Altered expression and activity of phase I and II biotransformation enzymes in human liver cells by perfluorooctanoate (PFOA) and perfluorooctane sulfonate (PFOS).

Human exposure assessments for perfluorooctanoate (PFOA) and perfluorooctane sulfonate (PFOS) have been mostly limited to the quantification of these chemicals in different environmental matrices, but only a few studies have addressed toxicological aspects associated with them. It has been suggested that both PFOA and PFOS are highly stable chemicals that are not metabolized, yet previous reports have described abnormal activity of important biotransformation pathways. Therefore, the goal of the present study was to investigate the effects of PFOA and PFOS on phase I and II biotransformation enzymes at the gene expression and activity levels, and by using the well-established human liver HepaRG cell line. Cells were exposed to a wide range of PFOA and PFOS concentrations for 24 or 48 h, prior to cytotoxicity measurements, and quantification of expression and activity of three cytochrome P450 enzymes (CYP1A2, CYP2C19 and CYP3A4) and two conjugation enzymes (glutathione-S-transferase (GST-M1) and UDP-glucuronosyltransferase (UGT-1A1)). Expression of all CYP enzymes was significantly reduced from exposure to both PFOA and PFOS after 48 h and from concentrations as low as 40-50 ng/L, with CYP3A4 also presenting the lowest activity. Among the conjugation enzymes, the expression of UGT was significantly reduced only by PFOA after 48 h of exposure, yet no significant alterations in its activity were observed. While the specific chemico-biological interactions of these compounds with gene expression and biotransformation pathways is not clear, the results from this study suggest that the interference of PFOA and PFOS with phase I and II biotransformation enzymes could potentially lead to adverse outcomes resulting from the inability of biotransformation pathways to function as needed.

Oxidative Potential of Chemical Mixtures Extracted from Contaminated Galveston Bay, TX Seafood Using a Human Cell Co-culture Model.

Increasing levels of pollution in Galveston Bay, TX, are of significant concern for populations that directly depend on fishing activities. Efforts to evaluate contaminant levels in commercial fish have been largely limited to the quantification of chemical mixtures in fish tissue, but little information exists about the toxicological potential of these chemicals on consumption of contaminated seafood. The present study makes use of a human cell co-culture model, mimicking the digestive system, to address the oxidative potential of chemical mixtures in seafood. Chemical extractions were performed on fillets from three fish species and oysters collected from different areas in Galveston Bay. The resulting extracts were used to expose intestinal and liver cells before the measurement of cytotoxicity and activity of antioxidant enzymes. The pesticide 4,4'-DDE was found in nearly all samples from all sites in concentrations ranging from 0.23-9.4 µg/kg. Similarly, total PCBs found in fish and oyster tissue ranged from 0.68-65.65 µg/kg, with PCB-118 being the most common congener measured. In terms of cytotoxicity, oyster extracts led to significant cell mortality, contrary to observations for fish extracts. Antioxidant enzymes, while not directly related to the presence of chemical mixtures in tissue, presented evidence of potential increases in activity from spotted trout extracts. Observations from this study suggest the need to evaluate toxicological aspects of contaminated seafood and support the use of in vitro models for the screening of accumulated chemicals.

Evaluating toxicity risk in sediments after remediation at a Superfund megasite using a Triad approach.

The Pine River downstream of the Velsicol Superfund site has been contaminated with various hydrophobic organic pollutants for more than 50 years. Remediation and sediment dredging near the site began in spring of 1999, and was completed in 2006. In 2011, the Michigan Department of Environmental Quality completed a baseline assessment report long-term monitoring plan for the Pine River. However, there has been limited assessment of the benthic community since this evaluation. The objective of this research was to evaluate the risk of Pine River sediment to aquatic macroinvertebrates downstream from the Superfund site after decades of degradation and dredging using the Triad approach. Three sites were selected downstream from the Superfund site, and an upstream reference site was used. At each site, macroinvertebrates surveys were conducted and sediments were collected for chemical analysis of DDT (1,1,1-trichloro-2,2-bis (4-chlorophenyl) ethane) and its degradation products and for laboratory toxicity testing for mortality and sublethal effects using Hyalella azteca and Chironomus dilutus. Sediment concentrations of DDT, DDD, and DDE were below levels expected to cause toxicity, and there was no observed toxicity in laboratory tests. Additionally, there were no statistically significant differences in richness, richness of Ephemeroptera, Plecoptera, and Trichoptera (EPT) species, total EPT abundance, percent EPT, or percent dominant taxa between the reference site and the downstream sites. There was an observed decrease in abundance of macroinvertebrate taxa at all downstream sites and a shift in macroinvertebrate structure when comparing the reference with most impaired sites. Although the sites downstream of the Superfund site remain different than the upstream control, there are improvements in species composition and abundance. However, more research is needed to evaluate the potential effects on ecosystem function.

Applicability of a human cell co-culture model to evaluate antioxidant responses triggered by chemical mixtures in fish and oyster homogenates.

The accumulation of chemical compounds in fish tissue represents significant health concerns for seafood consumers, but little is known about the risks to human health associated with such substances. The identification of adverse biological responses upon exposure to contaminants has been facilitated by the development of in vitro systems resembling the human dietary pathway. The present study explores the applicability of an organotypic co-culture system, using intestinal (Caco-2) and hepatic (HepaRG) cell lines, to provide insight into the toxicity of chemical mixtures found in commercially available seafood. Chemical extractions were conducted utilizing fish and oyster standard reference material (SRM) from the U.S. National Institute of Standards and Technology (NIST). Cells were seeded in monoculture and co-culture systems and exposed to SRM extracts before measurements of cytotoxicity and antioxidant responses. Exposure to oyster extracts led to significant cell mortality in monocultures. HepaRG cells in monoculture expressed lower levels of glutathione peroxidase and superoxide dismutase than HepaRG cells in co-culture, upon exposure to both oyster and fish extracts. These observations illustrate the importance of organotypic co-culture models to explore biological responses that could be otherwise difficult to evaluate in monocultures, and the adverse effects associated with the consumption of contaminated seafood.

Xenobiotic metabolism in the fish hepatic cell lines Hepa-E1 and RTH-149, and the gill cell lines RTgill-W1 and G1B: Biomarkers of CYP450 activity and oxidative stress.

The use of fish cell cultures has proven to be an effective tool in the study of environmental and aquatic toxicology. Valuable information can be obtained from comparisons between cell lines from different species and organs. In the present study, specific chemicals were used and biomarkers (e.g. 7-Ethoxyresorufin-O-deethylase (EROD) activity and reactive oxygen species (ROS)) were measured to assess the metabolic capabilities and cytotoxicity of the fish hepatic cell lines Hepa-E1 and RTH-149, and the fish gill cell lines RTgill-W1 and G1B. These cell lines were exposed to ß-naphthoflavone (BNF) and benzo[a]pyrene (BaP), the pharmaceutical tamoxifen (TMX), and the organic peroxide tert-butylhydroperoxide (tBHP). Cytotoxicity in gill cell lines was significantly higher than in hepatic cells, with BNF and TMX being the most toxic compounds. CYP1-like associated activity, measured through EROD activity, was only detected in hepatic cells; Hepa-E1 cells showed the highest activity after exposure to both BNF and BaP. Significantly higher levels of CYP3A-like activity were also observed in Hepa-E1 cells exposed to TMX, while gill cell lines presented the lowest levels. Measurements of ROS and antioxidant enzymes indicated that peroxide levels were higher in gill cell lines in general. However, levels of superoxide were significantly higher in RTH-149 cells, where no distinctive increase of superoxide-related antioxidants was observed. The present study demonstrates the importance of selecting adequate cell lines in measuring specific metabolic parameters and provides strong evidence for the fish hepatocarcinoma Hepa-E1 cells to be an excellent alternative in assessing metabolism of xenobiotics, and in expanding the applicability of fish cell lines for in vitro studies.