Date of Award

Summer 8-2017

Level of Access Assigned by Author

Open-Access Thesis

Degree Name

Master of Science (MS)

Department

Biomedical Sciences

Advisor

Carol H. Kim

Second Committee Member

Paul J. Millard

Third Committee Member

Melissa S. Maginnis

Abstract

Cystic fibrosis (CF) is a genetic disorder that affects 30,000 people in the United States and currently has no cure. Although CF affects all of the body’s systems, it is largely characterized as a lung disease. CF is caused by a mutation in both copies of the gene for cystic fibrosis transmembrane conductance regulator (CFTR). A mutation in the CFTR gene leads to improper movement of chloride ions and water into the airways, which dysregulates the airway surface liquid volume and composition. Individuals with CF are prone to lung infections due to inefficient bacterial clearance and by the age of 18, eighty percent of patients with CF harbor Pseudomonas aeruginosa, with it being the most prevalent respiratory microorganism at that age.

P. aeruginosa is a gram negative, opportunistic bacterium that is found all over the planet. It is a serious threat to people with CF due to its ability to form large colonies known as mucoid pseudomonas. The mucoid strain of P. aeruginosa provides increased resistance to not only antibiotics but also opsonization and phagocytosis. Even if P. aeruginosa infection can be managed there is a delicate balance between controlling the infection and causing exacerbated neutrophilia and tissue damage due to increased inflammation. Many factors can affect the balance of the immune system including environmental toxicants such as arsenic.

Arsenic exposure is a health risk to many people worldwide through contaminated drinking water. Arsenic is able to leech out of granite and other sediments into groundwater and subsequently become included in drinking water. Arsenic has been shown to affect the immune response to pathogens in the lungs and degrade CFTR, which can lead to increased infection and exacerbated CF.

It is important to research the effects of CF, P. aeruginosa, and arsenic to understand how they are modulating the immune system and to potentially discover novel therapies to combat CF and bacterial infection. The Kim lab recently completed RNA sequencing analysis to measure the dysregulation of genes in response to these factors in zebrafish embryos. To understand how the immune system is being modulated by these factors the role of non-coding RNAs (ncRNA) were investigated. ncRNA have remained largely uncharacterized and, until recently, were thought to serve little to no purpose in our genome. Recent studies have shown ncRNA to be involved in many important biological processes such as embryonic development, transcriptional regulation, apoptosis, and immunity. The Kim lab is particularly interested in the role of micro-RNAs (miRNA) and long intergenic non-coding RNAs (lincRNAs).

In order to better understand and characterize ncRNAs it is critical to study their mechanisms in vivo. The zebrafish (Danio rerio) is regarded as an excellent research model, because of the ease with which it can be genetically manipulated, the availability of transgenic lines, and its sole reliance on the innate immune system during the first 4-6 weeks of development. In addition, the zebrafish’s optical clarity allows for the use of fluorescently-labeled bacterial strains to track and image infection in live zebrafish embryos. With the use of the zebrafish and high-throughput sequencing, the lab aims to aid in the discovery of novel therapies for people suffering from cystic fibrosis and bacterial infection.

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