Mercury causes degradation of spatial cognition in a model songbird species.

Mercury is a widespread pollutant of increasing global concern that exhibits a broad range of deleterious effects on organisms, including birds. Because the developing brain is well-known to be particularly vulnerable to the neurotoxic insults of mercury, many studies have focused on developmental effects such as on the embryonic brain and resulting behavioral impairment in adults. It is not well understood how the timing of exposure, for example exclusively in ovo versus throughout life, influences the impact of mercury. Using dietary exposure to environmentally relevant methylmercury concentrations, we examined the role that timing and duration of exposure play on spatial learning and memory in a model songbird species, the domesticated zebra finch (Taeniopygia guttata castanotis). We hypothesized that developmental exposure was both necessary and sufficient to disrupt spatial memory in adult finches. We documented profound disruption of memory for locations of hidden food at two spatial scales, cage- and room-sized enclosures, but found that both developmental and ongoing adult exposure were required to exhibit this behavioral impairment. Methylmercury-exposed birds made more mistakes before mastering the spatial task, because they revisited unrewarded locations repeatedly even after discovering the rewarded location. Contrary to our prediction, hippocampal volume was not affected in birds exposed to methylmercury over their lifetimes. The disruption of spatial cognition that we detected is severe and would likely have implications for survival and reproduction in wild birds; however, it appears that individuals that disperse or migrate from a contaminated site might recover later in life if no longer exposed to the toxicant.

Developmental arsenic exposure induces dysbiosis of gut microbiota and disruption of plasma metabolites in mice.

Arsenic is a notorious environmental pollutant. Of note, developmental arsenic exposure has been found to increase the risk of developing a variety of ailments later in life, but the underlying mechanism is not well understood. Many elements of host health have been connected to the gut microbiota. It is still unclear whether and how developmental arsenic exposure affects the gut microbiota. In the present study, we found that developmental arsenic exposure changed intestinal morphology and increased intestinal permeability and inflammation in mouse pups at weaning. These alterations were accompanied by a significant change in gut microbiota, as evidenced by considerably reduced gut microbial richness and diversity. In developmentally arsenic-exposed pups, the relative abundance of Muribaculaceae was significantly decreased, while the relative abundance of Akkermansia and Bacteroides was significantly enhanced at the genus level. Metabolome and pathway enrichment analyses indicated that amino acid and purine metabolism was promoted, while glycerophospholipid metabolism was inhibited. Interestingly, the relative abundance of Muribaculaceae and Akkermansia showed a strong correlation with most plasma metabolites significantly altered by developmental arsenic exposure. These data indicate that gut microbiota dysbiosis may be a critical link between developmental arsenic exposure and metabolic disorders and shed light on the mechanisms underlying increased susceptibility to diseases due to developmental arsenic exposure.

Bone manganese is a sensitive biomarker of ongoing elevated manganese exposure, but does not accumulate across the lifespan.

Studies have established associations between environmental and occupational manganese (Mn) exposure and executive and motor function deficits in children, adolescents, and adults. These health risks from elevated Mn exposure underscore the need for effective exposure biomarkers to improve exposure classification and help detect/diagnose Mn-related impairments. Here, neonate rats were orally exposed to 0, 25, or 50 mg Mn/kg/day during early life (PND 1-21) or lifelong through âˆ¼ PND 500 to determine the relationship between oral Mn exposure and blood, brain, and bone Mn levels over the lifespan, whether Mn accumulates in bone, and whether elevated bone Mn altered the local atomic and mineral structure of bone, or its biomechanical properties. Additionally, we assessed levels of bone Mn compared to bone lead (Pb) in aged humans (age 41-91) living in regions impacted by historic industrial ferromanganese activity. The animal studies show that blood, brain, and bone Mn levels naturally decrease across the lifespan without elevated Mn exposure. With elevated exposure, bone Mn levels were strongly associated with blood Mn levels, bone Mn was more sensitive to elevated exposures than blood or brain Mn, and Mn did not accumulate with lifelong elevated exposure. Elevated early life Mn exposure caused some changes in bone mineral properties, including altered local atomic structure of hydroxyapatite, along with some biomechanical changes in bone stiffness in weanlings or young adult animals. In aged humans, blood Mn ranged from 5.4 to 23.5 ng/mL; bone Mn was universally low, and decreased with age, but did not vary based on sex or female parity history. Unlike Pb, bone Mn showed no evidence of accumulation over the lifespan, and may not be a biomarker of cumulative long-term exposure. Thus, bone may be a useful biomarker of recent ongoing Mn exposure in humans, and may be a relatively minor target of elevated exposure.

Persistent autism-relevant behavioral phenotype and social neuropeptide alterations in female mice offspring induced by maternal transfer of PBDE congeners in the commercial mixture DE-71.

Polybrominated diphenyl ethers (PBDEs) are ubiquitous persistent organic pollutants (POPs) that are known neuroendocrine disrupting chemicals with adverse neurodevelopmental effects. PBDEs may act as risk factors for autism spectrum disorders (ASD), characterized by abnormal psychosocial functioning, although direct evidence is currently lacking. Using a translational exposure model, we tested the hypothesis that maternal transfer of a commercial mixture of PBDEs, DE-71, produces ASD-relevant behavioral and neurochemical deficits in female offspring. C57Bl6/N mouse dams (F0) were exposed to DE-71 via oral administration of 0 (VEH/CON), 0.1 (L-DE-71) or 0.4 (H-DE-71) mg/kg bw/d from 3 wk prior to gestation through end of lactation. Mass spectrometry analysis indicated in utero and lactational transfer of PBDEs (in ppb) to F1 female offspring brain tissue at postnatal day (PND) 15 which was reduced by PND 110. Neurobehavioral testing of social novelty preference (SNP) and social recognition memory (SRM) revealed that adult L-DE-71 F1 offspring display deficient short- and long-term SRM, in the absence of reduced sociability, and increased repetitive behavior. These effects were concomitant with reduced olfactory discrimination of social odors. Additionally, L-DE-71 exposure also altered short-term novel object recognition memory but not anxiety or depressive-like behavior. Moreover, F1 L-DE-71 displayed downregulated mRNA transcripts for oxytocin (Oxt) in the bed nucleus of the stria terminalis (BNST) and supraoptic nucleus, and vasopressin (Avp) in the BNST and upregulated Avp1ar in BNST, and Oxtr in the paraventricular nucleus. Our work demonstrates that developmental PBDE exposure produces ASD-relevant neurochemical, olfactory processing and behavioral phenotypes that may result from early neurodevelopmental reprogramming within central social and memory networks.

Gene expression profiles of multiple brain regions in rats differ between developmental and postpubertal exposure to valproic acid.

We have previously reported that the valproic acid (VPA)-induced disruption pattern of hippocampal adult neurogenesis differs between developmental and 28-day postpubertal exposure. In the present study, we performed brain region-specific global gene expression profiling to compare the profiles of VPA-induced neurotoxicity between developmental and postpubertal exposure. Offspring exposed to VPA at 0, 667, and 2000 parts per million (ppm) via maternal drinking water from gestational day 6 until weaning (postnatal day 21) were examined, along with male rats orally administered VPA at 0, 200, and 900 mg/kg body weight for 28 days starting at 5 weeks old. Four brain regions-the hippocampal dentate gyrus, corpus callosum, cerebral cortex, and cerebellar vermis-were subjected to expression microarray analysis. Profiled data suggested a region-specific pattern of effects after developmental VPA exposure, and a common pattern of effects among brain regions after postpubertal VPA exposure. Developmental VPA exposure typically led to the altered expression of genes related to nervous system development (Msx1, Xcl1, Foxj1, Prdm16, C3, and Kif11) in the hippocampus, and those related to nervous system development (Neurod1) and gliogenesis (Notch1 and Sox9) in the corpus callosum. Postpubertal VPA exposure led to the altered expression of genes related to neuronal differentiation and projection (Cd47, Cyr61, Dbi, Adamts1, and Btg2) in multiple brain regions. These findings suggested that neurotoxic patterns of VPA might be different between developmental and postpubertal exposure, which was consistent with our previous study. Of note, the hippocampal dentate gyrus might be a sensitive target of developmental neurotoxicants after puberty.

Transcriptional pathways linked to fetal and maternal hepatic dysfunction caused by gestational exposure to perfluorooctanoic acid (PFOA) or hexafluoropropylene oxide-dimer acid (HFPO-DA or GenX) in CD-1 mice.

Per- and polyfluoroalkyl substances (PFAS) comprise a diverse class of chemicals used in industrial processes, consumer products, and fire-fighting foams which have become environmental pollutants of concern due to their persistence, ubiquity, and associations with adverse human health outcomes, including in pregnant persons and their offspring. Multiple PFAS are associated with adverse liver outcomes in adult humans and toxicological models, but effects on the developing liver are not fully described. Here we performed transcriptomic analyses in the mouse to investigate the molecular mechanisms of hepatic toxicity in the dam and its fetus after exposure to two different PFAS, perfluorooctanoic acid (PFOA) and its replacement, hexafluoropropylene oxide-dimer acid (HFPO-DA, known as GenX). Pregnant CD-1 mice were exposed via oral gavage from embryonic day (E) 1.5-17.5 to PFOA (0, 1, or 5 mg/kg-d) or GenX (0, 2, or 10 mg/kg-d). Maternal and fetal liver RNA was isolated (N = 5 per dose/group) and the transcriptome analyzed by Affymetrix Array. Differentially expressed genes (DEG) and differentially enriched pathways (DEP) were obtained. DEG patterns were similar in maternal liver for 5 mg/kg PFOA, 2 mg/kg GenX, and 10 mg/kg GenX (R2: 0.46-0.66). DEG patterns were similar across all 4 dose groups in fetal liver (R2: 0.59-0.81). There were more DEGs in fetal liver compared to maternal liver at the low doses for both PFOA (fetal = 69, maternal = 8) and GenX (fetal = 154, maternal = 93). Upregulated DEPs identified across all groups included Fatty Acid Metabolism, Peroxisome, Oxidative Phosphorylation, Adipogenesis, and Bile Acid Metabolism. Transcriptome-phenotype correlation analyses demonstrated > 1000 maternal liver DEGs were significantly correlated with maternal relative liver weight (R2 >0.92). These findings show shared biological pathways of liver toxicity for PFOA and GenX in maternal and fetal livers in CD-1 mice. The limited overlap in specific DEGs between the dam and fetus suggests the developing liver responds differently than the adult liver to these chemical stressors. This work helps define mechanisms of hepatic toxicity of two structurally unique PFAS and may help predict latent consequences of developmental exposure.

Mouse Strain and Sex-Dependent Differences in Long-term Behavioral Abnormalities and Neuropathologies after Developmental Zika Infection.

Exposure of the developing fetus to Zika virus (ZIKV) results in a set of brain abnormalities described as the congenital Zika syndrome. Although microcephaly is the most obvious outcome, neuropathologies, such as intracranial calcifications and polymicrogyria, can occur in the absence of microcephaly. Moreover, the full impact of exposure on motor, social, and cognitive skills during development remains uncharacterized. We examined the long-term neurobehavioral consequences of neonatal ZIKV exposure in four genetically divergent inbred mouse strains (C57BL/6J, 129S1/SvImJ, FVB/NJ, and DBA/2J). Male and female mice were infected on postnatal day 1, considered comparable with exposure late in the second trimester of humans. We demonstrate strain differences in early susceptibility to the virus and the time course of glial reaction in the brain. These changes were associated with strain- and sex-dependent differences in long-term behavioral abnormalities that include hyperactivity, impulsiveness, and motor incoordination. In addition, the adult brains of susceptible mice exhibited widespread calcifications that may underlie the behavioral deficits observed. Characterization of the neuropathological sequelae of developmental exposure to the Zika virus in different immunocompetent mouse strains provides a foundation for identifying genetic and immune factors that contribute to long-term neurobehavioral consequences in susceptible individuals.SIGNIFICANCE STATEMENT Developmental Zika virus (ZIKV) infection is now known to cause brain abnormalities in infants that do not display microcephaly at birth, and the full impact of these more subtle neuropathologies has yet to be determined. We demonstrate in a mouse model that long-lasting behavioral aberrations occur after developmental ZIKV exposure. We compare four divergent mouse strains and find that the effects of Zika infection differ greatly between strains, in terms of behavioral changes, sex differences, and the intracranial calcifications that develop in the brains of susceptible mice. These findings provide a foundation for identifying susceptibility factors that lead to the development of abnormal behaviors secondary to ZIKV infection early in life.

Early postnatal manganese exposure causes arousal dysregulation and lasting hypofunctioning of the prefrontal cortex catecholaminergic systems.

Studies have reported associations between environmental manganese (Mn) exposure and impaired cognition, attention, impulse control, and fine motor function in children. Our recent rodent studies established that elevated Mn exposure causes these impairments. Here, rats were exposed orally to 0, 25, or 50 mg Mn kg-1  day-1 during early postnatal life (PND 1-21) or lifelong to determine whether early life Mn exposure causes heightened behavioral reactivity in the open field, lasting changes in the catecholaminergic systems in the medial prefrontal cortex (mPFC), altered dendritic spine density, and whether lifelong exposure exacerbates these effects. We also assessed astrocyte reactivity (glial fibrillary acidic protein, GFAP), and astrocyte complement C3 and S100A10 protein levels as markers of A1 proinflammatory or A2 anti-inflammatory reactive astrocytes. Postnatal Mn exposure caused heightened behavioral reactivity during the first 5-10 min intervals of daily open field test sessions, consistent with impairments in arousal regulation. Mn exposure reduced the evoked release of norepinephrine (NE) and caused decreased protein levels of tyrosine hydroxylase (TH), dopamine (DA) and NE transporters, and DA D1 receptors, along with increased DA D2 receptors. Mn also caused a lasting increase in reactive astrocytes (GFAP) exhibiting increased A1 and A2 phenotypes, with a greater induction of the A1 proinflammatory phenotype. These results demonstrate that early life Mn exposure causes broad lasting hypofunctioning of the mPFC catecholaminergic systems, consistent with the impaired arousal regulation, attention, impulse control, and fine motor function reported in these animals, suggesting that mPFC catecholaminergic dysfunction may underlie similar impairments reported in Mn-exposed children.

Environmental Endocrine Disruptors and Endometriosis.

As a consequence of industrialization, thousands of man-made chemicals have been developed with few undergoing rigorous safety assessment prior to commercial use. Ubiquitous exposure to these compounds, many of which act as endocrine-disrupting chemicals (EDCs), has been suggested to be one factor in the increasing incidence of numerous diseases, including endometriosis. Endometriosis, the presence of endometrial glands and stroma outside the uterus, is a common disorder of reproductive-age women. Although a number of population-based studies have suggested that exposure to environmental EDCs may affect a woman's risk of developing this disease, results of epidemiology assessments are often equivocal. The development of endometriosis is, however, a process occurring over time; thus, a single assessment of toxicant body burden cannot definitively be linked to causation of disease. For this reason, numerous investigators have utilized a variety of rodent models to examine the impact of specific EDCs on the development of experimental endometriosis. These studies identified multiple chemicals capable of influencing physiologic processes necessary for the establishment and/or survival of ectopic tissues in rodents, suggesting that these compounds may also be of concern for women. Importantly, these models serve as useful tools to explore strategies that may prevent adverse outcomes following EDC exposure.

Postnatal exposure to finasteride causes different effects on the prostate of male and female gerbils.

The development and maintenance of prostate function depend on a fine balance between oestrogen and androgen levels. Finasteride inhibits 5α-reductase, which is responsible for the conversion of testosterone into its most active form, dihydrotestosterone. Enzymes that metabolize these hormones have a highly relevant role in both the normal prostate metabolism and in the occurrence of pathological conditions. There are few studies on the impact of finasteride on male prostate development and fewer studies on the female prostate and possible intersexual differences. Therefore, we treated male and female gerbils from 7 to 14 days in postnatal life with a high dose of finasteride (500 µg/kg/day); the prostate complexes were then removed and submitted to immunohistochemistry, immunofluorescence and three-dimensional reconstruction. In addition, hormonal serum dosages were administered. Treatment with finasteride resulted in an increased thickness of the periductal smooth musculature in the prostate of both male and female gerbils, such as well as a reduction in the thickness of developing prostate alveoli in both sexes. In addition, intersexual differences were observed as increased epithelial proliferation and decreases in the number of developing alveoli in females. Together, the data indicate that postnatal exposure to finasteride causes greater changes in the female gerbil prostate than in the male.

Differences in larval pesticide tolerance and esterase activity across honey bee (Apis mellifera) stocks.

Honey bee populations in North America are an amalgamation of diverse progenitor ecotypes experiencing varying levels of artificial selection. Genetic differences between populations can result in variable susceptibility towards environmental stressors, and here we compared pesticide tolerances across breeding stocks using a mixture of seven pesticides frequently found in colonies providing pollination services. We administered the pesticide mixture chronically to in vitro reared larvae at four concentrations of increasing Hazard Quotient (HQ, or cumulative toxicity) and measured mortality during larval development. We found that different stocks had significantly different tolerances to our pesticide mixture as indicated by their median lethal toxicity (HQ50). The intensively selected Pol-Line stock exhibited the greatest pesticide sensitivity while Old World (progenitor) and putatively feral stocks were the most pesticide-tolerant. Furthermore, we found that activity of the detoxification enzyme esterase was positively correlated with pesticide tolerance when measured using two different substrate standards, and confirmed that larvae from the Pol-Line stock had generally lower esterase activity. Consistent with an increased pesticide tolerance, the Old World and putatively feral stocks had higher esterase activities. However, esterases and other detoxification enzymes (CYP450s and GSTs) were found in similar abundances across stocks, suggesting that the differences in enzyme activity we observed might arise from stock-specific single nucleotide polymorphisms or post-translational modifications causing qualitative variation in enzyme activity. These results suggest that selective breeding may inadvertently increase honey bees' sensitivity to pesticides, whereas unselected, putatively feral and Old World stocks have larvae that are more tolerant.

Prenatal exposure to bisphenol-A altered miRNA-224 and protein expression of aromatase in ovarian granulosa cells concomitant with elevated serum estradiol levels in F1 adult offspring.

This study was aimed to predict bisphenol-A (BPA)-responsive miRNA's using an in silico approach and to study their expression in granulosa cells of animals exposed prenatally to BPA. Pregnant Wistar rats were exposed to BPA through water (25 µg/L, 250 µg/L, and 2.5 mg/L) during gestation. The expression of miRNA-133b, miRNA-378 and miRNA-224 were analyzed in ovarian granulosa cells. BPA affected the postnatal developmental landmarks such as weight of the pups at birth and reduced anogenital distance. BPA exposed animals showed elevated serum estradiol (E2) levels, while follicle-stimulating hormone levels were reduced. The expression of miRNA-224 and aromatase protein levels were found to be increased. This preliminary finding reveals the impact of early life exposure to BPA on the long-term ovarian functions that may be mediated through miRNA-based granulosa cell response. Besides, it is also a compelling indicator for the subclinical response that could have important consequences on female fertility.

Organophosphorus pesticides can influence the development of obesity and type 2 diabetes with concomitant metabolic changes.

Widespread use and the bioaccumulation of pesticides in the environment lead to the contamination of air, water, soil and agricultural resources. A huge body of evidence points to the association between the pesticide exposure and increase in the incidence of chronic diseases, e.g. cancer, birth defects, reproductive disorders, neurodegenerative, cardiovascular and respiratory diseases, developmental disorders, metabolic disorders, chronic renal disorders or autoimmune diseases. Organophosphorus compounds are among the most widely used pesticides. A growing body of evidence is suggesting the potential interdependence between the organophosphorus pesticides (OPs) exposure and risk of obesity and type 2 diabetes mellitus (T2DM). This article reviews the current literature to highlight the latest in vitro and in vivo evidences on the possible influence of OPs on obesity and T2DM development, as well as epidemiological evidence for the metabolic toxicity of OPs in humans. The article also draws attention to the influence of maternal OPs exposure on offspring. Summarized studies suggest that OPs exposure is associated with metabolic changes linked with obesity and T2DM indicated that such exposures may increase risk or vulnerability to other contributory components.

Effects of cross-fostering and developmental exposure to mixtures of environmental contaminants on hepatic gene expression in prepubertal 21 days old and adult male Sprague-Dawley rats.

The notion that adverse health effects produced by exposure to environmental contaminants (EC) may be modulated by the presence of non-chemical stressors is gaining attention. Previously, our lab demonstrated that cross-fostering (adoption of a litter at birth) acted as a non-chemical stressor that amplified the influence of developmental exposure to EC on the glucocorticoid stress-response in adult rats. Using liver from the same rats, the aim of the current study was to investigate whether cross-fostering might also modulate EC-induced alterations in hepatic gene expression profiles. During pregnancy and nursing, Sprague-Dawley dams were fed cookies laced with corn oil (control, C) or a chemical mixture (M) composed of polychlorinated biphenyls (PCB), organochlorine pesticides (OCP), and methylmercury (MeHg), at 1 mg/kg/day. This mixture simulated the contaminant profile reported in maternal human blood. At birth, some control and M treated litters were cross-fostered to form two additional groups with different biological/nursing mothers (CC and MM). The hepatic transcriptome was analyzed by DNA microarray in male offspring at postnatal days 21 and 78-86. Mixture exposure altered the expression of detoxification and energy metabolism genes in both age groups, but with different sets of genes affected at day 21 and 78-86. Cross-fostering modulated the effects of M on gene expression pattern (MM vs M), as well as expression of energy metabolism genes between control groups (CC vs C). In conclusion, while describing short and long-term effects of developmental exposure to EC on hepatic transcriptomes, these cross-fostering results further support the consideration of non-chemical stressors in EC risk assessments.

Developmental Exposure to Endocrine Disruptors Expands Murine Myometrial Stem Cell Compartment as a Prerequisite to Leiomyoma Tumorigenesis.

Despite the high prevalence and major negative impact of uterine fibroids (UFs) on women's health, their pathogenesis remains largely unknown. While tumor-initiating cells have been previously isolated from UFs, the cell of origin for these tumors in normal myometrium has not been identified. We isolated cells with Stro1/CD44 surface markers from normal myometrium expressing stem cell markers Oct-4/c-kit/nanog that exhibited the properties of myometrial stem/progenitor-like cells (MSCs). Using a murine model for UFs, we showed that the cervix was a hypoxic "niche" and primary site (96%) for fibroid development in these animals. The pool size of these MSCs also responded to environmental cues, contracting with age and expanding in response to developmental environmental exposures that promote fibroid development. Translating these findings to women, the number of MSCs in unaffected human myometrium correlated with risk for developing UFs. Caucasian (CC) women with fibroids had increased numbers of MSCs relative to CC women without fibroids, and African-American (AA) women at highest risk for these tumors had the highest number of MSCs: AA-with fibroids > CC-with fibroids > AA-without fibroids > CC-without fibroids. These data identify Stro1+ /CD44+ MSCs as MSC/progenitor cell for UFs, and a target for ethnic and environmental factors that increase UF risk. Stem Cells 2017;35:666-678.

The environmental neurotoxin ß-N-methylamino-L-alanine inhibits melatonin synthesis in primary pinealocytes and a rat model.

The environmental neurotoxin ß-N-methylamino-L-alanine (BMAA) is a glutamate receptor agonist that can induce oxidative stress and has been implicated as a possible risk factor for neurodegenerative disease. Detection of BMAA in mussels, crustaceans, and fish illustrates that the sources of human exposure to this toxin are more abundant than previously anticipated. The aim of this study was to determine uptake of BMAA in the pineal gland and subsequent effects on melatonin production in primary pinealocyte cultures and a rat model. Autoradiographic imaging of 10-day-old male rats revealed a high and selective uptake in the pineal gland at 30 minutes to 24 hours after 14 C-L-BMAA administration (0.68 mg/kg). Primary pinealocyte cultures exposed to 0.05-3 mmol/L BMAA showed a 57%-93% decrease in melatonin synthesis in vitro. Both the metabotropic glutamate receptor 3 (mGluR3) antagonist Ly341495 and the protein kinase C (PKC) activator phorbol-12-myristate-13-acetate prevented the decrease in melatonin secretion, suggesting that BMAA inhibits melatonin synthesis by mGluR3 activation and PKC inhibition. Serum analysis revealed a 45% decrease in melatonin concentration in neonatal rats assessed 2 weeks after BMAA administration (460 mg/kg) and confirmed an inhibition of melatonin synthesis in vivo. Given that melatonin is a most important neuroprotective molecule in the brain, the etiology of BMAA-induced neurodegeneration may include mechanisms beyond direct excitotoxicity and oxidative stress.

Low-dose developmental exposure to bisphenol A induces sex-specific effects in bone of Fischer 344 rat offspring.

BACKGROUND: Bisphenol A (BPA) is a component of polycarbonate plastics to which humans are regularly exposed at low levels, and an endocrine disruptor with effects on several hormonal systems. Bone is a sensitive hormone target tissue, and we have recently shown that in utero and lactational exposure to 25µg BPA/kg BW/day alters femoral geometry in rat offspring. OBJECTIVE: To investigate bone effects in rat offspring after developmental exposure to a BPA dose in the range of human daily exposure (0.1-1.5µg/kg BW/day) as well as a dose to corroborate previous findings. METHODS: Pregnant Fischer 344 rats were exposed to BPA via drinking water corresponding to 0.5µg/kg BW/day: [0.5], (n=21) or 50µg/kg BW/day: [50], (n = 16) from gestational day 3.5 until postnatal day 22, while controls were given only vehicle (n = 25). The offspring was sacrificed at 5 weeks of age. Bone effects were analyzed using peripheral quantitative computed tomography (pQCT), the 3-point bending test, plasma markers of bone turnover, and gene expression in cortical bone and bone marrow. RESULTS: Compared to controls, male offspring developmentally exposed to BPA had shorter femurs. pQCT analysis revealed effects in the [0.5] group, but not in the [50] group; BPA reduced both trabecular area (-3.9%, p < 0.01) and total cross sectional area (-4.1%, p < 0.01) of femurs in the [0.5] group, whereas no effects were seen on bone density. Conversely, bone length and size were not affected in female offspring. However, the procollagen type I N-terminal propeptide (P1NP), a peptide formed during type 1 collagen synthesis, was increased in plasma (42%: p < 0.01) in female offspring exposed to [0.5] of BPA, although collagen gene expression was not increased in bone. The biomechanical properties of the bones were not altered in either sex. Bone marrow mRNA expression was only affected in male offspring. CONCLUSIONS: Developmental low-dose exposure to BPA resulted in sex-specific bone effects in rat offspring. A dose approximately eight times lower than the current temporary EFSA human tolerable daily intake of 4µg/kg BW/day, reduced bone length and size in male rat offspring. Long-term studies are needed to clarify whether the increased plasma levels of P1NP in female offspring reflect development of fibrosis.

Developmental toxicity and induction of vitellogenin in embryo-larval stages of zebrafish (Danio rerio) exposed to methyl Paraben.

MethylParaben (MP), a methyl ester of p-hydroxybenzoic acid, is used as an anti-microbial preservative in foods, drugs and cosmetics for decades. It enters the aquatic environment, and can have toxic effects on aquatic organisms. Little is known on the developmental toxicity of MP exposure to zebrafish during early life stages. In this study, the developmental effects of MP were evaluated in embryo-larval zebrafish (at concentrations ranging from 100µM, 200µM, 400µM, 800µM and 1000µM for 96h post fertilization (hpf). The survival, hatching, heart beat rate and developmental abnormalities were observed in the embryos exposed to MP. MP exposure resulted in decreased heart rate and hatching rate. Defects including pericardial edema blood cell accumulation and bent spine were observed in all the treated concentration, except at 100µM. With increasing concentrations, the frequency of these defects increased. The 96 hpf LC50 of MP was calculated to be 428µM (0.065mg/L). Furthermore, RT-PCR result showed that in larval zebrafish exposed to 100µM (0.015mg/L) of MP till 96 hpf, expression of vitellogenin I (Vtg -I) was significantly upregulated compared to the control group. This data suggest that even though lower concentrations of MP do not cause phenotypic malformations, it leads to dysregulated expression of estrogenic biomarker gene Vtg-I.

Prenatal Exposure to Bisphenol A Disrupts Mouse Fetal Liver Maturation in a Sex-Specific Manner.

Bisphenol A (BPA) is one of the most prevalent endocrine disrupting chemicals in the environment. Developmental exposure to BPA is known to be associated with liver dysfunction and diseases, such as hepatic steatosis, liver tumors, metabolic syndrome, and altered hepatic gene expression, and DNA methylation profiles. However, the effects of BPA on rodent liver development are unknown. The present study was undertaken to address this important question using the mouse as an experimental model. Pregnant mice were exposed to BPA via diet from embryonic day 7.5 (E7.5) to E18.5. At E18.5, fetal livers were collected, and analyzed for changes in the expression of key hepatocyte maturation markers. We found the following significant alterations in BPA-exposed female but not male fetal livers: (a) levels of the mature hepatocyte markers, albumin and glycogen synthase proteins, were decreased (-65% and -40%, respectively); (b) levels of the immature hepatocyte marker, α-fetoprotein, were increased (+43%); (c) the level of C/EBP-α protein, the master transcription factor essential for hepatocyte maturation, was down-regulated (-50%); and (d) the level of PCNA protein (marker of proliferation) was elevated (+40%), while that of caspase-3 protein and activity (markers of apoptosis) was reduced (-40% and -55%, respectively), suggestive of a perturbed balance between cell proliferation and apoptosis in BPA-exposed female fetuses. Taken together, these findings demonstrate that prenatal exposure to BPA disrupts the mouse fetal liver maturation in a sex-specific manner, and suggest a fetal origin for BPA-induced hepatic dysfunction and diseases.

Mammary Gland Evaluation in Juvenile Toxicity Studies: Temporal Developmental Patterns in the Male and Female Harlan Sprague-Dawley Rat.

There are currently no reports describing mammary gland development in the Harlan Sprague-Dawley (HSD) rat, the current strain of choice for National Toxicology Program (NTP) testing. Our goals were to empower the NTP, contract labs, and other researchers in understanding and interpreting chemical effects in this rat strain. To delineate similarities/differences between the female and male mammary gland, data were compiled starting on embryonic day 15.5 through postnatal day 70. Mammary gland whole mounts, histology sections, and immunohistochemically stained tissues for estrogen, progesterone, and androgen receptors were evaluated in both sexes; qualitative and quantitative differences are highlighted using a comprehensive visual timeline. Research on endocrine disrupting chemicals in animal models has highlighted chemically induced mammary gland anomalies that may potentially impact human health. In order to investigate these effects within the HSD strain, 2,3,7,8-tetrachlorodibenzo-p-dioxin, diethylstilbestrol, or vehicle control was gavage dosed on gestation day 15 and 18 to demonstrate delayed, accelerated, and control mammary gland growth in offspring, respectively. We provide illustrations of normal and chemically altered mammary gland development in HSD male and female rats to help inform researchers unfamiliar with the tissue and may facilitate enhanced evaluation of both male and female mammary glands in juvenile toxicity studies.