This article is the first to describe the induction of numerous abnormalities, including both benign and malignant lesions, in reproductive tissues of aged female mice exposed prenatally to a broad range of BPA doses (0.1–1,000 μg/kg maternal body weight). The profile of reproductive tract lesions observed in this study is similar to what we observed following neonatal treatment with BPA (Newbold et al. 2007
). The BPA doses were low and within the range of human exposure, comparable with 1–13 μg/kg estimated intake levels of formula-fed infants and 0.043–14.7 μg/kg in young children (NTP 2008
); further, these doses have been reported to cause preneoplastic and neoplastic changes in perinatally exposed male (Ho et al. 2006
) and female (Durando et al. 2007
; Murray 2007
; Newbold et al. 2007
) experimental animal models. The present study adds to the growing body of literature that reports adverse effects following developmental exposure to low doses of BPA.
Among the benign abnormalities was an elevated incidence of ovarian cysts (67% in the BPA-1 group). Although ovarian cysts are histologically similar to those seen in our aged controls, the incidence is significantly higher than the controls in this study (25%) and in our historical controls; interestingly, the incidence is similar to that reported following neonatal BPA exposure (70%) (Newbold et al. 2007
). The incidence following either pre natal or neonatal BPA is higher than we have observed in mice developmentally exposed to 0.001 mg/kg DES (58%) (Newbold et al. 1990
), 50 mg/kg genistein (41%) (Newbold et al. 2001
), or tamoxifen (60%) (Newbold et al. 1997
), suggesting that the ovary may be a particularly sensitive target for the effects of BPA. Ongoing studies in our laboratory are investigating this possibility as well as mechanisms involved in the formation of ovarian cysts.
In one BPA-10 mouse, we observed prominent paraovarian cysts of mesonephric (Wolffian) duct origin similar to those reported in neonatally exposed mice (Newbold et al. 2007
). This lesion, combined with the finding of prominent cystic Wolffian duct remnants in the uterine wall of BPA mice, also suggests that the mesonephric duct system (Wolffian duct) may be a target of BPA because both cystic structures have the same fetal tissue origin. Mesonephric-derived tissues have been shown to be sensitive to the effects of perinatal DES exposure in both male (Newbold et al. 1985a
) and female mice (Haney et al. 1986
Another BPA-induced abnormality found in all dose groups is PPL of the oviduct. PPL has been described in mice perinatally treated with DES. DES was shown to interfere with the normal differentiation of the Mullerian duct (the precursor of the oviduct), resulting in structural (prenatal exposure) (Newbold et al. 1983
) and cellular (neonatal exposure) alterations (Newbold et al. 1984, 1985b
). The molecular mechanism likely involves altered HOX
gene expression in the differentiation of the reproductive tract (Taylor et al. 1997
) because prenatal DES delays the expression of these genes (Ma et al. 1998
). Subsequent studies suggest that DES works through multiple gene pathways (Miller et al. 1998
; Pavlova et al. 1994
). Thus, molecular “misprogramming” is mostly likely responsible for DES, as well as BPA-induced, oviductal alterations. Whether these compounds cause these effects through the classical ER-α or ER-β, or the newly identified ncmER (Alonso-Magdalena et al. 2005
) pathways, or some other pathway, remains to be determined.
The benign lesions (CEH and adenomyosis) also occurred in the uterus of mice prenatally treated with BPA, but the incidence was not statistically different from controls. Although these lesions were histologically similar to those in aged controls, the lesions were more severe and their involvement in the uterine horns was more extensive in the BPA-treated groups compared with controls in this study and with our historical controls.
Of particular significance in this study is the occurrence of more severe ovarian lesions (cystadenoma) in the three highest BPA dose groups. In the uterus, adenocarcinoma was not observed in this study, although atypical hyperplasia, its premalignant lesion, was present. We were not suprised by the lack of uterine adenocarcinoma because BPA has weak ER-α binding and because the induction of uterine epithelial tumors is associated with a chemical’s binding affinity for this particular receptor during neonatal life (Newbold and Liehr 2000
; Newbold et al. 2006
). In the uterus, we also found an increased incidence of stromal polyps in the BPA-100 group. These lesions are considered preneoplastic/neoplastic in experimental rodent models because they are often the site for the development of endometrial stromal sarcoma (Davis et al. 1999
; Maronpot 1999
). Historically, we have rarely seen stromal polyps in CD-1 mice, although we did observe this lesion in one control mouse in a previous study (Newbold et al. 2007
). Interestingly, in the present study, we identified a large invasive stromal sarcoma of the cervix after prenatal BPA-100 exposure; we have never seen this malignant lesion in any of our historical controls. Taking the stromal polyps and the stromal sarcoma together, these lesions suggest that stromal tissue is a target for BPA exposure, especially if exposure occurs during critical periods of differentiation of the reproductive tract. A similar finding has been shown in mice exposed to low-dose DES (Newbold et al. 2002
We identified adverse effects in the reproductive tract in all BPA-treated groups, but it is interesting that the lowest dose (BPA-0.1) was the most affected (). Non linear dose–response curves have been commonly reported in endocrinology studies (vom Saal et al. 1997
). One explanation for these effects can be found in DNA microarray studies (Coser et al. 2003
; Shioda et al. 2006
), where increasing doses of estrogens, from low to higher levels, result in entirely different arrays of genes that are turned on or off. Thus, the idea that there should be only a quantitative change in end points as the dose increases is not supported by these studies; instead, entirely different types of effects could occur as the high dose range is reached (for example, the changes we observed in the BPA-1000 group in the present study). This requires further investigation; however, the pattern of nonmonotonic effects is similar to what we observed in our neonatal BPA study (Newbold et al. 2007
In a previous NTP carcinogenesis bioassay, the NTP (1982)
reported that adult exposure to BPA was associated with cancers of the hematopoietic system. In the present study, we found one BPA-exposed mouse (BPA-1; 7 months of age) had a poorly differentiated sarcoma that infiltrated one ovary and the entire reproductive tract; we consider this lesion to be hematopoietic in origin. Although this study was designed to address only long-term, mainly carcinogenic, effects of BPA on reproductive tissues, certainly, the effects on the hematopoietic system warrants further follow-up.
In the present study, body weights were not different between BPA-treated mice and control mice. This lack of difference is most likely due to the advanced age of the mice in the study. Our laboratory and others have previously reported that developmental exposure to BPA, DES, and other environmental chemicals with endocrine-disrupting effects is associated with obesity in mice after they reach puberty and throughout maturity (Grun and Blumberg 2006
; Grun et al. 2006
; Howdeshell et al. 1999
; Miyawaki et al. 2007
; Newbold et al. 2005
; Rubin et al. 2001
); however, the animals in those studies were not examined at 18 months of age. Also, we have shown that significant differences in body weight in DES-treated mice compared with controls at 6–8 months of age become more difficult to detect as the animals age because of increased individual variability among all mice and because of increased disease and tumors (Newbold et al. 2007
); thus, this variability probably accounts for lack of detection of body weight differences in this study. More important, we found no apparent correlation of body weight and tumor occurrence in either individual animals or groups, although a few BPA-exposed mice (but no controls) died before the completion of the study.
In summary, the findings of the present study raise concerns about widespread exposure to BPA and, in particular, exposure to fetuses, infants, and children. In vitro
studies showing that BPA transforms SHE cells (Tsutsui et al. 2000
) and induces aneuploidy (Tsutsui et al. 1998
), and previous in vivo
studies showing that BPA causes mammary tumors (Durando et al. 2007
; Murray 2007
) and preneoplastic prostatic lesions (Ho et al. 2006
), along with evidence of BPA carcinogenicity following adult exposure (Huff 2001
) together indicate that the body of literature merits serious consideration. Although studies are needed to determine the potential adverse effects to humans exposed to BPA during critical stages of neo natal or early development, the potential risks and benefits should be thoroughly assessed to determine the appropriate balance of exposures of this chemical during development and the permanent effects that may follow.