Bisphenol A shapes children’s brain and behavior: towards an integrated neurotoxicity assessment including human data

The authors gratefully acknowledge editorial assistance provided by Richard Davies. VM is under contract within the Human Biomonitoring for Europe Project (European Union Commission H2020-EJP-HBM4EU). The authors acknowledge the funding received from the Biomedical Research Networking Center-CIBER de Epidemiología y Salud Pública (CIBERESP), and the Instituto de Salud Carlos III (ISCIII) (FIS-PI16/01820 and FIS-PI16/01812). The funders had no role in the study design, data.Concerns about the effects of bisphenol A (BPA) on human brain and behavior are not novel; however, Grohs and colleagues have contributed groundbreaking data on this topic in a recent issue of Environmental Health. For the first time, associations were reported between prenatal BPA exposure and differences in children’s brain microstructure, which appeared to mediate the association between this exposure and children’s behavioral symptoms. Findings in numerous previous mother-child cohorts have pointed in a similar worrying direction, linking higher BPA exposure during pregnancy to more behavioral problems throughout childhood as assessed by neuropsychological questionnaires. Notwithstanding, this body of work has not been adequately considered in risk assessment. From a toxicological perspective, results are now available from the CLARITY-BPA consortium, designed to reconcile academic and regulatory toxicology findings. In fact, the brain has consistently emerged as one of the most sensitive organs disrupted by BPA, even at doses below those considered safe by regulatory agencies such as the European Food Safety Authority (EFSA). In this Commentary, we contextualize the results of Grohs et al. within the setting of previous epidemiologic and CLARITY-BPA data and express our disquiet about the “all-or-nothing” criterion adopted to select human data in a recent EFSA report on the appraisal methodology for their upcoming BPA risk assessment. We discuss the most relevant human studies, identify emerging patterns, and highlight the need for adequate assessment and interpretation of the increasing epidemiologic literature in this field in order to support decision-making. With the aim of avoiding a myopic or biased selection of a few studies in traditional risk assessment procedures, we propose a future reevaluation of BPA focused on neurotoxicity and based on a systematic and comprehensive integration of available mechanistic, animal, and human data. Taken together, the experimental and epidemiologic evidence converge in the same direction: BPA is a probable developmental neurotoxicant at low doses. Accordingly, the precautionary principle should be followed, progressively implementing stringent preventive policies worldwide, including the banning of BPA in food contact materials and thermal receipts, with a focus on the utilization of safer substitutes.European Union (EU): H2020-EJP-HBM4EUBiomedical Research Networking Center-CIBER de Epidemiologia y Salud Publica (CIBERESP)Instituto de Salud Carlos III FIS-PI16/01820 FIS-PI16/0181

Horizontal DNA transfer mechanisms of bacteria as weapons of intragenomic conflict

Horizontal DNA transfer (HDT) is a pervasive mechanism of diversification in many microbial species, but its primary evolutionary role remains controversial. Much recent research has emphasised the adaptive benefit of acquiring novel DNA, but here we argue instead that intragenomic conflict provides a coherent framework for understanding the evolutionary origins of HDT. To test this hypothesis, we developed a mathematical model of a clonally descended bacterial population undergoing HDT through transmission of mobile genetic elements (MGEs) and genetic transformation. Including the known bias of transformation toward the acquisition of shorter alleles into the model suggested it could be an effective means of counteracting the spread of MGEs. Both constitutive and transient competence for transformation were found to provide an effective defence against parasitic MGEs; transient competence could also be effective at permitting the selective spread of MGEs conferring a benefit on their host bacterium. The coordination of transient competence with cell-cell killing, observed in multiple species, was found to result in synergistic blocking of MGE transmission through releasing genomic DNA for homologous recombination while simultaneously reducing horizontal MGE spread by lowering the local cell density. To evaluate the feasibility of the functions suggested by the modelling analysis, we analysed genomic data from longitudinal sampling of individuals carrying Streptococcus pneumoniae. This revealed the frequent within-host coexistence of clonally descended cells that differed in their MGE infection status, a necessary condition for the proposed mechanism to operate. Additionally, we found multiple examples of MGEs inhibiting transformation through integrative disruption of genes encoding the competence machinery across many species, providing evidence of an ongoing "arms race." Reduced rates of transformation have also been observed in cells infected by MGEs that reduce the concentration of extracellular DNA through secretion of DNases. Simulations predicted that either mechanism of limiting transformation would benefit individual MGEs, but also that this tactic's effectiveness was limited by competition with other MGEs coinfecting the same cell. A further observed behaviour we hypothesised to reduce elimination by transformation was MGE activation when cells become competent. Our model predicted that this response was effective at counteracting transformation independently of competing MGEs. Therefore, this framework is able to explain both common properties of MGEs, and the seemingly paradoxical bacterial behaviours of transformation and cell-cell killing within clonally related populations, as the consequences of intragenomic conflict between self-replicating chromosomes and parasitic MGEs. The antagonistic nature of the different mechanisms of HDT over short timescales means their contribution to bacterial evolution is likely to be substantially greater than previously appreciated

ADRB2 Arg16Gly Polymorphism, Lung Function, and Mortality: Results from the Atherosclerosis Risk in Communities Study

BACKGROUND: Growing evidence suggests that the Arg16Arg genotype of the beta-2 adrenergic receptor gene may be associated with adverse effects of beta-agonist therapy. We sought to examine the association of beta-agonist use and the Arg16Gly polymorphism with lung function and mortality among participants in the Atherosclerosis Risk in Communities study. METHODOLOGY AND PRINCIPAL FINDINGS: We genotyped study participants and analyzed the association of the Arg16Gly polymorphism and beta-agonist use with lung function at baseline and clinical examination three years later and with all-cause mortality during 10 years of follow-up. Lung function was characterized by percent-predicted forced expiratory volume in 1 second. Associations were examined separately for blacks and whites. Black beta-agonist users with the Arg/Arg genotype had better lung function at baseline and at the second clinical visit than those with Arg/Gly and Gly/Gly genotypes. Adjusted mean percent-predicted FEV(1) was 21% higher in Arg/Arg subjects compared to Gly/Gly at baseline (p = 0.01) and 20% higher than Gly/Gly at visit 2 (p = 0.01). Arg/Gly subjects had adjusted percent-predicted FEV(1) 17% lower than Arg/Arg at baseline but were similar to Arg/Arg subjects at visit 2. Although black beta-agonist users with the Arg/Arg genotype appeared to have better crude survival rates, the association between genotype and all-cause mortality was inconclusive. We found no difference in lung function or mortality by genotype among blacks who did not use beta-agonists or among whites, regardless of beta-agonist use. CONCLUSIONS: Black beta-agonist users with the ADRB2 Arg16Arg genotype had better lung function, and, possibly, better overall survival compared to black beta-agonist users with the Gly16Gly genotype. Our findings highlight the need for additional studies of sufficient size and statistical power to allow examination of outcomes among beta-agonist users of different races and genotypes

Mucosal Targeting of a BoNT/A Subunit Vaccine Adjuvanted with a Mast Cell Activator Enhances Induction of BoNT/A Neutralizing Antibodies in Rabbits

We previously reported that the immunogenicity of Hcβtre, a botulinum neurotoxin A (BoNT/A) immunogen, was enhanced by fusion to an epithelial cell binding domain, Ad2F, when nasally delivered to mice with cholera toxin (CT). This study was performed to determine if Ad2F would enhance the nasal immunogenicity of Hcβtre in rabbits, an animal model with a nasal cavity anatomy similar to humans. Since CT is not safe for human use, we also tested the adjuvant activity of compound 48/80 (C48/80), a mast cell activating compound previously determined to safely exhibit nasal adjuvant activity in mice.New Zealand White or Dutch Belted rabbits were nasally immunized with Hcβtre or Hcβtre-Ad2F alone or combined with CT or C48/80, and serum samples were tested for the presence of Hcβtre-specific binding (ELISA) or BoNT/A neutralizing antibodies.Hcβtre-Ad2F nasally administered with CT induced serum anti-Hcβtre IgG ELISA and BoNT/A neutralizing antibody titers greater than those induced by Hcβtre + CT. C48/80 provided significant nasal adjuvant activity and induced BoNT/A-neutralizing antibodies similar to those induced by CT.Ad2F enhanced the nasal immunogenicity of Hcβtre, and the mast cell activator C48/80 was an effective adjuvant for nasal immunization in rabbits, an animal model with a nasal cavity anatomy similar to that in humans

Longitudinal Evaluation of an N-Ethyl-N-Nitrosourea-Created Murine Model with Normal Pressure Hydrocephalus

Normal-pressure hydrocephalus (NPH) is a neurodegenerative disorder that usually occurs late in adult life. Clinically, the cardinal features include gait disturbances, urinary incontinence, and cognitive decline.Herein we report the characterization of a novel mouse model of NPH (designated p23-ST1), created by N-ethyl-N-nitrosourea (ENU)-induced mutagenesis. The ventricular size in the brain was measured by 3-dimensional micro-magnetic resonance imaging (3D-MRI) and was found to be enlarged. Intracranial pressure was measured and was found to fall within a normal range. A histological assessment and tracer flow study revealed that the cerebral spinal fluid (CSF) pathway of p23-ST1 mice was normal without obstruction. Motor functions were assessed using a rotarod apparatus and a CatWalk gait automatic analyzer. Mutant mice showed poor rotarod performance and gait disturbances. Cognitive function was evaluated using auditory fear-conditioned responses with the mutant displaying both short- and long-term memory deficits. With an increase in urination frequency and volume, the mutant showed features of incontinence. Nissl substance staining and cell-type-specific markers were used to examine the brain pathology. These studies revealed concurrent glial activation and neuronal loss in the periventricular regions of mutant animals. In particular, chronically activated microglia were found in septal areas at a relatively young age, implying that microglial activation might contribute to the pathogenesis of NPH. These defects were transmitted in an autosomal dominant mode with reduced penetrance. Using a whole-genome scan employing 287 single-nucleotide polymorphic (SNP) markers and further refinement using six additional SNP markers and four microsatellite markers, the causative mutation was mapped to a 5.3-cM region on chromosome 4.Our results collectively demonstrate that the p23-ST1 mouse is a novel mouse model of human NPH. Clinical observations suggest that dysfunctions and alterations in the brains of patients with NPH might occur much earlier than the appearance of clinical signs. p23-ST1 mice provide a unique opportunity to characterize molecular changes and the pathogenic mechanism of NPH

Mucin Secretion Induced by Titanium Dioxide Nanoparticles

Nanoparticle (NP) exposure has been closely associated with the exacerbation and pathophysiology of many respiratory diseases such as Chronic Obstructive Pulmonary Disease (COPD) and asthma. Mucus hypersecretion and accumulation in the airway are major clinical manifestations commonly found in these diseases. Among a broad spectrum of NPs, titanium dioxide (TiO2), one of the PM10 components, is widely utilized in the nanoindustry for manufacturing and processing of various commercial products. Although TiO2 NPs have been shown to induce cellular nanotoxicity and emphysema-like symptoms, whether TiO2 NPs can directly induce mucus secretion from airway cells is currently unknown. Herein, we showed that TiO2 NPs (<75 nm) can directly stimulate mucin secretion from human bronchial ChaGo-K1 epithelial cells via a Ca2+ signaling mediated pathway. The amount of mucin secreted was quantified with enzyme-linked lectin assay (ELLA). The corresponding changes in cytosolic Ca2+ concentration were monitored with Rhod-2, a fluorescent Ca2+ dye. We found that TiO2 NP-evoked mucin secretion was a function of increasing intracellular Ca2+ concentration resulting from an extracellular Ca2+ influx via membrane Ca2+ channels and cytosolic ER Ca2+ release. The calcium-induced calcium release (CICR) mechanism played a major role in further amplifying the intracellular Ca2+ signal and in sustaining a cytosolic Ca2+ increase. This study provides a potential mechanistic link between airborne NPs and the pathoetiology of pulmonary diseases involving mucus hypersecretion

Parkinson's disease induced pluripotent stem cells with triplication of the α-synuclein locus

A major barrier to research on Parkinson's disease is inaccessibility of diseased tissue for study. One solution is to derive induced pluripotent stem cells from patients and differentiate them into neurons affected by disease. Triplication of SNCA, encoding α-synuclein, causes a fully penetrant, aggressive form of Parkinson's disease with dementia. α-Synuclein dysfunction is the critical pathogenic event in Parkinson's disease, multiple system atrophy and dementia with Lewy bodies. Here we produce multiple induced pluripotent stem cell lines from an SNCA triplication patient and an unaffected first-degree relative. When these cells are differentiated into midbrain dopaminergic neurons, those from the patient produce double the amount of α-synuclein protein as neurons from the unaffected relative, precisely recapitulating the cause of Parkinson's disease in these individuals. This model represents a new experimental system to identify compounds that reduce levels of α-synuclein, and to investigate the mechanistic basis of neurodegeneration caused by α-synuclein dysfunction

Lkb1 and Pten Synergise to Suppress mTOR-Mediated Tumorigenesis and Epithelial-Mesenchymal Transition in the Mouse Bladder

The AKT/PI3K/mTOR pathway is frequently altered in a range of human tumours, including bladder cancer. Here we report the phenotype of mice characterised by deletion of two key players in mTOR regulation, Pten and Lkb1, in a range of tissues including the mouse urothelium. Despite widespread recombination within the range of epithelial tissues, the primary phenotype we observe is the rapid onset of bladder tumorigenesis, with median onset of approximately 100 days. Single deletion of either Pten or Lkb1 had no effect on bladder cell proliferation or tumour formation. However, simultaneous deletion of Lkb1 and Pten led to an upregulation of the mTOR pathway and the hypoxia marker GLUT1, increased bladder epithelial cell proliferation and ultimately tumorigenesis. Bladder tissue also exhibited characteristic features of epithelial-mesenchymal transition, with loss of the epithelial markers E-cadherin and the tight junction protein ZO-1, and increases in the mesenchymal marker vimentin as well as nuclear localization of epithelial-mesenchymal transition (EMT) regulator Snail. We show that these effects were all dependent upon mTOR activity, as rapamycin treatment blocked both EMT and tumorigenesis. Our data therefore establish clear synergy between Lkb1 and Pten in controlling the mTOR pathway within bladder epithelium, and show that loss of this control leads to the disturbance of epithelial structure, EMT and ultimately tumorigenesis