Tributyltin impairs the reproductive cycle in female rats

Triorganotins are environmental contaminants, commonly used in antifouling agents for boats, that bioaccumulate and thus are found in mammals and humans due to ingestion of contaminated seafood diets. The importance of triorganotins as environmental endocrine disruptors and consequent reproductive toxicity in different animal models is well known; however, the adverse effects on reproductive cycle are less well understood. The potential reproductive toxicity of tributyltin (TBT) on regular reproductive cycling of female rats was examined. Wistar female rats (12 wk old, weighing approximately 230 g) were divided into two groups: control (vehicle, ethanol 0.4%) and tributyltin (100 ng/kg/d, 7 d/wk, for 16 d by gavage). Tributyltin significantly decreased the cycle regularity (%), duration of the reproductive cycle, the proestrus and diestrus phases, and number of epithelial cell in proestrus phase. TBT also increased the duration of metestrus and the number of cornified cells in this phase. Ovary weight and serum 17β-estradiol levels decreased markedly, accompanied by a significant increase in progesterone levels. Histological analysis showed apoptotic cells in corpus luteum and granulosa cells layer, with cystic follicles after TBT exposure. Tributyltin also elevated number of atretic follicles and corpoa lutea. The micronucleus (MN) test, using Chinese hamster ovary cells, demonstrated a concentration-dependent mutagenic effect of TBT, and at 2.0?×?10(-2)ng/ml most of the cells were nonviable. The toxic potential of TBT over the reproductive cycle may be attributed to changes found in the ovarian weight, unbalanced levels of sexual female hormones, and number of ovarian follicles and corpora lutea.     

Contaminant-associated disruption of vitamin A and its receptor (retinoic acid receptor alpha) in free-ranging harbour seals (Phoca vitulina)

Polychlorinated biphenyls (PCBs) have been associated with a number of toxic effects in marine mammals such as endocrine disruption and immunotoxicity that, in turn, are widely thought to have contributed to population level impacts including reproductive failure and outbreaks of disease. In this study, the dietary hormone vitamin A and expression levels of one of its receptors, retinoic acid receptor alpha (RARalpha), were used as biomarkers of PCB-associated health effects in harbour seals. Harbour seal pups (n=24) were live-captured in coastal British Columbia, Canada, and Washington State, USA, and sampled for whole blood (to obtain peripheral blood mononuclear cells, PBMCs) and blood plasma, as well as biopsies of blubber and skin. Concentrations of circulatory vitamin A (retinol) in plasma and stored vitamin A in blubber were negatively associated with blubber PCB concentrations (R=-0.518, p=0.013 and R=-0.645, p=0.009, respectively). However, vitamin A concentrations in skin, an important target tissue, remained constant, which likely reflects a compensatory transfer from blubber to maintain physiological functions. In addition, we characterized the harbour seal RARalpha, and investigated its expression levels as a potential biomarker in seals. RARalpha expression in blubber, but not on PBMCs, was elevated in more contaminated animals (R=0.580, p=0.009). This may represent a direct contaminant-related effect, or, a compensation for the contaminant-related disruption of (circulatory and/or blubber) hormone levels. Since vitamin A is critical to developmental, reproductive and immunological health, our observations of a contaminant-related disruption of its physiology in free-ranging seals may portend population level consequences. Vitamin A concentrations and RARalpha expression levels can therefore represent relevant and sensitive biomarkers of PCB-associated toxic effects in toxicological studies of marine mammals.     

In vitro fertilization of oocytes from polyovular follicles in mouse ovaries exposed neonatally to diethylstilbestrol.

In 35-day-old female ICR/JCL mice given 5 daily injections of 1 microgram diethylstilbestrol (DES) from the day of birth, a significantly higher incidence of polyovular follicles was found in the ovaries than in those of age-matched control mice. Gap junctions of granulosa cells of mature follicles in neonatally DES-exposed mice were larger than those of the controls. When stimulated by gonadotropins (PMSG and hCG) and caged with males, the number of tubal embryos in DES-exposed mice was less, but the rate of fertilization and development was not different compared to the controls. Division of oocytes collected from the ovaries of 40-day-old DES-exposed and control mice after stimulation of gonadotropins was examined 24 to 72 h after in vitro insemination to ascertain whether fertilization had occurred in oocytes from polyovular follicles. Seventy-seven % of oocytes from uniovular follicles of control mice developed up to 8-cell stage embryos following in vitro insemination; 66% of those from similar follicles of DES-exposed mice developed into the same stage. By contrast, only 47% of oocytes from polyovular follicles of DES-exposed mice showed the division up to 8-cell stage 72 h after insemination, indicating a significantly lower fertilization rate compared to the oocytes from uniovular follicles of control and DES-exposed mice. Without insemination, oocytes taken from the same pools in these experiments never divided during the period of manipulation and incubation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Lack of effect of dietary diethylstilbestrol on reproductive performance.

Groups of male or female mice were pretreated for 2 wk and 1 wk, respectively, with flesh (liver or muscle) diets prepared from steers. In one experiment diethylstilbestrol (DES) was added to the diet at 0.5 or 5.0 ppb. In a second experiment diets prepared from DES-implanted steer flesh (liver or muscle) were fed. Tissues used in the control diet and DES-added diets were from DES-free steers. The animals were allowed to mate and diets continued until the first litter was delivered. Increasing DES levels in either liver or muscle diets or flesh from DES-implanted steers resulted in no significant differences either in litter size or in the number of fertile male or female mice between the control group and experimental groups. The offspring from each litter were mated and showed no significant difference in their reproductive performance. No abnormalities were noted in any offspring.     

Effect of a bradykinin potentiating fraction, from venom of the Egyptian scorpion, Buthus occitanus, on the ovaries and endometrium of mice

An isolated fraction with bradykinin potentiating activity, from the venom of the Egyptian scorpion Buthus occitanus, was injected i.p. in female mice (35 days old). Five days after the injection of a single sublethal dose (1 micrograms/g), the number of primary multilaminar and secondary follicles of the ovaries was increased. Graafian follicles were observed in 50% of the treated mice but in none of the control mice. The number and size of the uterine glands and endometrium were increased in treated mice. Concomitantly, estradiol was increased in the circulating blood, while progesterone was within the normal limits. The enhancement of cellular growth by the isolated venom fraction with bradykinin potentiating activity, could be attributed to an enhancement of some growth factor(s) responsible for stimulating the ovarian follicles and uterus.     

MULTIGENERATIONAL STUDY OF THE EFFECTS OF CONSUMPTION OF PCB-CONTAMINATED CARP FROM SAGINAW BAY,LAKE HURON,ON MINK. 3. ESTROGEN RECEPTOR AND PROGESTERONE RECEPTOR CONCENTRATIONS,AND POTENTIAL CORRELATION WITH DIETARY PCB CONSUMPTION

Mink (Mustela vison) were fed diets containing ocean fish (control diet, 0.0 ppm polychlorinated biphenyls, PCBs) or Saginaw Bay carp to provide 0.25, 0.5, or 1.0 ppm PCBs to examine the effect of PCBs on homeostasis of binding sites for ovarian steroid hormones. Ranch-raised mink fed Great Lakes fish contaminated with PCBs, or treated with PCBs directly, have demonstrated reproductive impairment including anovulation, fetal resorption, delayed ovulation, increased gestation, and decreased litter size. Previous studies have demonstrated that estrogen and progesterone levels are unaltered in mink treated with PCBs, suggesting that the effect of PCBs on reproduction is not medi ated through alterations in hormone homeostasis. In vitro studies have demonstrated that the most likely means by which PCBs exert antiestrogenic ability is through a downregulation of the estrogen receptor in normally estrogen-responsive tissues such as liver and uterus. Hepatic and uterine estrogen binding site concentrations were measured in female mink consuming diets containing PCBs for up to 18 mo at up to 1 ppm. Hepatic estrogen binding site concentrations generally decreased with increasing dietary PCB concentrations. Uterine estrogen binding site concentration did not decrease in these animals. Uterine progesterone receptor concentration also did not change with increasing PCB consumption. In total, the response of hepatic and uterine estrogen and uterine progesterone binding sites in mink fed diets containing Saginaw Bay carp suggests that concentrations of PCBs available to uterine tissue may not have been sufficient to decrease uterine estrogen receptor, despite their effect on hepatic estrogen receptor.     

Selenium accumulation,reproductive status,and histopathological changes in environmentally exposed redear sunfish

Comparisons were made of the accumulation of selenium, histopathological damage, and reproductive status of redear sunfish (Lepomis microlophus) collected in July 1986 from Martin Lake (a contaminated site) and Lake Tyler (a reference site). Hepatic concentrations of selenium were four times higher in Martin Lake sunfish (7.6±0.5 ppm) than in fish from the reference lake (2.1±0.2 ppm). Redears collected from the contaminated lake had lower condition factors than individuals collected from the reference site. Sunfish with elevated levels of hepatic selenium had substantial alterations in the liver including necrosis, cytoplasmic vacuolation, and Kupffer cell proliferation. The ovaries of mature fish collected from Martin Lake frequently had atretic follicles, abnormally shaped follicles, connective tissue hypertrophy, asynchronous oocyte development, and an overall reduction in the number of developing oocytes. These histopathological changes in the ovaries of Martin Lake sunfish were not accompanied by alterations in gonadal steroid titers in the blood. No histopathological lesions could be detected in the testes of Martin Lake fish. Most of the males collected from the contaminated site were immature and had lower circulating levels of sex steroid hormones than reference males. The results show that tissue burdens of selenium have declined by 25% since this sunfish population was sampled last in 1981. Further, the results of this study indicate that the overall health and reproductive status of selenium-contaminated fish collected from Martin Lake is still seriously impaired.     

Chronic effects of agar,guar gum,gum arabic,locust-bean gum,or tara gum in F344 rats and B6C3F1 mice

Diets containing 25,000 (2.5%) or 50,000 ppm (5.0%) agar, guar gum, gum arabic, locust-bean gum or tara gum were fed to groups of 50 male and 50 female F344 rats and B6C3F₁ mice for 103 wk. Separate groups of 50 rats and 50 mice of each sex served as controls for each study. There were no significant differences in survival between any of the dosed groups of rats or mice and their respective control groups. Depressions in body-weight gain greater than 10% for dosed groups relative to their respective control groups were observed for male (low dose only) and female mice fed diets containing agar, female mice fed diets containing guar gum (high dose only), male mice fed diets containing locust-bean gum (high dose only) and male and female mice fed diets containing tara gum (high dose only). Depressions in body-weight gain greater than 5% were observed for female rats fed diets containing agar, guar gum or gum arabic. There were no histopathological effects associated with the administration of the test materials. Under the conditions of these bioassays, none of the five polysaccharides was carcinogenic for F344 rats or B6C3F₁ mice of either sex.     

Studies on the adaptive response in mouse female germ cells X-irradiated in vitro at two different stages of maturation

BACKGROUND: Radioadaptation is a phenomenon whereby cells exposed to a low dose of ionizing radiation are more resistant to a much higher dose delivered some time thereafter. This phenomenon could result from the activation of damage repair and/or antioxidant defense systems by the low dose. MATERIALS AND METHODS: The existence of a cytogenetic adaptive response in female germ cells was investigated using a recently developed in vitro system. Mouse ovarian follicles were cultured from an early preantral stage up to ovulation. The follicles were X-irradiated with either 2 or 4 Gy ("challenge dose") preceded or not by 50 mGy ("conditioning dose", 5 h earlier), on days 0 or 12 of the culture. Ovulated oocytes were collected on day 13, fixed and analyzed for the presence of chromosome aberrations. RESULTS: Irradiation with 2 or 4 Gy on days 0 or 12 did not influence ovulation but had dose-dependent effects on the germinal vesicle breakdown of the oocytes. It also caused dose-dependent chromosome damage, with a greater sensitivity of oocytes to this effect when irradiation occurred on day 12 than on day 0. Prior irradiation of oocytes with the dose of 50 mGy led to a reduction in the yield of chromosome aberrations when irradiation occurred on day 12 but not on day 0. CONCLUSION: These results suggest that pre-irradiation of mouse pre-ovulatory oocytes with a low conditioning dose could confer on them some protection against radiation-induced chromosomal damage by a subsequent challenge dose of a few Gy.     

The reversible effects of raloxifene on luteinizing hormone levels and ovarian morphology in mice

Raloxifene is a selective estrogen receptor modulator that has estrogen agonist effects on bone and serum lipids and estrogen antagonist effects on breast and uterine tissues. This study assessed the effects of raloxifene hydrochloride (HCl) treatment on circulating luteinizing hormone (LH) levels and ovarian morphology in sexually mature, 15-week-old, female CD-1 mice. Mice were maintained on diets providing average daily doses of 0 or 233 mg/kg raloxifene for 2 weeks (Study 1) or 0, 7.9, or 236 mg/kg raloxifene for 4 weeks (Study 2). At the end of the treatment period, blood samples were collected every 2 hours for 24 h in Study 1 (5 mice per group) and at 10:00 a.m. and 10:00 p.m. in Study 2 (8 mice per group). Serum LH levels were measured by radioimmunoassay. Ovarian histomorphology was evaluated in the 10 mice per group (Study 1) and the 8 mice per group (Study 2). For the reversibility phase (Study 2), mice were fed untreated diets for 3 weeks; serum LH levels and ovarian histomorphology were then assessed. Raloxifene treatment at 233 mg/kg/day for 2 weeks (Study 1) significantly elevated circulating LH levels by 4- to 7-fold compared with control. Raloxifene-treated mice had elevated LH levels sustained over the 24-h sampling period and did not exhibit the preovulatory LH surge evident in some control mice at the 4:00 p.m., 6:00 p.m., and 8:00 p. m. time points. Mice treated with 236 mg/day raloxifene for 4 weeks (Study 2) had elevated LH levels (4.4-fold compared to control), whereas mice exposed to 7.9 mg/kg/day raloxifene had a slight, nonsignificant increase in LH (2-fold compared to control). In both dose groups, LH levels were indistinguishable from controls 3 weeks after raloxifene treatment was discontinued. The ovaries in six of the eight mice treated with 7.9 mg/kg/day raloxifene had dilated and/or anovulatory follicles. One mouse in this group had a single hemorrhagic follicle; however, corpora lutea distribution was normal, indicating that ovulation was occurring. Raloxifene-treated mice in Study 1 and mice treated with a comparable raloxifene dose (236 mg/day) in Study 2 had histomorphological changes in the ovary indicative of arrested follicular maturation, including anovulatory hemorrhagic follicles, some developing follicles, and very few corpora lutea. At the end of the reversibility phase, hemorrhagic follicles were no longer evident and follicular maturation and corpora lutea distribution were normal. Raloxifene treatment in mice produces a dose-dependent, sustained elevation in serum LH levels and is associated with changes in ovarian follicular morphology. These changes are reversible upon discontinuation of raloxifene treatment.     

Genotoxic and embryotoxic effects of gonadotropin-hyperstimulated ovulation of murine oocytes, preimplantation embryos, and term fetuses.

Compared to spontaneous ovulation, gonadotropin-hyperstimulated ovulation (superovulation) in mice resulted in a fourfold increase in the number of preimplantation embryos 3 days post coitum, 50% of which died before term. Both in vitro development of embryos during the preimplantation period and transfer of morulae from superovulated females to pseudopregnant untreated foster mothers indicate that the prenatal loss occurring shortly before implantation up to term is due to maternal factors rather than to direct hormonal effects on oocytes or early embryos. Indeed, no genotoxic events could be observed in 4-cell to blastocyst stage embryos from superovulated female mice as revealed by the chromosomal aberration test and the sister chromatid exchange assay. Chromosome analysis of the pronuclei from mouse zygotes showed an increased rate of aberrations in oocyte-derived nuclei after superovulation in comparison to spontaneous ovulation. The present data suggest that aberrant murine oocytes may be fertilized, but they do not survive the first cleavage stages. The result is discussed with respect to the high incidence of chromosomal abnormalities found in human oocytes after gonadotropin-hyperstimulated ovulation.     

Chronic exposure to dibromoacetic acid, a water disinfection byproduct, diminishes primordial follicle populations in the rabbit.

To determine if dibromoacetic acid (DBA) affects ovarian folliculogenesis, four groups of female Dutch-belted rabbits were exposed daily to 0, 1, 5, or 50 mg DBA/kg body weight in drinking water beginning in utero from gestation day 15 throughout life. Functionality of the endocrine axis was assessed by measuring serum concentrations of gonadotropins following an im injection of 10 microg GnRH at 12 (prepubertal; n = 6/dose group) and 24 (postpubertal; n = 10/dose group) weeks of age. A day after GnRH challenge, number of ovulation sites and ovarian weights were determined at necropsy. Left ovaries were processed for histopathology, serially sectioned at 6 microm, and every twelfth section stained with hematoxylin and eosin was evaluated. All healthy follicles were categorized as primordial, primary, small preantral, large preantral, or small antral follicles. The area of each section evaluated was measured and the number of follicles in each category expressed per mm2 unit area. In prepubertal animals, DBA caused a reduction in number of primordial follicles (p < 0.05) and total healthy follicles (p < 0.05) at 50 mg/kg dose level. In adult animals, there were fewer primordial follicles in both the 5 (p < 0.01) and 50 (p = 0.1) mg/kg dose groups. No profound changes in gonadotropin profiles were observed. Although chronic exposure to DBA did not appear to have an effect on late follicular development or ovulation, DBA did reduce the population of primordial follicles. The long-term health consequences of diminished primordial follicles are unknown, but it is very likely that reproductive senescence would occur earlier.     

Evaluation of ovarian toxicity of mono-(2-ethylhexyl) phthalate (MEHP) using cultured rat ovarian follicles

Mono-(2-ethylhexyl) phthalate (MEHP) is the most toxic metabolite of di-(2-ethylhexyl) phthalate (DEHP). It has been reported that DEHP causes abnormal reproductive development in women, and suppresses estradiol synthesis and ovulation in female rats with diminished size of preovulatory follicles. The present study was conducted to evaluate the ovarian toxicity of MEHP using cultured rat ovarian follicles. Secondary follicles were isolated from the ovaries of 14-day-old female rats and cultured for 48 hr with MEHP (0, 10, 30, and 100 μg/ml). At 0, 24, and 48 hr of MEHP treatment, follicular diameters were measured. After the culture, viability and apoptosis of follicles were assessed, and progesterone, androstenedione, testosterone, and estradiol levels in culture media were measured. At 100 μg/ml, suppression of follicular development was observed, which is associated with decreased viability of follicles and apoptosis of granulosa cells. At this concentration, progesterone level increased markedly, whereas androstenedione, testosterone, and estradiol levels decreased. At 10 and 30 μg/ml, follicular development was not suppressed, no apoptotic change was observed, and the levels of all measured steroid hormones tended to increase. The combined levels of all steroid hormones increased at all concentrations of MEHP, and the increase implies that MEHP activates the synthetic pathway from cholesterol to estradiol including de novo synthesis of cholesterol. However, the progesterone/androstenedione ratio increased extremely at 100 μg/ml, and the increase implies that MEHP inhibits the conversion of progesterone to androstenedione. In conclusion, MEHP induces ovarian toxicity via suppression of follicular development and abnormal steroid hormone synthesis in cultured rat ovarian follicles.     

Effects of estrogenic compounds on neonatal oocyte development

In the mouse, oocytes develop in germline cysts that undergo breakdown resulting in primordial follicles, consisting of a single oocyte surrounded by granulosa cells. During this process, approximately two-thirds of the oocytes die. Exposure of female mice to environmental estrogens can alter oocyte development, limiting the number of primordial follicles that can be used for reproduction. Here we asked whether exposure to synthetic estrogens, diethylstilbestrol, ethinyl estradiol and bisphenol A affected perinatal oocyte development. Neonatal mice were injected with a low or high dose of each compound on postnatal days (PND) 1-4 and ovaries analyzed on PND5. Cyst breakdown, oocyte survival and follicle development were altered. The percentage of single oocyte was reduced from 84% in controls to 50-75%. The oocyte number per section was increased from 8 to 12-16. Follicle activation was reduced with 62% primordial follicles in controls to over 80% in most cases.     

Subchronic Feeding Study of the Mycotoxin Fumonisin B1 in B6C3F1 Mice and Fischer 344 Rats

Fumonisin B1 (FB1) is a mycotoxin produced by Fusarium moniliforme,a common fungus which occurs naturally on corn, and other Fusariumspecies. FB1 and other fumonisins are now recognized as havingpotentially important animal and human health implications.However, few toxicological data are currently available. Maleand female B6C3F1 mice and Fischer 344 rats were fed diets containing0, 1, 3, 9, 27, or 81 ppm FB1 (98% purity) for 13 weeks. Nodifferences in behavior or appearance, body weight or food consumptionbetween control and FB1-fed groups were found. In mice, hepatopathyand altered serum chemical profiles indicative of hepatotoxicitywere found in females fed the 81 ppm diet. No adverse effectswere found in female mice fed 27 ppm FB1 or in male mice atany dietary level studied. In rats, nephrosis involving theouter medulla was found in males fed 9 ppm and, to a lesserdegree, in females fed 81 ppm FB1, while decreased kidney weightwas found in both sexes at dietary levels 9 ppm FB1. Althoughthe liver is a target organ of FB1 in rats, hepatotoxicity wasnot found in rats fed diets containing up to 81 ppm FB1 for90 days. Thus, FB1 was toxic to both species following subchronicoral exposure, although significant interspecies differencesin the no observed effect levels and organ-specific responseswere found.     

Effects of a low oral dose of diethylstilbestrol (DES) on reproductive tract development in F1 female CD-1 mice

The synthetic estrogen diethylstilbestrol (DES) is a model to study the effects on female reproductive tract of endocrine disrupting chemicals interacting with estrogen receptors. Pregnant CD-1 mice were given daily by gavage 10microg/kg bw of DES (the lower range of therapeutic exposure) during gestational days 9-16, critical period for reproductive tract development. Parameters of sexual development were recorded after weaning and at sexual maturation. No signs of general toxicity were observed in dams. In DES-treated group, reduced litter weight during lactation and earlier vaginal patency was observed. Uterus weight was increased in F1 treated females at weaning. Histological analysis showed reduced endometrium thickness and increased polyovular follicles, irregular and oocytes with condensed chromatin in the ovary at sexual maturity. Prenatal DES oral administration induces subtle but significant effects on puberty onset, uterine and ovary morphology.     

Melatonin has dose-dependent effects on folliculogenesis, oocyte maturation capacity and steroidogenesis

Chemo and/or radiotherapy applied to young cancer patients most often have severe effects upon female fertility. Today, few options are available to protect ovarian function in females. However, these options are either ineffective, belong to the field of experimental research or/and are not applicable to all patients. Drugs that could protect the oocyte and its surrounding feeder cells from damage can be of great importance. Melatonin, being an important indirect antioxidant and a powerful direct free radical scavenger could be such a reagent. This paper reports the direct effects of different melatonin concentrations (range: 1 nM to 2 mM) on folliculogenesis and oogenesis of in vitro cultured mouse ovarian follicles. Early secondary mouse follicles were cultured in vitro for 12 days under different melatonin regimes. Every fourth day, survival rates were scored, follicles were morphologically evaluated and medium was collected for steroid analyses. On day 12, in vitro ovulation was induced by hCG/EGF. Eighteen hours later, oocytes were measured, oocyte maturation was evaluated and normality of spindle and chromosomes ascertained. Results obtained in this study indicated that 2mM melatonin is toxic. One mM negatively influenced oocyte maturation capacity. In the presence of 100 microM melatonin, androstenedione and progesterone were increased whereas estradiol was not influenced. Lower melatonin concentrations had no effect on the evaluated parameters. These data indicate an effect of melatonin on theca cell steroidogenesis. For prophylactic use, a dose of 10 microM could be suitable to reduce oxidative stress in cultured follicles.     

Effects of 3,3′,4,4′-Tetrachlorobiphenyl on Foetal Germ Cells in Two Mouse Strains after Repeated Treatment of the Dams During and After Pregnancy

Female mice from the C57/B1- and CBA/S-strain, respectively, were administered 3,3′4,4′-tetrachlorobiphenyl once a week for two weeks prior to mating, during mating, gestation and lactation, altogether at seven weekly occasions. The orally administered amounts were 9 or 15 mg/kg b.wt. per occasion. Male and female offspring (F1), exposed in utero were collected for histological analysis of the testes and ovaries. Other C57/Bl-females (F1) were tested for their reproductive capacity. In the C57/Bl-males tetrachlorobiphenyl only had a temporary effect on germ cell numbers, and spermatogenesis was normal again at 56 days after birth. In the ovaries 28 days after birth, an increased number of oocytes and follicles were observed, the effect being more pronounced in the C57/Bl-strain. The reproductive capacity expressed as mean litter size seemed not to be affected by the histologically observed changes. Concerning the number of litters per female there were indications of an enhanced productivity in tetrachlorobiphenyl-exposed F1-females. Malformations of any kind were not observed.     

2-Bromopropane causes ovarian dysfunction by damaging primordial follicles and their oocytes in female rats.

Ovarian dysfunction induced by 2-bromopropane (2-BP) has been described in female factory workers and experimental animals. However, the underlying mechanism is still unclear. To establish the reproductive target site and define mechanisms of 2-BP toxicity in adult female rats, we examined the effects of different doses and duration of exposure to 2-BP in female rats. In the dose-dependent experiments, female rats were exposed to 2-BP at 100, 300, or 1000 ppm or fresh air (n = 9 each) in exposure chambers for 8 h/day for 9 weeks. In the time-course experiments, female rats were exposed to 2-BP at 3000 ppm for 8 h (n = 7 each). The rats were then euthanized 1, 3, 5, and 17 days after exposure. Differential follicle counts and in situ terminal deoxynucleotidyl transferase assay were used to evaluate 2-BP effect on primordial, growing, and antral follicles. Exposure to 2-BP at 300 and 1000 ppm produced a significant reduction in the percentage of primordial, growing, and antral follicles in a dose-dependent manner. Significant reduction in the percentage of primordial follicles at 17 days after exposure was observed in time-course experiments. Exposure to 2-BP at 3000 ppm for 8 h resulted in histological changes in primordial follicles complex at 5 and 17 days after exposure. These changes consisted of distortion of the symmetry of oocytes and their nuclei at Day 5 after exposure and appearance of eccentric pyknotic cells and shrinkage of oocyte nuclei at Day 17 after exposure. In situ end labeling showed increased numbers of apoptotic oocytes and granulosa cells in primordial follicles at Days 5 and 17 after exposure. Our results suggested that ovarian dysfunction induced by 2-BP was caused by the destruction of primordial follicle and its oocyte due to the induction of apoptosis. Our studies also show that the follicle differential count is a more sensitive method than the vaginal smear in monitoring the female reproductive disorders induced by 2-BP.