John G. Vandenbergh

NTP-CERHR expert panel report on the reproductive and developmental toxicity of bisphenol A.

Robert E. Chapin, Jane Adams, Kim Boekelheide, L. Earl Gray Jr, Simon W. Hayward, Peter S.J. Lees, Barry S. McIntyre, Kenneth M. Portier, Teresa M. Schnorr, Sherry G. Selevan, John G. Vandenbergh, and Susan R. Woskie Pfizer, Inc., Groton, CT University of Massachusetts, Boston, MA Brown University, Providence, RI U.S. Environmental Protection Agency, Research Triangle Park, NC Vanderbilt University Medical Center, Nashville, TN Johns Hopkins University, Baltimore, MD Schering Plough Research Institute, Summit, NJ American Cancer Society, Atlanta, GA National Institute for Occupational Safety and Health, Cincinnati, OH U.S. Public Health Service (Ret), Silver Spring, MD North Carolina State University, Raleigh, NC University of Massachusetts, Lowell, MA

Intrauterine position effects.

A review of the literature suggests that individual variability in sex-related traits may be influenced by variations in hormonal exposure during fetal development. In litter-bearing mammals, fetuses develop in utero and may be subjected to differing hormonal environments based upon the sex of neighboring fetuses. Female fetuses developing between two males tend to show masculinized anatomical, physiological and behavioral traits as adults. Female fetuses developing without adjacent males, on the other hand, tend to show more feminized traits as adults. These traits include permanently altered hormone levels, reproductive organs, aggressive behaviors, secondary sex ratios and susceptibility to endocrine disruption. This intrauterine effect is due to the transfer of testosterone from male fetuses to adjacent fetuses. While these effects have been most clearly demonstrated in mice, other rodents and swine also show intrauterine position (IUP) effects. Some of these effects are similar to the influence of prenatal stress on adult phenotypes. A few reports on human twins suggest that variability in some masculine and feminine traits may be due to intrauterine hormonal signals. IUP effects may impact a number of scientific fields of research such as endocrine disruption, toxicology, population biology, animal production and health.

Why public health agencies cannot depend on good laboratory practices as a criterion for selecting data: The case of Bisphenol A

Background In their safety evaluations of bisphenol A (BPA), the U.S. Food and Drug Administration (FDA) and a counterpart in Europe, the European Food Safety Authority (EFSA), have given special prominence to two industry-funded studies that adhered to standards defined by Good Laboratory Practices (GLP). These same agencies have given much less weight in risk assessments to a large number of independently replicated non-GLP studies conducted with government funding by the leading experts in various fields of science from around the world. Objectives We reviewed differences between industry-funded GLP studies of BPA conducted by commercial laboratories for regulatory purposes and non-GLP studies conducted in academic and government laboratories to identify hazards and molecular mechanisms mediating adverse effects. We examined the methods and results in the GLP studies that were pivotal in the draft decision of the U.S. FDA declaring BPA safe in relation to findings from studies that were competitive for U.S. National Institutes of Health (NIH) funding, peer-reviewed for publication in leading journals, subject to independent replication, but rejected by the U.S. FDA for regulatory purposes. Discussion Although the U.S. FDA and EFSA have deemed two industry-funded GLP studies of BPA to be superior to hundreds of studies funded by the U.S. NIH and NIH counterparts in other countries, the GLP studies on which the agencies based their decisions have serious conceptual and methodologic flaws. In addition, the U.S. FDA and EFSA have mistakenly assumed that GLP yields valid and reliable scientific findings (i.e., “good science”). Their rationale for favoring GLP studies over hundreds of publically funded studies ignores the central factor in determining the reliability and validity of scientific findings, namely, independent replication, and use of the most appropriate and sensitive state-of-the-art assays, neither of which is an expectation of industry-funded GLP research. Conclusions Public health decisions should be based on studies using appropriate protocols with appropriate controls and the most sensitive assays, not GLP. Relevant NIH-funded research using state-of-the-art techniques should play a prominent role in safety evaluations of chemicals.

Developmental exposure to environmental estrogens alters anxiety and spatial memory in female mice

Humans and wildlife are exposed to numerous anthropogenic drugs and pollutants. Many of these compounds are hormonally active, and recent evidence suggests that the presence of these endocrine disruptors permanently alters normal development and physiology in a variety of vertebrate species. Here, we report on the effects of developmental exposure to two common estrogenic pollutants, bisphenol A and ethinyl estradiol on sexually dimorphic, non-reproductive behavior. Mice (Mus musculus domesticus) were exposed to environmentally relevant levels of these chemicals (2 and 200 microg/kg/day for bisphenol A and 5 microg/kg/day for ethinyl estradiol) throughout prenatal and early postnatal development. As adults, the animals were observed in a variety of tests measuring sexually dimorphic behaviors including short-term spatial memory (in a radial-arm maze and a Barnes maze) and anxiety (in an elevated-plus maze and a light/dark preference chamber). Developmental exposure to ethinyl estradiol was found to masculinize behavior in all of the assays used. Bisphenol A increased anxious behavior in a dose-dependent fashion but had no effect on spatial memory. These results indicate that non-reproductive, sexually dimorphic behavior is sensitive to endocrine disruption. In addition, these experiments suggest that both humans and wildlife are being exposed to levels of these endocrine disrupting compounds that are sufficient to disrupt the development of the nervous system and that may have permanent consequences on sexually dimorphic behaviors.

A mixture of the "antiandrogens" linuron and butyl benzyl phthalate alters sexual differentiation of the male rat in a cumulative fashion.

Abstract Prenatal exposure to environmental chemicals that interfere with the androgen signaling pathway can cause permanent adverse effects on reproductive development in male rats. The objectives of this study were to 1) determine whether a documented antiandrogen butyl benzyl phthalate (BBP) and/or linuron (an androgen receptor antagonist) would decrease fetal testosterone (T) production, 2) describe reproductive developmental effects of linuron and BBP in the male, 3) examine the potential cumulative effects of linuron and BBP, and 4) investigate whether treatment-induced changes to neonatal anogenital distance (AGD) and juvenile areola number were predictive of adult reproductive alterations. Pregnant rats were treated with either corn oil, 75 mg/kg/day of linuron, 500 mg/kg/day of BBP, or a combination of 75 mg/kg/day linuron and 500 mg/kg/day BBP from gestational Day 14 to 18. A cohort of fetuses was removed to assess male testicular T and progesterone production, testicular T concentrations, and whole-body T concentrations. Male offspring from the remaining litters were assessed for AGD and number of areolae and then examined for alterations as young adults. Prenatal exposure to either linuron or BBP or BBP + linuron decreased T production and caused alterations to androgen-organized tissues in a dose-additive manner. Furthermore, treatment-related changes to neonatal AGD and infant areolae significantly correlated with adult AGD, nipple retention, reproductive malformations, and reproductive organ and tissue weights. In general, consideration of the dose-response curves for the antiandrogenic effects suggests that these responses were dose additive rather than synergistic responses. Taken together, these data provide additional evidence of cumulative effects of antiandrogen mixtures on male reproductive development.

The effects of gonadal hormones on the aggressive behaviour of adult golden hamsters (Mesocricetus auratus)

Abstract Aggressive behaviour in unisexual pair encounters between adult male hamsters decreases after castration and is restored by injections of either testosterone propionate or oestradiol benzoate. In similar encounters between females, gonadectomy fails to reduce aggression and injections of testosterone or oestrogen do not appear to change the levels of aggression. Aggression in females virtually disappears during oestrus indicating that progesterone, in the presence of oestrogen, reduces female aggression. The high level of aggressive behaviour of female hamsters apparently is not induced by gonadal hormones but can only be reduced by the endocrine state during oestrus. Flank marking behaviour in both sexes is under gonadal control, and, in females, lordosis is dependent on the presence of ovaries.

Effects of central and peripheral anosmia on reproduction of female mice

Abstract Anosmia, induced either by removal of the olfactory bulbs or by a ZnSO 4 flush of the nasal epithelium, resulted in lengthened and irregular estrous cycles in mice as determined by vaginal lavage. When mated with males, anosmic females conceived but at a rate lower than sham controls. Both central and peripheral anosmia disrupted nest building and maternal care but the severity of the effect was greater among centrally anosmic females. These results indicate that the sense of smell is essential for normal ovarian rhythmicity in the mouse and that bulbectomy produced more severe disruption of nest building and maternal care than anosmia alone.

Predictors of dominance in the male golden hamster (Mesocricetus auratus)

The outcome of social interactions between four male hamsters was significantly related to body weight (rs=0·66) and to the size and pigmentation of their lateral flank glands (rs=0·82). Weight was held constant in a second experiment and variations in gland size and pigmentation remained significantly related to the outcome of social encounters (rs=0·77). In a third experiment, castrate hamsters of uniform weight receiving either no hormone, 0·1 mg, 0·50 mg, or 1·00 mg testosterone propionate exhibited flank gland variations significantly related to social rank (rs=0·81). These experiments demonstrate that the state of the flank gland, which is related to endogenous androgen levels, can be used as a predictor of social rank in male hamsters.

Endocrine coordination in monkeys: Male sexual responses to the female

Abstract Free-ranging rhesus monkeys on island colonies near Puerto Rico breed with a distinct seasonal rhythm. Observations on adult monkeys in one such colony, La Parguera, during the non-breeding season showed that sexually quiescent male monkeys can be returned to a sexually active state by exposure to females artificially brought into estrus. Males responded both behaviorally by showing copulatory activities and by increased duration of grooming, and physiologically by showing evidence of increased testicular activity. Such behavioral and endocrine coordination points out: (1) the control hormones exert over primate sexual behavior, (2) that females communicate their endocrine state to males, and (3) the existence of a system for the synchronization of mating activities between the sexes, especially at the onset of the breeding season.

Reproductive coordination among free-ranging Rhesus monkeys ☆

Two female rhesus monkeys (Macaca mulatta) in one free-ranging social group were brought into sexual receptivity with estradiol benzoate during the nonmating season of the year. The presence of sexually active females resulted in increased sexual behavior and reddening of sex skin among the adult males in the experimental group. The effects on males were most intense on those of high social rank. These changes occurred while two other social groups were sexually quiescent and two months before the normal onset of mating activity in these groups. In the birth season following this wave of induced sexual activity, untreated females in the experimental group delivered infants one month earlier than in control groups.