Katerine S. Saili

Low dose effects of bisphenol A

In 2007, a group of experts critically analyzed hundreds of publications on bisphenol A (BPA), including the evidence for low dose effects. Here, we have updated these evaluations to determine the strength of the evidence for low dose effects of BPA. Based on the cut-offs for “low doses” established previously (i.e., the lowest observed adverse effect level [LOAEL], or 50 mg/kg/day for mammalian studies), we identified more than 450 low dose studies. Using an integrative approach, we examined five endpoints in depth that had evidence from two or more study types (in vitro, in vivo laboratory animal, and human). Based on all available studies, we are confident that consistent, reproducible, low dose effects have been demonstrated for BPA. We conclude that the doses that reliably produce effects in animals are 1–4 magnitudes of order lower than the current LOAEL of 50 mg/kg/day and many should be considered adverse.

Neurodevelopmental low-dose bisphenol A exposure leads to early life-stage hyperactivity and learning deficits in adult zebrafish

Developmental bisphenol A (BPA) exposure has been implicated in adverse behavior and learning deficits. The mode of action underlying these effects is unclear. The objectives of this study were to identify whether low-dose, developmental BPA exposure affects larval zebrafish locomotor behavior and whether learning deficits occur in adults exposed during development. Two control compounds, 17β-estradiol (an estrogen receptor ligand) and GSK4716 (a synthetic estrogen-related receptor gamma ligand), were included. Larval toxicity assays were used to determine appropriate BPA, 17β-estradiol, and GSK4716 concentrations for behavior testing. BPA tissue uptake was analyzed using HPLC and lower doses were extrapolated using a linear regression analysis. Larval behavior tests were conducted using a ViewPoint Zebrabox. Adult learning tests were conducted using a custom-built T-maze. BPA exposure to <30μM was non-teratogenic. Neurodevelopmental BPA exposure to 0.01, 0.1, or 1μM led to larval hyperactivity or learning deficits in adult zebrafish. Exposure to 0.1μM 17β-estradiol or GSK4716 also led to larval hyperactivity. This study demonstrates the efficacy of using the zebrafish model for studying the neurobehavioral effects of low-dose developmental BPA exposure.

Persistent Adult Zebrafish Behavioral Deficits Results from Acute Embryonic Exposure to Gold Nanoparticles

As the number of products containing nanomaterials increase, human exposure to nanoparticles (NPs) is unavoidable. Presently, few studies focus on the potential long-term consequences of developmental NP exposure. In this study, zebrafish embryos were acutely exposed to three gold NPs that possess functional groups with differing surface charge. Embryos were exposed to 50 μg/mL of 1.5 nm gold nanoparticles (AuNPs) possessing negatively charged 2-mercaptoethanesulfonic acid (MES) or neutral 2-(2-(2-mercaptoethoxy)ethoxy)ethanol (MEEE) ligands or 10 μg/mL of the AuNPs possessing positively charged trimethylammoniumethanethiol (TMAT). Both MES- and TMAT-AuNP exposed embryos exhibited hypo-locomotor activity, while those exposed to MEEE-AuNPs did not. A subset of embryos that were exposed to 1.5 nm MES- and TMAT-AuNPs during development from 6 to 120 h post fertilization was raised to adulthood. Behavioral abnormalities and the number of survivors into adulthood were evaluated at 122 days post fertilization. We found that both treatments induced abnormal startle behavior following a tap stimulus. However, the MES-AuNPs exposed group also exhibited abnormal adult behavior in the light and had a lower survivorship into adulthood. This study demonstrates that acute, developmental exposure to 1.5 nm MES- and TMAT-AuNPs, two NPs differing only in the functional group, affects larval behavior, with behavioral effects persisting into adulthood.

The Anti-Inflammatory Drug Leflunomide Is an Agonist of the Aryl Hydrocarbon Receptor

Background The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that mediates the toxicity and biological activity of dioxins and related chemicals. The AhR influences a variety of processes involved in cellular growth and differentiation, and recent studies have suggested that the AhR is a potential target for immune-mediated diseases. Methodology/Principal Findings During a screen for molecules that activate the AhR, leflunomide, an immunomodulatory drug presently used in the clinic for the treatment of rheumatoid arthritis, was identified as an AhR agonist. We aimed to determine whether any biological activity of leflunomide could be attributed to a previously unappreciated interaction with the AhR. The currently established mechanism of action of leflunomide involves its metabolism to A771726, possibly by cytochrome P450 enzymes, followed by inhibition of de novo pyrimidine biosynthesis by A771726. Our results demonstrate that leflunomide, but not its metabolite A771726, caused nuclear translocation of AhR into the nucleus and increased expression of AhR-responsive reporter genes and endogenous AhR target genes in an AhR-dependent manner. In silico Molecular Docking studies employing AhR ligand binding domain revealed favorable binding energy for leflunomide, but not for A771726. Further, leflunomide, but not A771726, inhibited in vivo epimorphic regeneration in a zebrafish model of tissue regeneration in an AhR-dependent manner. However, suppression of lymphocyte proliferation by leflunomide or A771726 was not dependent on AhR. Conclusions These data reveal that leflunomide, an anti-inflammatory drug, is an agonist of the AhR. Our findings link AhR activation by leflunomide to inhibition of fin regeneration in zebrafish. Identification of alternative AhR agonists is a critical step in evaluating the AhR as a therapeutic target for the treatment of immune disorders.

Experimental Evidence for Evolved Tolerance to Avian Malaria in a Wild Population of Low Elevation Hawai‘i ‘Amakihi (Hemignathus virens)

Introduced vector-borne diseases, particularly avian malaria (Plasmodium relictum) and avian pox virus (Avipoxvirus spp.), continue to play significant roles in the decline and extinction of native forest birds in the Hawaiian Islands. Hawaiian honeycreepers are particularly susceptible to avian malaria and have survived into this century largely because of persistence of high elevation refugia on Kaua‘i, Maui, and Hawai‘i Islands, where transmission is limited by cool temperatures. The long term stability of these refugia is increasingly threatened by warming trends associated with global climate change. Since cost effective and practical methods of vector control in many of these remote, rugged areas are lacking, adaptation through processes of natural selection may be the best long-term hope for recovery of many of these species. We document emergence of tolerance rather than resistance to avian malaria in a recent, rapidly expanding low elevation population of Hawai‘i ‘Amakihi (Hemignathus virens) on the island of Hawai‘i. Experimentally infected low elevation birds had lower mortality, lower reticulocyte counts during recovery from acute infection, lower weight loss, and no declines in food consumption relative to experimentally infected high elevation Hawai‘i ‘Amakihi in spite of similar intensities of infection. Emergence of this population provides an exceptional opportunity for determining physiological mechanisms and genetic markers associated with malaria tolerance that can be used to evaluate whether other, more threatened species have the capacity to adapt to this disease.

Global gene expression analysis reveals pathway differences between teratogenic and non-teratogenic exposure concentrations of bisphenol A and 17β-estradiol in embryonic zebrafish

Transient developmental exposure to 0.1μM bisphenol A (BPA) results in larval zebrafish hyperactivity and learning impairments in the adult, while exposure to 80μM BPA results in teratogenic responses, including craniofacial abnormalities and edema. The mode of action underlying these effects is unclear. We used global gene expression analysis to identify candidate genes and signaling pathways that mediate BPAs developmental toxicity in zebrafish. Exposure concentrations were selected and anchored to the positive control, 17β-estradiol (E2), based on previously determined behavioral or teratogenic phenotypes. Functional analysis of differentially expressed genes revealed distinct expression profiles at 24h post fertilization for 0.1μM versus 80μM BPA and 0.1μM versus 15μM E2 exposure, identification of prothrombin activation as a top canonical pathway impacted by both 0.1μM BPA and 0.1μM E2 exposure, and suppressed expression of several genes involved in nervous system development and function following 0.1μM BPA exposure.

Nonmammalian Models in Toxicology Screening

With the advent of industrialization around the world, increasing numbers of chemicals are being produced at an alarming rate before undergoing appropriate safety testing. One of the solutions to overcome this problem has been testing chemicals for toxicological effects in mammalian species such as rodents and primates. A major hurdle with the use of mammalian models is the time and cost it takes to complete toxicity testing. Therefore, it is practically impossible to cover the virtually limitless number of chemicals that require toxicity testing using mammalian models. Nonmammalian species such as the nematode worm, fruit fly, zebrafish, and others have been proposed as viable alternatives because of their usability in terms of providing fast and cost-effective assays that are comparable to those of the mammalian species. Each of these species has a relatively small life span, is cheap and easy to maintain in the laboratory, and offers endpoints for which assays can be automated to achieve high throughput capabilities. Here we compare the three most common nonmammalian model species for their use in screening for toxicological effects of different chemicals.