Yeh 2017 Thesis
|Yeh A (2017) Modulation of mitochondrial function as an indicator for sublethal effects of contaminants of emerging concern. Dissertation p117.|
Abstract: Effluent from wastewater treatment plants (WWTPs) are a primary source of a wide range of structurally-diverse compounds into Puget Sound, WA. These contaminants of emerging concern (CECs) include personal care products and pharmaceuticals (PPCPs), perfluorinated compounds, natural and synthetic hormones, alkylphenol and alkylphenol ethoxylate surfactants, current-use pesticides, and polybrominated diphenyl ether (PBDE) flame retardants. Currently, the types and concentrations of these emerging contaminants entering Puget Sound, and their effects on aquatic receptors, are poorly understood. Recent reports in numerous species of fish indicate exposure to CECs can cause sublethal, detrimental effects on growth, reproduction, and behavior, warranting development of biomarkers of exposure and effect of CECs in relevant biomonitoring species. There is evidence to suggest that numerous CECs negatively affect mitochondrial function in humans and laboratory animal models. Hence, the overarching hypothesis of my doctoral research was that mitochondrial dysfunction can serve as an indicator of sublethal effect, and exposure to, emerging contaminants.
Both human health- and aquatic health-related effects of CECs on mitochondrial function were addressed in my dissertation project. Aim 1 of my project was an oceans and human health study concerning the safety of salmon consumption by humans. The tested hypothesis was that the antioxidant effects of omega-3 polyunsaturated fatty acids found in salmon chemoprotect against mitochondrial and oxidative injury from a common “emerged” contaminant detected in Pacific salmon, the PBDE flame retardant 2,2’,4,4’-tetrabromodiphenyl ether (BDE 47). HepG2 cells were treated for 12 h with a mixture of omega-3s relevant to salmon consumption (oxEPA/oxDHA), followed by exposure to BDE 47 (100 μM) for 24 h. Pretreatment with oxEPA/oxDHA prevented BDE 47-induced production of reactive oxygen species (ROS) and depletion of GSH, significantly increased expression of protective cellular antioxidant response genes, and protected against BDE 47-induced loss of viability and mitochondrial membrane potential. Analysis of mitochondrial electron transport system (ETS) revealed extensive inhibition of state 3 respiration and maximum respiratory capacity by BDE 47 were partially reversed by treatment with oxEPA/oxDHA. These findings indicated that the antioxidant effects of salmon omega-3s protected against short exposures to BDE 47, including a protective role of these compounds on maintaining cellular and mitochondrial function.
The subsequent goal of my research was to examine the potential impacts of CECs in Puget Sound aquatic organisms. Thus, studies in Aim 2 involved an analytical field study to determine the specific types of CECs present in Puget Sound, and their potential to bioaccumulate in representative biomonitoring species. Samples of estuary water, WWTP effluent, and whole-bodies of Chinook salmon and staghorn sculpin were collected from three sites: two estuaries that receive effluent from WWTPs and a minimally-polluted estuarine reference site. Samples were analyzed for 150 chemical analytes by HPLC/MS/MS. In total, 81 analytes were detected in effluent samples in the ng/L to low μg/L range, 25 analytes were detected in estuary water samples at ng/L concentrations, and 42 analytes were detected in fish whole-bodies at ng/g concentrations. Noteworthy findings included high concentrations of CECs measured in WWTP effluent relative to other major U.S. WWTPs, greater-than-expected contamination of the reference site, and preferential bioaccumulation of CECs in Chinook salmon relative to sculpin. These results can inform environmental risk assessment or ecological studies at other sites in Puget Sound.
The analytical field study provided crucial data that was required to conduct subsequent experiments in Aim 3 that investigated the association between exposure to CECs and liver mitochondrial dysfunction in fish. Mitochondrial function was assessed in feral juvenile Chinook from the analytical field study, and in hatchery-reared juvenile Chinook exposed in the laboratory to a dietary mixture of CECs representative of the predominant contaminants detected in the field. Liver mitochondrial content was reduced in fish exposed to CECs in the laboratory, which occurred concomitantly with a reduction in expression of peroxisome proliferator-activated receptor (PPAR)-γ coactivator-1a (pgc-1α), a positive transcriptional regulator of mitochondrial biogenesis. The laboratory exposures also caused elevation of state 4 respiration per unit mitochondria, which drove a reduction of efficiency of oxidative phosphorylation relative to controls. The mixture-induced elevation of respiration was associated with increased oxidative injury as evidenced by increased mitochondrial protein carbonyls, elevated expression of glutathione (GSH) peroxidase 4 (gpx4), a mitochondrial-associated GSH peroxidase that protects against lipid peroxidation, and reduction of mitochondrial GSH. Juvenile Chinook sampled in from a WWTP effluent-impacted estuary with demonstrated releases of CECs showed similar trends toward reduced liver mitochondrial content and elevated respiratory activity per mitochondria (including state 3 and uncoupled respiration). Interestingly, respiratory control ratios were greater in fish from the contaminated site relative to fish from the reference site, which may have been due to differences in the timing of exposure to CECs under laboratory and field conditions. Collectively, these results demonstrate that exposure to environmentally-relevant mixtures of CEC can cause sublethal impacts on the bioenergetics of fish, including modulation of both quantity and quality of liver mitochondria.
Labels: MiParea: Respiration, Comparative MiP;environmental MiP, Pharmacology;toxicology
Organism: Human, Fishes Tissue;cell: Liver Preparation: Permeabilized cells, Permeabilized tissue
Coupling state: LEAK, OXPHOS, ET Pathway: N, CIV, NS, ROX HRR: Oxygraph-2k