Publication Date

2018

Document Type

Thesis

Committee Members

Saber Hussain (Advisor), Mark Nelson (Committee Member), Terry Oroszi (Committee Member), Richard Salisbury (Committee Member), Jeffrey Travers (Committee Member)

Degree Name

Master of Science (MS)

Abstract

Platelet Activating Factor (PAF) and its associated receptor, the PAF Receptor (PAFR), are important mediators of intercellular communication during an immune response. Once a physiological stimulus triggers an inflammatory response, epithelial, endothelial and immune cells synthesize and release PAF. PAF mediates the recruitment of immune cells, platelets, angiogenesis, expression of various genes, and increased PAF biosynthesis (Brown, 2006; Han, 2006; Whatley, 1988; Axelrod, 1988). In this study, we utilized HaCaT cells and a well characterized KB cell line derived from nasopharyngeal cells, which do not natively express the PAFR. KB cells had previously been transfected with a PAF receptor (KBP) or a mock transfection (KBM), yielding two cell lines. The KBM and KBP cell lines allow for a mechanistic in vitro assessment of charge-dependent nanoparticle (NP) mediated PAFR activation, where KBP cells should show more robust Interleukin-8 (IL-8) secretion than KBM cells. Furthermore, this cell model serves to identify PAFR agonist formation, which has been reported to be generated from cigarette smoke, ultraviolet B radiation, jet fuel, and other stimuli that result in oxidative stress. The PAFR is involved in clathrin-mediated endocytosis, yet the PAFR role in NP uptake has not been investigated. The ability of NPs to generate PAF agonists is not known and the mechanisms by which they are internalized have yet to be fully understood. To investigate whether NP charge contributes to direct PAF-like lipid formation, KBM, KBP and HaCaT cell lines were exposed to 1 to 100[micro]g/mL of 40nm Ag-NPs that were functionalized with either branched polyethyleneimine (BPEI) or citrate to give them a positive or negative surface charge, respectively. We demonstrated; a charge-dependent increase in NP-induced IL-8 production in KBP and HaCaT cells; indirectly measured an increased activation of the PAFR after 24 hours of exposure in KBP and HaCaT cells; dose-dependent uptake of NPs in all cell lines and increased reactive oxygen species (ROS) generation in KB cell lines only. Further analysis of NP-induced PAFR activation may grant us a better understanding of dermal inflammation and the potential role of PAF-like lipids in NP-mediated wound healing therapies.

Page Count

55

Department or Program

Department of Pharmacology and Toxicology

Year Degree Awarded

2018

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