Biotransformation and endocrine disruptive effects of contaminants in ringed seals - implications for monitoring and risk assessment

Marine mammals are exposed to persistent organic pollutants (POPs), which may be biotransformed to metabolites some of which are highly toxic. Both POPs and their metabolites may lead to adverse health effects, which have been studied using various biomarkers. Changes in endocrine homeostasis have been suggested to be sensitive biomarkers for contaminant-related effects. The overall objective of this doctoral thesis was to investigate biotransformation capacity of POPs and their potential endocrine disruptive effects in two contrasting ringed seal populations from the low contaminated Svalbard area and from the highly contaminated Baltic Sea. Biotransformation capacity was studied by determining the relationships between congener-specific patterns and concentrations of polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs), polybrominated diphenyl ethers (PBDEs) and their hydroxyl (OH)- and/or methylsulfonyl (MeSO2)-metabolites, and catalytic activities of hepatic xenobiotic-metabolizing phase I and II enzymes. The results suggest that the biotransformation of PCBs, PBDEs and toxaphenes in ringed seals depends on the congener-specific halogen-substitution pattern. Biotransformation products detected in the seals included OH-PCBs, MeSO2-PCBs and –DDE, pentachlorophenol, 4-OHheptachlorostyrene, and to a minor extent OH-PBDEs. The effects of life history state (moulting and fasting) on contaminant status and potential biomarkers for endocrine disruption, including hormone and vitamin homeostasis, were investigated in the low contaminated ringed seal population from Svalbard. Moulting/fasting status strongly affected thyroid, vitamin A and calcitriol homeostasis, body condition and concentrations of POPs and their OH-metabolites. In contrast, moulting/fasting status was not associated with variations in vitamin E levels. Endocrine disruptive effects on multiple endpoints were investigated in the two contrasting ringed seal populations. The results suggest that thyroid, vitamin A and calcitriol homeostasis may be affected by the exposure of contaminants and/or their metabolites in the Baltic ringed seals. Complex and non-linear relationships were observed between the contaminant levels and the endocrine variables. Positive relationships between circulating free and total thyroid hormone concentration ratios and OH-PCBs suggest that OH-PCBs may mediate the disruption of thyroid hormone transport in plasma. Species differences in thyroid and bone-related effects of contaminants were studied in ringed and grey seals from low contaminated references areas and from the highly contaminated Baltic Sea. The results indicate that these two species living at the same environment approximately at the same trophic level respond in a very different way to contaminant exposure. The results of this thesis suggest that the health status of the Baltic ringed seals has still improved during the last decade. PCB and DDE levels have decreased in these seals and the contaminant-related effects are different today than a decade ago. The health of the Baltic ringed seals is still suggested to be affected by the contaminant exposure. At the present level of the contaminant exposure the Baltic ringed seals seem to be at a zone where their body is able to compensate for the contaminant-mediated endocrine disruption. Based on the results of this thesis, several recommendations that could be applied on monitoring and assessing risk for contaminant effects are provided. Circulating OH-metabolites should be included in monitoring and risk assessment programs due to their high toxic potential. It should be noted that endogenous variables may have complex and highly variable responses to contaminant exposure including non-linear responses. These relationships may be further confounded by life history status. Therefore, it is highly recommended that when using variables related to endocrine homeostasis to investigate/monitor or assess the risk of contaminant effects in seals, the life history status of the animal should be carefully taken into consideration. This applies especially when using thyroid, vitamin A or calcitriolrelated parameters during moulting/fasting period. Extrapolations between species for assessing risk for contaminant effects in phocid seals should be avoided.Siirretty Doriast

Concentrations and endocrine disruptive potential of phthalates in marine mammals from the Norwegian Arctic

This study investigated concentrations of phthalates (diesters of phthalic acids) in blubber/adipose tissue of blue whales (Balaenoptera musculus), fin whales (Balaenoptera physalus), bowhead whales (Balaena mysticetus) and polar bears (Ursus maritimus) sampled in the Svalbard Archipelago (extending westward in the case of bowhead whales). Additionally, total concentrations (free and conjugated forms) of eight phthalate monoester metabolites were analysed in plasma of polar bears. Bis(2-ethylhexyl) phthalate (DEHP) was the only phthalate quantified among the 12 phthalates investigated. This compound was present in 6/7 fin whale samples, 4/7 blue whale samples, 2/5 bowhead whale samples and 1/12 polar bear samples. DEHP concentrations ranged from <20–398 ng/g wet weight. Phthalate metabolites, mono-n-butyl phthalate and monoisobutyl phthalate, were found in low concentrations (<1.2 ng/mL) in some of the polar bear samples. In vitro reporter gene assays were used to assess transcriptional activity of fin whale peroxisome proliferator-activated receptor gamma (PPARG), glucocorticoid receptor (GR) and the thyroid hormone receptor beta (THRB) by DEHP and diisononyl phthalate (DiNP). Due to the high degree of similarity of the ligand binding domain in the THRB and PPARG among whales, polar bears and humans, the transactivation results also apply for these species. DEHP showed both agonistic and antagonistic effects towards whale THRB at considerably higher concentrations than measured in the study animals; DiNP was a weak agonist of whale THRB. No significant agonistic or antagonistic effects were detected for DEHP or DiNP for whale PPARG, whereas DEHP and DiNP decreased basal luciferase activity mediated by whale GR at several test concentrations. In conclusion, DEHP was detected in the blubber of marine mammals from the Norwegian Arctic and it appears to have potential to modulate the transcriptional activity of whale THRB, but current DEHP concentrations do not modulate the function of the studied nuclear receptors in adipose tissue of blue whales, fin whales, bowhead whales or polar bears sampled from the Norwegian Arctic.publishedVersio

Establishment of killer whale (Orcinus orca) primary fibroblast cell cultures and their transcriptomic responses to pollutant exposure

Populations of killer whale (Orcinus orca) contain some of the most polluted animals on Earth. Yet, the knowledge on effects of chemical pollutants is limited in this species. Cell cultures and in vitro exposure experiments are pertinent tools to study effects of pollutants in free-ranging marine mammals. To investigate transcriptional responses to pollutants in killer whale cells, we collected skin biopsies of killer whales from the Northern Norwegian fjords and successfully established primary fibroblast cell cultures from the dermis of 4 out of 5 of them. Cells from the individual with the highest cell yield were exposed to three different concentrations of a mixture of persistent organic pollutants (POPs) that reflects the composition of the 10 most abundant POPs found in Norwegian killer whales (p,p’-DDE, trans-nonachlor, PCB52, 99, 101, 118, 138, 153, 180, 187). Transcriptional responses of 13 selected target genes were studied using digital droplet PCR, and whole transcriptome responses were investigated utilizing RNA sequencing. Among the target genes analysed, CYP1A1 was significantly downregulated in the cells exposed to medium (11.6 µM) and high (116 µM) concentrations of the pollutant mixture, while seven genes involved in endocrine functions showed a non-significant tendency to be upregulated at the highest exposure concentration. Bioinformatic analyses of RNA-seq data indicated that 13 and 43 genes were differentially expressed in the cells exposed to low and high concentrations of the mixture, respectively, in comparison to solvent control. Subsequent pathway and functional analyses of the differentially expressed genes indicated that the enriched pathways were mainly related to lipid metabolism, myogenesis and glucocorticoid receptor regulation. The current study results support previous correlative studies and provide cause-effect relationships, which is highly relevant for chemical and environmental management.publishedVersio

Supporting Information: Influence of Carbon and Lipid Sources on Variation of Mercury and Other Trace Elements in Polar Bears (\u3ci\u3eUrsus maritimus\u3c/i\u3e)

MATERIALS AND METHODS Stable Carbon Isotope Analysis and Quality Control Fatty Acids Analysis and Quality Control Mercury and Other Trace Metal and Element Analysis and Quality Control REFERENCE SECTIO

MOSJ status report for environmental pollutants in 2011

English version of the NPI report no.137 (2012)