| Assaying Estrogenicity by Quantitating the Expression Levels of Endogenous Estrogen-Regulated Genes Marianne Jørgensen, Brian Vendelbo, Niels E. Skakkebæk, and Henrik Leffers Department of Growth and Reproduction, Rigshospitalet, Copenhagen, Denmark 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 ß3 (TGFß3) , monoamine oxidase A, and  1-antichymotrypsin--and we evaluated estrogenic activity for 17ß-estradiol (E2) , diethylstilbestrol, -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 (E2, diethylstilbestrol, and -zearalanol) , whereas the potency of the other chemicals was 103- to 106-fold lower than that of E2. There were some marker gene-dependent differences in the relative potencies of the tested chemicals. TGFß3 was equally sensitive to the three high-potency estrogens, whereas the sensitivity to -zearalanol was approximately 10-fold lower than the sensitivity to E2 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 TGFß3, but 10- to 100-fold higher/lower with monoamine oxidase A and 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. Key words: 1-antichymotrypsin, competitive PCR, differential display, endocrine disruptors, estrogen, estrogenicity assay, gene expression, monoamine oxidase A, pS2, TGFß3. Environ Health Perspect 108:403-412 (2000) . [Online 17 March 2000] http://ehpnet1.niehs.nih.gov/docs/2000/108p403-412jorgensen/ abstract.html Address correspondence to M. Jørgensen, Department of Growth and Reproduction, Section GR-5064, Rigshospitalet, Blegdamsvej 9, DK-2100 Copenhagen, Denmark. Telephone: 45 35455127. Fax: 45 35456054. E-mail: marj@biobase.dk Funding for the work was provided by The Danish Strategic Environmental Initiative, The Danish Research Councils, and The European Commission. Received 16 August 1999 ; accepted 1 November 1999.
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