Year of Award

2014

Document Type

Thesis

Degree Type

Master of Science (MS)

Degree Name

Toxicology

Department or School/College

Department of Biomedical and Pharmaceutical Sciences

Committee Chair

Yoon Hee Cho

Commitee Members

Andrij Holian, Kent Pinkerton

Keywords

Epigenetic Changes, Prenatal, Environmental, Tobacco Smoke Exposure, Asthma

Subject Categories

Immunology of Infectious Disease | Other Pharmacology, Toxicology and Environmental Health | Toxicology

Abstract

Allergic asthma affects over 300 million people worldwide. Multiple factors have been shown to alter an individuals' susceptibility to this allergic airway disease such as exposure to polycyclic aromatic hydrocarbons (PAHs), endotoxin, and environmental tobacco smoke (ETS). Prenatal exposure to these contaminants, especially ETS, has been shown to increase risk for an individuals’ asthma risk, however, the mechanism by which this happens is still unknown. Epigenetic alterations, particularly DNA methylation patterns, have been introduced as a proposed mechanism. The goals of this project were to use a house dust mite (HDM) murine model of asthma to determine the methylation alterations of asthma-associated genes and the immunological effects after prenatal exposure to a low dose of ETS. The asthma-associated genes used in this experiment are genes encoding for: Th2 cytokines, Il-4 and Il-13, Th1 cytokine, Ifn-γ, and regulatory molecule, Foxp3. Male C57BL/6 mice were exposed prenatally to 1.0 mg/m3 of ETS and later challenged with HDM at 6 weeks old. Promoter regions of the asthma-associated genes were analyzed via bisulfite sequencing, using a Qiagen® PyroMark Q96MD pyrosequencer. Cytokines in bronchoalveolar lavage fluid (BALF) were analyzed using Quantikine ELISA (R&D) for IL-13 and Mesoscale Discovery® Pro-inflammatory Panel 1 kit for IFN-γ, IL-4 and IL-5. Serum IgE levels were determined using a BioLegend IgE ELISA kit. We concluded that prenatal exposure to low doses of ETS causes a severe increase in the immune response when challenged with HDM. We also found Ifn-γ to be hypermethylated in the HDM group and further hypermethylated after prenatal ETS exposure and Il-4 to be hypomethylated in the HDM group and further hypomethylated after ETS exposure. This suggests that prenatal ETS exposure increases the susceptibility to an allergic and asthmatic response when exposed to allergen, with promoter methylation as a proposed mechanism.

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© Copyright 2014 Sonja Christensen