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

are a primary (or secondary) effect or are the result of
cytotoxicity. Calcium levels were also affected and
collectively these changes may be the result of apoptosis
initiated by some other mechanism.
Utility (Adequacy) for CERHR Evaluation Process:
This study was not considered useful for the evaluation
process.
Tabuchi et al. (2006), supported in part by the Japanese
Ministry of Education, Culture, Sports, Science, and
Technology, examined the effects of bisphenol A on gene
expression in mouse Sertoli cell cultures. TTE3 cells were
incubated in media containing bisphenol A [purity not
reported] at 0 (DMSO vehicle) or 200 mM [46 mg/L] for up
to 12 hr [culture ware type not discussed]. Cells were
examined for viability using dye exclusion assays and for
apoptosis by formation of DNA ladders. RNA was
extracted from cells, and gene expression was determined
by PCR and microarray analyses. Data were
analyzed by Student t-test. Cell viability was decreased
in a time-related manner between 3–12 hr of bisphenol A
exposure, but there was no evidence of apoptosis. PCR
analysis indicated that bisphenol A exposure significantly
and time-dependently increased mRNA transcripts
for 2 endoplasmic reticulum stress markers,
hspa5 and ddit3. Microarray analysis demonstrated that
661 sets of genes were downregulated and 604 sets of
genes were upregulated 42-fold following bisphenol A
exposure. Pathway analysis of decreased gene clusters
revealed 2 significant genetic networks associated with
the cell cycle or cell growth and proliferation. In
increased gene clusters, two genetic networks were
associated with cell death, DNA replication, recombination
and repair, or injuries and abnormalities. The study
authors concluded that the genes, genetic clusters, and
genetic networks identified in this study are likely
involved in Sertoli cell injury following bisphenol A
exposure.
Strengths/Weaknesses: State-of-the-art technology
was used in this study to examine gene expression
changes after in vitro bisphenol A exposure of a Sertoli
cell line. Only one dose level was examined. The use of
hormone rich fetal bovine serum in the media may be a
confounder. The absence of DNA laddering is not
conclusive evidence of the absence of apoptosis (e.g.,
adherent cells undergoing apoptosis often are released
into the culture media). Moreover, it is not surprising that
given this ‘‘high’’ bisphenol A concentration, ‘‘novel’’
and likely non-specific gene changes were noted.
Utility (Adequacy) for CERHR Evaluation Process:
This study was not considered useful for the evaluation
process.
4.2.3 Male and female.
4.2.3.1 Rat: Two unpublished studies performed by
the International Research and Development Corporation
for General Electric (General Electric, 1976, 1978)
provided some information on reproductive toxicity in
rats orally exposed to bisphenol A. The studies are
described in detail in Section 3.2.3.1. There was no effect
on fertility in male and female rats given feed containing
up to 9000 ppm bisphenol A (B650 mg/kg bw/day in
males and 950 mg/kg bw/day in females) for an
unspecified period before mating (General Electric,
1976). A second study reported no effects on estrus
cyclicity or gestation length [data not shown by study
authors] or male or female fertility in rats given feed
Birth Defects Research (Part B) 83:157–395, 2008
BISPHENOL A
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containing bisphenol A at up to 1000 ppm (B60 mg/kg
bw/day in males and 100 mg/kg bw/day in females) for
B70 days before mating (General Electric, 1978).
Ema et al. (2001), supported by the Japanese Ministry
of Health and Welfare, conducted a multigeneration
reproductive toxicity study of bisphenol A in CD rats.
Animals were housed in suspended stainless steel cages
at the beginning of the study. From GD 17, wood chips
were used as bedding. Rats were fed CRF-1 chow
(Oriental Yeast Co). In the study that was conducted
according to GLP, F 0 male rats and female rats with 4–5day
estrous cycles were randomly assigned to groups of
25/sex. Five-week-old males and 10-week-old females
were gavaged with 0 (distilled water vehicle), 0.0002,
0.002, 0.020, or 0.200 mg/kg bw/day bisphenol A (99.9%
purity). Males were dosed for 10 weeks before mating and
during the mating period, which lasted up to 2 weeks.
Females were dosed from 2 weeks before mating, and
during the mating, gestation, and lactation periods. Doses
were based on results of studies by Nagel et al. (1997) and
vom Saal et al. (1998). Stability and concentration of dosing
solutions were verified. Dams delivered and nursed their
pups. At weaning on PND 22 (day of birth defined as
PND 0), 1 or 2 F1 weanlings/litter/sex (25/sex/group)
were selected to continue in the study. Dosing of F 1
animals began on PND 23 and continued for 10 weeks
before mating and through the mating period, which
lasted up to 3 weeks. Dosing was continued through the
gestation and lactation periods. Twenty-five F2 weanlings/sex/group
were selected on PND 22. Beginning on
PND 22, male F2 rats were dosed for 4 weeks and females
were dosed for 11 weeks before being killed.
Endpoints examined in adult rats included clinical
signs, body weight, and food intake. Fertility, copulation,
and gestational indices were examined in mating rats.
Vaginal smears were evaluated for 2 weeks before mating
in F0 and F1 females and at 9–11 weeks of age in F2
females. Dams were killed and necropsied following
weaning of their pups, and uterine implantation sites
were examined. Males were killed following mating.
Organs were weighed and histopathology examinations
were conducted in control and high-dose animals. Sperm
endpoints were measured in F 0 and F 1 adult males.
Serum hormone levels were measured in 6 adult F0 and
F1 males and proestrous females. At birth, pups were
counted, sexed, and examined for viability and external
malformations. On PND 4, litters were culled to 4 male
and 4 female pups. At weaning, 1 male and female F1
and F 2 weanling was killed for organ weight measurement;
histopathology exams were conducted in seminal
vesicles and coagulating glands of F 2 weanlings. Survival
and growth were monitored during the postnatal period.
Pups were examined for developmental landmarks and
attainment of vaginal opening or preputial separation.
Anogenital distance in pups was examined at numerous
time points during the lactation period and through
adulthood. Behavioral testing was conducted at 5–7
weeks of age. The litter was considered the experimental
unit in data obtained before weaning. Statistical analyses
included Bartlett test for homogeneity of variance,
ANOVA, and/or Dunnett multiple comparison, Kruskal–Wallis,
Mann–Whitney U, w 2 , or Fisher exact tests.
In F0 and F1 adult animals, there were no treatmentrelated
effects on clinical signs, body weight gain, or death.
The only significant reproductive effects reported in adult