Printer Friendly

The need to decide if all estrogens are intrinsically similar.

We used gene expression profiling m investigate whether the molecular effects induced by estrogens of different provenance are intrinsically similar. In this article we show that the physiologic estrogen 17[beta]-estradiol, the phytoestrogen genistein, and the synthetic estrogen diethylstilbestrol alter the expression of the same 179 genes in the intact immature mouse uterus under conditions where each chemical has produced an equivalent gravimetric and histologic uterotrophic effect, using the standard 3-day assay protocol. Data are also presented indicating the limitations associated with comparison of gene expression profiles for different chemicals at times before the uterotrophic effects are fully realized. We conclude that the case has yet to be made for regarding synthetic estrogens as presenting a unique human hazard compared with phytoestrogens and physiologic estrogens. Key words: diethylstilbestrol, estrogen, gene expression, genistein, microarray, phytoestrogen, toxicogenomics, uterus. Environ Health Perspect 112:1137-1142 (2004). doi:10.1289/ehp.7028 available via http://dx.doi.org/[Online 19 May 2004]

**********

The question of whether phytoestrogens and synthetic estrogens are toxicologically similar, or intrinsically different, presents a challenge to all involved in human hazard and risk assessments. Although there is a general concern that exposure to nanogram or microgram amounts of environmental estrogens may be associated with adverse health effects, in the public mind there is a widespread belief that foods and dietary supplements containing milligram quantities of phytoestrogens confer only health benefits. An implicit distinction therefore seems to have been drawn between synthetic and plant-derived estrogens--a belief sustained in the public mind by the assumption that natural is good and synthetic is bad--but an untested and potentially misleading notion for those involved with science-based human hazard/risk assessments.

Phytoestrogens and synthetic estrogens are generally considered separately in the literature. For example, Howdeshell et al. (1999) suggested a possible association between the advance in first estrus observed in mice exposed in utero to 2.4 [micro]g/kg of the synthetic environmental estrogen bisphenol A and reports of an increased incidence of hypospadias in boys (Paulozzi et al. 1997) and the earlier sexual maturation of girls (Herman-Giddens et al. 1997)--the implication being that synthetic estrogens present a greater hazard than the much higher levels of phytoestrogens being consumed by those same children. In contrast, there are reports of an increased incidence of hypospadias in boys born to vegetarians (North and Golding 2000), of alterations in the menstrual cycle (Cassidy et al. 1994), and of reduced breast cancer incidences (Messina 1999) among women eating diets rich in phytoestrogens. Support for these epidemiologic observations comes from experimental studies indicating that advances in sexual development in rodents can be induced by their exposure to phytoestrogens (Casanova et al. 1999; Cassidy and Fanghnan 2000; Safe et al. 2002). In contrast to these separate lines of inquiry, Newbold and colleagues have evaluated potential similarities between natural and synthetic estrogens. In seminal studies, they demonstrated that neonatal exposure of female mice to equipotent uterotrophic doses of the phytoestrogen genistein (GEN; Figure 1) or the synthetic estrogen diethylstilbestrol (DES) leads to an identical incidence of uterine adenomas at 18 months of age (Newbold et al. 2001). However, in attempting to draw parallels, or distinctions, between phytoestrogens and synthetic estrogens, it is imperative to consider growing awareness of the complexity of estrogen signaling pathway and the pleuripotential biologic activities of most organic chemicals--irrespective of their origin.

[FIGURE 1 OMITTED]

Estrogen signaling in mammalian cells is primarily mediated at the molecular level by two members of the nuclear receptor superfamily--estrogen receptors alpha (ER-[alpha]) and beta (ER-[beta]). Ligand-activated ER-[alpha] and ER-[beta] function as transcription factors, in conjunction with numerous coregulatory proteins, in order to activate or repress the transcription of ER-responsive genes (Hall et al. 2001; Moggs and Orphanides 2001). There is considerable variation in the binding affinity of ER-[alpha] and ER-[beta] among different estrogens (Kuiper et al. 1998). In the case of the chemicals studied here, the physiologic estrogen 17[beta]-estradiol ([E.sub.2]) and DES bind with a similar affinity to ER-[alpha] and ER-[beta], whereas GEN binds with approximately 20-fold higher affinity to ER-[beta] than to ER-[alpha] (Kuiper et al. 1998). Concerning nonhormonal properties of the test chemicals (most of which have only be defined in vitro), GEN inhibits a range of enzymes, including tyrosine kinases (Akiyama et al. 1987), nitric oxide synthase (Duarte et al. 1997), and topoisomerase II (Okura et al. 1988), and also decreases calcium-channel activity (Potier and Rovira 1999), lipid peroxidation (Arora et al. 1998), and diacylglycerol synthesis (Dean et al. 1989). Likewise, DES is reported to induce aneuploidy in mammalian cells (Aardema et al. 1998) and to bind to rat liver DNA (Williams et al. 1993). More recently, some phytoestrogens were reported to inhibit the aromatase-mediated conversion of testosterone to [E.sub.2] in vitro (Almstrup et al. 2002), and equol, the major circulating estrogenic metabolite associated with the dietary ingestion of phytoestrogens, is reported to selectively sequester dihydrotestosterone and thereby to act as a functional antiandrogen in vivo (Lund et al. 2004).

In order to advance understanding in this area, we decided to compare the genes expressed in the immature mouse uterus when it had grown in response to treatment with the estrogens [E.sub.2], DES, and GEN. The immature mouse uterus was selected for our analysis because it is a major estrogen-responsive organ and forms the basis for a reference assay of estrogenic activity (Owens and Ashby 2002), including carcinogenesis (Newbold et al. 2001). Furthermore, it expresses both ER-[alpha] and ER-[beta] (Weihua et al. 2000) and the androgen receptor (Frasor et al. 2003). We initially conducted a global analysis of gene expression in the mouse uterus at 1, 2, 4, 8, 24, 48, and 72 hr after exposure to a single high dose of either GEN (250 mg/kg) or [E.sub.2] (400 [micro]g/kg). These single high doses yielded a sustained uterotrophic response over 72 hr (Figure 2A) and were selected to avoid the complex transcriptional program that may result from the standard uterotrophic assay exposure regime in which each test compound is dosed by repeated administration on 3 consecutive days (Odum et al. 1997). Groups of 10 sexually immature mice [Alpk:APfCD-1; 19/20 days of age; maintained on RM1 diet (Special Diets Services Ltd., Witham, Essex, UK)] received a single subcutaneous injection of each compound or the test vehicle [arachis oil (AO); 5 mL/kg], and uterine RNA was isolated and pooled by group at each of the seven time points to determine gene expression levels among the 12,488 mouse genes represented on the Affymetrix MG-U74Av2 GeneChip (Affymetrix, High Wycombe, UK). Transcript profiling was performed using MG-U74Av2 GeneChip and Microarray Analysis Suite 5.0 (Affymetrix). Normalization and hierarchical clustering were performed with GeneSpring 6.0 (Silicon Genetics, Redwood City, CA, USA). MIAME (Minimum Information About a Microarray Experiment)-compliant microarray data are available as supplementary information and submitted to the Gene Expression Omnibus (GEO) database (GEO 2004). These data were analyzed using unsupervised hierarchical clustering and yielded temporal relationships between the expression profiles of 3,450 genes that were either up- or down-regulated (> 1.5-fold) by [E.sub.2] and/or GEN (Figure 2B). Each chemical induced a similar, multistage transcriptional response (Figure 2B), although it is noteworthy that we observed variations in the magnitude and timing of both early (e.g., c-fos) and late (e.g., lactotransferrin) ER-responsive genes during the uterotrophic responses induced by [E.sub.2] and GEN (Figure 2C).

[FIGURE 2 OMITTED]

A detailed description of the molecular functions of the genes affected, together with their association with physiologic changes during uterine growth, has been reported (Orphanides et al. 2003) and will be described in more detail in a future publication (Moggs et al., unpublished data).

These observations suggest that GEN does not induce "off-target" ER-independent transcriptional responses, that is, those associated with the properties of GEN other than estrogenicity. Furthermore, there was no evidence for the topoisomerase II-inhibiting properties of GEN in the bone marrow of the present mice despite demonstration of the sensitivity of that tissue to the potent micronucleus-inducing activity of the topoisomerase II inhibitor etoposide (data not shown). Together, these data led us to question whether a synthetic estrogen such as DES would also induce similar transcriptional responses in the immature mouse uterus.

In order to avoid temporal vagaries in gene expression (e.g., Figure 2C), we decided to anchor our transcript profiling data to the phenotype of the grown uterus by employing equipotent uterotrophic doses of [E.sub.2], GEN, and DES. We compared the global gene expression profiles in the uteri of intact immature mice stimulated with three daily low doses of either GEN, DES, or [E.sub.2], with an exposure regimen the same as that used in a standard 3-day uterotrophic assay (Odum et al. 1997). The route of administration and the doses of GEN and DES used were as described by Newbold et al. (2001) in their equivalent-outcome carcinogenicity bioassays of these two chemicals. Three independent replicates of four groups of sexually immature mice (Alpk:APfCD-1; 19/20 days of age; maintained on RM1 diet) received three daily subcutaneous injections of GEN (50 mg/kg), [E.sub.2] (2.5 [micro]g/kg), or DES (2 lag/kg). Control animals received the vehicle, AO (5 mL/kg). These doses elicited similar uterotrophic responses (72 hr after the initial dose; Figure 3A, Table 1) and identical histologic changes in the uteri of the treated animals (Table 1). Uterine RNA was isolated and pooled for each of the 12 groups and analyzed for changes in gene expression levels using the same Affymetrix microarray of 12,488 mouse genes. The data were analyzed using two independent statistical methods. First, unsupervised hierarchical clustering defined the global relationships (Euclidean distances) between the 12 gene expression profiles (Figure 3B). The three control groups clustered under one node, whereas the chemical treatment groups formed a separate node of compound-independent clusters, indicating equal similarity within and between the transcriptional responses induced by the three estrogens (Figure 3B). One-way analysis of variance (ANOVA), with Bonferroni (Holm 1979) correction (familywise error rate < 0.05) to minimize false positives, identified 179 genes where expression levels were altered by one or more chemical treatments (Figure 3C). Remarkably, Tukey post hoc testing revealed that all of these genes were affected in all nine compound treatment groups.

[FIGURE 3 OMITTED]

Table 2 highlights the high degree of similarity between the transcriptional responses to each of the three estrogens. These include established estrogen-responsive genes such as lactotransferrin, complement component 3, c-fos, small proline-rich protein 2A, and keratoepithelin (Hewitt et al. 2003; Naciff et al. 2003), together with many genes that have not previously been associated with estrogenicity (Table 2).

Although these three estrogens can alter the expression of some genes with different magnitudes [e.g., peptidyl arginine deiminase II is up-regulated to a lesser extent by [E.sub.2] (1.86-fold [+ or minus] 0.27) relative to GEN (9.11-fold [+ or -] 0.33) and DES (5.15-fold [+ or -] 1.53); Table 2], the present data show that the same genes are affected during equivalent uterotrophic responses. Previous studies have revealed both similarities and differences between transcriptional responses induced at a single time point after exposure to [E.sub.2] and DES in the uteri of immature ovariectomized mice (Watanabe et al. 2003) and after exposure to either GEN, bisphenol A, or 170[alpha]-ethynyl estradiol in the reproductive tract of intact adult rats (Naciff et al. 2002). We suggest that these reported differences most probably arise from dose-dependent variations in the magnitude and kinetics of gene expression (Figure 2C), rather than from the operation of distinct mechanisms of estrogenic action.

Our data indicate that estrogens of differing provenance may have in common the potential for both beneficial and adverse health effects. This highlights the need for an holistic approach to hazard assessment wherein preconceptions are replaced by an objective assessment of the likely perturbations of physiologic functions caused by combined exposures to physiologic, synthetic, and plant-derived estrogens. This need is reinforced by data showing that plasma concentrations of isoflavones in infants fed soy formula are approximately 200 times higher than for those fed human milk (Setchell et al. 1997), by the estimated daily intake of approximately 29 mg of phytoestrogens for individuals taking dietary supplements (Committee on Toxicity of Chemicals in Food, Consumer Products and the Environment 2003), and by the demonstration that estrogens of different provenance can act additively in the rodent uterus (Tinwell and Ashby 2004).
Table 1. Blotted uterine weights and endometrial and epithelial
cell heights (mean [+ or -] SDI after exposure to [E.sub.2],
GEN, or DES for 3 consecutive days. (a)

 Dose Blotted uterine
Compound (per kg) weight (m g)

AD 5 mL 13.0 [+ or -] 2.4
[E.sub.2] 2.5 [micro]g 45.3 [+ or -] 8.6 *
GEN 50 [micro]g 39.8 [+ or -] 5.3 *
DES 2.0 [micro]g 49.8 [+ or -] 13.0 *

 Cell height ([micro]m)

Compound Endometrium Epithelium

AD 159.0 [+ or -] 23.1 (11) 11.4 [+ or -] 1.1
[E.sub.2] 246.1 [+ or -] 52.4 * (9) 23.3 [+ or -] 1.4 *
GEN 273.7 [+ or -] 63.3 * (12) 23.7 [+ or -] 3.1 *
DES 273.2 [+ or -] 55.9 * (10) 22.6 [+ or -] 4.0 *

There were 12 animals/group, but not all of the histopathology
samples were suitable for analyses; numbers in parentheses
indicate the number of animals per group from which the histology
data were generated.

(a) Data were assessed for statistical significance using a
two-sided Student t-test: * p < 0.01.

Table 2. Quantitative data for 179 differentially expressed genes
(from Figure 3C) regulated in the mouse uterus by all three
estrogens ([E.sub.2], GEN, and DES). (a)

 Fold change in
 expression (mean
 [+ or -] SD)

 GenBank
 accession
Gene name no. [E.sub.2]

Up-regulated genes
 Solute carrier family U74079 1.8 [+ or -] 0.01
 9a3r1
 Keratin complex 2-8 X15662 2.6 [+ or -] 0.2
 Laminin beta 3 U43298 4.3 [+ or -] 0.1
 Claudin 7 AF087825 4.5 [+ or -] 0.5
 bHLH-Zip transcription U49507 2.6 [+ or -] 0.3
 factor
 RIKEN cDNA 1200008D14 AW208938 3.0 [+ or -] 0.3
 Basic HLH-domain Y07836 5.9 [+ or -] 1.0
 containing, class B2
 RIKEN cDNA 9930104H07 AW122310 3.0 [+ or -] 0.3
 Fucosyltransferase 2 AF064792 27.5 [+ or -] 1.2
 Deleted in polyposis 1 U28168 1.8 [+ or -] 0.1
 Microsomal glutathione AI843448 2.9 [+ or -] 0.2
 S-transferase 3
 Tumor-associated Ca Y08830 4.0 [+ or -] 0.3
 signal transducer 2
 Calpain 5 Y10656 5.5 [+ or -] 0.4
 Mitochondrial creatine Z13969 9.7 [+ or -] 1.1
 kinase
 ATPase 6v1a1 AW123765 2.0 [+ or -] 0.1
 Tumor-associated Ca AI563854 8.0 [+ or -] 0.4
 signal transducer 2
 Lymphocyte antigen 6 X04653 7.8 [+ or -] 0.9
 complex, locus A
 Chloride channel AV373378 26.4 [+ or -] 3.4
 calcium-activated 3
 Small proline-rich AJ005567 23.9 [+ or -] 1.5
 protein 21
 Oncoprotein induced AA615075 19.0 [+ or -] 3.1
 transcript 1
 Small proline-rich AJ005564 59.8 [+ or -] 8.4
 protein 2F
 Small proline-rich AJ005563 12.0 [+ or -] 1.0
 protein 2E
 Mucin 1 M84683 8.3 [+ or -] 0.6
 Lipoocalin 2 X81627 150.3 [+ or -] 15.0
 RIKEN cDNA 2210409B01 AF109906 3.5 [+ or -] 0.6
 Interferon-activated M31418 7.9 [+ or -] 1.0
 gene 202A
 Nuclear ankyrin-repeat AA614971 3.7 [+ or -] 0.6
 protein
 RIKEN cDNA 5730469M10 AI850090 22.0 [+ or -] 5.8
 RIKEN cDNA 1110034C02 AI837104 1.5 [+ or -] 0.1
 IMAGE cDNA 4988271 AV373294 8.0 [+ or -] 2.5
 RIKEN cDNA 5730493B19 AW122413 12.7 [+ or -] 0.3
 Peptidoglycan AV092014 13.4 [+ or -] 1.5
 recognition protein
 Inhibin beta-B X69620 13.6 [+ or -] 3.1
 CEA-related cell AF101164 11.9 [+ or -] 1.6
 adhesion molecule 2
 Keratin complex 1-19 M36120 4.4 [+ or -] 0.4
 CEA-related cell M77196 15.9 [+ or -] 2.4
 adhesion molecule 1
 SRC family-associated AB014485 2.7 [+ or -] 0.04
 phosphoprotein 2
 Peptidoglycan AF076482 7.7 [+ or -] 1.9
 recognition protein
 CEA-related cell M77196 19.5 [+ or -] 3.9
 adhesion molecule 1
 CEA-related cell X67279 6.4 [+ or -] 0.7
 adhesion molecule 1
 Spermidine N1-acetyl L10244 8.3 [+ or -] 0.9
 transferase
 RIKEN cDNA 0610007007 AI851762 2.7 [+ or -] 0.1
 Arginase 1 U51805 79.4 [+ or -] 9.8
 Acetyl-coenzyme A
 synthetase 2 AW125884 2.2 [+ or -] 0.2
 v-erb-b2 homolog 3 AI006228 3.4 [+ or -] 0.4
 Phospholipase D3 AF026124 2.6 [+ or -] 0.2
 RIKEN cDNA 0610031J06 AW122935 1.9 [+ or -] 0.1
 Complement component 1q X58861 2.1 [+ or -] 0.1
 Scotin AW123754 2.0 [+ or -] 0.1
 CD24a antigen M58661 3.2 [+ or -] 0.1
 Argininosuccinate M31690 2.7 [+ or -] 0.3
 synthetase 1
 ATPase 6v1a1 U13837 2.1 [+ or -] 0.1
 Gelsolin-like actin- X54511 3.6 [+ or -] 0.5
 capping protein
 Golgi phosphoprotein 2 AW125446 4.5 [+ or -] 0.5
 Aldolase 1A Y00516 2.3 [+ or -] 0.2
 Cathepsin L X06086 6.4 [+ or -] 0.8
 CD14 antigen X13333 3.0 [+ or -] 0.1
 Decay accelerating L41365 4.0 [+ or -] 0.1
 factor 2
 Actin-related protein AW212775 2.1 [+ or -] 0.2
 2/3 complex 1B
 Protective protein for J05261 2.0 [+ or -] 0.1
 [beta]-galactosidase
 Elastase 1 M27347 2.7 [+ or -] 0.1
 Connexin 26 M81445 10.6 [+ or -] 1.0
 Ceruloplasmin U49430 15.1 [+ or -] 2.8
 Cathepsin H U06119 3.0 [+ or -] 0.2
 Basigin Y16258 1.6 [+ or -] 0.1
 Peptidylprolyl isomerase X67809 2.2 [+ or -] 0.2
 C-associated
 Glutathione reductase 1 AI851983 2.3 [+ or -] 0.2
 START domain- X82457 1.5 [+ or -] 0.1
 containing 3
 CD68 antigen X68273 4.6 [+ or -] 0.6
 RIKEN cDNA E030027H19 AW211760 2.7 [+ or -] 0.3
 cDNA sequence B0004044 AI461767 3.1 [+ or -] 0.2
 E74-like factor 3 AF016294 5.1 [+ or -] 0.8
 Glutathione S- AI843119 5.0 [+ or -] 1.1
 transferase omega 1
 Interferon-stimulated AW122677 4.2 [+ or -] 0.1
 protein 20
 Clusterin D14077 3.6 [+ or -] 0.7
 Galectin 3 X16834 7.4 [+ or -] 1.3
 Small proline-rich AJ005559 51.1 [+ or -] 4.0
 protein 2A (b)
 Complement component K02782 14.8 [+ or -] 1.8
 3 (b)
 Small proline-rich AJ005561 220.3 [+ or -] 31.0
 protein 2C
 Small proline-rich AJ005565 9.4 [+ or -] 0.9
 protein 2G
 Prominin AF039663 3.5 [+ or -] 0.5
 Lactotransferrin (b) J03298 88.7 [+ or -] 18.4
 Carbonic anhydrase 2 M25944 7.9 [+ or -] 0.5
 Complement component U47810 36.5 [+ or -] 4.4
 factor 1
 Mannosidase 2alphaB1 U87240 2.0 [+ or -] 0.2
 Small proline-rich AJ005560 32.9 [+ or -] 3.7
 protein 2B
 Small proline-rich AJ005559 269.8 [+ or -] 23.7
 protein 2A (b)
 RIKEN cDNA 5830413E08 AI849939 3.3 [+ or -] 0.4
 RIKEN cDNA 1110029F20 AW125508 4.1 [+ or -] 0.1
 Annexin A3 AJ001633 2.7 [+ or -] 0.5
 Peptidase 4 U51014 2.0 [+ or -] 0.1
 Laminin gamma 2 U43327 6.3 [+ or -] 1.3
 Ubiquitin-like 3 AW120725 1.5 [+ or -] 0.1
 Urate oxidase M27695 23.8 [+ or -] 9.5
 Amiloride binding AI197481 3.5 [+ or -] 1.0
 protein 1
 Keratin complex 1-19 AU040563 4.5 [+ or -] 1.0
 Activated leukocyte cell L25274 3.6 [+ or -] 0.8
 adhesion molecule
 CCAAT/enhancer binding M61007 2.3 [+ or -] 0.1
 protein [beta]
 Peptidyl arginine AB013848 8.6 [+ or -] 0.7
 deiminase, type I
 Enolase 1 [alpha] AI841389 2.5 [+ or -] 0.3
 p53 apoptosis effector AI854029 2.9 [+ or -] 0.3
 related to Pmp22
 [beta]-Glucuronidase M19279 1.9 [+ or -] 0.1
 Leucine-rich AW230891 9.3 [+ or -] 1.1
 [alpha]-2-glycoprotein
 Quiescin 06 AW123556 3.7 [+ or -] 0.2
 GADD45a U00937 1.9 [+ or -] 0.2
 Alkaline phosphatase 2 J02980 9.2 [+ or -] 0.4
 Immediate early X67644 5.5 [+ or -] 0.8
 response 3
 Progressive ankylosis AW049351 2.2 [+ or -] 0.1
 RAS p21 protein AA163960 6.8 [+ or -] 0.9
 activator 4
 Tumor-associated calcium M76124 2.1 [+ or -] 0.2
 signal transducer 1
 Hydroxysteroid (17-beta) AA822174 1.9 [+ or -] 0.1
 dehydrogenase 11
 Platelet-activating U57746 1.9 [+ or -] 0.1
 factor acetylhydrolase
 1ba1
 Branched chain U42443 2.4 [+ or -] 0.2
 aminotransferase 1
 RIKEN cDNA 2400004E04 AI846720 1.7 [+ or -] 0.1
 Myeloblastosis oncogene M12848 2.8 [+ or -] 0.4
 [K.sup.+] conductance AF042487 3.1 [+ or -] 0.2
 calcium-activated
 channel N4
 ATPase 6v1b2 AI843029 1.7 [+ or -] 0.1
 Cystic fibrosis M60493 3.4 [+ or -] 0.5
 transmembrane
 regulator
 RIKEN cDNA 1110008P14 AI839839 4.3 [+ or -] 0.4
 Fused toes Z67963 2.6 [+ or -] 0.2
 Solute carrier family AW124340 3.5 [+ or -] 0.5
 39a8
 Cytochrome b-561 AI846517 2.2 [+ or -] 0.2
 Secreted X13986 30.2 [+ or -] 3.3
 phosphoprotein 1
 Ion transport regulator X93038 5.3 [+ or -] 0.4
 Fxyd3
 Janus kinase 3 L40172 2.1 [+ or -] 0.2
 Cytochrome b-245alpha AW046124 2.9 [+ or -] 0.4
 RIKEN cDNA A430096B05 AI465965 6.3 [+ or -] 1.0
 Small proline-rich AJ005568 8.6 [+ or -] 2.1
 protein 2J
 Cathepsin B M65270 2.2 [+ or -] 0.1
 RIKEN cDNA 1600025H15 AI842734 2.2 [+ or -] 0.1
 c-fos oncogene (b) V00727 3.2 [+ or -] 0.4
 Guanine nucleotide AI843937 1.6 [+ or -] 0.1
 binding protein
 [gamma]5
 Serine palmitoyl- U27455 1.6 [+ or -] 0.1
 transferase Ic2
 Cystatin B U59807 1.5 [+ or -] 0.1
 Villin 2 X60671 1.9 [+ or -] 0.2
 RIKEN cDNA 0610010012 AI849011 1.9 [+ or -] 0.1
 Matrix metallo- L36244 47.8 [+ or -] 18.6
 proteinase 7
 RIKEN cDNA 4930422J18 AV376312 2.0 [+ or -] 0.3
 RIKEN cDNA 1700017B05 AW049360 1.6 [+ or -] 0.1
 Galactosidase beta 1 M57734 1.8 [+ or -] 0.1
 Cathepsin C U74683 2.6 [+ or -] 0.1
 Interferon-stimulated X56602 3.6 [+ or -] 0.6
 protein 15
 MAP kinase--interacting Y11092 1.8 [+ or -] 0.1
 kinase 2
 Glutathione S- X98056 3.5 [+ or -] 0.4
 transferase theta 2
 Homeobox B6 M18401 1.5 [+ or -] 0.02
 Procollagen Vlalpha 3 AF064749 2.1 [+ or -] 0.2
 Interferon regulatory U73037 11.7 [+ or -] 0.9
 factor 7
 Scavenger receptor class AB008553 2.7 [+ or -] 0.1
 B2
 Polyimmunoglobulin AB001489 8.1 [+ or -] 0.7
 receptor
 Proteasome subunit Y10875 2.1 [+ or -] 0.04
 [beta]10
 RIKEN cDNA 0610010E05 AV312736 2.9 [+ or -] 0.3
 RIKEN cDNA 0610010E05 AI854839 3.7 [+ or -] 0.5
 Xanthine dehydrogenase X75129 12.2 [+ or -] 2.2
 Prominin AF039663 3.5 [+ or -] 0.2
 Interferon-induced U43084 17.5 [+ or -] 2.9
 protein IFIT1
 Interferon-induced U43086 8.1 [+ or -] 2.3
 protein IFIT3
 Proteasome subunit U22033 2.0 [+ or -] 0.1
 [beta]8
 RIKEN cDNA 1600023A02 AW121336 1.9 [+ or -] 0.1
 Small proline-rich AF057156 11.1 [+ or -] 3.7
 protein 1A
 MAP kinase-interacting AI845732 2.0 [+ or -] 0.1
 kinase 2
 Lymphocyte antigen 6 U47737 2.0 [+ or -] 0.01
 complex, locus E
 Guanylate nucleotide AJ007970 3.0 [+ or -] 0.1
 binding protein 2
 Peptidyl arginine D16580 1.9 [+ or -] 0.3
 deiminase, type II (b)
Down-regulated genes
 Solute carrier family AI838274 2.0 [+ or -] 0.2
 29a1
 Lymphocyte specific 1 D49691 1.6 [+ or -] 0.1
 Claudin 5 U82758 2.0 [+ or -] 0.2
 Potassium channel td12 AI842065 1.6 [+ or -] 0.04
 Zinc finger homeobox 1a D76432 1.5 [+ or -] 0.1
 Monoamine oxidase A AI848045 2.3 [+ or -] 0.2
 Histidine decarboxylase X57437 4.8 [+ or -] 0.8
 [alpha]-2 Adrenergic M97516 3.0 [+ or -] 0.3
 receptor
 Transcription factor 21 AF035717 1.8 [+ or -] 0.2
 Homeobox D8 X56561 2.2 [+ or -] 0.1
 Carboxypeptidase X2 AF017639 4.1 [+ or -] 0.7
 RIKEN cDNA A230106A15 AI848841 3.8 [+ or -] 0.2
 Reduced expression 3 AA790008 3.1 [+ or -] 0.2
 TGF-[beta] binding AA838868 1.8 [+ or -] 0.1
 protein 4
 Keratoepithelin (b) L19932 11.5 [+ or -] 2.5
 GLI-Kruppel family AB025922 11.6 [+ or -] 0.8
 member GLI

 Fold change in
 expression (mean [+ or -] SD)

Gene name GEN DES

Up-regulated genes
 Solute carrier family 2.0 [+ or -] 0.1 2.0 [+ or -] 0.2
 9a3r1
 Keratin complex 2-8 3.1 [+ or -] 0.2 3.1 [+ or -] 0.3
 Laminin beta 3 5.5 [+ or -] 1.1 5.3 [+ or -] 0.7
 Claudin 7 6.5 [+ or -] 1.0 5.8 [+ or -] 0.6
 bHLH-Zip transcription 3.1 [+ or -] 0.3 2.9 [+ or -] 0.1
 factor
 RIKEN cDNA 1200008D14 3.5 [+ or -] 0.1 3.3 [+ or -] 0.3
 Basic HLH-domain 6.6 [+ or -] 0.9 6.6 [+ or -] 0.8
 containing, class B2
 RIKEN cDNA 9930104H07 3.2 [+ or -] 0.4 3.3 [+ or -] 0.1
 Fucosyltransferase 2 34.6 [+ or -] 8.5 36.7 [+ or -] 5.5
 Deleted in polyposis 1 2.0 [+ or -] 0.02 2.0 [+ or -] 0.1
 Microsomal glutathione 3.3 [+ or -] 0.6 3.3 [+ or -] 0.1
 S-transferase 3
 Tumor-associated Ca 4.6 [+ or -] 0.9 4.6 [+ or -] 0.3
 signal transducer 2
 Calpain 5 6.3 [+ or -] 1.0 6.6 [+ or -] 0.6
 Mitochondrial creatine 12.2 [+ or -] 2.1 13.1 [+ or -] 1.8
 kinase
 ATPase 6v1a1 2.1 [+ or -] 13.2 2.1 [+ or -] 0.2
 Tumor-associated Ca 9.2 [+ or -] 1.0 8.5 [+ or -] 0.4
 signal transducer 2
 Lymphocyte antigen 6 8.8 [+ or -] 0.3 8.5 [+ or -] 0.4
 complex, locus A
 Chloride channel 26.7 [+ or -] 1.0 26.2 [+ or -] 3.9
 calcium-activated 3
 Small proline-rich 24.7 [+ or -] 1.3 23.6 [+ or -] 1.6
 protein 21
 Oncoprotein induced 20.0 [+ or -] 1.2 18.9 [+ or -] 2.5
 transcript 1
 Small proline-rich 65.8 [+ or -] 1.1 60.6 [+ or -] 2.6
 protein 2F
 Small proline-rich 12.9 [+ or -] 0.8 12.1 [+ or -] 0.9
 protein 2E
 Mucin 1 8.6 [+ or -] 0.3 8.5 [+ or -] 0.5
 Lipoocalin 2 175.7 [+ or -] 10.5 162.8 [+ or -] 6.5
 RIKEN cDNA 2210409B01 4.0 [+ or -] 0.3 3.8 [+ or -] 0.8
 Interferon-activated 9.8 [+ or -] 2.5 8.8 [+ or -] 0.7
 gene 202A
 Nuclear ankyrin-repeat 4.3 [+ or -] 0.7 4.1 [+ or -] 0.9
 protein
 RIKEN cDNA 5730469M10 30.5 [+ or -] 9.1 27.0 [+ or -] 6.5
 RIKEN cDNA 1110034C02 1.6 [+ or -] 0.1 1.6 [+ or -] 0.03
 IMAGE cDNA 4988271 10.6 [+ or -] 1.6 9.2 [+ or -] 1.1
 RIKEN cDNA 5730493B19 19.0 [+ or -] 4.1 15.7 [+ or -] 0.9
 Peptidoglycan 18.3 [+ or -] 3.0 14.5 [+ or -] 2.1
 recognition protein
 Inhibin beta-B 19.4 [+ or -] 4.7 16.0 [+ or -] 1.4
 CEA-related cell 17.8 [+ or -] 5.3 14.2 [+ or -] 1.7
 adhesion molecule 2
 Keratin complex 1-19 5.5 [+ or -] 1.1 4.8 [+ or -] 0.5
 CEA-related cell 23.9 [+ or -] 5.9 19.0 [+ or -] 3.8
 adhesion molecule 1
 SRC family-associated 3.2 [+ or -] 0.4 2.9 [+ or -] 0.3
 phosphoprotein 2
 Peptidoglycan 10.4 [+ or -] 2.7 9.0 [+ or -] 2.0
 recognition protein
 CEA-related cell 30.4 [+ or -] 8.8 22.2 [+ or -] 2.7
 adhesion molecule 1
 CEA-related cell 8.4 [+ or -] 1.1 7.1 [+ or -] 1.1
 adhesion molecule 1
 Spermidine N1-acetyl 11.2 [+ or -] 0.8 9.3 [+ or -] 0.6
 transferase
 RIKEN cDNA 0610007007 3.0 [+ or -] 0.3 2.8 [+ or -] 0.1
 Arginase 1 131.9 [+ or -] 20.0 99.6 [+ or -] 14.5
 Acetyl-coenzyme A
 synthetase 2 2.0 [+ or -] 0.1 2.2 [+ or -] 0.2
 v-erb-b2 homolog 3 3.1 [+ or -] 0.6 3.4 [+ or -] 0.4
 Phospholipase D3 2.4 [+ or -] 0.2 2.6 [+ or -] 0.2
 RIKEN cDNA 0610031J06 1.8 [+ or -] 0.1 1.8 [+ or -] 0.1
 Complement component 1q 2.0 [+ or -] 0.1 2.0 [+ or -] 0.2
 Scotin 2.0 [+ or -] 0.2 2.0 [+ or -] 0.2
 CD24a antigen 3.1 [+ or -] 0.1 3.3 [+ or -] 0.3
 Argininosuccinate 2.7 [+ or -] 0.3 2.8 [+ or -] 0.4
 synthetase 1
 ATPase 6v1a1 2.1 [+ or -] 0.2 2.2 [+ or -] 0.2
 Gelsolin-like actin- 3.7 [+ or -] 0.5 3.7 [+ or -] 0.2
 capping protein
 Golgi phosphoprotein 2 4.6 [+ or -] 0.5 4.7 [+ or -] 0.1
 Aldolase 1A 2.3 [+ or -] 0.1 2.4 [+ or -] 0.1
 Cathepsin L 6.3 [+ or -] 1.1 6.9 [+ or -] 0.4
 CD14 antigen 2.8 [+ or -] 0.1 3.1 [+ or -] 0.2
 Decay accelerating 3.8 [+ or -] 0.8 3.8 [+ or -] 0.2
 factor 2
 Actin-related protein 2.1 [+ or -] 0.1 2.1 [+ or -] 0.2
 2/3 complex 1B
 Protective protein for 2.0 [+ or -] 0.1 2.0 [+ or -] 0.1
 [beta]-galactosidase
 Elastase 1 2.5 [+ or -] 0.2 2.6 [+ or -] 0.1
 Connexin 26 9.8 [+ or -] 0.5 10.4 [+ or -] 0.8
 Ceruloplasmin 15.0 [+ or -] 5.5 14.5 [+ or -] 2.1
 Cathepsin H 3.0 [+ or -] 0.3 3.0 [+ or -] 0.3
 Basigin 1.5 [+ or -] 0.1 1.7 [+ or -] 0.1
 Peptidylprolyl isomerase 2.2 [+ or -] 0.1 2.4 [+ or -] 0.3
 C-associated
 Glutathione reductase 1 2.3 [+ or -] 0.1 2.6 [+ or -] 0.3
 START domain- 1.4 [+ or -] 0.03 1.5 [+ or -] 0.01
 containing 3
 CD68 antigen 4.2 [+ or -] 0.6 5.0 [+ or -] 0.7
 RIKEN cDNA E030027H19 2.7 [+ or -] 0.2 2.9 [+ or -] 0.1
 cDNA sequence B0004044 3.4 [+ or -] 0.1 3.8 [+ or -] 0.5
 E74-like factor 3 5.9 [+ or -] 0.5 6.5 [+ or -] 0.5
 Glutathione S- 4.6 [+ or -] 0.6 4.1 [+ or -] 0.7
 transferase omega 1
 Interferon-stimulated 4.3 [+ or -] 0.7 3.4 [+ or -] 0.4
 protein 20
 Clusterin 3.9 [+ or -] 0.9 3.4 [+ or -] 0.4
 Galectin 3 8.7 [+ or -] 0.7 6.9 [+ or -] 0.4
 Small proline-rich 78.3 [+ or -] 15.7 44.2 [+ or -] 5.2
 protein 2A (b)
 Complement component 18.8 [+ or -] 1.0 14.8 [+ or -] 0.8
 3 (b)
 Small proline-rich 340.5 [+ or -] 37.1 214.1 [+ or -] 41.1
 protein 2C
 Small proline-rich 11.0 [+ or -] 0.3 9.6 [+ or -] 0.6
 protein 2G
 Prominin 3.6 [+ or -] 0.6 3.4 [+ or -] 0.3
 Lactotransferrin (b) 99.2 [+ or -] 13.9 76.9 [+ or -] 21.8
 Carbonic anhydrase 2 8.2 [+ or -] 0.9 7.3 [+ or -] 0.4
 Complement component 38.4 [+ or -] 5.6 32.9 [+ or -] 4.2
 factor 1
 Mannosidase 2alphaB1 2.0 [+ or -] 0.1 1.9 [+ or -] 0.1
 Small proline-rich 39.2 [+ or -] 1.9 30.7 [+ or -] 5.0
 protein 2B
 Small proline-rich 329.1 [+ or -] 42.9 59.1 [+ or -] 40.8
 protein 2A (b)
 RIKEN cDNA 5830413E08 3.3 [+ or -] 0.5 3.0 [+ or -] 0.3
 RIKEN cDNA 1110029F20 4.1 [+ or -] 0.4 3.7 [+ or -] 0.1
 Annexin A3 4.2 [+ or -] 0.7 3.2 [+ or -] 0.6
 Peptidase 4 2.9 [+ or -] 0.3 2.3 [+ or -] 0.2
 Laminin gamma 2 17.3 [+ or -] 6.8 10.2 [+ or -] 1.0
 Ubiquitin-like 3 1.8 [+ or -] 0.1 1.7 [+ or -] 0.03
 Urate oxidase 143.9 [+ or -] 62.9 43.8 [+ or -] 17.7
 Amiloride binding 10.1 [+ or -] 0.8 6.0 [+ or -] 1.2
 protein 1
 Keratin complex 1-19 7.2 [+ or -] 1.0 5.6 [+ or -] 0.4
 Activated leukocyte cell 5.1 [+ or -] 0.9 4.4 [+ or -] 0.5
 adhesion molecule
 CCAAT/enhancer binding 2.8 [+ or -] 0.3 2.6 [+ or -] 0.1
 protein [beta]
 Peptidyl arginine 15.8 [+ or -] 3.1 12.0 [+ or -] 1.7
 deiminase, type I
 Enolase 1 [alpha] 3.2 [+ or -] 0.5 3.0 [+ or -] 0.3
 p53 apoptosis effector 4.1 [+ or -] 0.8 3.7 [+ or -] 0.5
 related to Pmp22
 [beta]-Glucuronidase 2.3 [+ or -] 0.2 2.2 [+ or -] 0.1
 Leucine-rich 17.6 [+ or -] 4.1 14.5 [+ or -] 2.6
 [alpha]-2-glycoprotein
 Quiescin 06 5.5 [+ or -] 1.2 4.8 [+ or -] 0.7
 GADD45a 2.6 [+ or -] 0.3 2.3 [+ or -] 0.1
 Alkaline phosphatase 2 22.6 [+ or -] 6.2 15.8 [+ or -] 2.0
 Immediate early 10.8 [+ or -] 2.2 8.9 [+ or -] 2.0
 response 3
 Progressive ankylosis 2.9 [+ or -] 0.4 2.8 [+ or -] 0.4
 RAS p21 protein 14.2 [+ or -] 2.5 12.1 [+ or -] 1.7
 activator 4
 Tumor-associated calcium 2.7 [+ or -] 0.3 2.6 [+ or -] 0.2
 signal transducer 1
 Hydroxysteroid (17-beta) 2.3 [+ or -] 0.3 2.4 [+ or -] 0.1
 dehydrogenase 11
 Platelet-activating 2.2 [+ or -] 0.2 2.3 [+ or -] 0.1
 factor acetylhydrolase
 1ba1
 Branched chain 3.4 [+ or -] 0.2 3.4 [+ or -] 0.01
 aminotransferase 1
 RIKEN cDNA 2400004E04 2.4 [+ or -] 0.2 2.3 [+ or -] 0.2
 Myeloblastosis oncogene 5.6 [+ or -] 1.1 5.0 [+ or -] 0.2
 [K.sup.+] conductance 4.2 [+ or -] 0.6 3.5 [+ or -] 0.8
 calcium-activated
 channel N4
 ATPase 6v1b2 1.8 [+ or -] 0.1 1.8 [+ or -] 0.1
 Cystic fibrosis 4.5 [+ or -] 0.8 3.9 [+ or -] 0.3
 transmembrane
 regulator
 RIKEN cDNA 1110008P14 6.0 [+ or -] 1.0 5.1 [+ or -] 0.2
 Fused toes 3.2 [+ or -] 0.3 2.8 [+ or -] 0.1
 Solute carrier family 4.5 [+ or -] 0.7 3.8 [+ or -] 0.3
 39a8
 Cytochrome b-561 2.5 [+ or -] 0.2 2.3 [+ or -] 0.2
 Secreted 47.4 [+ or -] 7.4 31.1 [+ or -] 2.7
 phosphoprotein 1
 Ion transport regulator 6.8 [+ or -] 1.1 5.5 [+ or -] 0.6
 Fxyd3
 Janus kinase 3 2.5 [+ or -] 0.2 2.2 [+ or -] 0.2
 Cytochrome b-245alpha 3.6 [+ or -] 0.4 2.9 [+ or -] 0.2
 RIKEN cDNA A430096B05 8.6 [+ or -] 0.03 6.3 [+ or -] 0.6
 Small proline-rich 13.8 [+ or -] 3.2 8.5 [+ or -] 0.7
 protein 2J
 Cathepsin B 2.6 [+ or -] 0.2 2.2 [+ or -] 0.1
 RIKEN cDNA 1600025H15 2.7 [+ or -] 0.3 2.2 [+ or -] 0.2
 c-fos oncogene (b) 4.7 [+ or -] 0.9 3.7 [+ or -] 0.8
 Guanine nucleotide 1.8 [+ or -] 0.1 1.6 [+ or -] 0.03
 binding protein
 [gamma]5
 Serine palmitoyl- 2.0 [+ or -] 0.2 1.7 [+ or -] 0.1
 transferase Ic2
 Cystatin B 1.7 [+ or -] 0.1 1.5 [+ or -] 0.02
 Villin 2 2.4 [+ or -] 0.3 1.9 [+ or -] 01
 RIKEN cDNA 0610010012 2.5 [+ or -] 0.3 1.9 [+ or -] 0.03
 Matrix metallo- 208.6 [+ or -] 83.3 48.2 [+ or -] 11.9
 proteinase 7
 RIKEN cDNA 4930422J18 2.9 [+ or -] 0.3 2.0 [+ or -] 0.2
 RIKEN cDNA 1700017B05 2.0 [+ or -] 0.1 1.6 [+ or -] 0.1
 Galactosidase beta 1 2.0 [+ or -] 0.1 1.7 [+ or -] 0.1
 Cathepsin C 3.3 [+ or -] 0.2 2.3 [+ or -] 0.3
 Interferon-stimulated 2.0 [+ or -] 0.2 3.8 [+ or -] 0.7
 protein 15
 MAP kinase--interacting 1.4 [+ or -] 0.1 1.9 [+ or -] 0.1
 kinase 2
 Glutathione S- 2.9 [+ or -] 0.4 3.6 [+ or -] 0.2
 transferase theta 2
 Homeobox B6 1.5 [+ or -] 0.1 1.6 [+ or -] 0.03
 Procollagen Vlalpha 3 1.9 [+ or -] 0.2 2.1 [+ or -] 0.02
 Interferon regulatory 8.1 [+ or -] 0.7 12.6 [+ or -] 1.5
 factor 7
 Scavenger receptor class 2.4 [+ or -] 0.3 2.6 [+ or -] 0.2
 B2
 Polyimmunoglobulin 6.2 [+ or -] 0.8 7.9 [+ or -] 1.0
 receptor
 Proteasome subunit 1.9 [+ or -] 0.04 2.1 [+ or -] 0.1
 [beta]10
 RIKEN cDNA 0610010E05 2.5 [+ or -] 0.2 2.7 [+ or -] 0.4
 RIKEN cDNA 0610010E05 3.0 [+ or -] 0.1 3.4 [+ or -] 0.3
 Xanthine dehydrogenase 8.9 [+ or -] 1.2 10.1 [+ or -] 1.5
 Prominin 2.9 [+ or -] 0.2 3.1 [+ or -] 0.3
 Interferon-induced 9.9 [+ or -] 1.5 14.0 [+ or -] 2.1
 protein IFIT1
 Interferon-induced 4.7 [+ or -] 0.2 6.7 [+ or -] 0.6
 protein IFIT3
 Proteasome subunit 1.8 [+ or -] 0.2 2.1 [+ or -] 0.1
 [beta]8
 RIKEN cDNA 1600023A02 1.7 [+ or -] 0.1 2.0 [+ or -] 0.04
 Small proline-rich 8.6 [+ or -] 0.8 14.7 [+ or -] 0.8
 protein 1A
 MAP kinase-interacting 1.7 [+ or -] 0.1 2.0 [+ or -] 0.2
 kinase 2
 Lymphocyte antigen 6 1.7 [+ or -] 0.1 2.0 [+ or -] 0.1
 complex, locus E
 Guanylate nucleotide 2.0 [+ or -] 0.2 2.6 [+ or -] 0.1
 binding protein 2
 Peptidyl arginine 9.1 [+ or -] 0.3 5.2 [+ or -] 1.5
 deiminase, type II (b)
Down-regulated genes
 Solute carrier family 2.9 [+ or -] 0.3 2.5 [+ or -] 0.1
 29a1
 Lymphocyte specific 1 2.3 [+ or -] 0.2 1.8 [+ or -] 0.1
 Claudin 5 2.8 [+ or -] 0.1 2.7 [+ or -] 0.5
 Potassium channel td12 2.0 [+ or -] 0.04 2.1 [+ or -] 0.1
 Zinc finger homeobox 1a 1.7 [+ or -] 0.1 1.8 [+ or -] 0.01
 Monoamine oxidase A 2.7 [+ or -] 0.2 2.5 [+ or -] 0.4
 Histidine decarboxylase 7.1 [+ or -] 0.6 5.4 [+ or -] 0.8
 [alpha]-2 Adrenergic 4.2 [+ or -] 1.1 3.6 [+ or -] 0.3
 receptor
 Transcription factor 21 2.2 [+ or -] 0.2 2.0 [+ or -] 0.1
 Homeobox D8 2.8 [+ or -] 0.03 2.5 [+ or -] 0.2
 Carboxypeptidase X2 5.2 [+ or -] 1.0 4.2 [+ or -] 0.2
 RIKEN cDNA A230106A15 4.7 [+ or -] 0.5 4.2 [+ or -] 0.7
 Reduced expression 3 3.5 [+ or -] 0.3 3.2 [+ or -] 0.4
 TGF-[beta] binding 2.1 [+ or -] 0.1 1.8 [+ or -] 0.2
 protein 4
 Keratoepithelin (b) 12.6 [+ or -] 0.9 9.7 [+ or -] 3.6
 GLI-Kruppel family 12.2 [+ or -] 2.6 8.2 [+ or -] 2.6
 member GLI

Abbreviations: CEA, carcinoembrionary antigen; SRC, steroid receptor
coactivator; TGF, transforming growth factor.

(a) Gene names (derived from the NetAffx database; Liu et al. 2003),
GenBank accession numbers (GenBank 2004), and mean ([+ or -] SDI fold
induction/repression of gene expression are shown in the same order
as the gene cluster in Figure 3C. (b) Genes mentioned in the text.


REFERENCES

Aardema MJ, Albertini S, Arni P, Henderson LM, Kirsch-Volders M, Mackay JM, et al. 1998. Aneuploidy: a report of an ECETOC task force. Mutat Res 410:3-79.

Akiyama T, Ishida J, Nakagawa S, Ogawara H, Watanabe S, Itoh N, et al. 1997. Genistein, a specific inhibitor of tyrosine-specific protein kinases. J Biol Chem 282:5592-5595.

Almstrup K, Fernandez MF, Petersen JH, Olea N, Skakkebaek NE, Leffers H. 2002. Dual effects of phytoestrogens result in U-shaped dose--response curves. Environ Health Perspect 110:743-748.

Arora A, Nair MG, Strasburg GM. 1998. Antioxidant activities of isoflavones and their biological metaboiites in a liposomal system. Arch Biochem Biophys 356:133-141.

Casanova M, You L, Gaido KW, Archibeque-Engie S, Janszen DB, Heck HA. 1999. Developmental effects of dietary phytoestrogens in Sprague-Dawley rats and interactions of genistein and daidzein with rat estrogen receptors [alpha] and [beta] in vitro. Toxicol Sci 51:236-244.

Cassidy A, Bingham S, Setchell KDR. 1994. Biological effects of a diet of soy protein rich in isoflavones on the menstrual cycle of premenopausal women. Am J Clin Nutr 60:333-340.

Cassidy A, Faughnan M. 2000. Phyto-oestrogens through the life cycle. Proc Nutr Soc 59:489-496.

Committee on Toxicity of Chemicals in Food, Consumer Products and the Environment. 2003. Phytoestrogens and Health. London:Food Standards Agency. Available: http://www. foodstandards.gov.uk/multimedia/pdfs/phytorepor0503 [accessed 2 June 2004].

Dean NM, Kanemitsu M, Boynton AL. 1989. Effects of the tyrosine-kinase inhibitor genistein on DNA synthesis and phospholipid-derived second messenger generation in mouse 10T1/2 fibroblasts and rat liver T51B cells. Biochem Biophys Res Commun 165:795-801.

Duarte J, Ocete MA, Perez-Vizcaino F, Zarzuelo A, Tamargo J. 1997. Effect of tyrosine kinase and tyrosine phosphatase inhibitors on aortic contraction and induction of nitric oxide synthase. Eur J Pharmacol 338:25-33.

Frasor J, Barnett DH, Danes JM, Hess R, Parlow AF, Katzenellenbogen BS. 2003. Response-specific and ligand dose-dependent modulation of estrogen receptor (ER) [alpha] activity by ER[beta] in the uterus. Endocrinology 144:3159-3166.

Genbank. 2004. Bethesda, MD:National Center for Biotechnology Information, National Library of Medicine. Available: http:// www.ncbi.nlm.nih.gov/Genbank/GenbankOverview.html [accessed 2 June 2004].

GEO. 2004. Gone Expression Omnibus Homepage. Bethesda, MD:National Center for Biotechnology Information, National Library of Medicine. Available: http://www.ncbi.nlm.nih. gov/geo/[accessed 2 June 2004].

Hall JM, Couse JF, Korach KS. 2001. The multifaceted mechanisms of estradiol and estrogen receptor signaling. J Biol Chem 276:36869-36872.

Herman-Giddens ME, Slora EJ, Wasserman RC, Bourdony CJ, Bhapkar MV, Koch GG, et al. 1997. Secondary sexual characteristics and menses in young girls seen in office practice: a study from the Pediatric Research in Office Settings network. Pediatrics 99:505-512.

Hewitt SC, Deroo B J, Hansen K, Collins J, Grissom S, Afshari CA, et al. 2003. Estrogen receptor-dependent genomic responses in the uterus mirror the biphasic physiological response to estrogen. Mol Endocrinol 17:2070-2083.

Holm S. 1979. A simple sequentially rejective Bonferroni test procedure. Scand J Stat 6:65-70.

Howdeshell KL, Hotchkiss AK, Thayer KA, Vandenbergh JG, vom Saal FS. 1999. Exposure to bisphenol A advances puberty. Nature 401:763-764.

Kuiper GG, Lemmen JG, Carlsson B, Corton JC, Safe SH, van der Saag PT, et al. 1998. Interaction of estrogenic chemicals and phytoestrogens with estrogen receptor [beta]. Endocrinology 139:4252-4263.

Liu G, Loraine AE, Shigeta R, Cline M, Cheng J, Valmeekam V, et al. 2003. NetAffx: Affymetrix probesets and annotations. Nucleic Acids Res 31:82-86.

Lund TD, Munson DJ, Haldy ME, Setchell KD, Lephart ED, Handa RJ. 2004. Equol is a novel anti-androgen that inhibits prostate growth and hormone feedback. Biol Reprod 70:1188-1195.

Messina MJ. 1999. Legumes and soybeans: overview of their nutritional profiles and health effects. Am J Clin Nutr 70:439S-450S.

Moggs JG, Orphanides G. 2001 Estrogen receptors: orchestrators of pleiotropic cellular responses. EMBO Rep 9:775-781.

Naciff JM, Jump ML, Torontali SM, Carr GJ, Tiesman JP, Overmann GJ, et al. 2002. Gene expression profile induced by 17[alpha]-ethynyl estradiol, bisphenol A, and genistein in the developing female reproductive system of the rat. Toxicol Sci 68:184-199.

Naciff JM, Overmann GJ, Torontali SM, Carr GJ, Tiesman JP, Richardson BD, et al. 2003. Gene expression profile induced by 17[alpha]-ethynyl estradiol in the prepubertal female reproductive system of the rat. Toxicol Sci 72:314-330.

Newbold RR, Banks EP, Bullock B, Jefferson WN. 2001. Uterine adenocarcinoma in mice treated neonatally with genistein. Cancer Res 61:4325-4328.

North K, Golding J. 2000. A maternal vegetarian diet in pregnancy is associated with hypospadias. Br J Urol Int 85:107-113.

Odum J, Lefevre PA, Tittensor S, Paton D, Routledge ED, Beresford NA, et al. The rodent uterotrophic assay: critical protocol features, studies with nonyl phenols, and comparison with a yeast estrogenicity assay. Regul Toxicol Pharmacol 25:176-188.

Okura A, Arakawe H, Oka H, Yoshinari T, Monden Y. 1988. Effect of genistein on topoisomerase activity and on the growth of [Val 12]Ha-ras-transformed NIH 3T3 cells. Biochem Biophys Res Commun 157:183-189.

Orphanides G, Moggs JG, Spurway T, Tinwell H, Ashby J, Kimber I. 2003. Use of gene expression profiling to understand the transcriptional programme associated with estrogen-induced uterine growth: implications for the use of surrogate molecular markers in toxicology [Abstract]. Toxicol Sci 72:12.

Owens JW, Ashby J. 2002. Critical review and evaluation of the uterotrophic bioassay for the identification of possible estrogen agonists and antagonists: in support of the validation of the OECD uterotrophic protocols for the laboratory rodent. Organisation for Economic Co-operation and Development. Crit Rev Toxicol 32:445-520.

Paulozzi LJ, Erickson JD, Jackson RJ. 1997. Hypospadias: trends in two US surveillance systems. Pediatrics 100:831-834.

Potier B, Rovira C. 1999. Protein tyrosine kinase inhibitors reduce high-voltage activating calcium currents in CA1 pyramidal neurones from rat hippocampal slices. Brain Res 816:587-597.

Safe SH, Pallaroni L, Yoon K, Gaido K, Ross S, McDonnell D. 2002. Problems for risk assessment of endocrine-active estrogenic compounds. Environ Health Perspect 110:925-929.

Setchell KDR, Zimmer-Nechemias L, Cai J, Heubi JE. 1997. Exposure of infants to phyto-oestrogens from soy-based infant formula. Lancet 350:23-27.

Tinwell H, Ashby J. 2004. Sensitivity of the immature rat uterotrophic assay to mixtures of estrogens. Environ Health Perspect 112:575-582.

Watanabe H, Suzuki A, Kobayashi M, Lubahn DB, Handa H, Iguchi T. 2003. Similarities and differences in uterine gene expression patterns caused by treatment with physiological and non-physiological estrogens. J Mol Endocrinol 3:487-497.

Weihua Z, Saji S, Makinen S, Cheng G, Jensen EV, Warner M, et al. 2000. Estrogen receptor (ER) [beta], a modulator of ER[alpha] in the uterus. Proc Natl Acad Sci USA 97:5936-5941.

Williams GM, latropoulos M, Cheung R, Radi L, Wang CX. 1993. Diethylstilbestrol liver carcinogenicity and modification of DNA in rats. Cancer Lett 68:193-198.

Jonathan G. Moggs, John Ashby, Helen Tinwell, Fei Ling Lim, David J. Moore, Ian Kimber, and George Orphanides

Syngenta CTL, Alderley Park, Cheshire, United Kingdom

Address correspondence to J.G. Moggs, Syngenta CTL, Alderley Park, Cheshire, SK10 4TJ UK. Telephone: 44-1625-519315. Fax: 44-1625-585715. E-mail: jonathan.moggs@syngenta.com

We thank A. Soames for the histopathology data, I. Kupershmidt and E. Hunter (Silicon Genetics) for advice on statistical analysis of microarray data, and T. Barlow (Food Standards Agency) for critical comments.

This work was partially supported by the U.K. Food Standards Agency.

The authors declare they have no competing financial interests.

Received 12 February 2004; accepted 19 May 2004.
COPYRIGHT 2004 National Institute of Environmental Health Sciences
No portion of this article can be reproduced without the express written permission from the copyright holder.
Copyright 2004, Gale Group. All rights reserved. Gale Group is a Thomson Corporation Company.

Article Details
Printer friendly Cite/link Email Feedback
Title Annotation:Research / Commentary
Author:Orphanides, George
Publication:Environmental Health Perspectives
Date:Aug 1, 2004
Words:7640
Previous Article:Spraying on a summer night: a safer way to stop West Nile virus.
Next Article:Using human disease outbreaks as a guide to multilevel ecosystem interventions.


Related Articles
Estrogens exonerated in breast cancer.
A new breadth to estrogen's bisexuality.
Weighing hormone therapy's benefits.
Estrogen flips testosterone gene switch.
Progestin adds to breast cancer risk.
Soy and Breast Cancer.
Detection of xenoestrogens in serum after immunoprecipitation of endogenous steroidal estrogens. (Articles).
Uranium, the newest 'hormone'.
Breast cancer & weight.
The feed factor: estrogenic variability in lab animal diets.

Terms of use | Privacy policy | Copyright © 2021 Farlex, Inc. | Feedback | For webmasters