Endocrine effects of hexabromocyclododecane (HBCD) in a one-generation reproduction study in Wistar rats.

The brominated flame retardant (BFR) hexabromocyclododecane was tested in a one-generation reproduction assay in Wistar rats, enhanced for endocrine parameters. A solution of the compound in corn oil was mixed in the feed, targeting at dietary exposure of 0-0.1-0.3-1-3-10-30-100 mg/kg body weight/day (mkd) in parental rats during 10 (males) or 2 (females) weeks premating, during gestation and lactation, and in their F1 offspring from weaning until final necropsy. Effects were assessed in F1 animals. Livers of these animals showed increased HBCD concentrations, in a dose-dependent way. The trabecular bone mineral density of the tibia was dose-dependently decreased in females (BenchMark Dose Lower confidence bound, BMDL=0.056 mkd). The IgG response after immunization with sheep red blood cells (SRBC) was increased in males (BMDL=0.46 mkd). Further sensitive effects were decreased weight of the testis (BMDL=1.5 mkd), increased fraction of neutrophilic granulocytes (BMDL=7.7 mkd), decreased concentration of apolar retinoids in female livers (BMDL=1.3 mkd), and decreased plasma alkaline phosphatase in females (BMDL=8.6 mkd). CYP19/aromatase activity in the ovary was correlated to the concentration of gamma-HBCD in the liver. A developmental origin of these effects is considered, and this is also true for sensitive effects observed in neurobehavioural testing in littermates from the same experiment, i.e. in the brainstem auditory evoked potentials and in a catalepsy test [Lilienthal, H., Van der Ven, L.T.M., Piersma, A.H., Vos, J.G. Neurobehavioral effects of the brominated flame retardant hexabromocyclododecane (HBCD) in rats after pre- and postnatal exposure, in press]. The low BMDLs of these effects may raise concern for human health, particularly when based on body burdens of HBCD, which leads to critical margins of exposure particularly for the occupational setting.

Effects of the brominated flame retardant hexabromocyclododecane (HBCD) on dopamine-dependent behavior and brainstem auditory evoked potentials in a one-generation reproduction study in Wistar rats.

Hexabromocyclododecane (HBCD) is a widely used brominated flame retardant which has been recently detected in many environmental matrices. Data from a subacute toxicity study indicated dose-related effects particularly on the pituitary thyroid-axis and retinoids in female rats. Brominated and chlorinated aromatic hydrocarbons are also reported to exert effects on the nervous system. Several investigations revealed a pronounced sensitivity of the dopaminergic system and auditory functions to polychlorinated biphenyls. Therefore, the present experiment should examine, whether or not HBCD affects these targets. Rats were exposed to 0, 0.1, 0.3, 1, 3, 10, 30 or 100 mg HBCD/kg body weight via the diet. Exposure started before mating and was continued during mating, gestation, lactation, and after weaning in offspring. Haloperidol-induced catalepsy and brainstem auditory evoked potentials (BAEPs) were used to assess dopamine-dependent behavior and hearing function in adult male and female offspring. On the catalepsy test, reduced latencies to movement onset were observed mainly in female offspring, indicating influences on dopamine-dependent behavior. The overall pattern of BAEP alterations, with increased thresholds and prolonged latencies of early waves, suggested a predominant cochlear effect. Effects were dose-dependent with lower bounds of benchmark doses (BMDL) between < or =1 and 10 mg/kg body weight for both catalepsy and BAEP thresholds. Tissue concentrations at the BMDL values obtained in this study were 3-4 orders of magnitude higher than current exposure levels in humans.

A 28-day oral dose toxicity study in Wistar rats enhanced to detect endocrine effects of decabromodiphenyl ether (decaBDE).

Decabromodiphenyl ether (decaBDE) is a widely used brominated flame retardant, considered to be of low toxicity. However, previous toxicity studies applied exposure methods with low bioavailability of this compound, and the actual hazard of decaBDE for humans, which are environmentally exposed to decaBDE, may thus be underestimated in current risk assessments. The present 28 days oral toxicity study in Wistar rats was designed to facilitate detection of endocrine and immune modulating effects of decaBDE using an exposure protocol with improved bioavailability. A technical preparation of high purity decaBDE was thus tested by daily exposure through gavage with an emulsion of soy phospholipon/lutrol as a carrier. Most sensitive effect in males were increased weight of seminal vesicle/coagulation gland with BMDL of 0.2mg/kg bw/day and increased expression of hepatic CYP1A and CYP2B (BMDLs 0.5-0.7 mg/kg bw/day). In females the most sensitive effect was decreased activity of P450c17 (CYP17), which is a key enzyme in the androgen synthesis pathway, in adrenals (BMDL 0.18 mg/kg bw/day). These results suggest that decaBDE may represent an as yet unreported hazard for reproductive health.

A 28-day oral dose toxicity study enhanced to detect endocrine effects of a purified technical pentabromodiphenyl ether (pentaBDE) mixture in Wistar rats.

A 28-day subacute oral toxicity study was performed in Wistar rats with a purified preparation of the commercial pentabromodiphenyl ether (pentaBDE), DE-71. The applied OECD407 protocol was enhanced for endocrine and immune parameters, and to enable benchmark dose analysis. A vehicle control group and 7 dose groups were included, which received 0.27, 0.82, 2.47, 7.4, 22.2, 66.7 or 200 mg pentaBDE/kg bw/d (mkd). The liver appeared to be a key target organ, showing a marked increase of weight and centrilobular hepatocellular hypertrophy, probably due to the observed induction of P450 enzymes, notably CYP1A and CYP2B. A marked decrease of circulating total thyroxine (TT4) and an increase of plasma cholesterol were probably secondary to the liver effects. Furthermore, dose-dependently decreased weight of epididymis, seminal vesicles, and prostate, as well as sperm head deformities in males, and induction of CYP17 activity in adrenals in females were observed, all possibly related to anti-androgenic activity. Finally, we observed a substantial increase of large unstained cells in the blood and a decrease of apolar retinoids in the liver. All these effects had benchmark doses at the lower confidence bound (BMDL) in the low- or mid-dose range, but particular sensitive, potentially adverse effects were TT4 decrease (BMDLs 1.1 in males and 1.8 mkd in females), and decrease of hepatic apolar retinoids (BMDLs 0.5 mkd in males and 2.3 mkd in females). These results contribute to refinement of the hazard identification of pentaBDE and improved risk assessment of human exposure to this industrial chemical and environmental pollutant.

Exposure to tetrabromobisphenol A (TBBPA) in Wistar rats: neurobehavioral effects in offspring from a one-generation reproduction study.

Within the framework of an EU project on risk assessment of brominated flame retardants, TBBPA was studied for neurobehavioral effects in rats. To permit benchmark dose analysis, eight dose levels were chosen ranging from 0 to 3000mg/kg body weight. Exposure of parental rats started 10 and 2 weeks before mating in males and females, respectively, and was continued throughout mating, gestation and lactation. After weaning, exposure was continued in the offspring throughout life. Previous studies had indicated TBBPA-induced effects on thyroid hormones. Because of the known implication of thyroid hormones in neurodevelopment, the present experiments tested if TBBPA exposure affects thyroid-dependent neurobehavioral functions in offspring, such as auditory responses and conditioned fear. Sweet preference was included because of sex-specific effects in littermates. No statistically significant effects were found on context or cue conditioned fear or sweet preference. Auditory responses were examined with brainstem auditory evoked potentials (BAEPs) at approximately 50-110 days of age. BAEP thresholds and wave IV latency were increased in exposed female rats in the low frequency range. In male rats, thresholds were unaffected, but absolute latency of wave IV and interpeak latencies II-IV showed exposure-related increases at low frequencies. The outcome pattern suggests a predominant cochlear effect of TBBPA in females while in males neural effects are more apparent. According to benchmark analysis, the critical effect doses (CED) for prolongations of wave IV latency at 0.5kHz were in the range of 35-70mg/kg body weight with lower bounds (BMDL) of approximately 8mg/kg in males and females. The BMDL values for elevation of hearing thresholds in females were in the range of 1-40mg/kg body weight, depending on frequency. The benchmark doses for effects on the BAEP were similar to values for decreases in circulating thyroid hormones. The comparison of the exposure level at which the most sensitive effect was found with current human exposure levels yielded a margin of exposure of about 5, according to a recent risk assessment. Further investigations are needed to examine exposure pathways, fate in the body and effects of TBBPA.

Endocrine effects of tetrabromobisphenol-A (TBBPA) in Wistar rats as tested in a one-generation reproduction study and a subacute toxicity study.

Endocrine effects of the brominated flame retardant tetrabromobisphenol-A (TBBPA) were studied in a one-generation reproduction assay in Wistar rats via repeated dietary exposure, applying eight dose groups at 0-3-10-30-100-300-1,000-3,000 mg/kg body weight/day (mkd). This design enables dose-response analysis and calculation of benchmark doses (BMDL). This reproduction study was preceded by a 28-day repeat dose subacute toxicity study, at 0-30-100-300 mkd. Major effects in the reproduction study included decreased circulating thyroxine (T4) with BMDLs of 31 (m) and 16 (f) mkd, and increased weight of testis and male pituitary (BMDLs of 0.5 and 0.6 mkd). The hypothyroxinemia correlated to a cluster of developmental parameters including delayed sexual development in females, decreased pup mortality, and effects on brainstem auditory evoked potentials [Lilienthal, H., Verwer, C.M., Van der Ven, L.T.M., Piersma, A.H., Vos, J.G., 2008. Neurobehavioral effects of tetrabromobisphenol A (TBBPA) in rats after pre- and postnatal exposure. Toxicology]. A second cluster of parameters in F1 animals was correlated to increased testis weight, and included female gonad weight, endometrium height, CYP19/aromatase activity in the ovary, and plasma testosterone levels in males. These two correlation clusters suggest a dual action of TBBPA. The only effects in the subacute study were decreased circulating T4 and increased T3 levels in males (BMDLs 48 and 124mkd), and non-significant trends for these parameters in females, suggesting that the other effects in the reproduction study were induced during development. Combined with data of human exposure to environmental TBBPA, the margin of exposure for highly exposed populations can be calculated at 2.6, and current use of TBBPA may therefore be a matter of concern for human health.

A 28-day oral dose toxicity study enhanced to detect endocrine effects of hexabromocyclododecane in Wistar rats.

A 28-day repeated dose study in rats (OECD407) enhanced for endocrine and immune parameters was performed with hexabromocyclododecane (HBCD). Rats were exposed by daily gavage to HBCD dissolved in corn oil in 8 dose groups with doses ranging between 0 and 200 mg/kg bw per day (mkd). Evaluation consisted of dose-response analysis with calculation of a benchmark dose at the lower 95% one-sided confidence bound (BMDL) at predefined critical effect sizes (CESs) of 10-20%. The most remarkable findings were dose-related effects on the thyroid hormone axis, that is, decreased total thyroxin (TT4, BMDL 55.5 mkd at CES--10%), increased pituitary weight (29 mkd at 10%) and increased immunostaining of TSH in the pituitary, increased thyroid weight (1.6 mkd at 10%), and thyroid follicle cell activation. These effects were restricted to females. Female rats also showed increased absolute liver weights (22.9 mkd at 20%) and induction of T4-glucuronyl transferase (4.1 mkd at 10%), suggesting that aberrant metabolization of T4 triggers feedback activation of the thyroid hormone system. These effects were accompanied by possibly secondary effects, including increased cholesterol (7.4 mkd at 10%), increased tibial bone mineral density (> 49 mkd at 10%), both in females, and decreased splenocyte counts (0.3-6.3 mkd at 20%; only evaluated in males). Overall, female rats appeared to be more sensitive to HBCD than male rats, and an overall BMDL is proposed at 1.6 mkd, based on a 10% increase of the thyroid weight, which was the most sensitive parameter in the sequence of events.