Logo of envhperEnvironmental Health PerspectivesBrowse ArticlesAbout EHPGeneral InformationAuthorsMediaProgramsPartnerships
Environ Health Perspect. May 2000; 108(5): 403–412.
PMCID: PMC1638061
Research Article

Assaying estrogenicity by quantitating the expression levels of endogenous estrogen-regulated genes.

Abstract

Scientific evidence suggests that humans and wildlife species may experience adverse health consequences from exposure to environmental chemicals that interact with the endocrine system. Reliable short-term assays are needed to identify hormone-disrupting chemicals. In this study we demonstrate that the estrogenic activity of a chemical can be evaluated by assaying induction or repression of endogenous estrogen-regulated "marker genes" in human breast cancer MCF-7 cells. We included four marker genes in the assay--pS2, transforming growth factor beta3 (TGFbeta3), monoamine oxidase A, and [alpha]1-antichymotrypsin--and we evaluated estrogenic activity for 17beta-estradiol (E(2)), diethylstilbestrol, [alpha]-zearalanol, nonylphenol, genistein, methoxychlor, endosulphan, o,p-DDE, bisphenol A, dibutylphthalate, 4-hydroxy tamoxifen, and ICI 182.780. All four marker genes responded strongly to the three high-potency estrogens (E(2), diethylstilbestrol, and [alpha]-zearalanol), whereas the potency of the other chemicals was 10(3)- to 10(6)-fold lower than that of E(2). There were some marker gene-dependent differences in the relative potencies of the tested chemicals. TGFbeta3 was equally sensitive to the three high-potency estrogens, whereas the sensitivity to [alpha]-zearalanol was approximately 10-fold lower than the sensitivity to E(2) and diethylstilbestrol when assayed with the other three marker genes. The potency of nonylphenol was equal to that of genistein when assayed with pS2 and TGFbeta3, but 10- to 100-fold higher/lower with monoamine oxidase A and [alpha]1-antichymotrypsin, respectively. The results are in agreement with results obtained by other methods and suggest that an assay based on endogenous gene expression may offer an attractive alternative to other E-SCREEN methods.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (4.0M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Kavlock RJ, Daston GP, DeRosa C, Fenner-Crisp P, Gray LE, Kaattari S, Lucier G, Luster M, Mac MJ, Maczka C, et al. Research needs for the risk assessment of health and environmental effects of endocrine disruptors: a report of the U.S. EPA-sponsored workshop. Environ Health Perspect. 1996 Aug;104 (Suppl 4):715–740. [PMC free article] [PubMed]
  • Toppari J, Larsen JC, Christiansen P, Giwercman A, Grandjean P, Guillette LJ, Jr, Jégou B, Jensen TK, Jouannet P, Keiding N, et al. Male reproductive health and environmental xenoestrogens. Environ Health Perspect. 1996 Aug;104 (Suppl 4):741–803. [PMC free article] [PubMed]
  • Carlsen E, Giwercman A, Keiding N, Skakkebaek NE. Evidence for decreasing quality of semen during past 50 years. BMJ. 1992 Sep 12;305(6854):609–613. [PMC free article] [PubMed]
  • Adami HO, Bergström R, Möhner M, Zatoński W, Storm H, Ekbom A, Tretli S, Teppo L, Ziegler H, Rahu M, et al. Testicular cancer in nine northern European countries. Int J Cancer. 1994 Oct 1;59(1):33–38. [PubMed]
  • Sharpe RM, Skakkebaek NE. Are oestrogens involved in falling sperm counts and disorders of the male reproductive tract? Lancet. 1993 May 29;341(8857):1392–1395. [PubMed]
  • Henderson BE, Ross RK, Pike MC, Casagrande JT. Endogenous hormones as a major factor in human cancer. Cancer Res. 1982 Aug;42(8):3232–3239. [PubMed]
  • Giwercman A, Skakkebaek NE. The human testis--an organ at risk? Int J Androl. 1992 Oct;15(5):373–375. [PubMed]
  • Davis DL, Bradlow HL, Wolff M, Woodruff T, Hoel DG, Anton-Culver H. Medical hypothesis: xenoestrogens as preventable causes of breast cancer. Environ Health Perspect. 1993 Oct;101(5):372–377. [PMC free article] [PubMed]
  • Soto AM, Sonnenschein C, Chung KL, Fernandez MF, Olea N, Serrano FO. The E-SCREEN assay as a tool to identify estrogens: an update on estrogenic environmental pollutants. Environ Health Perspect. 1995 Oct;103 (Suppl 7):113–122. [PMC free article] [PubMed]
  • Jobling S, Reynolds T, White R, Parker MG, Sumpter JP. A variety of environmentally persistent chemicals, including some phthalate plasticizers, are weakly estrogenic. Environ Health Perspect. 1995 Jun;103(6):582–587. [PMC free article] [PubMed]
  • Zava DT, Blen M, Duwe G. Estrogenic activity of natural and synthetic estrogens in human breast cancer cells in culture. Environ Health Perspect. 1997 Apr;105 (Suppl 3):637–645. [PMC free article] [PubMed]
  • Taylor CM, Blanchard B, Zava DT. A simple method to determine whole cell uptake of radiolabelled oestrogen and progesterone and their subcellular localization in breast cancer cell lines in monolayer culture. J Steroid Biochem. 1984 May;20(5):1083–1088. [PubMed]
  • Shafie S, Brooks SC. Characteristics of the dextran-coated charcoal assay for estradiol receptor in breast cancer preparations. J Lab Clin Med. 1979 Nov;94(5):784–798. [PubMed]
  • Klein KO, Baron J, Colli MJ, McDonnell DP, Cutler GB., Jr Estrogen levels in childhood determined by an ultrasensitive recombinant cell bioassay. J Clin Invest. 1994 Dec;94(6):2475–2480. [PMC free article] [PubMed]
  • Lan NC, Katzenellenbogen BS. Temporal relationships between hormone receptor binding and biological responses in the uterus: studies with short- and long-acting derivatives of estriol. Endocrinology. 1976 Jan;98(1):220–227. [PubMed]
  • Gray LE, Jr, Ostby J, Ferrell J, Rehnberg G, Linder R, Cooper R, Goldman J, Slott V, Laskey J. A dose-response analysis of methoxychlor-induced alterations of reproductive development and function in the rat. Fundam Appl Toxicol. 1989 Jan;12(1):92–108. [PubMed]
  • Sumpter JP, Jobling S. Vitellogenesis as a biomarker for estrogenic contamination of the aquatic environment. Environ Health Perspect. 1995 Oct;103 (Suppl 7):173–178. [PMC free article] [PubMed]
  • Andersen HR, Andersson AM, Arnold SF, Autrup H, Barfoed M, Beresford NA, Bjerregaard P, Christiansen LB, Gissel B, Hummel R, et al. Comparison of short-term estrogenicity tests for identification of hormone-disrupting chemicals. Environ Health Perspect. 1999 Feb;107 (Suppl 1):89–108. [PMC free article] [PubMed]
  • Jørgensen M, Hummel R, Bévort M, Andersson AM, Skakkebaek NE, Leffers H. Detection of oestrogenic chemicals by assaying the expression level of oestrogen regulated genes. APMIS. 1998 Jan;106(1):245–251. [PubMed]
  • VanderKuur JA, Hafner MS, Christman JK, Brooks SC. Effects of estradiol-17 beta analogues on activation of estrogen response element regulated chloramphenicol acetyltransferase expression. Biochemistry. 1993 Jul 13;32(27):7016–7021. [PubMed]
  • Meyer T, Koop R, von Angerer E, Schönenberger H, Holler E. A rapid luciferase transfection assay for transcription activation effects and stability control of estrogenic drugs in cell cultures. J Cancer Res Clin Oncol. 1994;120(6):359–364. [PubMed]
  • Vinggaard AM, Joergensen EC, Larsen JC. Rapid and sensitive reporter gene assays for detection of antiandrogenic and estrogenic effects of environmental chemicals. Toxicol Appl Pharmacol. 1999 Mar 1;155(2):150–160. [PubMed]
  • Andersson AM, Skakkebaek NE. Exposure to exogenous estrogens in food: possible impact on human development and health. Eur J Endocrinol. 1999 Jun;140(6):477–485. [PubMed]
  • Nishikawa J, Saito K, Goto J, Dakeyama F, Matsuo M, Nishihara T. New screening methods for chemicals with hormonal activities using interaction of nuclear hormone receptor with coactivator. Toxicol Appl Pharmacol. 1999 Jan 1;154(1):76–83. [PubMed]
  • Masamura S, Santner SJ, Heitjan DF, Santen RJ. Estrogen deprivation causes estradiol hypersensitivity in human breast cancer cells. J Clin Endocrinol Metab. 1995 Oct;80(10):2918–2925. [PubMed]
  • Gaido KW, Leonard LS, Lovell S, Gould JC, Babaï D, Portier CJ, McDonnell DP. Evaluation of chemicals with endocrine modulating activity in a yeast-based steroid hormone receptor gene transcription assay. Toxicol Appl Pharmacol. 1997 Mar;143(1):205–212. [PubMed]
  • Bolger R, Wiese TE, Ervin K, Nestich S, Checovich W. Rapid screening of environmental chemicals for estrogen receptor binding capacity. Environ Health Perspect. 1998 Sep;106(9):551–557. [PMC free article] [PubMed]
  • Petit F, Le Goff P, Cravédi JP, Valotaire Y, Pakdel F. Two complementary bioassays for screening the estrogenic potency of xenobiotics: recombinant yeast for trout estrogen receptor and trout hepatocyte cultures. J Mol Endocrinol. 1997 Dec;19(3):321–335. [PubMed]
  • Liang P, Pardee AB. Differential display of eukaryotic messenger RNA by means of the polymerase chain reaction. Science. 1992 Aug 14;257(5072):967–971. [PubMed]
  • Jørgensen M, Bévort M, Kledal TS, Hansen BV, Dalgaard M, Leffers H. Differential display competitive polymerase chain reaction: an optimal tool for assaying gene expression. Electrophoresis. 1999 Feb;20(2):230–240. [PubMed]
  • Darbre P, Yates J, Curtis S, King RJ. Effect of estradiol on human breast cancer cells in culture. Cancer Res. 1983 Jan;43(1):349–354. [PubMed]
  • Voss H, Nentwich U, Duthie S, Wiemann S, Benes V, Zimmermann J, Ansorge W. Automated cycle sequencing with Taquenase: protocols for internal labeling, dye primer and "doublex" simultaneous sequencing. Biotechniques. 1997 Aug;23(2):312–318. [PubMed]
  • Fields C, Adams MD, White O, Venter JC. How many genes in the human genome? Nat Genet. 1994 Jul;7(3):345–346. [PubMed]
  • Jakowlew SB, Breathnach R, Jeltsch JM, Masiakowski P, Chambon P. Sequence of the pS2 mRNA induced by estrogen in the human breast cancer cell line MCF-7. Nucleic Acids Res. 1984 Mar 26;12(6):2861–2878. [PMC free article] [PubMed]
  • Orlando C, Pinzani P, Pazzagli M. Developments in quantitative PCR. Clin Chem Lab Med. 1998 May;36(5):255–269. [PubMed]
  • Santagati S, Garnier M, Carlo P, Violani E, Picotti GB, Maggi A. Quantitation of low abundance mRNAs in glial cells using different polymerase chain reaction (PCR)-based methods. Brain Res Brain Res Protoc. 1997 Aug;1(3):217–223. [PubMed]
  • Rio MC. Quelle(s) fonction(s) pour les peptides dits "en feuille de trèfle"? Bull Cancer. 1997 Apr;84(4):443–446. [PubMed]
  • Taupin D, Wu DC, Jeon WK, Devaney K, Wang TC, Podolsky DK. The trefoil gene family are coordinately expressed immediate-early genes: EGF receptor- and MAP kinase-dependent interregulation. J Clin Invest. 1999 May;103(9):R31–R38. [PMC free article] [PubMed]
  • Jeltsch JM, Roberts M, Schatz C, Garnier JM, Brown AM, Chambon P. Structure of the human oestrogen-responsive gene pS2. Nucleic Acids Res. 1987 Feb 25;15(4):1401–1414. [PMC free article] [PubMed]
  • Berry M, Nunez AM, Chambon P. Estrogen-responsive element of the human pS2 gene is an imperfectly palindromic sequence. Proc Natl Acad Sci U S A. 1989 Feb;86(4):1218–1222. [PMC free article] [PubMed]
  • El-Ashry D, Chrysogelos SA, Lippman ME, Kern FG. Estrogen induction of TGF-alpha is mediated by an estrogen response element composed of two imperfect palindromes. J Steroid Biochem Mol Biol. 1996 Nov;59(3-4):261–269. [PubMed]
  • Wang F, Porter W, Xing W, Archer TK, Safe S. Identification of a functional imperfect estrogen-responsive element in the 5'-promoter region of the human cathepsin D gene. Biochemistry. 1997 Jun 24;36(25):7793–7801. [PubMed]
  • Kraus WL, Montano MM, Katzenellenbogen BS. Identification of multiple, widely spaced estrogen-responsive regions in the rat progesterone receptor gene. Mol Endocrinol. 1994 Aug;8(8):952–969. [PubMed]
  • Wahli W, Martinez E, Corthésy B, Cardinaux JR. cis- and trans-acting elements of the estrogen-regulated vitellogenin gene B1 of Xenopus laevis. J Steroid Biochem. 1989;34(1-6):17–32. [PubMed]
  • Ma ZQ, Bondiolotti GP, Olasmaa M, Violani E, Patrone C, Picotti GB, Maggi A. Estrogen modulation of catecholamine synthesis and monoamine oxidase A activity in the human neuroblastoma cell line SK-ER3. J Steroid Biochem Mol Biol. 1993 Dec;47(1-6):207–211. [PubMed]
  • Ma ZQ, Violani E, Villa F, Picotti GB, Maggi A. Estrogenic control of monoamine oxidase A activity in human neuroblastoma cells expressing physiological concentrations of estrogen receptor. Eur J Pharmacol. 1995 Sep 15;284(1-2):171–176. [PubMed]
  • Holschneider DP, Kumazawa T, Chen K, Shih JC. Tissue-specific effects of estrogen on monoamine oxidase A and B in the rat. Life Sci. 1998;63(3):155–160. [PubMed]
  • Klaiber EL, Broverman DM, Vogel W, Peterson LG, Snyder MB. Individual differences in changes in mood and platelet monoamine oxidase (MAO) activity during hormonal replacement therapy in menopausal women. Psychoneuroendocrinology. 1996 Oct;21(7):575–592. [PubMed]
  • Klaiber EL, Broverman DM, Vogel W, Peterson LG, Snyder MB. Relationships of serum estradiol levels, menopausal duration, and mood during hormonal replacement therapy. Psychoneuroendocrinology. 1997 Oct;22(7):549–558. [PubMed]
  • Chakravorty SG, Halbreich U. The influence of estrogen on monoamine oxidase activity. Psychopharmacol Bull. 1997;33(2):229–233. [PubMed]
  • Sarabia SF, Liehr JG. Induction of monoamine oxidase B by 17 beta-estradiol in the hamster kidney preceding carcinogenesis. Arch Biochem Biophys. 1998 Jul 15;355(2):249–253. [PubMed]
  • Bates SE, Davidson NE, Valverius EM, Freter CE, Dickson RB, Tam JP, Kudlow JE, Lippman ME, Salomon DS. Expression of transforming growth factor alpha and its messenger ribonucleic acid in human breast cancer: its regulation by estrogen and its possible functional significance. Mol Endocrinol. 1988 Jun;2(6):543–555. [PubMed]
  • Takahashi T, Eitzman B, Bossert NL, Walmer D, Sparrow K, Flanders KC, McLachlan J, Nelson KG. Transforming growth factors beta 1, beta 2, and beta 3 messenger RNA and protein expression in mouse uterus and vagina during estrogen-induced growth: a comparison to other estrogen-regulated genes. Cell Growth Differ. 1994 Sep;5(9):919–935. [PubMed]
  • Dannecker C, Possinger K, Classen S. Induction of TGF-beta by an antiprogestin in the human breast cancer cell line T-47D. Ann Oncol. 1996 Apr;7(4):391–395. [PubMed]
  • Knabbe C, Lippman ME, Wakefield LM, Flanders KC, Kasid A, Derynck R, Dickson RB. Evidence that transforming growth factor-beta is a hormonally regulated negative growth factor in human breast cancer cells. Cell. 1987 Feb 13;48(3):417–428. [PubMed]
  • Derynck R, Lindquist PB, Lee A, Wen D, Tamm J, Graycar JL, Rhee L, Mason AJ, Miller DA, Coffey RJ, et al. A new type of transforming growth factor-beta, TGF-beta 3. EMBO J. 1988 Dec 1;7(12):3737–3743. [PMC free article] [PubMed]
  • Massot O, Baskevitch PP, Capony F, Garcia M, Rochefort H. Estradiol increases the production of alpha 1-antichymotrypsin in MCF7 and T47D human breast cancer cell lines. Mol Cell Endocrinol. 1985 Oct;42(3):207–214. [PubMed]
  • Krøll J, Briand P. Estrogen-dependent release of serum proteins from MCF-7 breast carcinoma cells in vitro. Anticancer Res. 1988 Jan-Feb;8(1):89–91. [PubMed]
  • Blumberg B, Evans RM. Orphan nuclear receptors--new ligands and new possibilities. Genes Dev. 1998 Oct 15;12(20):3149–3155. [PubMed]
  • Dowsett M, Macaulay V, Gledhill J, Ryde C, Nicholls J, Ashworth A, McKinna JA, Smith IE. Control of aromatase in breast cancer cells and its importance for tumor growth. J Steroid Biochem Mol Biol. 1993 Mar;44(4-6):605–609. [PubMed]
  • Bowtell DD. Options available--from start to finish--for obtaining expression data by microarray. Nat Genet. 1999 Jan;21(1 Suppl):25–32. [PubMed]
  • Debouck C, Goodfellow PN. DNA microarrays in drug discovery and development. Nat Genet. 1999 Jan;21(1 Suppl):48–50. [PubMed]

Articles from Environmental Health Perspectives are provided here courtesy of National Institute of Environmental Health Science

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

  • Compound
    Compound
    PubChem Compound links
  • MedGen
    MedGen
    Related information in MedGen
  • PubMed
    PubMed
    PubMed citations for these articles
  • Substance
    Substance
    PubChem Substance links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...