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http://hdl.handle.net/11375/24164
Title: | Direct Effects of Pesticides on Brown Adipose Tissue |
Other Titles: | In vitro Effects Of Pesticides On Brown Adipose Tissue Function |
Authors: | Llanos, Andrea |
Advisor: | Holloway, Alison |
Department: | Health Sciences |
Publication Date: | 2018 |
Abstract: | Obesity and its associated comorbidities can occur when energy consumption exceeds energy expenditure. Brown adipose tissue (BAT) is an important contributor to energy expenditure as its mitochondria are rich in uncoupling protein 1 (UCP-1). UCP-1 uncouples oxidative phosphorylation from the generation of ATP to induce a futile cycle and consequently the generation of heat. Recent studies demonstrate that reduced BAT activity is correlated with many chronic metabolic diseases including obesity and type 2 diabetes, but the mechanisms mediating these effects are currently unclear. There are an increasing number of reports describing how pesticides, currently used to increase crop production, contribute to the development of metabolic syndrome including effects on adipogenesis, food intake, glucose and lipid homeostasis being reported. However, the effects of these compounds on BAT remain largely unknown. The objective of this study was to examine the direct effects of pesticides used to increase crop production on BAT function. Nine pesticides and their primary metabolites were chosen to enter a 3-tiered screening process. Compounds were selected based on widespread usage and/or reports of metabolic perturbations in epidemiological studies, or animal experiments. Immortalized BAT cells derived from UCP-1 luciferase reporter mice, or wild-type FVB/N mice were treated with compounds at 1pM, 1nM, and 1M doses. To elucidate the effects of selected pesticides on UCP-1 expression, UCP-1 promoter activity, mRNA and protein levels were quantified using a luciferase assay system, RT-qPCR, and western blotting respectively. The most consistent compound was further tested to determine its effects on BAT function by whole cell respiration assay and mitochondrial enzyme activity assays (i.e., citrate synthase, and cytochrome c oxidase assay). Of the 18 compounds that entered the primary screen, 8 compounds reduced UCP-1 promoter activity by >25%. RT-qPCR revealed that 5 compounds reduced UCP-1 mRNA levels at the same doses by 25% or more. Of these 5 remaining compounds, 3 also exhibited a reduction in UCP-1 protein content by >25%. Of these 3 compounds, chlorpyrifos was most consistent and further analyzed for its effects on BAT function. Chlorpyrifos reduced the expression of genes important for mitochondrial function, while also reducing maximal respiratory capacity and the activity of cytochrome c oxidase of treated brown adipocytes. These data demonstrate that the commonly used pesticide, chlorpyrifos, directly suppresses UCP-1 expression and BAT function of cultured brown adipocytes from mice at concentrations as low as 1pM. Furthermore, these data also show that its action might be in part, by perturbations in complex IV (cytochrome c oxidase) of the electron transport chain of BAT mitochondria, in addition to the suppression of UCP-1. |
URI: | http://hdl.handle.net/11375/24164 |
Appears in Collections: | Open Access Dissertations and Theses |
Files in This Item:
File | Description | Size | Format | |
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Llanos_Andrea_M_Sept2018_MSc(Master of Health Science).pdf | 1.68 MB | Adobe PDF | View/Open |
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