(9): 4128 - 4132. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. R. Ferreri Estrogen-TNF interactions and vascular inflammation Am J Physiol Heart Circ Physiol, June 1, 2007; 292(6): H2566 - H2569. [Full Text] [PDF]
|
|
|
|
 |
|
 R. J. Paul, P. S. Bowman, J. Johnson, and A. F. Martin Effects of sex and estrogen on myosin COOH-terminal isoforms and contractility in rat aorta Am J Physiol Regulatory Integrative Comp Physiol, February 1, 2007; 292(2): R751 - R757. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. P. V. Dantas and K. Sandberg Does 2-Methoxyestradiol Represent the New and Improved Hormone Replacement Therapy for Atherosclerosis? Circ. Res., August 4, 2006; 99(3): 234 - 237. [Full Text] [PDF]
|
|
|
|
 |
|
 A. Kher, K. K. Meldrum, M. Wang, B. M. Tsai, J. M. Pitcher, and D. R. Meldrum Cellular and molecular mechanisms of sex differences in renal ischemia-reperfusion injury Cardiovasc Res, September 1, 2005; 67(4): 594 - 603. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. A. Khalil Sex Hormones as Potential Modulators of Vascular Function in Hypertension Hypertension, August 1, 2005; 46(2): 249 - 254. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. F. Reckelhoff Sex Steroids, Cardiovascular Disease, and Hypertension: Unanswered Questions and Some Speculations Hypertension, February 1, 2005; 45(2): 170 - 174. [Full Text] [PDF]
|
|
|
|
 |
|
 R. K. Dubey, S. P. Tofovic, and E. K. Jackson Cardiovascular Pharmacology of Estradiol Metabolites J. Pharmacol. Exp. Ther., February 1, 2004; 308(2): 403 - 409. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. M. Orshal and R. A. Khalil Gender, sex hormones, and vascular tone Am J Physiol Regulatory Integrative Comp Physiol, February 1, 2004; 286(2): R233 - R249. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. L. Lattanzi, C. B. Santos, M. D. Mudry, and J. L. Baranao Exposure of Bovine Oocytes to the Endogenous Metabolite 2-Methoxyestradiol During In Vitro Maturation Inhibits Early Embryonic Development Biol Reprod, December 1, 2003; 69(6): 1793 - 1800. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Y. Shimoni and X.-F. Liu Sex differences in the modulation of K+ currents in diabetic rat cardiac myocytes J. Physiol., July 15, 2003; 550(2): 401 - 412. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. C. Reinhart, R. K. Dubey, B. Cometti, P. J. Keller, and M. Rosselli Differential Effects of Natural and Environmental Estrogens on Endothelin Synthesis in Bovine Oviduct Cells Biol Reprod, April 1, 2003; 68(4): 1430 - 1436. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Funke-Kaiser, F. Reichenberger, K. Kopke, S.-M. Herrmann, J. Pfeifer, H.-D. Orzechowski, W. Zidek, M. Paul, and E. Brand Differential binding of transcription factor E2F-2 to the endothelin-converting enzyme-1b promoter affects blood pressure regulation Hum. Mol. Genet., February 15, 2003; 12(4): 423 - 433. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T.-M. Lee, T.-F. Chou, and C.-H. Tsai Differential Role of KATP Channels Activated by Conjugated Estrogens in the Regulation of Myocardial and Coronary Protective Effects Circulation, January 7, 2003; 107(1): 49 - 54. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Liu, S.-H. Yang, E. Perez, K. D. Yi, S. S. Wu, K. Eberst, L. Prokai, K. Prokai-Tatrai, Z. Y. Cai, D. F. Covey, et al. Neuroprotective Effects of a Novel Non-Receptor-Binding Estrogen Analogue: In Vitro and In Vivo Analysis Stroke, October 1, 2002; 33(10): 2485 - 2491. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. L. Moreau, A. J. Donato, D. R. Seals, F. A. Dinenno, S. D. Blackett, G. L. Hoetzer, C. A. Desouza, and H. Tanaka Arterial intima-media thickness: site-specific associations with HRT and habitual exercise Am J Physiol Heart Circ Physiol, October 1, 2002; 283(4): H1409 - H1417. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. M. LaVallee, X. H. Zhan, C. J. Herbstritt, E. C. Kough, S. J. Green, and V. S. Pribluda 2-Methoxyestradiol Inhibits Proliferation and Induces Apoptosis Independently of Estrogen Receptors {alpha} and {beta} Cancer Res., July 1, 2002; 62(13): 3691 - 3697. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. C. Villablanca, K. A. Lewis, and J. C. Rutledge Time- and dose-dependent differential upregulation of three genes by 17beta -estradiol in endothelial cells J Appl Physiol, March 1, 2002; 92(3): 1064 - 1073. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Fischer, A. Baessler, and H. Schunkert Renin angiotensin system and gender differences in the cardiovascular system Cardiovasc Res, February 15, 2002; 53(3): 672 - 677. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. K. Dubey, S. Oparil, B. Imthurn, and E. K. Jackson Sex hormones and hypertension Cardiovasc Res, February 15, 2002; 53(3): 688 - 708. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. K. Dubey, D. G. Gillespie, and E. K. Jackson A2B Adenosine Receptors Stimulate Growth of Porcine and Rat Arterial Endothelial Cells Hypertension, February 1, 2002; 39(2): 530 - 535. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. F. Kuebler, D. Jarrar, B. Toth, K. I. Bland, L. Rue III, P. Wang, and I. H. Chaudry Estradiol Administration Improves Splanchnic Perfusion Following Trauma-Hemorrhage and Sepsis Arch Surg, January 1, 2002; 137(1): 74 - 79. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. P. Tofovic, R. K. Dubey, and E. K. Jackson 2-Hydroxyestradiol Attenuates the Development of Obesity, the Metabolic Syndrome, and Vascular and Renal Dysfunction in Obese ZSF1 Rats J. Pharmacol. Exp. Ther., December 1, 2001; 299(3): 973 - 977. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. K. Dubey and E. K. Jackson Genome and Hormones: Gender Differences in Physiology: Invited Review: Cardiovascular protective effects of 17{beta}-estradiol metabolites J Appl Physiol, October 1, 2001; 91(4): 1868 - 1883. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. L. David, M. H. C. Carvalho, A. L.N. Cobra, D. Nigro, Z. B. Fortes, N. A. Reboucas, and R. C.A. Tostes Ovarian Hormones Modulate Endothelin-1 Vascular Reactivity and mRNA Expression in DOCA-Salt Hypertensive Rats Hypertension, September 1, 2001; 38(3): 692 - 696. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||