Pubertal glyphosate-based herbicide exposure aggravates high-fat diet-induced obesity in female mice.

Glyphosate-based herbicides (GBH) are the most widely used pesticides globally. Studies have indicated that they may increase the risk of various organic dysfunctions. Herein, we verified whether exposure to GBH during puberty increases the susceptibility of male and female mice to obesity when they are fed a high-fat diet (HFD) in adulthood. From the 4th-7th weeks of age, male and female C57Bl/6 mice received water (CTL group) or 50 mg GBH /kg body weight (BW; GBH group). From the 8th-21st weeks of age, the mice were fed a standard diet or a HFD. It was found that pubertal GBH exposure exacerbated BW gains and hyperphagia induced by HFD, but only in female GBH-HFD mice. These female mice also exhibited high accumulation of perigonadal and subcutaneous fat, as well as reduced lean body mass. Both male and female GBH-HFD displayed hypertrophic white adipocytes. However, only in females, pubertal GBH exposure aggravated HFD-induced fat accumulation in brown adipocytes. Furthermore, GBH increased plasma cortisol levels by 80% in GBH-HFD males, and 180% in GBH-HFD females. In conclusion, pubertal GBH exposure aggravated HFD-induced obesity, particularly in adult female mice. This study provides novel evidence that GBH misprograms lipid metabolism, accelerating the development of obesity when individuals are challenged by a second metabolic stressor, such as an obesogenic diet.

Effects of bisphenol A on murine salivary glands and human tumor cell lines.

Bisphenol A (BPA) is an endocrine-disrupting chemical with a potential role in endocrine cancers. However, the effects of BPA on the salivary glands have been barely explored. We investigated the impact of in vivo sub-chronic exposure to BPA and its in vitro effects on human salivary gland mucoepidermoid carcinoma cell lines. Male and female mice were exposed to BPA (30 mg/kg/day). Sublingual and submandibular salivary glands from an estrogen-deficiency model were also analyzed. BPA concentration in salivary glands was evaluated by gas chromatography coupled to ion trap mass spectrometry. Immunohistochemical analysis using anti-p63 and anti-α-SMA antibodies was performed on mouse salivary gland tissues. Gene expression of estrogen receptors alpha and beta, P63 and α-SMA was quantified in mouse salivary gland and/or mucoepidermoid (UM-HMC-1 and UM-HMC-3A) cell lines. Cell viability, p63 and Ki-67 immunostaining were evaluated in vitro. BPA disrupted the tissue architecture of the submandibular and sublingual glands, particularly in female mice, and increased the expression of estrogen receptors and p63, effects that were accompanied by significant BPA accumulation in these tissues. Conversely, ovariectomy slightly impacted BPA-induced morphological changes. In vitro, BPA did not affect the proliferation of neoplastic cells, but augmented the expression of p63 and estrogen receptors. The present data highlight a potential harmful effect of BPA on salivary gland tissues, particularly in female mice, and salivary gland tumor cells. Our findings suggest that estrogen-dependent pathways may orchestrate the effects of BPA in salivary glands.

Environmental Health and Toxicology: Immunomodulation Promoted by Endocrine-Disrupting Chemical Tributyltin.

Tributyltin (TBT) is an environmental contaminant present on all continents, including Antarctica, with a potent biocidal action. Its use began to be intensified during the 1960s. It was effectively banned in 2003 but remains in the environment to this day due to several factors that increase its half-life and its misuse despite the bans. In addition to the endocrine-disrupting effect of TBT, which may lead to imposex induction in some invertebrate species, there are several studies that demonstrate that TBT also has an immunotoxic effect. The immunotoxic effects that have been observed experimentally in vertebrates using in vitro and in vivo models involve different mechanisms; mainly, there are alterations in the expression and/or secretion of cytokines. In this review, we summarize and update the literature on the impacts of TBT on the immune system, and we discuss issues that still need to be explored to fill the knowledge gaps regarding the impact of this endocrine-disrupting chemical on immune system homeostasis.

Efeitos do bisfenol A nas glândulas salivares de camundongos e em linhagem de células tumorais humanas / Effects of bisphenol A on murines salivary glands and human tumor cell lines

O Bisfenol A (BPA) é um produto químico desregulador endócrino com potencial papel em cânceres endócrinos. Entretanto, os efeitos do BPA sobre as glândulas salivares têm sido pouco explorados. Investigamos o impacto da exposição in vivo subcrônica ao BPA e seus efeitos in vitro nas linhagens celulares do carcinoma mucoepidermóide de glândula salivar. Camundongos machos e fêmeas foram expostos ao BPA (30.000 µg/kg/dia). Analisamos glândulas salivares sublinguais e submandibulares desses camundongos C56BL6 e camundongos fêmeas submetidos a ovariectomia. A concentração de BPA nas glândulas salivares foi avaliada por cromatografia gasosa acoplada à espectrometria de massa com armadilha de íons. A expressão dos receptores ERα e ERß foi avaliada em amostras de glândulas salivares de camundongos e linhagens celulares UM-HMC-1 e UM-HMC-3A pelo qRT-PCR, RNA mensageiro. Foram avaliadas in vitro, a viabilidade celular p63 e Ki-67, a imunoglobulina e a expressão de proteínas-alvo relacionadas às vias de sobrevivência/proliferação. O BPA prejudicou a morfologia das glândulas submandibulares e sublingual, particularmente nas fêmeas, e aumentou a expressão dos receptores de estrogênio, também acompanhado por um acúmulo significativo de BPA nestes tecidos. Ao contrário, a redução dos níveis de estrogênio pelo modelo de ovariectomia teve um leve impacto nas alterações morfológicas induzidas pela BPA. In vitro, a BPA não afetou a proliferação de células neoplásicas, mas aumentou a expressão dos receptores de p63 e estrogênio. A fosforilação ERK1/2 foi aumentada enquanto a fosforilação AKT e NF-κB foi reduzida. Os dados destacam um efeito nocivo do BPA sobre os tecidos das glândulas salivares e células tumorais. Os dependentes de estrogênio podem combinar o mecanismo de acúmulo de BPA, juntamente com a via de sinalização pró-sobrevivência e aumento da expressão da p63.

Bisphenol A (BPA) is an endocrine-disrupting chemical with potential role in endocrine cancers. However, BPA effects on the salivary glands have been barely explored. We investigated the impact of in vivo sub-chronic exposure to BPA and its in vitro effects on salivary gland mucoepidermoid carcinoma cell lines. Male and female mice were exposed to BPA (30,000 µg/kg/day). We analyzed sublingual and submandibular salivary glands from sham C56BL6 mice and that female ones underwent ovariectomy. Concentration of BPA in salivary glands was evaluated by gas chromatography coupled to ion trap mass spectrometry. Expression of ERα and ERß receptors was assessed in mouse salivary gland samples and cell lines UM-HMC-1 and UM-HMC3A by qRT-PCR, mARN. In vitro, cell viability p63 and Ki-67 immunostaining and expression of target proteins related to survival/proliferation pathways were evaluated. BPA impaired the architectural of the submandibular and sublingual glands, particularly in female mice, and increased the expression of estrogen receptors. This was accompanied by a significant accumulation of BPA in these tissues. Conversely, the reduction of estrogen levels by the ovariectomy model slightly impacted BPA-induced morphological changes. In vitro, BPA did not affect the proliferation of neoplastic cells, but increased the expression of p63 and estrogen receptors. ERK1/2 phosphorylation was increased while AKT and NF-κB phosphorylation was reduced. Data highlight a harmful effect of BPA on salivary gland tissues and tumorigenic cells. Estrogendependent may orchestrate the BPA-accumulation mechanism, along with pro-survival signaling pathway and increased p63 expression.

Effects of tributyltin (TBT) on the intermediate metabolism of the crab Callinectes sapidus.

This study investigated if the exposure to tributyltin (TBT), a chemical used worldwide in boat antifouling paints, could result in metabolic disturbances in the blue crab Callinectes sapidus. After the exposure to TBT 100 or 1000 ng.L-1 for 48 and 96 h, hemolymph and tissues were collected to determine the concentration of metabolites and lipid peroxidation. The levels of glucose, lactate, cholesterol, and triglycerides in the hemolymph were not affected by TBT exposure. Hemolymph protein and heart glycogen increased in the crabs exposed to TBT 1000 for 96 h. Anterior gills protein and lipoperoxidation decreased after 96 h in all groups. These results suggest that C. sapidus can maintain energy homeostasis when challenged by the TBT exposure for 48 h and that metabolic alterations initiate after 96 h.

Are the Effects of DES Over? A Tragic Lesson from the Past.

Diethylstilbestrol (DES), a transplacental endocrine-disrupting chemical, was prescribed to pregnant women for several decades. The number of women who took DES is hard to know precisely, but it has been estimated that over 10 million people have been exposed around the world. DES was classified in the year 2000 as carcinogenic to humans. The deleterious effects induced by DES are very extensive, such as abnormalities or cancers of the genital tract and breast, neurodevelopmental alterations, problems associated with socio-sexual behavior, and immune, pancreatic and cardiovascular disorders. Not only pregnant women but also their children and grandchildren have been affected. Epigenetic alterations have been detected, and intergenerational effects have been observed. More cohort follow-up studies are needed to establish if DES effects are transgenerational. Even though DES is not currently in use, its effects are still present, and families previously exposed and their later generations deserve the continuity of the research studies.

The endocrine disrupting effects of sodium arsenite in the rat testis is not mediated through macrophage activation.

Arsenic (As) is an endocrine disrupting chemical that can disturb the male reproductive system. In a previous study, it was suggested that testicular macrophages could display a role in endocrine disruption induced by As exposure. This work aimed to evaluate the effects of chronic As exposure in the testis function of Wistar rats and examine the participation of macrophage activation and inflammatory response in these processes. We examined gene expression of steroidogenic machinery and immunological markers by RT-QPCR, plasma testosterone concentrations, sperm count and morphology, and histomorphometrical parameters after 60-days exposure to 1 or 5 mg.kg-1.day-1 of sodium arsenite, combined or not with 50 µg.kg-1 of LPS administered one day before euthanasia. We have demonstrated that As exposure reduced the weight of androgen-dependent organs and induced changes in spermatogenesis, in particular at the highest dose. LPS and As co-exposure promoted a decrease in testosterone synthesis, but did not increase the overexpression of markers of macrophage activation seen in LPS-only rats. Our results suggest that As does not alter the testicular macrophage function, but under immunological challenges LPS and As can display a complex interaction, which could lead to endocrine disruption.

In Silico Analysis to Identify Molecular Targets for Chemicals of Concern: The Case Study of Flocoumafen, an Anticoagulant Pesticide.

Rodenticides are pesticides used worldwide, with little information available regarding health consequences in wildlife and humans. The aim of the present study was to use virtual screening to identify potential targets for flocoumafen, a superwarfarin rodenticide. Blind docking of more than 841 human proteins was carried out employing AutoDock Vina. The strength of the ligand interaction with the proteins was quantified based on the binding affinity score (kcal/mol). Results indicate that flocoumafen could be a promiscuous ligand for diversity of cellular protein targets. The best complexes were obtained for prostaglandin F synthase (-14.2 kcal/mol) and serum albumin (-14.0 kcal/mol) followed by glucocorticoid receptor 2, matrix metalloproteinase-9, nuclear receptor ROR-alpha, and activin receptor type-1, all with values equal or better than -13.5 kcal/mol. Docking method validation based on the root-mean-square deviation showed that flocoumafen had good capability to predict corresponding co-crystallized poses; and molecular dynamics simulations suggested the complex with greater binding affinity was thermodynamically stable. Protein-protein interaction networks built with main protein targets revealed that protein kinase B (AKT1), ribosomal protein S6 kinase B1 (RPS6KB1), phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), retinoid X receptor alpha (RXRA), and protein phosphatase 2 catalytic subunit alpha (PPP2CA) were major hub proteins, whereas the gene ontology analysis reported that cellular response to endogenous stimulus, protein binding, and the TOR complex were the biological processes, molecular function, and cell component enrichments, respectively. These results should motivate more ecotoxicity testing for flocoumafen and other superwarfarins, as well as precautionary legislation to minimize exposure to these highly toxic chemicals. Environ Toxicol Chem 2021;40:2034-2043. © 2021 SETAC.

Obesity and endocrine-disrupting chemicals.

Obesity is now a worldwide pandemic. The usual explanation given for the prevalence of obesity is that it results from consumption of a calorie dense diet coupled with physical inactivity. However, this model inadequately explains rising obesity in adults and in children over the past few decades, indicating that other factors must be important contributors. An endocrine-disrupting chemical (EDC) is an exogenous chemical, or mixture that interferes with any aspect of hormone action. EDCs have become pervasive in our environment, allowing humans to be exposed daily through ingestion, inhalation, and direct dermal contact. Exposure to EDCs has been causally linked with obesity in model organisms and associated with obesity occurrence in humans. Obesogens promote adipogenesis and obesity, in vivo, by a variety of mechanisms. The environmental obesogen model holds that exposure to obesogens elicits a predisposition to obesity and that such exposures may be an important yet overlooked factor in the obesity pandemic. Effects produced by EDCs and obesogen exposure may be passed to subsequent, unexposed generations. This "generational toxicology" is not currently factored into risk assessment by regulators but may be another important factor in the obesity pandemic as well as in the worldwide increases in the incidence of noncommunicable diseases that plague populations everywhere. This review addresses the current evidence on how obesogens affect body mass, discusses long-known chemicals that have been more recently identified as obesogens, and how the accumulated knowledge can help identify EDCs hazards.

Disruption of fertility, placenta, pregnancy outcome, and multigenerational inheritance of hepatic steatosis by organotin exposure from contaminated seafood in rats.

Early life exposure to endocrine-disrupting chemicals (EDCs) is an emerging risk factor for development of complications later in life and in subsequent generations. We previously demonstrated that exposure to the EDC organotin (OT), which is present in contaminated seafood, resulted in reproductive abnormalities in female rats. However, few studies have explored the effect of OT accumulation in seafood on pregnancy outcomes. This led us to consider the potential effects of the OT present in seafood on fertility, pregnancy, the placenta, and the offspring. In this investigation, we assessed whether exposure to the OT in contaminated seafood resulted in abnormal fertility and pregnancy features and offspring complications. OT in contaminated seafood (LNI) was administered to female rats, and their fertility, pregnancy outcomes, and fetal liver morphology were assessed. LNI caused abnormal fertility, a reduction in the total number of pups, and an increase in serum testosterone levels compared to controls. Furthermore, LNI exposure caused irregular uterine morphology with inflammation and fibrosis and led to a reduction in embryonic implantation. In pregnant rats, LNI caused abnormal lipid profiles and livers with steatosis features. LNI exposure also causes placental morpho-physiology disruption, a high presence of glycogen and inflammatory cells, and irregular lipid profiles. In addition, LNI exposure caused an increase in large amounts of carbohydrate and lipid delivery to the fetus via an increase in placental nutrient sensor protein expressions (GLUT1, IRß/mTOR and Akt). In both genders of offspring, LNI exposure led to an increase in body weights, liver megakaryocytes, lipid accumulation, and oxidative stress (OS) levels. Collectively, these data suggest that OT exposure from contaminated seafood in female rats leads to reduced fertility, uterine implantation failure, pregnancy and placental metabolic outcome irregularities, offspring adiposity, liver steatosis, and an increase in OS. Furthermore, some of the effects of OT may be the result of obesogenic and multigenerational effects of OT in adult female rats.

Profiling epigenetic changes in human cell line induced by atrazine exposure.

How environmental chemicals can affect and exert their toxic effect at a molecular level has gained significant interest in recent years, not only for understanding their immediate health implications over exposed individuals, but also for their subsequent progeny. Atrazine (ATZ) is a commonly used herbicide in the U.S. and a long-suspected endocrine disrupting chemical. The molecular mechanism conferring long-term adverse health outcomes, however, remain elusive. Here, we explored changes in epigenetic marks that arise after exposure to ATZ at selected doses using image-based analysis coupled with data clustering. Significant decreases in methylated CpG (meCpG) and histone 3 lysine 9 tri-methylated (H3K9me3) were observed in the selected human cell line with a clear spatial preference. Treating cells with ATZ leads to the loss of a subpopulation of cells with high meCpG levels as identified in our clustering and histogram analysis. A similar trend was observed in H3K9me3 potentially attributing to the cross-talking between meCpG and H3K9me3. Changes in meCpG are likely to be associated with alterations in epigenetic enzyme expression levels regulating meCpG and persist after the removal of ATZ source which collectively provide a plausible mechanism for long-term ATZ-induced toxicity.

Exposure to bisphenol A and diabetes risk in Mexican women.

Bisphenol A (BPA) is an endocrine-disrupting chemical widely used in the production of polycarbonate plastics and epoxy resins, which has been previously linked to diabetes among non-Hispanic populations. As part of a case control study for breast cancer, only controls with BPA information were included in this report. The final sample size comprises 70 self-reported diabetics and 334 non-diabetics. Urinary free bisphenol A (BPA-F) (µg/L) was determined by solid-phase extraction and HPLC/FLD analysis. Logistic regression models were used to evaluate the association between BPA-F and self-reported diabetes. After adjusting by age, urinary BPA-F (4.06-224.53 µg/g creatinine) was associated with diabetes exposure (OR = 1.85; 95% CI 1.04, 3.28) compared with women in the reference category (0.67-4.05 µg/g creatinine). BPA may be an environmental cofactor of diabetes. More studies are needed to confirm this result, especially in Hispanic populations.

Maternal bisphenol A exposure disrupts spermatogenesis in adult rat offspring.

Bisphenol A (BPA) is an endocrine-disrupting chemical (EDC) that is widely used in the manufacturing of plastics and inner linings of food cans. Previously, it was reported that BPA disturbed the sexual dimorphic nucleus of the hypothalamus and delaying the onset of puberty attributed to an estrogenic action. In addition, BPA during the perinatal period increased LH serum concentrations in male offspring of dams at doses below the reproductive NOAEL (No Observable Adverse Effect Level) based upon World Health Organization guidelines. Based upon these findings, the objective of this study was to (1) determine the effects of perinatal treatment with low doses of BPA on regulation of spermatogenesis in adult offspring and (2) elucidate molecular mechanisms involved in the pathogenesis of gonadal dysfunction. The expression of genes related to spermatogenesis was disrupted with adverse consequences on sperm production, reserves, and function. Both BPA treated groups exhibited reduction in sperm production and epithelial height of seminiferous tubules, accompanied by diminished integrity of the acrosome and plasma membrane, decreased mitochondrial activity and increased incidence of morphological abnormalities. The sperm transit time was also slower. However, only in the group receiving the higher BPA dose was transcript expression of genes affected (reduced Ar and increased Esr1). It is of interest that serum testosterone levels were elevated in the same group where Ar was decreased. Data suggest that exposure to low BPA doses during hypothalamic sexual differentiation period produces permanent deleterious effects on spermatogenesis in adulthood.

Acrylamide: A review about its toxic effects in the light of Developmental Origin of Health and Disease (DOHaD) concept.

The endocrine system is highly sensitive to endocrine-disrupting chemicals (EDC) which interfere with metabolism, growth and reproduction throughout different periods of life, especially in the embryonic and pubertal stages, in which gene reprogramming may be associated with impaired development and control of tissues/organs even in adulthood. Acrylamide is considered a potential EDC and its main source comes from fried, baked and roasted foods that are widely consumed by children, teenagers and adults around the world. This review aimed to present some aspects regarding the acrylamide formation, its toxicokinetics, the occurrence of acrylamide in foods, the recent findings about its effects on different systems and the consequences for the human healthy. The challenges to characterize the molecular mechanisms triggered by acrylamide and to establish safe levels of consumption and/or exposure are also discussed in the present review.

Occurrence and removal of estrogens in Brazilian wastewater treatment plants.

This paper evaluated the occurrence and removal efficiency of four estrogenic hormones in five biological wastewater treatment plants (WWTPs), located in the State of Ceará, Brazil. The five WWTPs comprised: two systems consisted of one facultative pond followed by two maturation ponds, one facultative pond, one activated sludge (AS) system followed by a chlorination step, and one upflow anaerobic sludge blanket (UASB) reactor followed by a chlorination step. Estrogen occurrence showed a wide variation among the analyzed influent and effluent samples. Estrone (E1) showed the highest occurrence in the influent (76%), whereas both 17ß-estradiol (E2) and 17α-ethynylestradiol (EE2) presented a 52% occurrence, and the compound 17ß-estradiol 17-acetate (E2-17A), a 32% one. The occurrence in the effluent samples was 48% for E1, 28% for E2, 12% for E2-17A, and 40% for EE2. The highest concentrations of E1 and EE2 hormones in the influent were 3050 and 3180 ng L(-1), respectively, whereas E2 and E2-17A had maximum concentrations of 776 and 2300 ng L(-1), respectively. The lowest efficiencies for the removal of estrogenic hormones were found in WWTP consisted of waste stabilization ponds, ranging from 54 to 79.9%. The high-rate systems (AS and UASB), which have chlorination as post-treatment, presented removal efficiencies of approximately 95%.