Gene expression changes in bottlenose dolphin, Tursiops truncatus, skin cells following exposure to methylmercury (MeHg) or perfluorooctane sulfonate (PFOS) (original) (raw)
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Marine Environmental Research, 2016
Chemicals discovered in water at levels that may be significantly different than expected are referred to as "contaminants of emerging concern" (CECs) because the risk to environmental health associated with their presence and frequency of occurrence is not known. The perfluorooctanoic acid (PFOA), used to make fluoropolymers, and the bisphenol A (BPA), a monomer used in epoxy resins and polycarbonate plastics present in many hard plastic bottles and metal-based food storage are worldwide distributed compounds considered CECs because of their agonist or antagonist effects on the endocrine receptors. We applied an ex vivo assay using skin biopsy slices from the bottlenose dolphin (Tursiops truncatus) to analyze global gene expression in response to PFOA or BPA exposure. Small slices of skin biopsy were cultured and independently treated with different concentrations of PFOA or BPA prior hybridization to a custom-made microarray. Results illustrate how the skin transcriptome holds information on the contaminant exposure, showing potential long-term effects on the dolphin health status. Most important is the potential of the novel speciesspecific approach described: the skin transcriptomic signature could be used as classifier for a specific contaminant and the significantly regulated genes used to develop a patented kit of biomarkers of exposure.
Environmental Science & Technology, 2019
Cetaceans in the Southern California Bight (SCB) are exposed to high levels of halogenated organic contaminants (HOCs), which have previously been linked to impaired reproductive health and immune responses. We used a combination of molecular tools to examine the potential physiological impacts of HOC exposure in two bottlenose dolphin (Tursiops truncatus) ecotypes in the SCB. We quantified 25 HOCs in the blubber of 22 biopsies collected from males between 2012 and 2016. We then analyzed genome-wide gene expression in skin using RNA-sequencing and measured blubber testosterone to compare HOC exposure with cellular and endocrine biomarkers. We found high levels of HOCs in both ecotypes with significantly higher total polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs), tris(4-chlorophenyl)methanol (TCPMOH), and chlordane-related compounds in the coastal ecotype versus the offshore ecotype. We found evidence of PBDE bioaccumulation in both ecotypes, however, the pattern of bioaccumulation or endocrine disruption for other HOCs was different between the ecotypes, suggesting potential endocrine disruption in the coastal ecotype. We also observed correlations between HOCs and gene coexpression networks enriched for xenobiotic metabolism, hormone metabolism, and immune response that could indicate cellular effects from HOC exposure. By integrating measurements of HOC load with both transcriptome profiling and endocrine biomarkers, our approach provides insight into HOC exposure and potential impacts on wild cetacean health in southern California.
Environmental Toxicology, 2013
The need for powerful new tools to detect the effects of chemical pollution, in particular of persistent organic pollutants (POPs) and polycyclic aromatic hydrocarbons (PAHs) on Mediterranean cetaceans led us to develop and apply a suite of sensitive biomarkers for integument biopsies of stranded and free-ranging animals. This multi-response ex vivo method has the aim to detect toxicological effects of contaminant mixtures. In the present study, we applied an ex vivo assay using skin biopsy and liver slices, combining molecular biomarkers [Western blot of Cytochrome P450 1A1 (CYP1A1) and Cytochrome P450 2B (CYP2B)] and gene expression biomarkers (Quantitative real-time PCR of CYP1A1, heat shock protein 70, estrogen receptor alpha and E2F transcription factor) in response to chemical exposure [organochlorines compounds (OCs), polybrominated diphenyl ethers (PBDEs), and PAHs] for stranded Mediterranean Stenella coeruleoalba. The main goal of this experiment was to identify the biomarker and/or a suite of biomarkers that could best detect the presence of a specific class of pollutants (OCs, PBDEs, and PAHs) or a mixture of them. This multi-response biomarker methodology revealed an high sensitivity and selectivity of responses (such as CYP1A and ER a mRNA variations after OCs and PAHs exposure) and could represent a valid future approach for the study of inter-and intra-species sensitivities to various classes of environmental contaminants. #
Aquatic Toxicology, 2008
The main effluent from oil and gas production is produced water (PW), a waste that contains low to moderate concentrations of oil-derived substances such as polycyclic aromatic hydrocarbons (PAHs) and alkylphenols (APs). PW components may be present in seawater at low concentrations over large areas in the vicinity of oil and gas production facilities. In this study, zebrafish (Danio rerio) were exposed to control and three treatments (high-, pulsed-, low-dose) of a synthetic PW mixture for 1, 7 and 13 weeks. The aim was to investigate the development of transcriptome and biomarker responses as well as relationships between early responses and population-relevant effects. The synthetic PW contained a mixture of low-molecular-weight PAHs (<5 ring) and short-chain APs (C1-C4). The water-borne exposure levels (sum PAH) ranged from 0.54 ppb (low dose) to 5.4 ppb (high dose). Bile pyrene metabolites ranged from 17-133 ng g −1 bile in the control group to 23-1081 ng g −1 bile in the high exposure group. Similar levels have been observed in wild fish, confirming an environmentally relevant exposure. The expression of mRNAs of hepatic genes was investigated in the high exposure group using the Zebrafish OligoLibrary TM from Compugen. Functional clustering analysis revealed effects in the reproductive system, the nervous system, the respiratory system, the immune system, lipid metabolism, connective tissue and in a range of functional categories related to cell cycle and cancer. The majority of differentially expressed mRNAs of genes were down-regulated, suggesting reduction in gene transcription to be as relevant as up-regulation or induction when assessing biological responses to PW exposure. Biomarkers for effects of PAHs (cytochrome P450 1A) and environmental estrogens (vitellogenin) did not appear to be affected by the chronic exposure to low concentration of PW components. Effects at the population level included a reduction in condition factor in male fish from all exposed groups and spinal column deformations in the F1 generation of exposed groups. The different exposure regimes did not produce any significant differences in reproduction or recruitment. The results from this study demonstrate that environmentally relevant concentrations of PW affect gene expression and population-relevant endpoints in zebrafish, although links between the two were not obvious.
Marine Environmental Research, 2008
The aim of the present study was to propose the immunofluorescence technique in cultured fibroblasts from Mediterranean cetaceans as a new ''in vitro" tool to explore the susceptibility of these marine mammals to different xenobiotic compounds. The cell lines were cultured from integument biopsies of freeranging and stranded cetaceans (dead within 12 h). Using the indirect immunofluorescence assay, we detected endogenous proteins induced by different contaminants. Here we present the method used for qualitative and quantitative evaluation of cytochromes P450 (CYP1A1 and CYP2B) induced by some POPs (DDTs and PCBs) and emerging contaminants (PBDEs) in fibroblast cell cultures of striped dolphin (Stenella coeruleoalba) and bottlenose dolphin (Tursiops truncatus). Immunofluorescence was quantified with a specially designed Olympus macro, DetectIntZ. A major result was the possibility of using this ''in vitro" assay to quantify induction of endogenous proteins.
Immunotoxic effects of environmental pollutants in marine mammals
Due to their marine ecology and life-history, marine mammals accumulate some of the highest levels of environmental contaminants of all wildlife. Given the increasing prevalence and severity of diseases in marine wildlife, it is imperative to understand how pollutants affect the immune system and consequently disease susceptibility. Advancements and adaptations of analytical techniques have facilitated marine mammal immunotoxicology research. Field studies, captive-feeding experiments and in vitro laboratory studies with marine mammals have associated exposure to environmental pollutants, most notable polychlorinated biphenyls (PCBs), organo-chlorine pesticides and heavy metals, to alterations of both the innate and adaptive arms of immune systems, which include aspects of cellular and humoral immunity. For marine mammals, reported immunotoxicology endpoints fell into several major categories: immune tissue histopathology, haematology/circulating immune cell populations, functional immune assays (lymphocyte proliferation, phagocytosis, respiratory burst, and natural killer cell activity), immunoglobulin production, and cytokine gene expression. Lymphocyte proliferation is by far the most commonly used immune assay, with studies using different organic pollutants and metals predominantly reporting immunosuppressive effects despite the many differences in study design and animal life history. Using combined field and laboratory data, we determined effect threshold levels for suppression of lymphocyte proliferation to be between b0.001–10 ppm for PCBs, 0.002–1.3 ppm for Hg, 0.009–0.06 for MeHg, and 0.1–2.4 for cadmium in polar bears and several pinniped and cetacean species. Similarly, thresholds for suppression of phagocytosis were 0.6–1.4 and 0.08–1.9 ppm for PCBs and mercury, respectively. Although data are lacking for many important immune endpoints and mechanisms of specific immune alterations are not well understood, this review revealed a systemic suppression of immune function in marine mammals exposed to environmental contaminants. Exposure to immunotoxic contaminants may have significant population level consequences as a contributing factor to increasing anthropogenic stress in wildlife and infectious disease outbreaks.
Activation of Transcription Factors in Zebrafish Cell Cultures by Environmental Pollutants
Archives of Biochemistry and Biophysics, 2000
Many classes of environmental pollutants are found at significant levels in the aquatic environment. We are designing a fish model as an inexpensive and efficient system for the assessment of aquatic pollution. Three classes of environmental pollutants-halogenated and nonhalogenated aromatic hydrocarbons, heavy metals, and potent electrophiles-are known to upregulate particular mammalian genes via the activation of specific DNA motifs called aromatic hydrocarbon (AHREs), heavy metal (MREs), and electrophile (EPREs) response elements, respectively. We have made plasmid constructs, using these mammalian or trout response elements to drive the luciferase reporter gene. Here we show that transient transfection of the zebrafish ZEM2S cell line with these reporter constructs imparts dose-dependent gene induction upon exposure to a variety of chemicals within each of these three classes of inducers: [a] (AHREmediated) 2,3,7,8-tetrachlorodibenzo-p-dioxin, 3-methylcholanthrene, 3,4,5,3,4