Abstract.To evaluate the effects of bisphenol A (BPA), a candidate endocrine disruptor (ED), on embryonic development, we examined the mRNA expression levels of the aryl hydrocarbon receptor (AhR; which binds with many EDs and plays crucial roles in their metabolism) and related factors [aryl hydrocarbon receptor repressor (AhRR) and AhR nuclear translocator (Arnt)], xenobiotic metabolizing enzymes [XMEs; cytochrome P450 1A1 (CYP1A1) and UDP-glucuronosyltransferase, and the glutathione S-transferase Ya subunit (GST)], in murine embryos exposed in utero to BPA (0.02, 2, 200, and 20,000 µg/kg/day) and 17β-estradiol (E2; 5 µg/kg/day, used as a positive control) at 6.5-13.5 or 6.5-17.5 days post coitum (dpc) using the quantitative real-time reverse transcriptionpolymerase chain reaction (RT-PCR) method. Protein levels of CYP1A1 and GST in embryonic livers were estimated by Western immunoblotting. Exposure in utero to BPA [0.02 (1/100 dose of environmental exposure), 2, 200, and 20,000 µg/kg/day] increased AhR mRNA expression in the cerebra, cerebella, and gonads (testes and ovaries) of male and female mid-and late-developmental stage (14.5-and 18.5-dpc, respectively) embryos. BPA dose-independently up-regulated the expression of AhRR and Arnt in mid-and late-stage embryos. BPA had no remarkable effect on the mRNA levels of XMEs in mid-stage embryos, but dose-dependently up-regulated the expression in late-stage embryos. Moreover, the protein levels of these enzymes in the livers of late-stage embryos were increased. The present findings revealed that exposure to BPA in utero disrupts the expression of AhR and related factors and of xenobiotic metabolizing enzymes, and that mid-stage embryos, in the organogenic stage, are sensitive to BPA. Key words: Aryl hydrocarbon receptor (AhR), AhR relating factor, Bisphenol A (BPA), Murine embryo, Xenobiotic metabolizing enzymes (XMEs) (J. Reprod. Dev. 51: [593][594][595][596][597][598][599][600][601][602][603][604][605] 2005) isphenol A [BPA; 2,2-bis (4-hydroxyphenyl) propane], which is composed of two benzene rings and in conformation resembles a synthetic estrogen, diethylstilbestrol (DES), is a common plasticizer for polycarbonate and epoxy resins and a candidate environmental endocrine disruptor (ED) [1,2]. BPA has toxicological effects on the reproductive, immunological, and nervous systems in mammals, binds with estrogen receptor-α (ERα; NR3A1) and ER-β (NR3A2), and induces estrogenic activity [3][4][5][6]. BPA affects the expression levels of ERs and ER-like nuclear receptor mRNAs in the rat uterus [6], but the mechanism of its actions has yet

Abstract.To evaluate the effects of bisphenol A (BPA), a candidate endocrine disruptor (ED), on embryonic development, we examined the mRNA expression levels of the arylhydrocarbon receptor (AhR), which binds with many EDs and plays crucial roles in xenobiotic metabolism, and of the retinoic acid receptor (RAR) α and retinoid X receptor (RXR) α, key factors in nuclear receptordependent retinoid signal transduction, in murine embryos exposed in utero to BPA (0.02, 2, 200, and 20,000 µg/kg/day) at 6.5-13.5 or 6.5-17.5 days post coitum (dpc), using the real-time reverse transcription-polymerase chain reaction (RT-PCR) method. Extremely low-dose BPA (0.02 µg/kg/ day; 1/100 the dose of environmental exposure) remarkably increased AhR mRNA expression in the cerebra, cerebella, and gonads (testes and ovaries) of male and female 14.5-and 18.5-dpc-embryos. In utero exposure to BPA at 2, 200, and 20,000 µg/kg/day also increased levels of AhR mRNA. In gonads of 14.5-dpc-embryos, AhR mRNA levels were elevated and showed diphasic (U) dose-response curves following exposure to BPA, but inverted U dose-response curves were obtained for 18.5-dpcembryos. Exposure to BPA increased expression levels of RARα and RXRα mRNAs in the cerebra, cerebella, and gonads of male and female 14.5-and 18.5-dpc-embryos. Extremely low-dose BPA (0.02 µg/kg/day) increased RARα mRNA expression in the cerebella of male and female 14.5-and 18.5-dpc-embryos and in the gonads of female 14.5-dpc-embryos, and significantly increased RXRα mRNA expression in the cerebra and cerebella of male and female 14.5-dpc-embryos. The present findings confirm that in utero exposure to an extremely low dose of BPA up-regulates the mRNA expression of AhR, RARα, and RXRα in murine embryos and disrupts the receptor-dependent signal transducing systems, and will contribute to the assessment of the toxic effects of BPA on xenobiotic metabolism and retinoid signals in embryogenesis. Key words: Arylhydrocarbon receptor (AhR), Bisphenol A, Murine embryo, Retinoic acid receptor (RAR) α, Retinoid X receptor (RXR) α (J. Reprod. Dev. 51: [315][316][317][318][319][320][321][322][323][324] 2005) isphenol A [BPA; 2,2-bis (4-hydroxyphenyl) p r o p a n e ] , a c o m m o n p l a s t i c i z e r a n d a candidate environmental endocrine disruptor (ED), has toxicological effects on the reproductive, immunological, and nervous systems of mammals [1][2][3][4]. BPA binds to estrogen receptor-α (ERα; NR3A1) and -β (ERβ; NR3A2), and induces

Bisphenol A (BPA), a candidate endocrine disruptor (ED), is considered to bind to estrogen receptors and to regulate expressions of estrogen responsive genes. It has also shown evidence of affecting the reproductive, immunological and nervous systems of mammalian embryos. However, the effects of BPA on placentae, a central organ of feto-maternal interlocution, are still unclear. To reveal the mechanisms of BPA effects on placentae in mammals, we compared the mRNA expression of 20 nuclear receptors between placentae of vehicle controls and those of orally BPA exposed pregnant mice by a DNA microarray technique. In murine placentae, mRNAs of 11 nuclear receptors were not detected. However, greater than 1.5 fold changes in mRNA expression of nine nuclear receptors between vehicle control and BPA treated mice were noted. Moreover, remarkable changes in mRNA expression of six non-nuclear receptor proteins were induced by BPA exposure. There were various differences in the effects of BPA on the expression of these mRNAs between the placentae with male embryos and those with female embryos. Such embryo-sex dependent differences are interesting and important pointers to understanding of the endocrine disrupting effect of BPA. The present data indicate that BPA affects the expression of nuclear receptor mRNAs in placentae and may disrupt the physiological functions of placentae.

Abstract. Retinoic acid receptor (RAR) α and retinoid X receptor (RXR) α are key factors in a nuclear receptor-dependent signal. To evaluate the effects of bisphenol A (BPA), a candidate endocrine disruptor (ED), on embryonic development, we examined the mRNA levels of RARα and RXRα in murine embryos, exposed in utero to BPA (2 µg/kg/day) at 6.5-17.5 days post-coitum (dpc), by the realtime reverse transcription-polymerase chain reaction (RT-PCR) method. Higher levels of RARα mRNA in cerebra of male and female embryos of control groups were detected at 14.5 dpc. In utero BPA reduced the RARα mRNA expression. Higher levels of RXRα mRNA in cerebra of male and female embryos were seen at 12.5 dpc. The exposure decreased RXRα mRNA expression in male but not female embryos. No remarkable change in the RAR α mRNA expression level was noted in cerebella of male or female embryos of the control group during embryonic development. Exposure to BPA increased expression levels of RARα mRNA in cerebella of male and female embryos at 12.5 dpc. Higher levels of RXRα mRNA in cerebella of male and female embryos were seen, but no remarkable changes were noted during embryonic development. BPA significantly decreased the expression levels of RXRα mRNA in cerebella of female embryos at 12.5, 14.5 and 18.5 dpc. RARα and RXRα mRNAs were expressed in gonads (testes and ovaries) of murine embryos from 12.5 to 18.5 dpc.In utero exposure to BPA decreased levels of RARα mRNA in testes of 14.5-and 18.5-dpc-embryos, levels of RXRα mRNA in testes of 14.5-dpc-embryos, and levels of RXRα mRNA in ovaries of 14.5-dpc-embryos. The present findings indicate that RARα and RXRα play crucial roles in organogenesis, and the growth and development of murine embryos, and will contribute to the assessment of the toxic effects of BPA on retinoid signals in embryogenesis. Key words: Bisphenol A, Embryo, Mouse, Retinoic acid receptor α, Retinoid X receptor α (J. Reprod. Dev. 49: [539][540][541][542][543][544][545] 2003) isphenol A [BPA; 2,2-bis (4-hydroxyphenyl) propane], a plasticizer and a cand idate endocrine disruptor (ED), has physiological effects on the reproductive, immunological and nervous systems in mammals [1][2][3]. BPA binds to estrogen receptor α (ERα; NR3A1) and ERβ (NR3A2) and induces estrogenic activity [1][2][3]. BPA affects the levels of ER and nuclear receptor mRNA in rat uterus [4], but no detailed information on the mechanism of its actions has been reported.Retinoic acid receptor α (RARα; NR1B1) and retinoid X receptor α (RXRα ; NR2B1) specifically bind with 9-cis-retinoic acid (RA), a bioactive metabolite of vitamin A (retinol), and all-trans-RA, r e s p e c t i v e l y . R A h a s m a r k e d e f f e c t s o n embryogenesis, and cell proliferation, growth, differentiation and death [5,6]. In mouse embryos, RA is endogenously synthesized from 7 days postcoitum (dpc) [7], and the expression of RARα and

Historical control data on rabbit prenatal developmental toxicity studies, performed between 1994-2010, were obtained from 20 laboratories, including 11 pharmaceutical and chemical companies and nine contract laboratories, in Japan. In this paper, data were incorporated from a laboratory if the information was based on 10 studies or more. Japanese White rabbits and New Zealand White rabbits were used for prenatal developmental toxicity studies. The data included maternal reproductive findings at terminal cesarean sections and fetal findings including spontaneous incidences of morphological alterations. No noticeable differences between strains or laboratories were observed in the maternal reproductive and fetal developmental data. The inter-laboratory variations in the incidences of fetal external, visceral, and skeletal alterations seem to be due to differences in the selection of observation parameters, observation criteria, and classification of the findings, and terminology of fetal alterations.

The goal of this study was to determine whether in the Japanese quail the male genital tract contains receptors for progesterone, androgen and estrogen (PR, AR and ER, respectively), which have significant roles in reproductive functions, and whether their localization changes during sexual maturation. The epididymis and ductus deferens (middle and ampulla regions) of immature (approximately 30-day-old) and mature male Japanese quail were collected and frozen sections of them were immunostained for PR, AR and ER. The immunoreaction products for AR and PR were found in the nuclei of epithelial cells in the efferent ductules, epididymal duct, and the middle and ampulla regions of the ductus deferens of mature and immature birds. In the mature birds, the epithelial cells of the efferent ductules, epididymal duct, and the middle and ampulla regions of the ductus deferens were positive for ER, although some of the cells in the ductus deferens were negative. The epithelial cells of the ductules in the epididymis stained positive for ER, but the immunoreactions were negligible in the ductus deferens of immature birds. These results suggest that the epididymis and ductus deferens in quail possesses PR, AR and ER receptors. Each receptor is expressed before sexual maturation, although enhancement of ER expression may occur during maturation.

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