This Article Abstract Full Text (PDF) Submit a related Letter to the Editor Purchase Article View Shopping Cart Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Copyright Permission Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Cho, J. Articles by Lee, Y. Search for Related Content PubMed PubMed Citation Articles by Cho, J. Articles by Lee, Y.

Ginsenoside-Rb1 from Panax ginseng C.A. Meyer Activates Estrogen Receptor- and -ß, Independent of Ligand Binding

College of Life Science (JY.C., W.P., YJ.L.), Institute of Biotechnology, Department of Bioscience and Biotechnology, Sejong University, Seoul 143-747, Korea; College of Pharmacy (SK.L.), Seoul National University, Seoul 151-742, Korea; and Department of Obstetrics and Gynecology (W.A.), Catholic Research Institutes of Medical Science, College of Medicine, The Catholic University of Korea, Seoul 137-701, Korea

Address all correspondence and requests for reprints to: YoungJoo Lee, Ph.D., Department of Bioscience and Biotechnology, Sejong University, Kwang-Jin-Gu, Seoul 143-747, Korea. E-mail: yjlee{at}sejong.ac.kr.

	Abstract

Top Abstract Introduction Materials and Methods Results Discussion References

 
We studied the estrogenic activity of a component of Panax ginseng, ginsenoside-Rb1. The activity of ginsenoside-Rb1 was characterized in a transient transfection system, using estrogen receptor isoforms and estrogen-responsive luciferase plasmids, in COS monkey kidney cells. Ginsenoside-Rb1 activated both and ß estrogen receptors in a dose-dependent manner with maximal activity observed at 100 µM, the highest concentration examined. Activation was inhibited by the estrogen receptor antagonist ICI 182,780, indicating that the effects were mediated through the estrogen receptor. Treatment with 17ß-estradiol or ginsenoside-Rb1 increased expression of the progesterone receptor, pS2, and estrogen receptor in MCF-7 cells and of AP-1-driven luciferase genes in COS cells. Although these data suggest that it is functionally very similar to 17ß-estradiol, ginsenoside-Rb1 failed to displace specific binding of [³H]17ß-estradiol from estrogen receptors in MCF-7 whole-cell ligand binding assays. Our results indicate that the estrogen-like activity of ginsenoside-Rb1 is independent of direct estrogen receptor association.

	Introduction

Top Abstract Introduction Materials and Methods Results Discussion References

 
GINSENG HAS BEEN used for over 2000 yr in oriental countries to enhance stamina and immune function, where it has been suggested to have pharmacological activities in the cardiovascular, endocrine, immune, and central nervous systems (1). Its use has expanded to Western countries and continues to rise with the increasing popularity of complementary and alternative medicine. It is one of the best-selling herbs in the United States (2). Although many studies have examined the pharmacological action of ginseng extracts, a detailed mechanism has yet to be determined. The major pharmacologically active components of ginseng are ginsenosides, which are steroidal saponins comprising 3–6% of ginseng (3). It has been shown that ginsenosides decrease the levels of total cholesterol and triglyceride via cAMP production and inhibit the accumulation of calcium ions in liver cells (4). Ginsenosides potentiate analgesia and inhibit analgesic tolerance (5). The cardioprotective action of ginsenosides is because of effects on vasodilation via nitric oxide (NO) release (6, 7). Other activities, such as anticarcinogenic and neurological effects, have also been reported for ginsenosides (8, 9).

In the United States, ginseng is used to alleviate menopausal symptoms, as are black cohosh (Cimicifuga racemosa), chaste tree berry (Vitex agnus-castus), dong quai (Angelica sinensis), evening primrose oil (Oenethera biennis), motherwort (Leonurus cardiaca), red clover (Trifolium pratense), and licorice (Glycyrrhiza glabra) (10). One recent randomized controlled clinical trial showed that only black cohosh had a beneficial effect on postmenopausal hot flashes (10). Other in vitro studies have measured estrogenic activity in red clover and licorice, which was not demonstrated in black cohosh (11). Various studies have indicated that ginseng has estrogenic activity, although no clinical trials have demonstrated real efficacy as an estrogen-replacement therapy (10). Ginseng extracts are able to stimulate the growth of estrogen receptor (ER)-positive cells (12). Ginsenoside-Rg1 and -Rh1 have been shown to contain estrogen-like activity (13, 14), but more comprehensive data are needed to adequately evaluate this activity.

Among 26 identified ginsenosides, ginsenoside-Rb1, -Ro, -Rg1, -Rc, and -Re are highly abundant. In particular, ginsenoside-Rb1 makes up 0.37–0.5% of ginseng extracts, depending on manufacturing and processing methods, and belongs to the protopanaxadiol class of ginsenosides (http://www.netnam.vn/icasia/english/products/redkogin/redkogind/redkogind.htm). We have previously reported that ginsenoside-Rb1 has estrogenic activity (15). In the present study, we aimed to characterize the differential activity of ginsenoside-Rb1 toward ER isoforms and ß. We examined its ability to induce endogenous ER-responsive genes and to act as an ER ligand. Our data show that ginsenoside-Rb1 activates both ERs and ß in the absence of receptor binding.

	Materials and Methods

Top Abstract Introduction Materials and Methods Results Discussion References

 
Reagents

Ginsenoside-Rb1 was provided by the Korea Ginseng and Tobacco Research Institute (Daejeon, Korea). 17ß-Estradiol (E2) was purchased from Sigma Chemical Co. (St. Louis, MO). ICI 182,780 (ICI) was obtained from ZENECA Pharmaceuticals (Tocris, UK). Ginsenoside-Rb1 was dissolved in 20% ethanol at a concentration of 15 mg/ml. E2 was dissolved in 100% ethanol. All the compounds were added to the medium such that the total ethanol concentration was never higher than 0.15%. An untreated group served as a control.

Plasmids

ERE2-tk81-luc, constructed by inserting the fragment of the herpes simplex thymidine kinase promoter and two copies of the vitellogenin estrogen-responsive element (ERE) into pA3luc (16), was a kind gift of Dr. Larry Jameson. Expression vector for ERs and ß were from Dr. Pierre Chambon and Dr. Vincent Giguere, respectively. pAP-1-LUC plasmid was purchased from Stratagene (La Jolla, CA).

Cell cultures

ER-positive human breast adenocarcinoma, MCF-7, cells were purchased from the Korea Cell Line Bank. MCF-7 cells were maintained in phenol red-free DMEM containing 1x antibiotic/antimycotic mix (Invitrogen, Gaithersburg, MD), 5 mM HEPES, and 0.37% sodium bicarbonate, supplemented with 10% fetal bovine serum (FBS) (HyClone, Logan, UT). COS cells were maintained under identical conditions except that 10% calf serum was used instead of FBS. Cells were grown at 37 C in a humidified atmosphere of 95% air/5% CO₂ and fed every 3–4 d. Before hormone induction, the cells were washed with PBS and cultured in DMEM/10% charcoal-dextran stripped FBS (CD-FBS) for 2 d to eliminate any estrogenic source before treatment. All treatments were done with DMEM/10% CD-FBS, and 10 nM E2 was used to maximize the response unless otherwise noted.

Transient transfection and luciferase assays

Cells were seeded in 24-well plates at a density of 7 x 10⁴ cells per well. After 24 h, plasmids were transiently transfected into the cell by a calcium phosphate-DNA coprecipitation method. A total of 0.5 µg of DNA in 25 µl of CaCl₂·H₂O (250 mM CaCl₂) was mixed with 25 µl of 2x HBS (280 mM NaCl, 10 mM KCl, 1.5 mM Na₂HPO₄·2H₂O, 12 mM dextrose, and 50 mM HEPES) with constant bubbling, and within 5–10 min this solution was added to each well. The next day, transfected cells were washed with PBS and treated with compounds. Luciferase activity was determined 24 or 48 h after drug treatments with an AutoLumaat LB953 luminometer using the luciferase assay system (Promega Corp., Madison, WI) and expressed as relative light units. The mean and SEs of triplicate or quadruplicate samples are shown for representative experiments. All transfection experiments were repeated three or more times with similar results.

RNA extraction and RT-PCR

MCF-7 cells were grown in six-well plates in phenol red-free DMEM containing 10% CD-FBS. Near-confluent monolayers were treated with the compounds for 24 h. The wells were rinsed in PBS, and total RNA was isolated by lysing the cells in guanidinium isothiocyanate using the TRIzol reagent (Invitrogen) according to the manufacturer’s instructions. After extraction, RNA was precipitated by recommended procedures and dissolved in diethylpyrocarbonate-treated water. To synthesize first-strand cDNA, 5 µl total RNA was incubated in 0.5 µg of oligo(dT)₁₈ primer (Invitrogen) and 5 µl deionized water at 70 C for 5 min. RT reactions were performed using 40 U of Moloney murine leukemia virus reverse transcriptase (Promega Corp.) in 5x reaction buffer [250 mM Tris-HCl (pH 8.3) at 25 C, 375 mM KCl, 15 mM MgCl₂, and 50 mM dithiothreitol] and 20 mM dNTP mixtures at 37 C for 60 min. The reaction was terminated by heating at 70 C for 10 min, followed by cooling at 4 C. The resulting cDNA was added to the PCR mixture containing 10x PCR buffer [100 mM Tris-HCl (pH 8.3), 500 mM KCl, and 15 mM MgCl₂], 25 U of rTaq polymerase (TaKaRa, Japan), 4 µl of 2.5 mM dNTP mixtures, and 10 pmol of primers each. The final volume was 50 µl. The sequences for the human progesterone receptor (PR) and pS2 primers were 5'-CCA TGT GGC AGA TCC CAC AGG AGT T-3' and 5'-TGG AAA TTC AAC ACT CAG TGC CCG G-3' for PR (17) and 5'-CAT GGA GAA CAA GGT GAT CTG -3' and 5'-CAG AAG CGT GTC TGA GGT GTC-3' for pS2 (18), and those of human ß-actin were 5'-CCT GAC CCT GAA GTA CCC CA-3' and 5'-CGT CAT GCA GCT CAT AGC TC-3' (19). The PCR product for PR is 271 bp and 365 bp for pS2 and 550 bp for ß-actin. The reactions were initiated by 3 min of denaturation at 94 C followed by amplification at 94 C for 45 sec, 55 C for 45 sec, and 72 C for 45 sec; 24 cycles for PR or pS2 and 20 cycles for ß-actin. The PCR was ended by elongation at 72 C for 5 min. The PCR products were analyzed by 2% agarose gel electrophoresis and visualized by ethidium bromide staining, quantified using a bio-imaging analyzer (Bio-Rad Laboratories, Inc., Hercules, CA), and band intensity was normalized to the intensity of ß-actin mRNA.

Western blotting

Protein was isolated by using radioimmune precipitation buffer [containing 150 mM NaCl, 50 mM Tris-HCl, 5 mM EDTA, 1% Nonidet P-40, 0.5% deoxycholate, 1% SDS with protease inhibitor cocktail (Sigma)] on ice for 1 h and then centrifuged for 20 min at 13,000 x g. Supernatant was collected, and protein concentrations were measured using the Bradford method (Bio-Rad). Fifty micrograms of protein were dissolved in sample buffer and boiled for 5 min before loading onto an 8% acrylamide gel. After SDS-PAGE, proteins were transferred to a polyvinylidene difluoride membrane, blocked with 5% nonfat dry milk in Tris-buffered saline/0.05% Tween, and incubated with rabbit anti-polyclonal antibody to ER (0.4 mg/ml; Santa Cruz Biotechnology, Inc., Santa Cruz, CA) for 2 h at 1:500. After washing with Tris-buffered saline/0.05% Tween, blots were incubated with goat antirabbit horseradish peroxidase-conjugated secondary antibody and visualized with enhanced chemiluminescence ECL kits (Amersham Bioscience, Little Chalfont, UK).

ER binding assay

MCF-7 cells were stripped of any estrogen by keeping them in phenol red-free DMEM supplemented with 10% CD-FBS for 2 d. Cells were incubated with 1 nM [2,4,6,7-³H]E2 (89 Ci/mmol; Amersham Bioscience) and a 100-fold excess of nonlabeled E2 (100 nM) or 25–50 µM ginsenoside-Rb1 for 1 h at 37 C in a humidified atmosphere of 95% air/5% CO₂. Aliquots of the medium were measured before and after the incubation with the cells to determine the total uptake of E2 into the cells. After removal of the medium, cells were washed with ice-cold PBS/0.1% methylcellulose twice, harvested by scraping and centrifugation, and lysed with 100% ethanol, 500 µl per 60 mm dish, for 10 min at room temperature (20). The radioactivity of extracts was measured by liquid scintillation counting.

Statistical analysis

Values shown represent mean ± SD. Statistical analysis was performed by Student’s t test with a P value of less than 0.05 being considered statistically significant.

	Results

Top Abstract Introduction Materials and Methods Results Discussion References

 
Ginsenoside-Rb1 activates estrogen-responsive luciferase genes in the presence of either ER or ERß

We have previously demonstrated that ginsenoside-Rb1 (Fig. 1) activates ER in MCF-7 cells (15). It has been shown that some phytoestrogens such as genistein and coumestrol differ in their activity on ER and ERß (21). In the present study, we investigated whether there is differential activation of these two receptor isoforms in response to ginsenoside-Rb1 by examining the transcription of an EREcontaining reporter plasmid in the presence of ER or ERß in ER-negative COS cells. Ginsenoside-Rb1 activated both ER and ERß in a dose-dependent manner (Fig. 2A). Luciferase gene activation was observed over two orders of ginsenoside-Rb1 concentration up to 100 µM, the highest concentration tested. Higher concentrations would have resulted in unacceptably high concentrations of ethanol in the cell media. In the presence of the ER isoform, ginsenoside-Rb1 and E2 activated ERE-driven luciferase expression to a similar extent (91% of E2 activity); approximately 60% of the E2 response was seen with the ERß isoform (Fig. 2A). These data indicate that ginsenoside-Rb1 acts as a dose-dependent agonist, stimulating transcription through both receptors, but that it has slightly higher affinity for ER than for ERß.

View larger version (15K): [in this window] [in a new window]   FIG. 1. Chemical structure of ginsenoside-Rb1.

View larger version (24K): [in this window] [in a new window]   FIG. 2. Ginsenoside-Rb1-induced transactivation was assessed by cotransfection of the (ERE)-luciferase with either ER or ERß expression plasmids in COS cells. Cell extracts were prepared and analyzed by a luciferase assay. A, Ginsenoside-Rb1 activated both ER and ERß in a dose-dependent manner. Cells were exposed to E2 (10 nM), ginsenoside-Rb1 (100–0.5 µM) for 24 h. *, P < 0.05; **, P < 0.005. B, Ginsenoside-Rb1 activation was inhibited by ICI. Cells were exposed to E2 (10 nM) or ginsenoside-Rb1 (100 µM) or in combination with ICI (1 µM) for 24 h. The data are representative of at least three independent experiments performed in triplicate with similar results, expressed as relative luciferase units and the SEM of triplicate samples. *, P < 0.05; **, P < 0.005.

Ginsenoside-Rb1 activates AP-1 luciferase reporter plasmids, PR and pS2 mRNA, and ER protein in an ER-dependent manner

To further characterize the effects of ginsenoside-Rb1, the effects on ER-mediated AP-1 responses, endogenous estrogen-responsive PR and pS2 mRNAs, and ER protein were examined. Ligand-occupied ERs are known to mediate gene transcription from AP-1 enhancer elements (22). AP-1 is implicated in the inducible expression of a variety of genes involved in the regulation of proliferation and apoptosis, as well as in cellular stress responses and inflammation (23). With transient ER expression in COS cells, both ginsenoside-Rb1 and E2 stimulated AP-1-driven luciferase plasmid transactivation (Fig. 3A). Ginsenoside-Rb1 did not affect luciferase production in the absence of ER (data not shown). After treatment of MCF-7 cells with the compounds for 24 h, steady-state mRNA levels were measured by RT-PCR of total RNA, as indicated in Fig. 3B. Constitutively expressed human ß-actin mRNA was used as an internal control. Ginsenoside-Rb1 increased steady-state mRNA levels of the PR and pS2 genes after 24 h of treatment, as did E2 (Fig. 3B). Coincubation with 1 µM ICI efficiently blocked the activation of PR and pS2 mRNA expression, indicating activation through ER. We also have examined the ER protein levels in MCF-7 cells by Western analysis. ER protein levels were down-regulated after 24 h of either E2 or ginsenoside-Rb1 treatments as compared with control (Fig. 3C). These data indicate that ginsenoside-Rb1 is capable of activating an ER-mediated pathway.

View larger version (45K): [in this window] [in a new window]   FIG. 3. Effects of ginsenoside-Rb1 on AP1mediated activation, PR and pS2 mRNA, and ER protein levels. A, Ginsenoside-Rb1-induced expression of AP1-driven reporter plasmids. COS cells were transiently transfected with an AP1-containing reporter and ER expression plasmids and treated with E2 (10 nM) or ginsenoside-Rb1 (50 µM) at the indicated concentrations for 24 h. Cell extracts were prepared and analyzed by a luciferase assay. The data are representative of at least three independent experiments performed in triplicate with similar results, expressed as relative luciferase units and the SEM of triplicate samples. B, The semiquantitative RT-PCR for PR and pS2 in the MCF-7 cells shown is a representative of three independent experiments. Cells were exposed to ginsenoside-Rb1 (50 µM) with and without ICI (1 µM) for 24 h. PR amplification product was detected at 271 bp (left) and 365 bp for pS2 (right). C, The Western blot analysis shown is a representative of two independent experiments. Cells were treated with E2 (10 nM) or ginsenoside-Rb1 (50 µM) alone for 24 h. ER protein was detected at 62 kDa (right). Equal loading of protein in each lane was confirmed by ß-actin protein (43 kDa). ER densitometry values are expressed as a percentage of the control (left).

To determine whether ginsenoside activation occurs via direct binding to ER, we examined the ability of ginsenoside-Rb1 to compete with ³H-labeled E2 for ER binding in MCF-7 cells (Fig. 4). Specific binding was calculated as total binding minus nonspecific binding (determined in the presence of unlabeled E2 or ginsenoside-Rb1 at the concentration as indicated in the figure). At 50 µM, ginsenoside-Rb1 did not inhibit ³H-labeled E2 binding to ER (Fig. 4). We also failed to observe any binding in a cell line that was stably transfected with ER (data not shown). In contrast, ginsenoside-Rh1, a weak phytoestrogen, displaced approximately 44% of ³H-labeled E2 binding to ER at a concentration of 50 µM in MCF-7 cells.

View larger version (11K): [in this window] [in a new window]   FIG. 4. Competitive binding of ginsenside-Rb1 to ER in MCF-7 cells. Ginsenoside-Rb1 failed to compete E2 occupancy of ER. Cells were incubated with 1 nM 3H-labeled E2 (89 Ci/mmol) plus vehicle, nonlabeled E2 (100 nM), ginsenoside-Rb1 (25–50 µM), or ginsenoside-Rh1 (50 µM) for 1 h. Radioactivity of ethanol extracts of cell lysates was measured by scintillation counting. Specific binding was calculated as total binding minus nonspecific binding. Data were expressed as percentage of ligand binding, which is (total binding – nonspecific binding)/total binding x 100. *, P < 0.05.

	Discussion

Top Abstract Introduction Materials and Methods Results Discussion References

In Asia, ginseng has been used for thousands of years as a tonic, to fight various aspects of stress, and to restore homeostasis (1). In Western countries, it is being used as an alternative herb for postmenopausal women. Accumulating evidence suggests that ginseng contains either direct or indirect estrogenic activity (29). Ginseng extracts activate estrogen-responsive genes and regulate the growth of human breast cancer cells (12). Recent studies by Chan et al. (13) showed that picomolar ginsenoside-Rg1 from Panax notoginseng activated ER-mediated transcription without direct receptor interaction. Our group has reported estrogenic activity of ginsenoside-Rh1 in the micromolar range (14). Although ginsenosides share structural similarities with estrogen and may activate ERs, more detailed studies are needed to fully characterize their activities (12, 29).

The results from other studies in different systems indirectly suggest the regulation of estrogen-responsive genes by ginsenoside-Rb1. It was shown to decrease cardiac contraction in adult rat ventricular myocytes, in part through an increase in NO production (6). Although a correlation between the increase in NO and ER activation was not evaluated, estrogen is known to enhance NO production (30). Ginsenoside-Rb1 also regulates adrenal tyrosine hydroxylase (31), which is known to be under estrogen regulation (32).

The data presented here show that ginsenoside-Rb1 activated both ER and ERß, leading to the transactivation of estrogen-responsive genes. However, this activation occurred in the absence of direct receptor binding, as examined using receptor competition assays. This indicates that ginsenoside-Rb1 activates ER via a mechanism or mechanisms other than that of classical, hormone-mediated activation. A variety of agents, including growth factors, neurotransmitters, and cAMP, activate ER in ligand-independent manners (33). Recent data on the pharmacokinetics of ginsenoside-Rb1 show that its absolute oral bioavailability is as low as 4.35% in rats (34). Based on a calculation using the figures in the report by Xu et al. (34), approximately 180 mg/kg ginsenoside-Rb1 should be taken orally to obtain a serum concentration of 50 µM, the concentration that activated estrogen receptors in our assays. The ginsenoside persists for 3 d, but because of its low absorption, initial administration or formulation changes are necessary before clinical application (34). It has been shown recently that ginsenoside-Rb1 is teratogenic in the rat embryo at concentrations over 30 µM (35). Potential toxicity should be kept in mind during the clinical development of this compound. The exact cause of the estrogenic effects of ginseng may not be ginsenside-Rb1, because of its low serum concentration despite its in vitro estrogenic activity. However, these approaches are essential to provide a scientific rationale for using ginseng for estrogen-related symptoms. In this report, we have addressed the in vitro mechanism of one of the main components of ginseng. Studies investigating the upstream targets of ginsenoside-Rb1 that lead to ER activation and in vivo studies will improve our understanding of the clinical application of ginseng.

	Footnotes

 
This work was supported in part by grants from the Korean Science and Engineering Foundation (2000-20700-005-3; SK.L.), Korea Tobacco and Ginseng Corp. R&D Program 2000 (YJ.L.), Korean Ministry of Health and Welfare (HMP-00-O-21600-0009 to YJ.L.), and BK21 program (YJ.L.).

JY.C. and W.P. contributed equally to this work.

Abbreviations: CD-FBS, Charcoal-dextran stripped fetal bovine serum; E2, 17ß-estradiol; ER, estrogen receptor; ERE, estrogen-responsive element; PR, progesterone receptor.

Received October 20, 2003.

Accepted March 18, 2004.

	References

Top Abstract Introduction Materials and Methods Results Discussion References

 

1. Attele AS, Wu JA, Yuan CS 1999 Ginseng pharmacology: multiple constituents and multiple actions. Biochem Pharmacol 58:1685–1693[CrossRef][Medline]
2. Yuan CS, Wu JA 2002 Ginsenoside variability in American ginseng samples. Am J Clin Nutr 75:600–601[Free Full Text]
3. Yoon M, Lee H, Jeong S, Kim JJ, Nicol CJ, Nam KW, Kim M, Cho BG, Oh GT 2003 Peroxisome proliferator-activated receptor is involved in the regulation of lipid metabolism by ginseng. Br J Pharmacol 138:1295–1302[CrossRef][Medline]
4. Park KH, Shin HJ, Song YB, Hyun HC, Cho HJ, Ham HS, Yoo YB, Ko YC, Jun WT, Park HJ 2002 Possible role of ginsenoside Rb1 on regulation of rat liver triglycerides. Biol Pharm Bull 25:457–460[CrossRef][Medline]
5. Nemmani KV, Ramarao P 2003 Ginsenoside Rf potentiates U-50,488Hinduced analgesia and inhibits tolerance to its analgesia in mice. Life Sci 72:759–768[CrossRef][Medline]
6. Scott GI, Colligan PB, Ren BH, Ren J 2001 Ginsenosides Rb1 and Re decrease cardiac contraction in adult rat ventricular myocytes: role of nitric oxide. Br J Pharmacol 134:1159–1165[CrossRef][Medline]
7. Chen X 1996 Cardiovascular protection by ginsenosides and their nitric oxide releasing action. Clin Exp Pharmacol Physiol 23:728–732[Medline]
8. Ernst E, Cassileth BR 1999 How useful are unconventional cancer treatments? Eur J Cancer 35:1608–1613
9. Choi S, Jung SY, Ko YS, Koh SR, Rhim H, Nah SY 2002 Functional expression of a novel ginsenoside Rf binding protein from rat brain mRNA in Xenopus laevis oocytes. Mol Pharmacol 61:928–935[Abstract/Free Full Text]
10. Kronenberg F, Fugh-Berman A 2002 Complementary and alternative medicine for menopausal symptoms: a review of randomized, controlled trials. Ann Intern Med 137:805–813[Abstract/Free Full Text]
11. Oerter Klein K, Janfaza M, Wong JA, Chang RJ 2003 Estrogen bioactivity in Fo-Ti and other herbs used for their estrogen-like effects as determined by a recombinant cell bioassay. J Clin Endocrinol Metab 88:4077–4079[Abstract/Free Full Text]
12. Duda RB, Zhong YZ, Navas V, Li MZC, Toy BR, Alavarez JG 1999 American ginseng and breast cancer therapeutic agents synergistically inhibit MCF-7 breast cancer cell growth. J Surg Oncol 72:230–239[CrossRef][Medline]
13. Chan RY, Chen WF, Dong A, Guo D, Wong MS 2002 Estrogen-like activity of ginsenoside Rg1 derived from Panax notoginseng. J Clin Endocrinol Metab 87:3691–3695[Abstract/Free Full Text]
14. Lee Y, Jin Y, Lim W, Ji S, Choi S, Jang S, Lee S 2003 A ginsenoside-Rh1, a component of ginseng saponin, activates estrogen receptor in human breast carcinoma MCF-7 cells. J Steroid Biochem Mol Biol 84:463–468[CrossRef][Medline]
15. Lee YJ, Jin YR, Lim WC, Park WK, Cho JY, Jang S, Lee SK 2003 Ginsenoside-Rb1 acts as a weak phytoestrogen in MCF-7 human breast cancer cells. Arch Pharm Res 26:58–63[Medline]
16. Gehm BD, McAndrews JM, Chien PY, Jameson JL 1997 Resveratrol, a polyphenolic compound found in grapes and wine, is an agonist for the estrogen receptor. Proc Natl Acad Sci USA 94:14138–14143[Abstract/Free Full Text]
17. Mercier G, Turque N, Schumacher M 2001 Rapid effects of triiodothyronine on immediate-early gene expression in Schwann cells. Glia 2001 35:81–89[CrossRef][Medline]
18. Liu J, Burdette JE, Xu H, Gu C, van Breemen RB, Bhat KP, Booth N, Constantinou AI, Pezzuto JM, Fong HH, Farnsworth NR, Bolton JL 2001 Evaluation of estrogenic activity of plant extracts for the potential treatment of menopausal symptoms. J Agric Food Chem 49:2472–2479[CrossRef][Medline]
19. Ren L, Marquardt MA, Lech JJ 1997 Estrogenic effect of nonylphenol on pS2, ER and MUC1 gene expression in human breast cancer cells-MCF-7. Chem Biol Interact 104:55–64[CrossRef][Medline]
20. Lee YJ, Gorski J 1996 Estrogen-induced transcription of the progesterone receptor gene does not parallel estrogen receptor occupancy. Proc Natl Acad Sci USA 93:15180–15184[Abstract/Free Full Text]
21. Kuiper GG, Lemmen JG, Carlsson B, Corton JC, Safe SH, Saag PT, Burg B Gustafsson JA 1998 Interaction of estrogenic chemicals and phytoestrogens with estrogen receptor ß. Endocrinology 139:4252–4263[Abstract/Free Full Text]
22. Paech K, Webb P, Kuiper GG, Nilsson S, Gustafsson J, Kushner PJ, Scanlan TS 1997 Differential ligand activation of estrogen receptors ER and ERß at AP1 sites. Science 277:1508–1510[Abstract/Free Full Text]
23. Maggi-Capeyron MF, Ceballos P, Cristol JP, Delbosc S, Le Doucen C, Pons M, Leger CL, Descomps B 2001 Wine phenolic antioxidants inhibit AP-1 transcriptional activity. J Agric Food Chem 49:5646–5652[CrossRef][Medline]
24. Woo J, Lau E, Ho SC, Cheng F, Chan C, Chan AS, Haines CJ, Chan TY, Li M, Sham A 2003 Comparison of Pueraria lobata with hormone replacement therapy in treating the adverse health consequences of menopause. Menopause 10:352–361[CrossRef][Medline]
25. Cos P, De Bruyne T, Apers S, Vanden Berghe D, Pieters L, Vlietinck AJ 2003 Phytoestrogens: recent developments. Planta Med 69:589–599[CrossRef][Medline]
26. Cassidy A 2003 Potential risks and benefits of phytoestrogen-rich diets. Int J Vitam Nutr Res 73:120–126[CrossRef][Medline]
27. Mishra SI, Dickerson V, Najm W 2003 Phytoestrogens and breast cancer prevention: what is the evidence? Am J Obstet Gynecol 188:66–70[CrossRef]
28. Glazier MG, Bowman MA 2001 A review of the evidence for the use of phytoestrogens as a replacement for traditional estrogen replacement therapy. Arch Intern Med 161:1161–1172[Abstract/Free Full Text]
29. Amato P, Christophe S, Mellon PL 2002 Estrogenic activity of herbs commonly used as remedies for menopausal symptoms. Menopause 9:145–150[CrossRef][Medline]
30. Farhat MY, Lavigne MC, Ramwell PW 1996 The vascular protective effects of estrogen. FASEB J 10:615–624[Abstract]
31. Kim HS, Zhang YH, Fang LH, Lee MK 1999 Effects of ginsenosides on bovine adrenal tyrosine hydroxylase. J Ethnopharmacol 66:107–111[CrossRef][Medline]
32. Ivanova T, Beyer C 2003 Estrogen regulates tyrosine hydroxylase expression in the neonate mouse midbrain. J Neurobiol 54:638–647[CrossRef][Medline]
33. Pasapera Limon AM, Herrera-Munoz J, Gutierrez-Sagal R, Ulloa-Aguirre A 2003 The phosphatidylinositol 3-kinase inhibitor LY294002 binds the estrogen receptor and inhibits 17ß-estradiol-induced transcriptional activity of an estrogen sensitive reporter gene. Mol Cell Endocrinol 200:199–202[CrossRef][Medline]
34. Xu QF, Fang Xl, Chen DF 2003 Pharmacokinetics and bioavailability of ginsenoside Rb1 and Rg1 from Panax notoginseng in rats. J Ethnopharmacol 84:187–192[CrossRef][Medline]
35. Chan LY, Chiu PY, Lau TK 2003 An in-vitro study of ginsenoside rb1-induced teratogenicity using a whole rat embryo culture model. Hum Reprod 18:2166–2168[Abstract/Free Full Text]

This article has been cited by other articles:

				M. L. King, S. R. Adler, and L. L. Murphy Extraction-Dependent Effects of American Ginseng (Panax quinquefolium) on Human Breast Cancer Cell Proliferation and Estrogen Receptor Activation. Integr Cancer Ther, September 1, 2006; 5(3): 236 - 243. [Abstract] [PDF]

				Y. Cui, X.-O. Shu, Y.-T. Gao, H. Cai, M.-H. Tao, and W. Zheng Association of Ginseng Use with Survival and Quality of Life among Breast Cancer Patients Am. J. Epidemiol., April 1, 2006; 163(7): 645 - 653. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Submit a related Letter to the Editor Purchase Article View Shopping Cart Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Copyright Permission Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Cho, J. Articles by Lee, Y. Search for Related Content PubMed PubMed Citation Articles by Cho, J. Articles by Lee, Y.