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

280 CHAPIN ET AL.
males/group. [It was not stated how data from the 2
control groups were handled for sperm analyses.]
Sperm data were analyzed by ANOVA. Organ weight
data were analyzed by ANCOVA, Pearson’s correlation
analysis, ANOVA, and least significant means test. [It
was not clear if the offspring or litter were considered
the statistical unit; only one randomly selected male
per litter was used per F. vom Saal, personal communication,
June 20, 2007.]
Statistically significant findings are summarized in
Table 76. Exposure to bisphenol A resulted in doserelated
reductions in daily sperm production efficiency
(i.e., per g testis) that attained statistical significance at
the highest dose level. Some significant but non-doserelated
effects were observed for body and organ
weights. Epididymal weights were reduced at both
doses. At the low dose, body and seminal vesicle weights
were reduced and preputial weight was increased. In
mice treated with octylphenol, daily sperm production
was reduced at the low dose but there was no effect on
reproductive organ weights. The study authors concluded
that exposure of the fetus to low doses of
endocrine-disrupting chemicals can affect the size and
function of reproductive organs.
[The NTP Statistics Subpanel (NTP, 2001) noted that
vom Saal et al. (1998) did not apparently require overall
differences by ANOVA to be significant before applying
the least significant difference test, which is prone to
false positive findings without the overall protection of
ANOVA. The NTP Subpanel was not able to confirm any
of the significant findings reported for bisphenol A. The
NTP Subpanel noted that in theory, their reanalysis of
organ weights was not necessarily in conflict with the
findings of the study authors because of the use of
different statistical methods (Dunnett test vs. Fisher least
significant difference test).]
Strengths/Weaknesses: Strengths are the use of oral
delivery and low dose levels. Weakness are the inability
to assume the genetic comparability and responsiveness
of CF-1 mice maintained in a closed colony for almost 20
years is comparable to other sources of CF-1 mice),
failure to weight-adjust the maternal dose daily, the lack
of information on testis weight (that is needed for
consideration of daily sperm production), small sample
size for sperm production measurement, and the questions
about the statistical analysis. An additional weakness
is the unusual/unexplained findings of low dose
only effect on weights.
Utility (Adequacy) for CERHR Evaluation Process:
The body weight data contained in this study are
adequate for the evaluation process, however overall
utility is limited because of sample size and statistical
concerns. Data on reproductive organ weights and sperm
production are considered inadequate for the evaluation.
Nagel et al. (1997), supported by NIH and the
University of Missouri-Columbia, examined the effect
of prenatal bisphenol A exposure on mouse prostate
weight. The mice used in this study were the same ones
used in the study by vom Saal et al. (1998), and
experimental details are provided in the above summary
of that study. CF-1 mice were fed Purina Laboratory
Chow 5001 and housed in polypropylene cages with corn
cob bedding. The mice (7/group) were dosed with
bisphenol A [purity not reported] at 0.002 and
0.020 mg/kg bw/day on GD 11–17. A control group of
6 mice was given the tocopherol-stripped corn oil vehicle
during the same time period. Vehicle and dosing
solutions were fed to the mice using a micropipette. A
second control group of 5 dams was unhandled. Because
there were no significant differences between the 2
control groups, data from the 2 groups were pooled.
Females were allowed to litter. Pups were weaned at 23
days of age and housed 3/cage. One male/litter was
selected and housed individually for 1 month. Body
weights of males were measured throughout the study.
Selected males were killed at 6 months of age for
measurement of prostate weight. Data for prostate
weight were analyzed by ANCOVA using body weight
as the covariate. If it was determined that body weight
did not account for differences in prostate weight, data
were reanalyzed by ANOVA without adjustment for
body weight. Selection of 1 male/litter controlled for
litter effects. Body weights were lower in males from the
0.002 mg/kg bw/day group than in controls. Statistical
analyses revealed that prostate weight was not related to
body weight. Compared to control values, prostate
weights were 30% higher in the 0.002 mg/kg bw/day
group and 35% higher in the 0.020 mg/kg bw/day
group. The study authors concluded that bisphenol A
alters the reproductive system of mice at doses near
reported ranges of human exposure.
[The NTP Statistics Subpanel (NTP, 2001) concluded
that Nagel et al. (1997) used appropriate statistical
methods, and the Subpanel reached essentially the
same conclusions as the study authors regarding
elevated prostate weight.]
Strengths/Weaknesses: Strengths are the use of the
same methods as vom Saal et al. (1998) and the use of
dose levels in the range of human exposure. The
independent confirmation of the data analysis by the
NTP Statistics Subpanel is another strength. The use of a
small sample size, closed mouse colony, and the failure to
present any histopathological analyses are weaknesses.
The Purina 5001 chow has high and variable levels of soy
phytoestrogens, and the corn cob bedding is known to be
problematic due to antiestrogenic constituents. This
study did not use a positive control, although there are
earlier reports from this laboratory using
diethylstilbestrol.
Utility (Adequacy) for CERHR Evaluation Process:
This study is adequate and useful for the evaluation
process.
Cagen et al. (1999a), support not indicated [authors
noted to work in industry], examined the effects of
prenatal bisphenol A exposure on the developing
reproductive system of male mice. The study attempted
to duplicate the findings by vom Saal et al. (1998) and
Nagel et al. (1997) by repeating their procedures.
Exceptions were: (1) use of larger group sizes to increase
statistical power; (2) use of 4 dose levels instead of 2; (3)
use of 2 methods to determine sperm counts; (4) killing of
male offspring at 90 instead of 180 days; (5) conducting
the study according to GLP; (6) obtaining mice from a
commercial source instead of an in-house bred colony;
and (7) housing males individually after weaning. In the
study by Cagen et al. (1999a), CF-1 mice gaining more
than 4.5 g weight from GD 0 to 10 were randomly
assigned to groups of 28 animals and administered
bisphenol A (499% pure) 0.0002, 0.002, 0.020, or 0.2 mg/
kg bw/day on GD 11–17. Two negative control groups
Birth Defects Research (Part B) 83:157–395, 2008