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Development of an in vitro method to study heterogeneous particle mimic exposures

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Thesis type
(Thesis) Ph.D.
Date created
Particulate matter, a heterogeneous mixture of solid and liquid particles suspended in the atmosphere, has been strongly linked to several respiratory diseases and cardiovascular disease. Due to its impact on human health, epidemiological, in vivo, and in vitro studies are used to assess toxicity of diverse types of particulate matter, and mechanisms behind disease pathogenesis. To enable insight on how particle composition affects cellular response, an in vitro dose-response apparatus and methodology were developed to deliver heterogeneous particles of controlled compositions onto human lung cell mono- and co-cultures. Silica particles, an ambient particle type sourced from crustal matter, were generated using the apparatus and were delivered onto cultures of alveolar epithelial cells (A549) up to an estimated 3.2 µg cm-2. No change was observed in membrane bound protein intercellular adhesion molecule-1 (ICAM-1) expression. In a proof of concept study, tumor necrosis factor (TNF)-α, a potent cytokine, was introduced as a secondary component to silica. This binary particle type was also delivered onto A549 monocultures, and ICAM-1 signal increased with an increase in dose. Silica particles were also delivered onto co-cultures containing A549 cells and macrophages differentiated from blood monocyte cells (THP-1). Cytokine interleukin (IL)-6 was significantly up-regulated in co-cultures, but IL-8 was not. Lipopolysaccharide (LPS), a component of the cell wall in Gram negative bacteria, and nickel, in the form of Ni(NO3)2·6H2O, were introduced as secondary components to silica. These binary particle types were generated at ratios representative of respirable agricultural soils and dosed onto co-cultures. It was observed that silica plus LPS particles caused an up-regulation of IL-6 and IL-8 response relative to silica particles, whereas silica plus nickel particles did not. LPS bioavailability was determined as 10±3% and 1.0±0.6% for silica plus LPS particles at atmospherically-relevant LPS to silica mass ratios of 91 ppm and 910 ppm, respectively. The reduction in bioavailability showed how interactions between compositions in a heterogeneous particle type can affect cellular response.
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Supervisor or Senior Supervisor
Thesis advisor: Agnes, George
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