Download Effects of Acute Exposure to PCBs 126 and 153 on Anterior Pituitary

47, 158 –169 (1999)
Copyright © 1999 by the Society of Toxicology
TOXICOLOGICAL SCIENCES
Effects of Acute Exposure to PCBs 126 and 153 on Anterior Pituitary
and Thyroid Hormones and FSH Isoforms in
Adult Sprague Dawley Male Rats
D. Desaulniers, 1 K. Leingartner, M. Wade, E. Fintelman, A. Yagminas, and W. G. Foster
Environmental and Occupational Toxicology Division, Bureau of Chemical Hazards, Environmental Health Directorate, Health Protection Branch,
Department of Health, Ottawa, Ontario, Canada
Received May 1, 1998; accepted October 5, 1998
3,3*,4,4*,5-Pentachlorobiphenyl (PCB 126) and 2,2*,4,4*,5,5*hexachlorobiphenyl (PCB 153) were administered to adult male
rats in order to identify sensitive indicators of endocrine disruption. We tested the hypothesis that PCB exposure modifies folliclestimulating hormone (FSH) pituitary isoforms, as well as the
pituitary and serum concentrations of FSH, luteinizing hormone
(LH), growth hormone, prolactin, and thyroid-stimulating hormone (TSH). Effects on serum levels of thyroxine (T4) and testosterone (T), and prostate androgen receptor content, were also
tested. In one experiment, 5 groups of 8 rats each received two ip
injections, one day apart, of either corn oil or 6.25, 25, 100 or 400
mg/kg/day of PCB 126. Decreases ( p < 0.05) in the serum concentrations of T4 and LH started at doses of 25 and 100 mg/kg/day,
respectively. Serum FSH concentrations were reduced ( p 5 0.07)
in the highest dose group. In contrast, pituitary content of FSH
and LH increased with PCB-126 doses ( p 5 0.004, p 5 0.002,
respectively). Despite changes in reproductive hormones, PCB-126
had no effect on the androgen receptor content of the prostate. The
effect of PCB-126 was tested in the hemicastrated rat, and suggested adverse effects on testosterone secretion. To test the effects
of PCB exposure on FSH pituitary isoforms, 4 groups of 10 male
rats received two ip injections, one day apart, of either corn oil,
PCB 153 (25 mg/kg/day), estradiol-17b (E2; 20 mg/kg/day), or
PCB 126 (0.1 mg/kg/day). Serum T4 levels were higher ( p < 0.01)
in the E2 and PCB 153 groups, and slightly reduced in the PCB
126-treated groups, compared to controls. Simultaneous purification of pituitary FSH and TSH isoforms was performed by HPLC,
using two chromatofocusing columns in series. In contrast to TSH
isoforms, the distribution of FSH isoforms over the chromatography run differed slightly between treatment groups; the amounts
of FSH isoform eluted during the pH gradient were lower ( p <
0.05) in E2 and PCB 153-treated rats than in control or PCB
126-treated rats. The similarity between the effects of E2 and PCB
153 on T4 and FSH isoforms supports the contention that PCB
153 possesses estrogenic properties. Serum LH and T4 concentrations were the most sensitive and practical endocrine indicators of
PCBs 126 and 153 exposure in male rats.
1
To whom correspondence should be addressed at Room 330, Environmental Health Centre, Bldg # 8, Tunney’s Pasture, Postal Locator 0803D,
Ottawa, Ontario, Canada K1A 0L2. Fax: 613–941– 4768. E-mail:
[email protected].
Key Words: 3,3*,4,4*,5-pentachlorobiphenyl (PCB 126);
2,2*,4,4*,5,5*-hexachlorobiphenyl (PCB 153); follicle-stimulating
hormone (FSH); thyroid-stimulating hormone (TSH); luteinizing
hormone (LH); rat.
The worldwide commercial production of PCBs essentially
ceased in the late 1970s, and yet, more than 70% of the global
production of PCBs is estimated to be still in use or in stock
(Hileman, 1993). Humans continue to be exposed to PCBs
because of their presence in “environmental hot spots,” their
accidental release from disposal sites (Phaneuf et al., 1995),
and their presence in our diet, due in part to the accumulation
of PCBs in certain species of fish and seafood collected in the
more contaminated areas (Hansen et al., 1995; Li and Hansen,
1997). Despite their gradual decline, PCBs remain the major
contaminants of human tissues (Newsome et al., 1995), and
thus, they still represent a major toxicological issue. Some
PCBs and their metabolites have estrogenic (Connor et al.,
1997; Soto et al., 1995) or antiestrogenic properties (Connor et
al., 1997; Krishnan and Safe, 1993) and alter gonadotropin
release from female rat pituitary cells in vitro (Jansen et al.,
1993). PCBs alter thyroid functions in many ways (Brouwer et
al., 1998). They indirectly favor thyroxine excretion by the
activation of glucuronidating enzymes and by displacing thyroxine from transthyretin, the major transporting protein in
rodents. They can also cause direct effects on the thyroid by
inhibiting proteolysis of thyroglobulin (van Birgelen et al.,
1995). PCB 126 (3,39,4,49,5-pentachlorobiphenyl), a CYP1A1
inducer, non-ortho substituted, is the most toxic PCB congener
and has antiestrogenic properties (Krishnan and Safe, 1993).
PCB 153 (2,29,4,49,5,59-hexachlorobiphenyl), is a diortho-substituted congener with CYP2B-inducing properties. It has been
shown to have estrogenic effects in the rat (Li et al., 1994) and
is one of the more highly concentrated PCB congeners found in
human tissues (Foster, 1995; Newsome et al., 1995).
Gonadotropins, luteinizing hormone, (LH); follicle-stimulating hormone (FSH), and thyroid-stimulating hormone (TSH),
are glycoproteins present in multiple forms within the pituitary
158
ENDOCRINE EFFECTS OF PCBs 126 AND 153
gland and in circulation. Hormonal isoforms differ in their
degree of glycosylation and sialylation (Baenziger and Green,
1988; Smith and Baenziger, 1988) and amino acid sequence
(Gharib et al., 1990; Matthews et al., 1993; Ryan et al., 1987;
Weiss et al., 1992; Wilson et al., 1990), which alter their
half-lives, affinity for the receptors (Stanton et al., 1996),
ability to activate different intracellular signal transduction
systems (FSH, Arey et al.1997; TSH, Schaaf et al., 1997) and
thus, their in vivo and in vitro biological activity (Beitins and
Padmanabhan, 1991; Dahl and Stone, 1992; Hassing et al.,
1993; Stanton et al., 1992; Wide and Bakos, 1993). Up to 20
pituitary isoforms of human FSH and 30 isoforms of human
LH have been purified (Stanton et al., 1992, 1993). Different
isoforms are secreted depending on the endocrine environment
of the pituitary gland, and isoforms vary according to age,
stage of reproductive cycle, and in certain types of infertility
(Beitins and Padmanabhan, 1991; Dahl and Stone, 1992; Simoni et al., 1992; Hassing et al., 1993; Wide and Bakos, 1993).
In the search for sensitive indicators of xenobiotic exposure,
FSH isoforms may reveal another level of endocrine disruption
where the quality, but not the quantity, of the hormone is
affected.
The measurement of circulating hormones is still an essential tool in diagnostic endocrinology. However, because of the
dynamic nature of hormone secretion, measuring reproductive
hormones during extreme endocrine conditions is useful for
permitting the observation of endocrine disorders that could
not be detected under normal conditions. Some examples include the comparison of endocrine events during the normal
estrous cycle and following superovulatory treatment in mature
animals (Desaulniers et al., 1995a,b) or in HCB-treated rats
(Foster et al., 1993a). Others used a gonadotropin-releasing
hormone treatment to reveal adverse effects of lead on LH and
testosterone secretion in monkeys (Foster et al., 1993b). We
tested the usefulness of the hemicastration model (Frankel and
Mock, 1982; Frankel and Wright, 1982) in PCB-treated rat,
with the objective of developing a sensitive in-vivo toxicological model to facilitate the detection of endocrine-disrupting
effects. This in vivo model, not requiring the administration of
exogenous hormones, provides insights to the integrity of the
mechanisms regulating testosterone production and FSH secretion. The removal of one testicle creates an unusual endocrine
compensatory effect, whereby the remaining testicle increases
its testosterone output to match normal levels in circulation. In
addition, hemicastration increases FSH concentrations in serum and pituitary, due to a reduction in inhibin levels (Brown
and Chakraborty, 1991; Brown et al., 1991; Medhamurthy et
al., 1995). Inhibin is secreted by both testes and inhibits FSH
secretion, so the removal of one gonad decreases inhibin output. Thus, this experimental model tests the functionality of the
hypothalamo-pituitary-testicular axis and should help to identify the primary site of toxic insults. Therefore, the acute
endocrine-disrupting effects of PCB 126 were tested in adult
male rats by measuring, in both the serum and the pituitary
159
gland, hormones that are involved in reproductive functions
(LH, FSH, prolactin (PRL), and testosterone), development,
metabolism (growth hormone (GH) and PRL), and thyroid
function (TSH and thyroxine). In addition, the data generated
by the PCB 126-treated, hemicastrated rat model, and the
analysis of FSH isoforms in PCBs 126- and 153-treated male
rats, were assessed as toxicological investigative tools.
MATERIALS AND METHODS
Animal treatments. Adult male Sprague Dawley rats (Charles River,
St-Constant, Québec, Canada), weighing 390 – 422 g, and at approximately 90
days of age, were used for all experiments. In a preliminary experiment, the
effects of anaesthetising rats using isoflurane inhalation (n 5 8) or sodium
pentobarbital ip injection (n 5 7) were compared by measuring concentrations
of LH, FSH, TSH, GH, and PRL in the serum and pituitary gland homogenates,
and also testosterone and thyroxine in the serum. Both serum TSH (1.8 6 0.4
ng/ml vs. 2.4 6 0.2, p 5 0.07) and thyroxine (3.1 6 0.2 vs 4.0 6 0.4 ng/ml,
p 5 0.05) were lower in the sodium pentobarbital group than in the isoflurane
group. All other serum hormone concentrations were also slightly lower in the
sodium pentobarbital group when compared to isoflurane-anesthetized rats.
The reverse situation occurred for hormone concentrations measured in the
pituitary gland homogenate. Although anesthesia, or the use of some anesthetic
agents, does not always induce detectable endocrine alterations (Cohen et al.,
1983; Praputpittaya and Kimura, 1987; Waddell and Bruce, 1984), isoflurane
anaesthesia was selected for all other experiments because (1) LH pulses
(Strutton and Coen, 1996) and testosterone (Bardin and Peterson, 1967; Cooper and Waites, 1974; Hirsh et al., 1981) could be suppressed by anesthesia,
and our results showed that some hormone levels are either significantly or
slightly higher in isoflurane- than in sodium pentobarbital-treated rats; (2)
increasing the duration of the anesthesia could decrease hormone levels
(Schultz et al., 1995; Strutton and Coen, 1996), and, in contrast to sodium
pentobarbital, isoflurane was found to induce a more rapid anesthesia. This
occurred within a similar time frame between rats, following initiation of the
anesthetic treatment, and thus subjected rats to less stress; and finally (3)
sodium pentobarbital can be metabolized by the liver enzymes that are induced
by PCBs, which could lead to variable anesthetic response among PCB-treated
rats.
In one experiment, the dose-response effects of acute exposure to PCB 126
were tested. On day 0 of the experiment, the rats received an ip injection of
either corn oil or PCB 126 in corn oil at different concentrations: 6.25, 25, 100
and 400 mg/kg bw (Table 2). After 24 h, the same dosing regime was repeated.
Rats were anesthetized two days after the second dose and exsanguinated via
the abdominal aorta (between 9:00 A.M. and noon). The pituitary gland, brain,
liver, a sample of fat, the testicles, and the thyroid gland were dissected out,
weighed, and frozen at 285°C. Since serum LH decreased at a PCB-126 dose
of 100 mg/kg/day (Table 2) supporting the contention of pituitary effects, this
dose was then used for all subsequent experiments.
In a second type of experiment, the ability of PCB 126 to induce alterations
of the hypothalamo-pituitary-testicular axis was further tested in hemicastrated
rats (Table 3), to determine if this model would be a more sensitive method for
detecting altered testosterone and FSH secretions. Intact rats were treated, as
previously described, using 100 mg/kg bw of PCB 126 per day for two days,
and then hemicastrated on day 3, to ensure that the compensatory endocrine
changes would be occurring under the influence of the PCB treatment. The rats
were hemicastrated under isoflurane anesthesia according to the protocol of
Waynforth and Flecknell (1992). Briefly, the protocol entailed exposing the
left testicle by making a midscrotal incision, and ligating the testicular and
epididymal vessels to remove the left testicle and epididymis. The rats were
necropsied and exsanguinated on the 4th and 6th days after hemicastration.
In a third type of experiment, pituitary FSH and TSH isoforms were purified
from PCB 126-treated rats, to test if they could provide sensitive indicators of
160
DESAULNIERS ET AL.
TABLE 1
Peptide Hormone Double Antibody Radioimmunoassay
Reagents a and Sensitivity b
Assay
LH
FSH
TSH
GH
PRL
Iodination
hormone
Standard
curve
hormone
rLH-I-9
AFP-10250C
rFSH-I-8
AFP-11454B
rTSH-I-9
AFP-11542B
rGH-I-6
AFP-5676B
rPRL-I-6
AFP-10505B
rLH-RP-3
AFP-7187B
rFSH-RP2
AFP-4621B
rTSH-RP3
AFP-5512B
rGH-RP-2
AFP-3190B
rPRL-RP-3
AFP-4459B
Primary
antiserum
Sensitivity
ng/mL
rLH-S-11
0.16
rFSH-S-11
AFP-C0972881
rTSH-RIA-6
AFP329691Rb
rGH-S-5
1.2
rPRL-S-9
AFP-131581570
0.47
0.8
0.7
a
The names of the reagents are preceded by NIDDK. The secondary
precipitating antibody was an anti-rabbit IgG developed in goat (Sigma Immunochemicals, St-Louis, MO) for all peptide hormone assays, except for GH,
which required a goat anti-monkey IgG (Antibodies Incorporated, Davis, CA).
b
Sensitivity is defined as the smallest dose of reference preparation always
three standard deviations apart from the nonspecific binding. The serum
volumes used in the assays were: LH and FSH 250 mL, PRL 100 mL, TSH 150
mL and GH 20 mL.
exposure. To ensure that this experiment would have detectable effects, two
additional groups of rats were given either 25 mg/kg bw of PCB 153 or 20
mg/kg bw of estradiol-17b in two doses over two days. These doses of estradiol
and PCB 153 were selected because they have been reported to cause uterotrophic effects (Li et al., 1994). In this experiment, the thyroid glands were
dissected out, fixed, and examined for histological effects.
Assays. Gonadotropin, PRL, GH and TSH concentrations were determined
by double antibody radioimmunoassays (RIA). The method of iodination
(Lussier et al., 1994) and assay procedures were as previously described
(Desaulniers et al., 1997). Assay reagents and sensitivities are described in
Table 1. Commercial double antibody, or coated tube, I 125 RIA kits (ICN
Biomedicals, Costa Mesa, CA) were used for the measurement of testosterone,
thyroxine, and triiodothyronine (T3). Ethoxyresorufin (EROD) and pentoxyresorufin o-deethylase (PROD) activities in 10 000 3g liver homogenate
supernatants were determined on a Perkin-Elmer model LS50B fluorescence
spectrometer equipped with a micro-plate reader, according to the method of
Burke et al., (1985). PROD and EROD hepatic activities were calculated from
the slope of the linear portion of the reaction.
Chromatofocusing on HPLC. All the buffers contained 1% glycerol
(Kojima et al., 1995; Stumpf et al., 1992). Pituitary gland homogenates were
prepared, on ice, just before HPLC analysis. The glands were sonicated in vials
containing 200 ml of 0.05 M phosphate-buffered saline, 1% BSA and several
protease inhibitors (1 mM EDTA, 1.0 mM pepstatin A, 10 mM E-64, 17 mM
2,3-dehydro-2-deoxy-n-acetylneuraminic acid and 1.5 mg/ml aprotinin, Sigma
Chemicals, St. Louis, MO). The homogenates were microcentrifuged at 1300 g
for 30 min at 4°C (Model 5415C, Brinkmann Instruments, Rexdale, Ontario,
Canada), and the supernatants were kept on ice. The sonication and centrifugation procedures were repeated two more times to obtain a final homogenate
volume of 0.6 mL. These steps were sufficient to recover 99.2 % of the FSH.
Preliminary experiments showed that additional sonication and recentrifugation of the pituitary pellet extracted only 0.8% of total immunoassayable FSH.
The homogenates were prepurified and at the same time reconstituted in 0.025
M methylpiperazine (pH 5.7) buffer by using gel filtration (PD-10 disposable
column, Pharmacia Biotech, Montréal, Quebec, Canada). Relative to the
amount of FSH applied to the PD-10 column, 78.4 6 1.8 % of the FSH (n 5
17) was recovered in a pool of 1.5 ml eluting after an initial volume of 2 mL.
Of this 1.5 ml eluate, 1 ml was applied to the HPLC column.
The HPLC system (System Gold, Beckman) was equipped with a UV
detector (model 166) set at 280 nm, a flow-through pH detector (Pharmacia
Biotech), a fraction collector (Isco, Foxy) and two chromatofocusing columns
(mono-p HR 5/5 and 5/20, Pharmacia Biotech) in series. The columns were
equilibrated with start buffer (0.025 M methylpiperazine/HCl at pH 5.7), and
the homogenate was injected. A pH gradient was developed for 65 min by
eluting the columns with polybuffer-74 (Pharmacia Biotech) diluted 1:8 with
water and adjusted to pH 4 with HCl. This was followed by a NaCl gradient
(0 to 2 M) for 30 min and then elution of the columns with 2 M NaCl for 10
min. Fractions (one per minute for 110 min) were collected in 133100-mm
borosilicate test tubes containing 200 ml of stabilizing buffer (0.5 M phosphate-buffered saline containing 3.5% BSA and the protease inhibitors already
described), and then kept at 215°C for assay within 3 days of their elution. The
total amount of FSH recovered from the HPLC fractions was determined to be
79.4 6 3.0% (n 5 18) of the amount applied to the HPLC column.
Histology of the thyroid gland. Thyroids were fixed in 10% neutral
buffered formalin and embedded in paraffin. Longitudinal sections (6 mm
thick) were cut at a depth of 450 mm into the gland. None of the histological
measurements were taken from the peripheral inactive follicles (Ness et al.,
1993). The colloid vacuolation was analysed (magnification 5 2003) by
categorizing 300 to 400 follicles per rat into five levels of vacuolation (1 5 no
vacuoles, 2 5 less than 20% of the colloid area covered with vacuoles, 3 5 20
to 50% covered, 4 5 more than 50% covered, and 5 5 foamy colloid). All
other histological analyses were performed objectively using the Optimas
computer-imaging system (Bioscan, Edmonds, WA). The colloid density was
analyzed (magnification 5 2003) using luminescence readings (0 5 white;
255 5 black). The colloid vacuolation and density were analyzed using
preparations treated with periodic acid Schiff’s reagent which stains for carbohydrates. The epithelial height was measured from an average of 34 follicles
per rat (magnification 5 10003), and follicle areas were calculated using the
gray binary system, from an average of 14 follicles per rat (magnification 5
4003). The epithelial heights and follicle areas were taken from preparations
treated with VanGieson’s stain for collagen.
Data analysis. To test for differences in the elution pattern of FSH and
TSH isoforms induced by the PCB or the estradiol treatments, the data was
analyzed for each rat and the peaks of hormone, easily identifiable among rats,
were used as markers of chromatographic regions containing isoforms with
similar retention times. These regions were called isoform groups 1 to 6 for
TSH, and 1 to 9 for FSH (Fig. 1). For the statistical analysis, the concentration
of each hormone per fraction was expressed as a percentage of the total amount
of hormone eluted during the whole chromatography run, and the percentages
were added for each isoform group. The large amount of FSH eluted in isoform
group 9 (see Fig. 2) prevented the data from reaching homogeneity of variance,
even after arcsin transformation. Thus, 2 two-way analysis of variance
(ANOVA) tests were used to analyze the isoform data. First, a two way
ANOVA was performed, excluding the data from isoform group 9, testing the
effects of treatment, isoform group, and treatment 3 isoform group interaction.
Then differences in the overall treatment means of isoform groups 1 to 8 were
assessed by the Duncan’s multiple range test. A second two-way ANOVA was
performed, testing for an effect of treatment, isoform groups, and their interaction, using the amount of FSH eluted from the combined isoform groups 1
to 8 against that eluted in isoform group 9 (inset Fig. 2). For all other analyses,
homogeneity of variance between dose groups was verified by O’Brien and
Brown-Forsythe tests (SAS Institute Inc, 1992), and the data was log transformed, if required. The data were analyzed using ANOVA, with the model
indicated in footnotes of the data table. Effects were considered significant at
p # 0.05 and a tendency was indicated by p # 0.1.
ENDOCRINE EFFECTS OF PCBs 126 AND 153
161
concentration (Table 2). There was a tendency for FSH concentrations to be lower in rats treated with the highest dose of
PCB 126 compared with controls ( p 5 0.07, Table 2). Serum
thyroxine concentrations decreased in rats treated with 25
mg/kg/day of PCB 126 or more ( p 5 0.0001), but this was not
associated with significant changes in TSH concentrations (Table 2). In contrast to the decreasing concentrations of LH and
FSH observed in the serum, the content of LH ( p 5 0.002) and
FSH ( p 5 0.004) increased in the pituitary gland of rats treated
with PCB-126 (Table 2). None of the doses of PCB 126 altered
the cytosolic or the nuclear androgen receptor content of the
prostate (Table 2).
Experiment 2
Alterations of the hypothalamo-pituitary-testicular axis induced by a dose of 100 mg/kg/day of PCB 126 was further
tested in hemicastrated rats (Table 3). In contrast to Experiment 1, body weight gain was decreased ( p 5 0.005) by the
PCB 126 (100 mg/kg/day)-treatment in hemicastrated rats
(PCB treated: 7 6 5 g; untreated: 27 6 4 g); note that the delay
FIG. 1. HPLC chromatofocusing run for the separation of TSH and FSH
isoforms from two pituitary glands. (A) Changes in pH detected by the flow
through electrode and the gradient of NaCl during the chromatography run. (B)
TSH and (C) FSH concentrations in each of the 0.5 mL fractions collected
every minute during the run. Isoforms with similar retention times, easily
identifiable between rats, were divided into 6 regions for TSH (B) and 9
regions for FSH (C). Rats 17 and 31, both treated with PCB 153, were selected
to show an inter-individual difference observed in 8/20 rats, regardless of the
treatment group. This difference is the presence of FSH eluting in the region
2 of the chromatogram in rat 31 (C). Effects of treatments are illustrated in
Figure 2.
RESULTS
Experiment 1
Body weight gain declined in rats that received 100 mg/kg/
day of PCB 126 but this effect was only significant in the rats
given a dose of 400 mg/kg/day (Table 2). Even at the highest
dose, this effect on growth could not be associated with a
statistically significant effect on GH, although the variability
and the mean concentration of serum GH decreased (Table 2).
The smallest dose of PCB 126 (6.25 mg/kg/day) significantly
increased hepatic EROD activity, which reached maximal induction at a dose of 100 mg/kg/day. Serum LH concentrations
decreased ( p 5 0.004) in rats treated with 100 mg/kg/day, but
it was not associated with a significant change in testosterone
FIG. 2. Amount of FSH eluted during intervals of the pH and NaCl
gradients of the HPLC-chromatofocusing run in relation to the treatment
groups. The amounts of FSH (mean 6 SE; n 5 5) are expressed as a
percentage of the total amount of FSH eluted during each chromatography run.
A two-way ANOVA on arcsine transformed data from the first 8 regions
revealed a significant effect of pH interval ( p 5 0.0001) and treatment ( p 5
0.0001) with no significant interaction. Asterisks beside the legends indicate
that the amount of FSH eluted during region 1– 8 in estradiol and PCB
153-treated rats, is smaller than in control rats ( p , 0.05, Duncan’s multiple
range test). This can also be observed in the inset which illustrates the effects
of a Treatment 3 Isoform group interaction ( p 5 0.02), the effects of
treatment (p . 0.05) and isoform group (groups 1– 8 vs 9; p , 0.0001), derived
from a second two-way ANOVA comparing the combined isoform groups 1– 8
to group 9.
162
DESAULNIERS ET AL.
TABLE 2
Effects of Acute Exposure to PCB 126 on Body Weight Gain and Hepatic Ethoxyresorufin-o-deethylase Activity, Hormone
Concentrations in the Serum and the Pituitary Gland, and Androgen Receptor Content of the Prostate
Intraperitoneal doses of PCB 126 (mg/kg/day)
0
6.25
25
Body weight gain and hepatic ethoxyresorufin-o-deethylase (EROD) activity:
Weight gain (g)
16.6 6 3.0 a
17.4 6 2.8 a
18.4 6 1.5 a
d
c
EROD (nm/min/mg)
0.03 6 0.00
1.34 6 0.1
2.80 6 0.32 b
Hormone concentrations in the serum:
LH (ng/mL)
0.75 6 0.15 a
0.62 6 0.12 a
0.62 6 0.1 a
T (ng/mL)
2.42 6 0.65
1.32 6 0.34
1.98 6 0.84
FSH (ng/mL)
11.88 6 0.87
9.85 6 0.84
11.11 6 0.94
TSH (ng/mL)
2.4 6 0.18
2.13 6 0.27
2.97 6 0.59
T4 (mg/dL)
3.95 6 0.35 a
3.48 6 0.31 ab
2.78 6 0.17 bc
T3** (ng/dL)
72.8 6 3.9 a
79.4 6 4.1 a
67.8 6 5.5 ab
GH (ng/mL)
29.9 6 14.4
24.1 6 6.2
32.8 6 19.0
PRL (ng/mL)
8.1 6 1.41
8.16 6 1.35
9.18 6 2.19
Hormone concentrations in the pituitary gland (relative to the wet weight of the gland):
LH (ng/mg)
753 6 80 b
1139 6 188 a
1439 6 103 a
c
bc
FSH (ng/mg)
337 6 46
416 6 47
580 6 36 a
TSH (ng/mg)
596 6 75
544 6 73
789 6 94
GH (mg/mg)
54.4 6 9.9
61.7 6 9.2
64.1 6 4.8
PRL (ng/mg)
414 6 67
494 6 87
603 6 94
Androgen receptor content (fmole/mg protein) of the prostate:
Cytosol
5.1 6 1.0
5.1 6 1.5
5.6 6 1.7
Nuclear
15.6 6 1.6
12.8 6 1.2
15.0 6 2.2
100
400
p value*
12.6 6 2.2 a
4.07 6 0.19 a
27.2 6 3.6 b
3.81 6 0.14 a
0.0001
0.0001 (log)
0.28 6 0.03 b
1.75 6 0.55
9.57 6 1.01
3.33 6 0.39
2.48 6 0.29 c
55.3 6 3.2 b
42.0 6 16.0
8 6 1.23
0.29 6 0.03 b
1.18 6 0.37
8.55 6 0.84
2.34 6 0.49
1.26 6 0.12 d
42.7 6 1.3 c
8.0 6 2.0
8.02 6 1.4
0.004
0.57
0.07
0.23 (log)
0.0001
0.0001 (log)
0.47
0.98
1374 6 156 a
528 6 56 ab
719 6 87
68.1 6 3.8
452 6 39
1508 6 101 a
513 6 34 ab
700 6 66
69.6 6 4.1
459 6 47
0.002
0.004
0.17
0.55
0.39
8.8 6 1.4
16.0 6 3.2
6.0 6 1.3
11.8 6 1.3
0.83
0.34
Note. T, testosterone; T4, thyroxine; T3, triiodothyronine.
* p value from a one way ANOVA. Mean 6 standard error of the mean, n 5 8 rats/group.
a,b,c,d
One way ANOVA followed by Duncan’s multiple range test; means with different letters are significantly different (p , 0.05).
** Analyzed during revision.
between PCB treatment and euthanasia was longer than in
Experiment 1. Except for the reduced concentration of thyroxine, no other significant hormonal differences between hemicastrated groups were induced by the PCB-126 treatment (Table 3). The testosterone, LH, FSH and TSH concentrations
were also compared with those of intact rats (see Table 2), not
treated or treated with the same dose of PCB 126. In all cases,
the data was compared using a two way ANOVA to test for
effects of hemicastration, PCB treatment, and their interaction,
followed by multiple comparison procedures (see Table 3). In
contrast to the reduced LH concentration induced in intact rats
by dosages of 100 mg/kg/day of PCB 126 compared to controls
( p , 0.05, Table 2), the PCB 126-treated, hemicastrated rats
had the highest LH concentration, which was statistically
higher than the PCB 126-treated intact rats but was not statistically higher than control intact rats (Table 2) or hemicastrated, untreated rats (Table 3). The elevated LH concentration
in the PCB 126-treated, hemicastrated rats is associated with
the lowest concentration of testosterone, which was statistically lower than in the normal rats (Table 2) but not in the
untreated, hemicastrated rats (Table 3). FSH concentrations in
hemicastrated rats (Table 3) and in the intact rats (Table 2)
were not affected by the PCB-126 treatment. Hemicastration
slightly increased FSH concentrations, since the PCB-treated
and -untreated, hemicastrated rats (n 5 24, 12.3 6 0.6 ng/mL,
Table 3) had a tendency for higher ( p 5 0.1) FSH concentrations than those measured in PCB treated and untreated intact
rats (n 5 16, 10.7 6 0.7 ng/mL; Table 2). Hemicastration had
no effect on TSH concentration (Table 3), whereas the 100
mg/kg/day PCB-126 treatment increased ( p 5 0.01) the TSH
concentrations in the combined intact (Table 2) and hemicastrated (Table 3) rats.
Experiment 3
Rats treated with PCBs 153 and 126 had elevated ( p 5
0.0001) PROD and EROD hepatic activity, respectively (Table
4). In all treatment groups, circulating levels of FSH, LH, TSH
and testosterone were lower than in controls, although the
difference was significant only for LH ( p 5 0.04). Thyroxine
concentrations were significantly higher in the estradiol- and
PCB 153-treated rats than in controls. The estradiol treatment
decreased pituitary concentrations of FSH but increased those
of prolactin (Table 4). Although thyroxine was not significantly lower in the PCB 126-treated group than in the control
group (Table 4), the colloid of the thyroid follicles was less
dense (colloid density: luminescence units higher) in the PCB
126-treated rats than in the controls (Table 5).
163
ENDOCRINE EFFECTS OF PCBs 126 AND 153
TABLE 3
Effects of Acute Exposure to PCB 126 on Hormonal Serum Concentrations in Hemicastrated Male Rats
Hormonal serum concentrations a
HC
HC 1 PCB 126
100 mg/kg/day
Testosterone b
(ng/mL)
LH c
(ng/mL)
FSH d
(ng/mL)
TSH e
(ng/mL)
Thyroxine
(mg/dL)
Prolactin
(ng/mL)
GH
(ng/mL)
1.42 6 0.45
0.6 6 0.07
12.18 6 0.8
2.31 6 0.34
3.68 6 0.38
11.85 6 2.2
8.97 6 2.65
0.76 6 0.15
0.83 6 0.13
12.49 6 1.1
3.14 6 0.34
1.62 6 0.17*
7.89 6 1.4
13.63 6 3.0
Note. HC, hemicastrated; mean 6 standard error of the mean, 12 rats/group.
* Significantly lower than hemicastrated rats (t-test, p 5 0.0001).
a
The rats were hemicasterated 48 h following the second i.p. injection of vehicle (corn oil) or PCB 126. Necropsy occurred 4 and 6 days following
hemicastration.
b
The testosterone, LH, FSH and TSH concentrations were also compared with those of intact rats (see Table 2), not treated or treated with the same dose of
PCB 126 as above. In all cases, the data was compared using two way anova to test for effects of hemicastration, PCB treatment and their interaction, followed
by all pairwise multiple comparison procedures (Student-Newman-Keuls method), unless indicated otherwise. Testosterone analysis using log transformed data:
effect of hemicastration ( p 5 0.04), and a tendency for an effect of PCB 126 ( p 5 0.09). The concentration in the hemicastrated-PCB treated group (0.76 ng/mL)
is lower than in the intact control group (2.42 ng/mL, Table 2, Mann-Whitney Rank Sum Test, p 5 0.01).
c
LH analysis using log transformed data: effect of hemicastration ( p 5 0.05), and an effect of hemicastration 3 PCB 126 interaction ( p 5 0.003). The intact
rats treated with PCB 126 (see Table 2) had a lower ( p , 0.05) concentration of LH than the untreated intact rats or both groups of hemicastrated rats.
d
FSH analysis on the raw data: there is a tendency ( p 5 0.10) in hemicastrated (PCB treated or not) rats to have higher FSH concentrations than in intact
(PCB treated or not, Table 2) rats.
e
TSH analysis on the raw data: effect of PCB 126 ( p 5 0.01) with higher concentrations measured in PCB treated rats.
TSH and FSH isoforms were separated into 6 and 9 chromatographic regions of similar retention times among rats (Fig.
1). The reproducibility of the technique is shown by the overlap of the pH, TSH and FSH profiles between rat samples (Fig.
1). A peak of FSH isoforms eluted between pH 5.25 and 4.5 in
8/20 rats, regardless of the treatment group, indicated interindividual differences in isoform distribution (Fig. 1). The
average amount of FSH eluted during separate chromato-
TABLE 4
Effects of Acute Exposure to Estradiol-17b, PCB 126 or PCB 153 on Body Weight Gain, Hepatic Ethoxy- and
Pentoxy-Resorufin-o-Deethylase Activity, and the Hormone Concentrations in the Serum and in the Pituitary Gland
Control (oil)
Estradiol-17b
PCB 153
Body weight gain* and hepatic ethoxy (EROD)** and pentoxyresorufin-o-deethylase (PROD)** activity:
Weight gain (g)
20.4 6 2.9
18.0 6 2.8
16.8 6 1.6
EROD (nm/min/mg)
0.19 6 0.01 c
0.18 6 0.01 c
0.46 6 0.05 b
PROD (nm/min/mg)
0.3 6 0.05 c
0.25 6 0.03 c
3.12 6 0.25 a
Hormone concentrations in the serum:**
LH (ng/mL)
0.55 6 0.1 a
0.31 6 0.08 b
0.28 6 0.04 b
T (ng/mL)
3.22 6 0.38
2.07 6 0.5
2.32 6 0.54
FSH (ng/mL)
9.54 6 0.55
7.63 6 0.92
8.4 6 0.49
TSH (ng/mL)
2.67 6 0.64
1.69 6 0.27
1.53 6 0.23
T4 (mg/dL)
4.43 6 0.36 b
5.6 6 0.26 a
5.27 6 0.33 a
Hormone concentrations in the pituitary gland (relative to the wet weight of the gland):****
LH (ng/mg)
849 6 119
857 6 80
984 6 52
FSH (ng/mg)
382 6 26 a
299 6 19 b
409 6 28 a
TSH (ng/mg)
792 6 97
648 6 110
724 6 78
GH (mg/mg)
48 6 10
60 6 6
72 6 7
PRL (ng/mg)
480 6 70 b
1160 6 152 a
626 6 58 b
PCB 126
p value***
17.3 6 1.4
12.8 6 1.04 a
0.48 6 0.04 b
0.71
0.0001 (log)
0.0001 (log)
0.25 6 0.04 b
2.26 6 0.48
8.27 6 0.65
1.42 6 0.12
3.81 6 0.2 b
0.04
0.37
0.26
0.08
0.0004
1070 6 155
438 6 36 a
785 6 59
56 6 7
796 6 134 b
0.37 (log)
0.005
0.67
0.14
0.002
Note. T, testosterone; T4, thyroxine. Mean 6 standard error of the mean.
*: n 5 4 rats/group.
**: n 5 10 rats/group.
***: p value for a one way anova.
****: Control n 5 6. Estradiol-17b, 0.02 mg/kg/day, n 5 10. PCB 153, 25 mg/kg/day, n 5 10. PCB 126, 0.1 mg/kg/day, n 5 7.
a,b,c
: One way ANOVA followed by Duncan’s multiple range test; means with different letters are significantly different ( p , 0.05).
164
DESAULNIERS ET AL.
TABLE 5
Histology of Thyroid Glands after Treatments
Treatment groups
Measurements
Epithelial height mm
Follicle area mm 2
Colloid density
(Luminescence unit)
CTRL (9) a
E2 (10)
PCB126 (9)
PCB153 (9)
11.1 6 0.2 b
(242) c
891.1 6 61.9
(144)
76.3 6 1.4
(899)
11.6 6 0.1
(346)
1140.6 6 121.4
(109)
79.9 6 1.3
(1043)
12.0 6 0.1
(311)
866.8 6 65.4
(128)
93.9 6 1.0*
(1049)
11.4 6 0.1
(370)
827.4 6 66.7
(131)
89.8 6 1.5
(895)
Percent area of
colloid occupied
with vesicles
Colloid vacuolation
0%:
,20%:
20–50%:
.50%:
Foamy follicles
Number of follicles (%)
50.6 6 6.3
13.4 6 2.4
5.2 6 1.3
4.5 6 1.4
26.4 6 4.6
56.8 6 5.6
12.3 6 2.2
4.1 6 1.3
1.7 6 0.7
25.2 6 3.9
56.4 6 10.2
11.8 6 2.7
2.3 6 0.8
1.2 6 0.5
28.2 6 9.9
56.2 6 5.5
17.2 6 3.1
1.5 6 0.5
3.0 6 2.1
22.0 6 4.7
* Significantly different from control ( p 5 0.048). Analyzed by ANOVA and Duncan’s multiple range test with the rat nested in the treatment group as random
error (Littell et al., 1991).
a
Number of rats per group.
b
Mean 6 standard error.
c
Number of observations.
graphic regions or isoform groups, is presented in Figure 2 for
each treatment group,. It indicates that approximately 70% of
the total FSH eluted between 1–2 M NaCl. Two way ANOVA
over the first eight isoform groups (treatment p 5 0.0001;
isoform p 5 0.0001; interaction p . 0.05), followed by a
Duncan’s multiple comparison test among treatment groups,
demonstrated that a lower amount of FSH eluted in E2 and
PCB 153-treated rats ( p , 0.05) than in control and PCB
126-treated rats (this can be observed from the asterisks beside
the legends and the inset of Fig. 2). The data is presented as a
percentage of the hormone eluted during the whole chromatographic run (Fig. 2); thus, a treatment decreasing the amount of
FSH over the first eight isoform groups will necessarily increase the amount of FSH in group 9. This effect is demonstrated by a significant treatment 3 isoform group interaction
( p 5 0.02) when the amount of FSH eluted over the first eight
isoform groups is compared with that of the ninth group (inset
Fig. 2). The other effects in the model were isoform group
( p 5 0.0001) and treatment ( p . 0.05). No effects could be
detected for the TSH isoforms; however, the chromatofocusing
technique was only optimized for the separation of FSH isoforms, not TSH isoforms.
DISCUSSION
The effects of acute exposure to PCB 126 were assessed by
measuring the dose response on serum and pituitary hormone
levels in intact rats. In order to search for more sensitive
indicators of endocrine effects, hormones were also measured
in hemicastrated rats, and the pituitary isoforms of FSH and
TSH were studied. Our results demonstrate that circulating LH
and thyroxine concentrations in the intact adult male rat were
the most easily obtainable and sensitive endocrine indicators of
PCBs 126 and 153 exposure. The approaches of studying
hemicastrated rats and the pituitary isoforms were useful in
further characterizing the effects of PCB 126 and PCB 153.
PCB-126 treatment decreased thyroxine (Tables 2, 3, and 4)
and T3 concentrations in a dose related manner (Table 2).
Despite this, there were no feedback responses leading to a
dose-response increase in TSH concentration (Table 2). This is
not an unusual observation in PCB-treated rats (Barter and
Klaassen, 1994; Liu et al., 1995; Morse et al., 1996). Therefore, this leads to the suggestion that, in addition to interfering
with the thyroid gland, thyroid hormone transport, and metabolism (Brower et al., 1998), PCBs would also interfere with the
hypothalamo-pituitary axis (Brower, 1998). Exposure to coplanar PCBs or to 2,3,7,8-tetrachlorodibenzodioxin is known to
alter TSH concentrations (Kohn et al., 1996; Seo et al., 1995)
but serum TSH response is not always predictable (Capen,
1995; Morse et al., 1996). The slight increase in TSH induced
by 100 mg/kg/day of PCB 126 (Tables 2 and 3) contrasts with
the non-significant reduction in TSH concentration observed in
Table 4. Studying the endocrine events through time following
ENDOCRINE EFFECTS OF PCBs 126 AND 153
acute exposure to PCBs could provide the information which
would explain the apparent inconsistencies (Tables 2 and 3 vs.
Table 4) in the trends of the TSH responses.
Both estradiol and PCB 153 increased thyroxine concentrations (Table 4). This effect of estradiol could be explained by
the fact that estradiol is known to regulate thyroid-releasing
hormone (TRH) gene expression (Croissandeau et al., 1996),
59-iodothyronine deiodinase activities (Lisboa et al., 1997),
up-regulates the TRH receptor (Kimura et al., 1994) and, with
striking efficiency, decreases the activity of the adenohypophyseal TRH-degrading ectoenzyme, which is restored in 24 – 48 h
following the injection (Schomburg and Bauer, 1997). Estrogenic xenobiotics are known to alter thyroid hormones in male
rats (Gray Jr. et al., 1989; Li and Hansen, 1997), in accordance
with the PCB 153-induced increase in thyroxine concentrations
(Table 4), which might be attributed to its estrogenic properties
(Li et al., 1994). In accordance with our observations, PCB 153
administration to pregnant rats (64 and 16 mg/kg/day on days
10 –16 of gestation) slightly increased thyroxine concentration
at weaning ( p 5 0.058, Ness et al., 1993). Also, 5 mg/kg bw
per day of Aroclor 1254 on Days 10 to 16 of gestation induced
a small, but statistically significant increase in thyroxine concentration in the adult male offspring on postnatal day 90
(Morse et al., 1996). In that study, PCB 153 was a predominant
congener measured in the maternal and fetal plasma and in the
fetal brain. In prepubertal female rats PCB 153 slightly increased thyroxine concentration at low dose of exposure preceded by a slight decline with increasing doses (Li et al.,
1994). Despite this, using experimental models different from
ours, PCB 153 has usually been found to decrease thyroxine
concentration (Ness et al., 1993; Morse et al., 1996; van
Birgelen et al., 1992).
The histological analysis of the thyroid glands only revealed
that the colloid is less dense in PCB 126-treated rats than in the
controls (Table 5), suggesting mobilization of glandular thyroid hormones to delay systemic hypothyroidism (Hansen et
al., 1995) facilitated by coplanar PCB activated hepatic enzymes (Chu et al., 1994; Desaulniers et al., 1997; Kohn et al.,
1996; Seo et al., 1995). Important changes in thyroid histology
are usually associated with significant changes in TSH levels
(Liu et al., 1995), so our histological observations are in
accordance with the absence of statistically significant changes
in TSH concentrations (Table 4). Collectively, the indicators of
thyroid function from the PCBs 126- and 153-treated rats
support the idea that measurement of the thyroxine concentration is the most sensitive endocrine indicator of PCB exposure
(Gray Jr. et al., 1993) and is more rapid than histological
assessment of the thyroid gland.
Although the hemicastrated rat model was unsuccessful at
increasing the sensitivity of hormone measurements to assess
endocrine-disrupting effects, the combined results from the
PCB 126-intact and -hemicastrated rats (Tables 2 and 3) suggest that PCB 126 affected the endocrinology of the reproductive system by acting at both the hypothalamo-pituitary axis
165
and the testicle. Although LH concentrations in intact PCB
126-treated rats were significantly reduced and testosterone
concentrations were not, both hormones showed a decrease.
This suggests that the hypothalamo-pituitary axis is a site of
PCB-126 toxic insult and that PCB 126 may have had little
effect on testicular responsiveness to LH stimulation. The low
testosterone concentrations despite slightly elevated LH levels
in hemicastrated PCB 126-treated rats, suggest a decrease in
testicular responsiveness to LH stimulation, similar to that
previously characterized in TCDD-treated male rats (Cooke et
al., 1998; Moore et al., 1985; Ruangwises et al., 1991; ). In
contrast to the absence of TSH response to the decrease in
thyroxine (Table 2), the hemicastrated rat model suggests that
the PCB-126 treatment did not inhibit the hormonal feedback
regulatory system of the hypothalamo-pituitary-testicular axis.
Indeed, the PCB-126 treatment decreased LH secretion in
intact rats (Table 2), but it was unable to reduce the LH serum
concentrations in PCB-treated, hemicastrated rats, probably
because this latter group had the lowest concentration of testosterone (Tables 2 vs 3), and thus, a reduced negative feedback regulation of LH secretion at the hypothalamo-pituitary
level. Also, the PCB-126 treatment did not prevent a compensatory increase in FSH, which resulted from the diminished
negative regulatory feedback induced by lower concentrations
of inhibin created by the removal of one testicle. The hemicastrated rat offers a useful in vivo model for the characterization of toxic insults to the hypothalamo-pituitary-testicular
axis. To improve the use of this in vivo model in reproductive
toxicology, detailed time course studies of the endocrine events
following hemicastration must be done to identify the important time points at which the relationship between FSH and
inhibin, and testosterone and LH, should be assessed.
The present study represents the first attempt to characterize
changes in isoforms of pituitary hormones in response to toxic
exposure. The small, but statistically significant, decreases in
the amount of FSH eluted during the pH gradient (Fig. 2;
isoform groups 1– 8) induced by estradiol and PCB 153, compared to control and PCB 126-treated rats, suggest that a small
percentage of the FSH molecules in the pituitary gland were
altered, causing them to attach more tightly to the HPLC
column. This led to an increase in the proportion of FSH
isoforms eluted under the high NaCl concentration gradient
(1–2 M), which were confirmed by other investigators to be the
more acidic isoforms (Keel and Schanbacher, 1987). In other
species, the effects of estradiol on circulating hormone and
FSH isoforms have been reported to be similar to those documented here. In men, estradiol treatment reduced circulating
LH and testosterone concentrations (Cemeroglu et al., 1997;
Veldhuis and Dufau, 1993) and in castrated rams, exogenous
estrogen induced a marked increase in the acidic forms of
ovine pituitary FSH eluting during the NaCl peak (Keel and
Schanbacher, 1987). Both PCBs 126 and 153 decreased circulating LH concentrations (Tables 2 and 4), and yet, only PCB
153 changed the distribution of FSH isoforms (Fig. 2). The
166
DESAULNIERS ET AL.
explanation might involve different mechanisms through
which PCBs exert their effects. PCB 153 and PCB 126 are
known to induce different tissue responses, in particular, on the
activation of hepatic enzymes (Table 3; Li and Hansen, 1997),
and also on intracellular Ca21 homeostasis (Seegal et al.,
1990; Shain et al., 1991; Kodavanti et al., 1993; Maier et al.,
1994; Wong et al., 1997), which highly controls gonadotropinreleasing hormone (GnRH) neurons (Conn, 1996). GnRH is a
major regulator of gonadotropin production (Evans et al.,
1996), and of the secretion of specific LH and FSH isoforms in
humans (Matikainen et al., 1992; Phillips and Wide, 1994;
Zambrano et al., 1995), but not in all animals (Hassing et al.,
1993). The approach of studying FSH isoforms, although labor
intensive, could be more sensitive than measuring the circulating and the pituitary levels of FSH, since they were not
significantly affected by PCB 153 exposure (Table 4). Alteration of the distribution of FSH pituitary isoforms suggests
another level of endocrine disruption where the quality, rather
than the quantity, of FSH was affected. Whether the slight
changes in the relative proportions of the FSH pituitary isoforms reflect changes in immunoreactive FSH fragments
within the pituitary, or if these changes lead to significant and
biologically relevant alterations in the circulating isoforms,
remains to be investigated.
The measurement of the pituitary gland content of the hormones provided additional insights for the mechanisms of the
hormonal effects. The increased pituitary content of LH (even
at the smallest dose of PCB 126) and FSH in PCB 126-treated
rats (Table 2), associated with lower circulating levels, supports PCB 126-induced alteration of the secretory mechanisms
of the gonadotroph cells. The lower pituitary content of FSH in
estradiol treated rats (Table 4), in the absence of an increased
circulatory level, suggests a decreased synthesis of FSH. Estradiol is also known to increase the transcription and secretion
of PRL (Lamberts and MacLeod, 1990) and indeed, estradiol
treatment increased pituitary PRL content in the present study
(Table 4), as did the synthetic estrogen, diethylstilbestrol, when
administered to immature female rats (Wade et al., 1997).
In conclusion, estradiol, PCB 126 and PCB 153 alter several
endocrine parameters, probably, through different mechanisms. All three compounds decreased LH secretion, whereas
only estradiol and PCB 153 slightly altered FSH isoforms. The
mechanism of action of estradiol could be dissociated from that
of PCB 153, since estradiol decreased FSH pituitary content
and increased pituitary prolactin, but, in contrast to both PCBs,
it had no effect on hepatic enzymes. In regard to thyroid
function, PCB 126 decreased thyroxine concentration, whereas
both estradiol and PCB 153 slightly increased thyroxine concentration. The use of the hemicastrated rat model and the
study of pituitary FSH isoforms were labor-intensive and not
useful in providing more sensitive indicators of endocrine
disruption, yet they permitted further characterization of the
effects of these xenobiotics. The PCB 126-treated, hemicastrated rat model supported a testicular deficiency in PCB 126-
treated rats, whereas the slight changes in the elution pattern of
FSH pituitary isoforms suggest another level of endocrine
disruption and an estrogen-like effect of PCB 153. Finally,
serum LH and thyroxine appeared to be the only parameters
altered by all three compounds. Thus, because of the regular
occurrence and the sensitivity of these easily obtainable measurements, these results further support the measurement of
serum LH and thyroxine as useful indicators of endocrine
dysfunction, following acute exposure to xenobiotics, in adult
male rats.
ACKNOWLEDGMENTS
The authors are grateful to Dr. V. Seligy and Dr. G. Cooke for their essential
comments in the preparation of this manuscript; to W. Phan and C. Ulloa who
contributed to some parts of this project as co-op students; to L. Casavant for
excellent technical assistance; and to the National Hormone and Pituitary
Program, the National Institute of Diabetes and Digestive and Kidney Diseases
and to Dr. A. F. Parlow for providing assay reagents. This project was funded
by Health Canada.
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