Monograph on the Potential Human Reproductive and ... - OEHHA

Strengths/Weaknesses: This study is a more detailed
follow-up of the previous study by these authors (Furuya
et al., 2002), and replication of these results is a strength.
Additional strengths are the use of multiple exposure
levels and the oral route of administration. The lack of
information on statistical methods is a weakness.
Utility (Adequacy) for CERHR Evaluation Process:
This study is not useful to the evaluation process. While
the in vitro studies are useful for mechanistic insights,
cellular evaluation, and endpoint identification, inter alia,
the studies as a group were considered not useful for the
evaluation process.
3.2.11 In vitro. Takai et al. (2000), supported by the
Japanese Ministry of Education, Science, and Culture, the
Ministry of Health and Welfare, and the Science and
Technology Agency, examined the effects of in vitro
bisphenol A exposure on preimplantation mouse embryos.
Two-cell embryos were obtained from B6C3F 1
mice and incubated for 48 hr in media containing
bisphenol A [purity not indicated] at concentrations
ranging from 100 pM to 100 mM [23 ng/L to 23 mg/L]
[culture ware not discussed]. A negative control group
was exposed to the ethanol vehicle and the effects of
tamoxifen were also tested. Cell numbers were counted,
and trophoblast spreading was evaluated in blastocysts.
Statistical analyses included w 2 , Fisher post-hoc, and
Student t-tests. The number of embryos or samples/
group ranged from 14–400 for each endpoint evaluated.
Significant effects observed with bisphenol A exposure
(percent change vs. control) included increased rate of
development from 2- to 8-cell embryos following 24 hr
exposure to 3 nM [0.68 lg/L] (94% vs. 88%), increased
development to the blastocyst stage following 48 hr
exposure to 1 and 3 nM [0.23 and 0.68 lg/L] (69% in both
dose groups vs. 58.7%), and decreased development to
the blastocyst stage following 48 hr exposure to 100 mM
[23 mg/L] bisphenol A (31.2 vs. 58.7%). No effects were
observed at concentrations between 10 nM and 10 mM
[23 lg/L and 2.3 mg/L] bisphenol A. [Data were not
shown by study authors.] Addition of 100 nM tamoxifen
to cultures decreased development to the blastocyst stage
at 1 and 3 nM [0.23 and 0.68 lg/L] bisphenol A and
increased development to blastocyst stage at 100 mM
[23 mg/L] bisphenol A. Trophoblast spreading was
increased in blastocysts exposed to 100 mM [23 mg/L]
bisphenol A. Bisphenol A exposure did not affect
morphology of or cell numbers in blastocysts. The study
authors concluded that environmentally relevant concentrations
of bisphenol A may affect early embryonic
development through the ER and may also affect
subsequent development.
Strengths/Weaknesses: The wide range of bisphenol A
concentrations is a strength. The postulated involvement
of the ER in bisphenol A activity could have been
more convincingly demonstrated with a positive
control such as 17b-estradiol and with a more
specific estrogen antagonist than tamoxifen. The
use of serum-free and phenol red-free media is an
appropriate way to avoid estrogenic contamination
but is an artificial environment compared to the
estrogen-rich milieu in which preimplantation embryos
normally develop.
Utility (Adequacy) for CERHR Evaluation Process:
This study provides some mechanistic information but is
not useful in the evaluation process.
Birth Defects Research (Part B) 83:157–395, 2008
BISPHENOL A
317
Takai et al. (2001), supported by the Japanese Ministry
of Education, Science, Sports, and Culture, the Ministry
of Health and Welfare, and the National Institute for
Environmental Studies, examined the effects of in vitro
preimplantation exposure of mice to bisphenol A. Twocell
embryos were obtained from B6C3F 1 mice and
incubated for 48 hr in media containing bisphenol A
[purity not indicated] at 0 (ethanol vehicle), 1 nM
[0.23 lg/L] or 100 mM [23 mg/L] [culture ware not
discussed]. Embryos were assessed for number developing
to the blastocyst stage, and then blastocysts were
transferred to uterine horns of pseudopregnant mice (7/
mouse). The dams were allowed to deliver and nurse the
litters until weaning on PND 21 (day of birth not
defined). Pups were randomly culled to maintain litter
sizes at no more than 6. Body weight of pups was
measured at birth and at weaning. Litters and pups were
considered the experimental unit for statistical analyses.
Statistical analyses included w 2 and Fischer protected
least significant difference tests. The number of embryos
developing to the blastocyst stage was significantly
increased by exposure to bisphenol A at 1 nM [0.23 lg/
L] but decreased by exposure to 100 mM [23 mg/L] (72.2
and 33.3% at each respective concentration vs. 62.1% in
controls). Developing embryos appeared morphologically
normal and there were no significant differences in
the numbers of cells. Birth weight, number of pups/litter,
and sex ratio were not affected by treatment. At
weaning, pups in both dose groups weighed more than
controls (34–39% greater) and the effect was significant
on a litter and pup basis. The study authors concluded
that bisphenol A may affect early embryonic and
postnatal development at low, environmentally relevant
concentrations.
Strengths/Weaknesses: This study was cleverly
designed as a follow-up to the previous study and
appears to show that a low concentration of bisphenol A
stimulates early embryo development while a high
concentration inhibits early embryo development.
This study did not evaluate the effect of exogenous
bisphenol A under physiologic conditions. The use of
serum-free and phenol red-free media is an appropriate
way to avoid estrogenic contamination but is an artificial
environment compared to the estrogen-rich milieu in
which preimplantation embryos normally develop. The
trophic effects of bisphenol A at low concentration may
have been compensating for the estrogen deprivation of
the control culture. It would have been interesting to
compare physiologic concentrations of 17b-estradiol to
the control culture conditions.
Utility (Adequacy) for CERHR Evaluation Process:
This study is not useful in the evaluation process.
Li et al. (2003), support not indicated, examined the
effect of in vitro bisphenol A exposure on postimplantation
mouse and rat embryos. A limited
amount of information was available for the study,
which was published in Chinese, but included an
abstract and data tables presented in English. GD 8.5
mouse embryos and GD 9.5 rat embryos were cultured
for 48 hr in media containing bisphenol A [purity not
indicated] at 0, 40, 60, 80, or 100 mg/L [culture
ware not discussed]. Exposure of rat embryos to
bisphenol A concentrations Z60 mg/L resulted in
reduced crown-rump length and yolk sac diameter
and affected yolk sac circulation and morphologic