HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Liu, H.-b. Articles by Voskuhl, R. R. Search for Related Content PubMed PubMed Citation Articles by Liu, H.-b. Articles by Voskuhl, R. R.

Estrogen Receptor Mediates Estrogen’s Immune Protection in Autoimmune Disease

Hong-biao Liu^*, Kyi Kyi Loo^*, Karen Palaszynski^*, Judith Ashouri^*, Dennis B. Lubahn and Rhonda R. Voskuhl^1,*

^* Department of Neurology, Reed Neurological Research Center, University of California, Los Angeles, CA 90095; and Department of Biochemistry, Child Health and Animal Sciences, University of Missouri, Columbia, MO 65211

	Abstract

Top Abstract Introduction Materials and Methods Results Discussion References

 
Estrogens are known to influence a variety of autoimmune diseases, but it is not known whether their actions are mediated through classic estrogen receptor (ER). The presence of a functional ER was demonstrated in secondary lymphoid tissues, then ER expression was shown at both the RNA and protein levels in these tissues. Use of ER knockout mice revealed that both the estrogen-induced disease protection and the estrogen-induced reduction in proinflammatory cytokines were dependent upon ER in the prototypic Th1-mediated autoimmune disease experimental autoimmune encephalomyelitis. These findings are central to the design of selective ER modifiers which aim to target biologic responses in specific organ systems.

	Introduction

Top Abstract Introduction Materials and Methods Results Discussion References

 
Estrogens have biological activity in numerous organ systems including the immune system, the reproductive system, the cardiovascular system, the CNS, bone, breast, colon, and prostate. An understanding of which estrogen receptor (ER)² mediates estrogen’s action in a given organ system is central to the development of selective ER modifiers (SERMs) which aim to target therapeutic effects and minimize toxic side effects.

Gender and hormonal influence play a role in a wide variety of human autoimmune diseases. Multiple sclerosis (MS), rheumatoid arthritis (RA), uveitis, and thyroiditis improve during pregnancy while other diseases such as systemic lupus erythematosis (SLE) tend to worsen (1). An immune shift from Th1 to Th2 that occurs during pregnancy as the maternal immune system attempts to avoid fetal rejection has been proposed as a mechanism underlying the improvement in Th1-mediated autoimmune diseases during pregnancy (1).

Pregnancy is a complex situation characterized by changes in numerous factors including a steady increase in levels of estrogens. Interestingly, increases in estrogens appear to recapitulate the effect of pregnancy on autoimmune diseases, at least in part. Estrogen treatment in mice decreases the severity of experimental autoimmune encephalomyelitis (EAE) and collagen-induced arthritis (CIA), while it worsens SLE (2, 3, 4, 5). Immune shifts from Th1 to Th2 have been observed during estrogen treatment in these animal models (2, 3, 6). There is also evidence in humans with MS and RA that estrogen treatment may be beneficial, while estrogen treatment of SLE worsens the disease (7, 8, 9, 10, 11, 12, 13).

Estrogens may act through genomic or nongenomic mechanisms. Genomic mechanisms were originally described and mediated through ER, the "classic" ER (14). However, since characterization of ER, other ERs have been described including ER (15). Most recently, ER has been detected, but thus far only in teleosts (16). Nongenomic mechanisms involve membrane effects (17) with a putative membrane associated "ER-X" being recently reported (18). Interestingly, the estrogen-mediated protection in CIA was recently shown to possibly be caused by genomic effects since treatment with ICI 182,780 blocked the protective effect of estrogen on disease when used at a dose known to block ER and ER, but not membrane effects (19). However, it was not determined which nuclear receptor was involved in the estrogen induced immune protection in CIA. In this manuscript the prototypic Th1-mediated autoimmune disease EAE is used to determine whether the protective effect of estrogen treatment is mediated through classic ER.

	Materials and Methods

Top Abstract Introduction Materials and Methods Results Discussion References

 
Animals and reagents

Homozygous male ER knockout mice were generated by backcrossing the knockout (20) onto the C57BL/6 strain for 16 generations. Littermates were used as wild-type (WT) controls. Animals were maintained in accordance with guidelines established by the National Institutes of Health and as mandated by the Univeristy of California, Los Angeles Office for the Protection of Research Subjects. Human ovary was purchased from BioChain Institute (Hayward, CA). Myelin oligodendrocyte glycoprotein (MOG) 35–55 peptide was synthesized to >98% purity by Chiron Mimotopes (San Diego, CA).

In vitro estrogen treatment and cytokine analysis

C57BL/6 mice were immunized with MOG_35–55 peptide (100 µg/mouse) and draining lymph node cells were harvested after 10 days. Cells were stimulated in vitro with autoantigen (25 µg/ml) in the presence or absence of estriol (28 ng/ml). After 6 h of stimulation, TNF- was detected by intracellular staining with TNF--specific Ab (BD PharMingen, San Diego, CA) and subsequent FACS analysis.

EMSA

Cytoplasmic or nuclear proteins from spleen and brain were prepared using NE-PER Nuclear and Cytoplasmic Extraction Reagents based on the manufacturer’s instruction (Pierce, Rockford, IL). Samples were preincubated with biotin-conjugated estrogen response element (ERE; Panomics, Redwood City, CA) then subjected to electrophoresis. Recombinant ER and ER receptors (Affinity, Golden, CO) were used as positive controls.

RT-PCR

Total RNA (2 µg) was reverse-transcribed to cDNA using the Reverse Transcription System (Applied Bioscience, Branchburg, NJ). For amplification, the outer primer set corresponded to nucleotides 461–480 (sense) and 1918–1937 (anti-sense) of the ER gene, while with the inner primer set corresponded to 659–679 (sense) and 981-1003 (anti-sense) as previously numbered (21). The primer sequences are as follows: mER outer set sense 461–480 5'-GCC GCC TTC AGT GCC AAC AG-3'; mER inner set sense 659–679 5'-AAT TCT GAC AAT CGA CGC CAG-3'; mER inner set anti-sense 1003–981 5'-GTG CTT CAA CAT TCT CCC TCC TC-3'; mER outer set anti-sense 1937–1918 5'-AGG AAT GTG CTG AAG TGG AG-3'.

Western blots

Intracellular proteins were prepared using NE-PER Nuclear and Cytoplasmic Extraction Reagents (as above) and Western blots run as described previously (22). To detect ER (67 kDa) a rabbit ER-specific primary Ab (Santa Cruz Biotechnology, Santa Cruz, CA) was used with a biotin-labeled goat anti-rabbit IgG secondary Ab. Alternatively, westerns were probed with E2-BSA (Sigma-Aldrich, St. Louis, MO) labeled with biotin using the EZ-link Sulfo-NHS-LC-Biotinylation kit (Pierce).

In vivo hormone treatment, induction of active EAE, and cytokine analysis

C57BL/6 male mice were treated with estriol (5 mg s.c. 90-day release pellets) or with placebo pellets (Innovative Research of America, Sarasota, FL) as described (2). One week later, active EAE was induced by immunizing with MOG_35–55 peptide in complete Freund’s adjuvant as described (23). Mice were monitored daily for signs of EAE on a standard EAE grading scale of 0–5 (0, unaffected; 5, moribund). Mice in each group were sacrificed at days 16–24 during EAE, splenocytes were harvested and stimulated with autoantigen (MOG_35–55 peptide at 25 µg/ml) as described (2). Supernatants were collected after 48 h and levels of TNF-, IFN-, IL-2, and IL-5 were determined by cytometric bead array (BD PharMingen).

	Results

Top Abstract Introduction Materials and Methods Results Discussion References

 
Functional ER are present in peripheral immune tissues

We first examined immune cells from secondary lymphoid tissues of mice for the presence of a functional ER. Lymph node cells from MOG peptide 35–55-immunized C57BL/6 mice were stimulated with the autoantigen in the presence or absence of estriol in vitro, and a significant decrease in TNF (TNF-) was observed, p = 0.036 (Fig. 1A). This reduction in TNF- production during an autoantigen-specific immune response had been previously reported during in vivo estrogen treatment of mice with EAE (3). These in vitro data indicated that estrogen’s effect can be mediated by a direct effect of an estrogen on murine immune cells during an autoantigen-specific immune response. Next, we assessed murine splenic tissue proteins for their ability to bind to the ERE. In the nuclear fraction, binding was evidenced by retardation of migration of bands in EMSA (Fig. 1B). The lack of well delineated bands in lanes containing tissue proteins, with clear single bands in lanes containing recombinant proteins, is likely due to heterogeneity in ER types within tissues. This is consistent with the fact that either ER or ER could bind the ERE and that variants of ERs have been demonstrated in nonimmune tissues previously (24, 25). Together the alteration in cytokine production during in vitro estriol treatment of immune cells and the binding of splenic protein to ERE indicated that functional ERs are expressed in murine secondary lymphoid tissues.

View larger version (45K): [in this window] [in a new window]   FIGURE 1. There are functional ERs expressed in secondary lymphoid tissues. A, Estrogen treatment was shown to have direct effects on cytokine production by immune cells. MOG35–55 peptide-specific lymph node cells were harvested from MOG35–55-immunized C57BL/6 mice. Cells were stimulated in vitro with autoantigen in the presence or absence of estriol at a dose physiologic with levels that occur during pregnancy (28 ng/ml) (46 ). TNF- was detected by intracellular staining and subsequent FACS analysis. Data are expressed as percent change of TNF--positive staining in placebo or estriol treated, each as compared with placebo treated. Data are pooled from four separate experiments. Error bars indicate variation between experiments. *, p < 0.05 when comparing estriol vs placebo treated. In vitro estriol treatment had no effect on cell viability as assessed by 72 h proliferation assays (data not shown). B, Splenic ER was shown to bind the estrogen response element (ERE). Cytoplasmic or nuclear proteins from spleen were prepared and samples were preincubated with biotin-conjugated ERE. Splenic nuclear (N) and cytoplasmic (C) proteins were shown to bind ERE as demonstrated by retardation of migration of bands in EMSA. Recombinant human ER, recombinant human ER, and murine brain nuclear (N) and cytoplasmic (C) proteins served as positive controls. Biotin-EAE alone served as the negative control.

ER expression at the RNA and protein levels

ER expression has been reported in bone marrow and thymus (26, 27, 28), but direct evidence of expression in murine peripheral immune tissues is lacking. While RT-PCR data detecting ER transcripts in peripheral immune tissues have been reported in mice, there has been no sequencing of these products (29, 30). Furthermore, expression of ER at the protein level in the peripheral immune system has remained poorly characterized since Western blot data were negative for specific bands of the correct size (31) while flow cytometry of immune cells was positive (29, 32, 33). Together these previous reports raised the question of whether the classic ER, vs another ER variant (24, 25), was detected in immune cells by flow cytometry. Thus, we characterized ER expression in secondary lymphoid tissues of mice at the RNA level by both nested RT-PCR and sequencing and at the protein level by Western blot using both Ab and ligand binding. Nested RT-PCR using ER-specific primers revealed bands of the expected size in both spleen (Fig. 2A) and lymph node (data not shown). The product from the inner primer set was then sequenced and confirmed to be ER. Western blots of splenic tissue using an ER-specific Ab detected expression at the protein level at the expected size of 67 kDa (Fig. 2B). ER expression at the protein level was then confirmed by probing Western blots with 17 estradiol-BSA whereby a band was identified in spleen which colocalized to the size of the band detected with ER-specific Ab probing (Fig. 2C). Together these data are conclusive evidence of the expression of classic ER in murine peripheral immune tissues.

View larger version (28K): [in this window] [in a new window]   FIGURE 2. ER is expressed at the RNA and protein levels in mouse spleen. A, ER gene expression was detected in murine spleen by nested RT-PCR. RNA from murine uterus tissue served as a positive control. No RNA served as a negative control. The inner primer PCR product was confirmed by sequencing. B, ER protein expression was demonstrated in spleen. Western blots of tissue proteins were probed with ER-specific primary Ab. A protein of the expected size for ER (67 kDa) was detected in spleen. Human ovary and murine brain served as positive control tissues. Rabbit IgG served as a primary Ab negative control. The amounts of protein loaded per lane were 100 µg for spleen and brain and 60 µg for ovary. C, ER expression at the protein level is confirmed by probing Western blots with estradiol-BSA. Western blots of tissue proteins were probed with E2-BSA-biotin instead of ER-specific Ab. A protein of the expected size for ER (67 kDa) was detected in spleen. Human ovary and murine brain served as positive control tissues. Probing with BSA-biotin alone served as a negative control.

The estrogen-mediated protection in EAE is dependent upon ER

While an estrogen-mediated protective effect had been previously reported in EAE and in CIA, the mechanism of the immune protection has remained unknown. Therefore, we used the EAE model to determine whether the estrogen induced immune protection in this prototypic Th1 autoimmune disease was mediated through ER. Since mechanisms of action of estrogens have been shown to differ depending upon whether physiologic or pharmacologic doses are used (34), we used a dose of estriol for EAE treatment which had previously been shown to induce estriol levels in sera of mice which were physiologic with natural murine pregnancy (2). Homozygous ER KO mice were not protected from disease during estriol treatment, while WT mice were protected as evidenced by a significant decrease in disease severity in estriol vs placebo treated, p < 0.0001 (Fig. 3). These data demonstrate that the protective effect of estriol in EAE is dependent upon the presence of an intact ER.

View larger version (19K): [in this window] [in a new window]   FIGURE 3. The estrogen induced immune protection in EAE is mediated through ER. A reduction in mean clinical EAE scores occurred in estriol-treated WT, but not ER KO, mice. C57BL/6 male mice were treated with estriol or with placebo (sustained release pellets). One week later, active EAE was induced by immunizing with MOG35–55 peptide. There were a total of 40 mice, with 10 mice in each of four groups: WT treated with placebo (WT Placebo), WT treated with estriol (WT Estriol), homozygous ER knockout treated with placebo (KO Placebo), and homozygous ER knockout treated with estriol (KO Estriol). Error bars represent variation in scores between mice within each group.

The estrogen-mediated immune modulation in EAE is dependent upon ER

Significant reductions in proinflammatory cytokine production (TNF-, p < 0.005; IFN-, p < 0.005; and IL-2, p < 0.05) were observed during autoantigen-specific immune responses of WT mice treated with estriol as compared with placebo (Fig. 4). Importantly, these reductions did not occur in ER knockout mice treated with estriol. Since not all cytokines were reduced by estriol treatment, apoptosis was an unlikely explanation for the decrease in the above cytokines in estriol-treated WT mice. Levels of the Th2 cytokine IL-5 were increased in estriol-treated WT mice as compared with placebo-treated WT mice. This estriol-induced increase in IL-5 production was less dramatic but still present in ER KO mice. Together these data demonstrate that both the disease protection and the reduction in proinflammatory cytokine production induced by estriol treatment in EAE are mediated through ER.

View larger version (18K): [in this window] [in a new window]   FIGURE 4. The estrogen-mediated immune modulation in EAE is dependent upon ER. A reduction in proinflammatory cytokine production occurred in estriol-treated WT, but not ER KO, mice. EAE mice in each group were sacrificed at days 16–24 during EAE, and splenocytes were harvested and stimulated with MOG35–55 peptide. Supernatants were collected and levels of TNF-, IFN-, IL-2, and IL-5 determined by cytometric bead array. Data are expressed as percent change in two to six pooled experiments for indicated cytokine in supernatants from estriol or placebo-treated EAE mice, each as compared with the placebo-treated EAE. Homozygous ER knockout = KO. Error bars indicate variation between experiments. *, p < 0.05; **, p < 0.005 when comparing cytokine levels in estriol vs placebo treated. Cytokine values in picograms per milliliter from a representative experiment are as follows: TNF-: WT Placebo = 862.7 ± 198.5, WT Estriol = 71.1 ± 387.4, KO Placebo = 497.7 ± 142.0, KO Estriol = 497.7 ± 32.9; IFN-: WT Placebo = 1545.9 ± 92.0, WT Estriol = 152.5 ± 160.4, KO Placebo = 1320.4 ± 16.3, KO Estriol = 1118.8 ± 265.2; IL-2: WT Placebo = 137.8 ± 74.5, WT Estriol = 10.9 ± 1.7, KO Placebo = 94.1 ± 23.7, KO Estriol = 95.7 ± 12.7; IL-5: WT Estriol = 97.0 ± 12.2, WT Placebo = 29.4 ± 5.5, KO Estriol = 39.8 ± 18.8, KO Placebo = 18.2 ± 2.9

	Discussion

Top Abstract Introduction Materials and Methods Results Discussion References

Regarding the effects of estrogen treatment on cytokine production, it is interesting to contrast our findings with those in a previous report in which estradiol was given in vivo and primary T cell responses to OVA were shown to be enhanced for Th1 cytokine production, with this effect dependent upon ER (35). While both studies agree that ER is important for immune efffects, the studies differ in that Th1 cytokine production was decreased by estrogen treatment in our paper while being increased in the previous paper. The differences between the two papers could be that estriol was used in our studies while estradiol was used in the previous study. Estriol and estradiol are clearly not the same. Estradiol has been shown to lead to hypertension, increased coagulation and to protect from osteoporosis more than estriol (36, 37, 38, 39, 40, 41). They can even have opposing effects. For example, estriol can protect from estradiol-mediated breast cancer (42, 43). However, since reports by three groups have shown that both estradiol and estriol are protective in both EAE and CIA (2, 3, 44, 45), there do not appear to be major differences between the two estrogen types in immune effects on disease. Indeed, the finding that estradiol treatment increases Th1 cytokine production during OVA-specific immune responses (35) is in direct contrast to previous reports by several groups that estradiol treatment ameliorates the prototypic Th1-mediated disease EAE. Perhaps differences between the effect estradiol treatment on OVA-specific responses in healthy mice and autoantigen-specific responses in EAE mice relates to either the strain or the age of the mice treated. F₅ generation C57BL/6 mice were used in the former study while F₁₆ C57BL/6 mice were used in our study. However, such strain differences also appear to be an unlikely explanation since estradiol has been shown to ameliorate EAE in numerous strains (C57BL/6, SJL, B10.PL, B10RIII). The age of the mice treated remains a consideration since the former report involved treatment of 4-wk-old mice, while all the EAE studies involved treatment of adult mice at least 8 wk of age. Differences in the nature of the immune response to OVA vs autoantigen is also a consideration.

While both estadiol and estriol have been shown to ameliorate EAE, we have focused considerable attention on estriol treatment of EAE for two reasons. First, unlike estradiol which is present during the menstrual cycle of the nonpregnant state, estriol is not present in appreciable amounts in the nonpregnant state. The nonpregnant female state is not a time of protection from autoimmune diseases such as MS and RA, since nonpregnant females are known to be highly susceptible. Estriol becomes detectable only during the first trimester of pregnancy and increases progressively throughout gestation, and pregnancy is a state of relative protection from MS and RA. Thus, there a correlation between disease protection and the presence of estriol, while the relationship between disease protection and the presence of estradiol appears more complex. Second, we focused on estriol treatment of EAE as a translational research strategy for MS in light of the fact that estriol is known to have less toxicity in humans than estradiol (36, 37, 38). The ultimate goal of the findings in this manuscript are to define the precise mechanism of action of the therapeutic efficacy of estriol treatment in EAE. These findings will then be contrasted to findings of the precise mechanism of action of estrogen related toxicities such as increased risk of breast cancer, endometrial cancer, and vascular disease. Our findings suggest that therapeutic efficacy may be achieved through stimulation of ER. If it is ultimately shown that some or all toxicities are mediated through ER or nongenomic effects, then a window of selectivity exists whereby one could treat disease with an ER agonist or other SERM to achieve efficacy and minimize toxicity. In contrast, if it is ultimately shown that all toxicities are mediated through ER, then selectivity will not be possible at the level of the ER. However, selectivity may still be achievable through modulation of tissue-specific cofactors involved in ER action. To pursue the latter, one must first define the ER used in a given tissue for a given biologic effect. Then one may begin to identify tissue-specific differences in cofactors for each of the biologic effects.

In conclusion, the protective effect of estrogen treatment on autoantigen-specific immune responses in a prototypic Th1-mediated autoimmune disease is mediated through ER. This does not however rule out additional nongenomic (membrane, ER-X) or genomic (ER, ER) effects on estrogen-mediated immune protection. The finding of an important role for ER carries far reaching implications for the design of SERMs for the treatment of a wide variety of autoimmune diseases which aim toward optimizing efficacy and minimizing toxicity. They are equally as relevant to the design of SERMs for treatment of nonautoimmune diseases with different target organs (the reproductive system, the cardiovascular system, bone, breast, colon, and prostate) which aim to avoid effects on the immune system.

	Footnotes

 
This work was supported by the National Institutes of Health Grants AI50839 and NS45443 (to R.R.V.) and the National Multiple Sclerosis Society Grant RD3407 (to R.R.V.).

¹ Address correspondence and reprint requests to Dr. Rhonda Voskuhl, Reed Neurological Research Center, Room A145, 710 Westwood Plaza, Los Angeles, CA 90095. E-mail address: rvoskuhl{at}ucla.edu

² Abbreviations used in this paper: ER, estrogen receptor; EAE, experimental autoimmune encephalomyelitis; SERM, selective ER modifier; ERE, estrogen response element; SLE, systemic lupus erythematosis; CIA, collagen-induced arthritis; MOG, myelin oligodendrocyte glycoprotein; WT, wild type; MS, multiple sclerosis; RA, rheumatoid arthritis.

Received for publication May 29, 2003. Accepted for publication October 10, 2003.

	References

Top Abstract Introduction Materials and Methods Results Discussion References

 

1. Whitacre, C. C., S. C. Reingold, P. A. O’Looney. 1999. A gender gap in autoimmunity. Science 283:1277.[Free Full Text]
2. Kim, S., S. M. Liva, M. A. Dalal, M. A. Verity, R. R. Voskuhl. 1999. Estriol ameliorates autoimmune demyelinating disease: implications for multiple sclerosis. Neurology 52:1230.[Abstract/Free Full Text]
3. Ito, A., B. F. Bebo, Jr, A. Matejuk, A. Zamora, M. Silverman, A. Fyfe-Johnson, H. Offner. 2001. Estrogen treatment down-regulates TNF- production and reduces the severity of experimental autoimmune encephalomyelitis in cytokine knockout mice. J. Immunol. 167:542.[Abstract/Free Full Text]
4. Jansson, L., R. Holmdahl. 1998. Estrogen-mediated immunosuppression in autoimmune diseases. Inflam. Res. 47:290.[Medline]
5. Carlsten, H., A. Tarkowski, R. Holmdahl, L. A. Nilsson. 1990. Oestrogen is a potent disease accelerator in SLE-prone MRL lpr/lpr mice. Clin. Exp. Immunol. 80:467.[Medline]
6. Carlsten, H., R. Holmdahl, A. Tarkowski. 1991. Analysis of the genetic encoding of oestradiol suppression of delayed-type hypersensitivity in (NZB x NZW) F₁ mice. Immunology 73:186.[Medline]
7. Sicotte, N. L., S. M. Liva, R. Klutch, P. Pfeiffer, S. Bouvier, S. Odesa, T. C. Wu, R. R. Voskuhl. 2002. Treatment of multiple sclerosis with the pregnancy hormone estriol. Ann. Neurol. 52:421.[Medline]
8. Hall, G. M., M. Daniels, E. C. Huskisson, T. D. Spector. 1994. A randomised controlled trial of the effect of hormone replacement therapy on disease activity in postmenopausal rheumatoid arthritis. Ann. Rheum. Dis. 53:112.[Abstract/Free Full Text]
9. Spector, T. D., E. Roman, A. J. Silman. 1990. The pill, parity, and rheumatoid arthritis. Arthritis Rheum. 33:782.[Medline]
10. van Zeben, D., J. M. Hazes, J. P. Vandenbroucke, B. A. Dijkmans, A. Cats. 1990. Diminished incidence of severe rheumatoid arthritis associated with oral contraceptive use. Arthritis Rheum. 33:1462.[Medline]
11. Jungers, P., M. Dougados, C. Pélissier, F. Kuttenn, F. Tron, P. Lesavre, J. F. Bach. 1982. Influence of oral contraceptive therapy on the activity of systemic lupus erythematosus. Arthritis Rheum. 25:618.[Medline]
12. Ostensen, M.. 1999. Sex hormones and pregnancy in rheumatoid arthritis and systemic lupus erythematosus. Ann. NY Acad. Sci. 876:131.[Medline]
13. Sánchez-Guerrero, J., M. H. Liang, E. W. Karlson, D. J. Hunter, G. A. Colditz. 1995. Postmenopausal estrogen therapy and the risk for developing systemic lupus erythematosus. Ann. Intern. Med. 122:430.[Abstract/Free Full Text]
14. Greene, G. L., P. Gilna, M. Waterfield, A. Baker, Y. Hort, J. Shine. 1986. Sequence and expression of human estrogen receptor complementary DNA. Science 231:1150.[Abstract/Free Full Text]
15. Kuiper, G. G., E. Enmark, M. Pelto-Huikko, S. Nilsson, J. A. Gustafsson. 1996. Cloning of a novel receptor expressed in rat prostate and ovary. Proc. Natl. Acad. Sci. USA 93:5925.[Abstract/Free Full Text]
16. Hawkins, M. B., J. W. Thornton, D. Crews, J. K. Skipper, A. Dotte, P. Thomas. 2000. Identification of a third distinct estrogen receptor and reclassification of estrogen receptors in teleosts. Proc. Natl. Acad. Sci. USA 97:10751.[Abstract/Free Full Text]
17. Guo, Z., J. Krucken, W. P. Benten, F. Wunderlich. 2002. Estradiol-induced nongenomic calcium signaling regulates genotropic signaling in macrophages. J. Biol. Chem. 277:7044.[Abstract/Free Full Text]
18. Toran-Allerand, C. D., X. Guan, N. J. MacLusky, T. L. Horvath, S. Diano, M. Singh, E. S. Connolly, Jr, I. S. Nethrapalli, A. A. Tinnikov. 2002. ER-X: a novel, plasma membrane-associated, putative estrogen receptor that is regulated during development and after ischemic brain injury. J. Neurosci. 22:8391.[Abstract/Free Full Text]
19. Jansson, L., R. Holmdahl. 2001. Enhancement of collagen-induced arthritis in female mice by estrogen receptor blockage. Arthritis Rheum. 44:2168.[Medline]
20. Lubahn, D. B., J. S. Moyer, T. S. Golding, J. F. Couse, K. S. Korach, O. Smithies. 1993. Alteration of reproductive function but not prenatal sexual development after insertional disruption of the mouse estrogen receptor gene. Proc. Natl. Acad. Sci. USA 90:11162.[Abstract/Free Full Text]
21. White, R., J. A. Lees, M. Needham, J. Ham, M. Parker. 1987. Structural organization and expression of the mouse estrogen receptor. Mol. Endocrinol. 1:735.[Abstract/Free Full Text]
22. Liu, H., A. J. MacKenzie-Graham, K. Palaszynski, S. Liva, R. R. Voskuhl. 2001. "Classic" myelin basic proteins are expressed in lymphoid tissue macrophages. J. Neuroimmunol. 116:83.[Medline]
23. Suen, W. E., C. M. Bergman, P. Hjelmstrom, N. H. Ruddle. 1997. A critical role for lymphotoxin in experimental allergic encephalomyelitis. J. Exp. Med. 186:1233.[Abstract/Free Full Text]
24. Kos, M., S. Denger, G. Reid, K. S. Korach, F. Gannon. 2002. Down but not out: a novel protein isoform of the estrogen receptor is expressed in the estrogen receptor knockout mouse. J. Mol. Endocrinol. 29:281.[Abstract]
25. Asaithambi, A., S. Mukherjee, M. K. Thakur. 1997. Expression of 112-kDa estrogen receptor in mouse brain cortex and its autoregulation with age. Biochem. Biophys. Res. Commun. 231:683.[Medline]
26. Igarashi, H., T. Kouro, T. Yokota, P. C. Comp, P. W. Kincade. 2001. Age and stage dependency of estrogen receptor expression by lymphocyte precursors. Proc. Natl. Acad. Sci. USA 98:15131.[Abstract/Free Full Text]
27. Kawashima, I., K. Seiki, K. Sakabe, S. Ihara, A. Akatsuka, Y. Katsumata. 1992. Localization of estrogen receptors and estrogen receptor-mRNA in female mouse thymus. Thymus. 20:115.[Medline]
28. Seiki, K., K. Sakabe. 1997. Sex hormones and the thymus in relation to thymocyte proliferation and maturation. Arch. Histol. Cytol. 60:29.[Medline]
29. Sakazaki, H., H. Ueno, K. Nakamuro. 2002. Estrogen receptor in mouse splenic lymphocytes: possible involvement in immunity. Toxicol. Lett. 133:221.[Medline]
30. Tornwall, J., A. B. Carey, R. I. Fox, H. S. Fox. 1999. Estrogen in autoimmunity: expression of estrogen receptors in thymic and autoimmune T cells. J. Gend. Specif. Med. 2:33.[Medline]
31. Tiffoche, C., C. Vaillant, D. Schausi, M. L. Thieulant. 2001. Novel intronic promoter in the rat ER gene responsible for the transient transcription of a variant receptor. Endocrinology 142:4106.[Abstract/Free Full Text]
32. Liu, H. Y., A. C. Buenafe, A. Matejuk, A. Ito, A. Zamora, J. Dwyer, A. A. Vandenbark, H. Offner. 2002. Estrogen inhibition of EAE involves effects on dendritic cell function. J. Neurosci. Res. 70:238.[Medline]
33. Grimaldi, C. M., J. Cleary, A. S. Dagtas, D. Moussai, B. Diamond. 2002. Estrogen alters thresholds for B cell apoptosis and activation. J. Clin. Invest. 109:1625.[Medline]
34. Dubal, D. B., H. Zhu, J. Yu, S. W. Rau, P. J. Shughrue, I. Merchenthaler, M. S. Kindy, P. M. Wise. 2001. Estrogen receptor , not , is a critical link in estradiol-mediated protection against brain injury. Proc. Natl. Acad. Sci. USA 98:1952.[Abstract/Free Full Text]
35. Maret, A., J. D. Coudert, L. Garidou, G. Foucras, P. Gourdy, A. Krust, S. Dupont, P. Chambon, P. Druet, F. Bayard, J. C. Guery. 2003. Estradiol enhances primary antigen-specific CD4 T cell responses and Th1 development in vivo: essential role of estrogen receptor expression in hematopoietic cells. Eur. J. Immunol. 33:512.[Medline]
36. Utian, W. H.. 1980. The place of oestriol therapy after menopause. Acta Endocrinol. Supplementum. 233:51.
37. Follingstad, A. H.. 1978. Estriol, the forgotten estrogen?. J. Am. Med. Assoc. 239:29.[Abstract/Free Full Text]
38. Head, K. A.. 1998. Estriol: safety and efficacy. Altern. Med. Rev. 3:101.[Medline]
39. Hayashi, T., I. Ito, H. Kano, H. Endo, A. Iguchi. 2000. Estriol (E3) replacement improves endothelial function and bone mineral density in very elderly women. J. Gerontol. A Biol. Sci. Med. Sci. 55:B183.[Abstract/Free Full Text]
40. Toy, J. L., J. A. Davies, G. P. McNicol. 1978. The effects of long-term therapy with oestriol succinate on the haemostatic mechanism in postmenopausal women. Br. J. Obstet. Gynaecol. 85:363.[Medline]
41. Erkkola, R., R. Lammintausta, R. Punnonen, L. Rauramo. 1978. The effect of estriol succinate therapy on plasma renin activity and urinary aldosterone in postmenopausal women. Maturitas 1:9.[Medline]
42. Lemon, H. M.. 1973. Oestriol and prevention of breast cancer. Lancet 1:546.
43. Lemon, H. M.. 1975. Estriol prevention of mammary carcinoma induced by 7,12-dimethylbenzanthracene and procarbazine. Cancer Res. 35:1341.[Abstract/Free Full Text]
44. Jansson, L., T. Olsson, R. Holmdahl. 1994. Estrogen induces a potent suppression of experimental autoimmune encephalomyelitis and collagen-induced arthritis in mice. J. Neuroimmunology 53:203.[Medline]
45. Subramanian, S., A. Matejuk, A. Zamora, A. A. Vandenbark, H. Offner. 2003. Oral feeding with ethinyl estradiol suppresses and treats experimental autoimmune encephalomyelitis in SJL mice and inhibits the recruitment of inflammatory cells into the central nervous system. J. Immunol. 170:1548.[Abstract/Free Full Text]
46. Gilmore, W., L. P. Weiner, J. Correale. 1997. Effect of estradiol on cytokine secretion by proteolipid protein-specific T cell clones isolated from multiple sclerosis patients and normal control subjects. J. Immunol. 158:446.[Abstract]

This article has been cited by other articles:

				D. Fairweather, S. Frisancho-Kiss, and N. R. Rose Sex Differences in Autoimmune Disease from a Pathological Perspective Am. J. Pathol., September 1, 2008; 173(3): 600 - 609. [Abstract] [Full Text] [PDF]

				S. Tiwari-Woodruff, L. B. J. Morales, R. Lee, and R. R. Voskuhl Differential neuroprotective and antiinflammatory effects of estrogen receptor (ER){alpha} and ERbeta ligand treatment PNAS, September 11, 2007; 104(37): 14813 - 14818. [Abstract] [Full Text] [PDF]

				R. H. Straub The Complex Role of Estrogens in Inflammation Endocr. Rev., August 1, 2007; 28(5): 521 - 574. [Abstract] [Full Text] [PDF]

				K. Dahlman-Wright, V. Cavailles, S. A. Fuqua, V. C. Jordan, J. A. Katzenellenbogen, K. S. Korach, A. Maggi, M. Muramatsu, M. G. Parker, and J.-A. Gustafsson International Union of Pharmacology. LXIV. Estrogen Receptors Pharmacol. Rev., December 1, 2006; 58(4): 773 - 781. [Full Text] [PDF]

				L. B. J. Morales, K. K. Loo, H.-b. Liu, C. Peterson, S. Tiwari-Woodruff, and R. R. Voskuhl Treatment with an estrogen receptor alpha ligand is neuroprotective in experimental autoimmune encephalomyelitis. J. Neurosci., June 21, 2006; 26(25): 6823 - 6833. [Abstract] [Full Text] [PDF]

				F. Hildebrand, W. J. Hubbard, M. A. Choudhry, B. M. Thobe, H.-C. Pape, and I. H. Chaudry Are the protective effects of 17{beta}-estradiol on splenic macrophages and splenocytes after trauma-hemorrhage mediated via estrogen-receptor (ER)-{alpha} or ER-{beta}? J. Leukoc. Biol., June 1, 2006; 79(6): 1173 - 1180. [Abstract] [Full Text] [PDF]

				K. C. Lambert, E. M. Curran, B. M. Judy, G. N. Milligan, D. B. Lubahn, and D. M. Estes Estrogen Receptor {alpha} (ER{alpha}) Deficiency in Macrophages Results in Increased Stimulation of CD4+ T Cells while 17{beta}-Estradiol Acts through ER{alpha} to Increase IL-4 and GATA-3 Expression in CD4+ T Cells Independent of Antigen Presentation J. Immunol., November 1, 2005; 175(9): 5716 - 5723. [Abstract] [Full Text] [PDF]

				A. Mao, V. Paharkova-Vatchkova, J. Hardy, M. M. Miller, and S. Kovats Estrogen Selectively Promotes the Differentiation of Dendritic Cells with Characteristics of Langerhans Cells J. Immunol., October 15, 2005; 175(8): 5146 - 5151. [Abstract] [Full Text] [PDF]

				K. M. Palaszynski, D. L. Smith, S. Kamrava, P. S. Burgoyne, A. P. Arnold, and R. R. Voskuhl A Yin-Yang Effect between Sex Chromosome Complement and Sex Hormones on the Immune Response Endocrinology, August 1, 2005; 146(8): 3280 - 3285. [Abstract] [Full Text] [PDF]

				H. H. van den Broek, J. G. Damoiseaux, M. H De Baets, and R. M. Hupperts The influence of sex hormones on cytokines in multiple sclerosis and experimental autoimmune encephalomyelitis: a review Multiple Sclerosis, June 1, 2005; 11(3): 349 - 359. [Abstract] [PDF]

				M M. Elloso, K. Phiel, R. A Henderson, H. A Harris, and S. J Adelman Suppression of experimental autoimmune encephalomyelitis using estrogen receptor-selective ligands J. Endocrinol., May 1, 2005; 185(2): 243 - 252. [Abstract] [Full Text] [PDF]

				L. Garidou, S. Laffont, V. Douin-Echinard, C. Coureau, A. Krust, P. Chambon, and J.-C. Guery Estrogen Receptor {alpha} Signaling in Inflammatory Leukocytes Is Dispensable for 17{beta}-Estradiol-Mediated Inhibition of Experimental Autoimmune Encephalomyelitis J. Immunol., August 15, 2004; 173(4): 2435 - 2442. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Liu, H.-b. Articles by Voskuhl, R. R. Search for Related Content PubMed PubMed Citation Articles by Liu, H.-b. Articles by Voskuhl, R. R.