Medical Sciences Commons^™

Secretion Of Salivary Proteins And Their Interactions, Karla Tonelli Bicalho Crosara

Electronic Thesis and Dissertation Repository

Saliva’s clinical application for disease diagnosis and monitoring is limited by incomplete knowledge of salivary protein interactions, the effect of stimulation on the salivary proteome, and if these factors impact protein identification. This thesis expands knowledge of the salivary interactome and effects of stimulation intensity and duration on parotid saliva’s proteome. First, previous in vitro studies identified 43 proteins in the histatin 1-protein network and demonstrated histatin 1’s increased stability in whole saliva when interacting with amylase. We hypothesized that protein-protein-interaction databases could enlarge the histatin 1-protein network. A comprehensive histatin 1-protein network was created using STRING database, merging previous …

A Characterization Of Rgnef Biophysical Properties And Interactome, Brooke E. Wile

Electronic Thesis and Dissertation Repository

ALS is a progressive and fatal neurodegenerative disorder whose pathologic hallmark is the presence of neuronal cytoplasmic inclusions (NCIs). In approximately 97% of ALS cases, NCIs are found to be TDP-43+. Rho-guanine nucleotide exchange factor (RGNEF) has recently been implicated in ALS pathophysiology through its co-localization and coimmunoprecipitation with TDP-43+. RGNEF has also been shown to harbour cytoprotective effects in the N-terminal region and is responsible for the regulation of low molecular weight-neurofilament (NFL), intimately involved neural structure, through its predicted RNA-binding domain (RBD). This study looked to purify constructs of RGNEF through nickel immobilized metal affinity chromatography (Ni-IMAC) and …

Creating Tools To Study The Signaling And Function Of The Adhesion Family Of Gpcrs, Victor M. Mirka

Electronic Thesis and Dissertation Repository

Adhesion GPCRs (aGPCRs) are difficult to study because they are activated by mechanical force. aGPCRs are autoproteolytically cleaved into N-terminal and C-terminal fragments. Mechanical force removes the N-terminal fragment revealing a tethered ligand activating the receptor. Proteinase Activated Receptors (PARs) are N-terminally cleaved by proteinases revealing a tethered ligand activating the receptor. We hypothesized the tethered ligand of aGPCRs could be revealed by replacing the N-terminal fragment with a PAR N-terminus. We fused the PAR2 N-terminus to the C-terminal fragments of four aGPCRs: CD97, EMR2, GPR56, and BAI1. PAR2-aGPCR chimeric receptors dose dependently recruited G-proteins and β-arrestins, supporting our hypothesis. …

Trna Regulation In Humans: The Cellular Effect Of A Pathological Hars Y454s Mutation, Rosan Kenana

Electronic Thesis and Dissertation Repository

tRNAs are the adapter molecules involved in translating the genetic code into functional protein in a living cell. tRNAs are charged with their cognate amino acids - by aminoacyl-tRNA synthetases (aaRS or ARS) - which are then transferred to a growing peptide in a process called mRNA translation. The efficiency of translation is dependent on the ratio of ARS enzymes to their cognate tRNAs and the availability of correctly amino acylated tRNAs. Disruptions of this process, caused by mutations in ARS genes, in particular, have been linked to complex inherited diseases. USH3B syndrome, a recessively inherited disorder among consanguineous families …

Characterizing The Role Of Tdg In Fxr-Dependent Signaling, Oladapo A. Onabote

Electronic Thesis and Dissertation Repository

Thymine DNA Glycosylase (TDG) plays a key role in active demethylation by excising intermediates of 5-methylcytosine. The function of TDG is required for embryonic development, as Tdg-null embryos die at E11.5. To bypass this embryonic lethality, our lab generated conditional Tdg knockout (TDG_CKO) mice. These mice develop late-onset hepatocellular carcinoma (HCC), partly due to impaired Farnesoid X Receptor (FXR) signaling. Interestingly, Fxr-knockout mice display a similar phenotype and transcriptional profile to TDG_CKO mice, prompting us to investigate a role for TDG in FXR signaling. To this end, we generated Tdg/Fxr double-knockout (DKO) mice. …

Discovery Of Novel Mechanisms Regulating Cancer Extravasation In The Chorioallantoic Membrane Model, Yohan Kim

Electronic Thesis and Dissertation Repository

Cancer metastasis is a multistep process that begins with the invasion of tumour cells into the stroma and migration towards the blood vessels. Tumour cells that have entered the bloodstream must then survive and leave by a process known as extravasation. Finally, extravasated cells proliferate and establish the secondary site in the metastatic cascade. Although extravasation encompasses key events during cancer cell invasion to aid in the development of effective treatments, an in vivo model that rapidly, reproducibly and economically recapitulates cancer cell extravasation is needed. Therefore, the objectives of my research were to 1) establish and validate an in …

Modulation Of Inflammation Driven Wound Healing After Glaucoma Surgery, James J. Armstrong

Electronic Thesis and Dissertation Repository

Dysregulated wound healing contributes to most currently unanswered ophthalmological morbidity. Opacification and structure altering contractures compromise the delicate ocular anatomy upon which ocular function and healthy vision are reliant. Glaucoma filtration surgery, corneal stromal injury, proliferative vitreoretinopathy and age-related macular degeneration are major contributors to ocular morbidity – all with myofibroblast transdifferentiation and pathognomonic scarring activity at their core.

This thesis aims to revaluate the means by which dysregulated ocular wound healing is combated with evidence describing a novel strategy to mitigate its effects. A translational approach was used. An initial retrospective analysis of over ten thousand glaucoma surgeries found …

Proteomic Characterization Of Human Multipotent Stromal Cells Secreted Proteins With Therapeutic Potential For Β-Cell Regeneration, Miljan Kuljanin

Electronic Thesis and Dissertation Repository

Novel strategies to stimulate the expansion of β-cell mass in situ are warranted for diabetes therapy. Cell-replacement therapies for the treatment of diabetes have become a focal point in recent years. Endogenous regeneration of β-cell mass has been demonstrated using human multipotent stromal cells (hMSC). However, the secretory factors responsible for initiating endogenous regeneration remain unknown. Successful large-scale proteomic applications to address these questions have been limited in part by difficulties in correctly selecting the appropriate methodologies. Thus, the goal of this thesis was a combination of assessing different proteomic workflows to facilitate investigation into hMSC biology, applying these methods …

Electrospun Collagen Nanofibers For Tissue Engineering, Nisha Sharma

Electronic Thesis and Dissertation Repository

Design and fabrication of the scaffold is an important part of the tissue engineering process. Nanofibrous scaffolds based on proteins are gaining increasing acceptance due to its structural similarity to the extracellular matrix. Making use of the electrospinning technique, rat tail collagen type I nanofibers were produced using a collagen in hexafluoroisopropanol (HFIP) solution. In addition to optimizing the electrospinning process parameters, the effect of humidity on fiber morphology and diameter was investigated for fiber size control for particular tissue engineering applications. A generalized humidity effect on polymer fiber diameter of the polymer solution electrospinning process was developed. The as …

The Role Of Bone Sialoprotein In The Tendon-Bone Insertion, Ryan M. Marinovich

Electronic Thesis and Dissertation Repository

Tendons and ligaments insert into bone through a transitional tissue termed the enthesis which is susceptible to injury and difficult to repair. Entheses contain a region of calcified fibrocartilage (CFC), however mineral-associated proteins in this tissue remain poorly characterized. Bone sialoprotein (BSP) is a phosphoprotein associated with mineralizing tissues. In these studies BSP was identified in the CFC of entheses by immunohistochemistry. Analysis of the entheses of Bsp^-/- mice indicate abnormalities in the CFC. Compared to controls, the CFC of the quadriceps tendon enthesis is 28% and 41 % longer in 15 week and 14 month old Bsp^{-/- …}

Inhibition Of Elongation Factor 1a-1 Activity And Hepatic Lipotoxicity, Alexandra Margaret Anne Hetherington

Electronic Thesis and Dissertation Repository

Elongation factor 1A-1 (eEF1A-1) was previously identified as a mediator of fatty acid-induced cell death (lipotoxicity) downstream of endoplasmic reticulum (ER) stress. Furthermore, inhibition of the peptide elongation activity of eEF1A-1 with the cyclic depsipeptide didemnin B (DB) diminishes ER stress and lipotoxicity in cultured hepatocytes. Since ER stress is involved in nonalcoholic fatty liver disease (NAFLD), it was hypothesized that administration of DB to obese mice with NAFLD would reduce hepatic lipotoxicity. Treatment with DB for one week improved several parameters associated with hepatic lipotoxicity and modestly decreased food intake without evidence of illness. Liver triglycerides and protein markers …

Nicotinamide Riboside Delivery Generates Nad+ Reserves To Protect Vascular Cells Against Oxidative Damage, Krista M. Hawrylyshyn

Electronic Thesis and Dissertation Repository

The ability of vascular cells to withstand oxidative insults is critical to vascular health. NAD⁺, which drives poly (ADP-ribose) polymerase (PARP) and sirtuin (SIRT) reactions, can be compromised and strategies for overcoming this limitation in the vasculature do not exist. This study determines if nicotinamide riboside (NR) delivery can augment NAD⁺ stores and fuel resistance to oxidative stress. I established that oxidative-stress insult on vascular cells decreased NAD⁺ levels, accompanied by a striking increase in nuclear PAR-chain accumulation. PARP inhibition abolished PAR-chain formation and preserved NAD⁺ levels, establishing PARP in NAD⁺ consumption in this …

The Effects Of Rhamm On Cell Division, Cell Motility, And Tumorigenesis, Sallie Elhayek

Electronic Thesis and Dissertation Repository

Overexpression of a RHAMM isoform (RHAMM^Δ163) transforms fibroblasts but the mechanisms underlying this oncogenic function are not well understood. RHAMM^Δ163 binds to the mitotic spindle and centrosomes via a C-terminal leucine zipper; these interactions are predicted to regulate genomic stability and cell polarity and proposed to account for the oncogenic function of RHAMM^Δ163. We hypothesized that RHAMM leucine zipper maintains mitotic spindle integrity and impacts directional cell migration through its interactions with microtubule and centrosome structures. The consequences of a mutated leucine zipper on cell division, cell motility, and tumorigenesis were assessed. Although mutant RHAMM …

Cortisol And Testosterone In Hair As Biological Markers Of Systolic Heart Failure, Justin Chan

Electronic Thesis and Dissertation Repository

Congestive heart failure (CHF) is associated with increased stress and alterations in metabolism, favoring catabolism over anabolism. Hormonal profiles of patients with heart failure have been assessed using serum and saliva as matrices, which are only point measurements and do not provide long-term information. Scalp hair is a novel matrix that allows for measurement of hormones over a period of several months. We aimed to evaluate whether levels of cortisol and testosterone and their ratio (C/T) in hair correlate with severity of heart failure. We conducted a prospective study in ambulatory male patients with a left ventricular ejection fraction (LVEF) …

The Role Of Oxygen Tension And Insulin-Like Growth Factor Signaling In The Placental Mesenchymal Stem Cell Fate, Amer Youssef

Electronic Thesis and Dissertation Repository

The human placenta of different gestational ages is a readily available source for isolation of adult mesenchymal stem cell (MSC) for potential use in regenerative therapies. The chorionic villous region, the largest component of a placenta that interfaces with the maternal circulation, is a rich source of placental MSCs (PMSCs). To remain multipotent, PMSCs are best maintained in culture conditions that mimic the in vivo microenvironment. Insulin like growth factors (IGFs, IGF-I and -II) and oxygen tension are two of the most important microenvironmental factors in the placenta. They are of low concentration or tension respectively, at early gestation, and …

Identification Of The Salivary Proteome In Children Throughout The Course Of Dental Eruption, Nada Tabbara

Electronic Thesis and Dissertation Repository

The salivary proteome is recognized as a valuable source of potential oral and systemic disease biomarkers. Major efforts in salivary research have been dedicated to identify and characterize salivary proteins present in saliva using both classical biochemical methods and proteomics approaches in adults. Despite considerable research on the salivary proteome, little attention has been given to the changes in the salivary proteome occurring in children, specifically from 0-3 years of age. Through the use of anionic PAGE, SDS PAGE, HPLC and MS/MS, salivary protein profiles in children before, during and after dental eruption were compared with edentulous adult controls. We …

The Cross Talk Between Trkb And Nmda Receptors Through Rasgrf1, Asghar Talebian

Electronic Thesis and Dissertation Repository

Introduction: Brain-derived neurotrophic factor (BDNF) facilitates neuronal differentiation by activation of the TrkB receptor-tyrosine-kinase. BDNF/TrkB also modulates the activity of the excitatory N-methyl-D-aspartate neurotransmitter receptor (NMDAR), thereby also regulating neural plasticity and LTP/LTD, two forms of synaptic plasticity that contribute to the storage of information inside the brain. How TrkB cross-talks to modulate NMDAR function is not known, although our working model involves the RasGrf1 signaling molecule, a nucleotide exchange factor for Ras/Rac-GTPases, which interacts with both receptors. In response to TrkB activation, Ras-Grf1 is tyrosine phosphorylated and mediates neurite outgrowth in PC12 cells. RasGrf1 also binds constitutively to …

The Differential Expression Of Egfl7 Transcripts During Angiogenesis In Human Fibrosarcoma, Navid Baktash

Electronic Thesis and Dissertation Repository

The growth of a tumor depends on de novo angiogenesis, which involves multiple signaling cascades and is also a target for many anti-angiogenic therapies. Epidermal growth factor like (EGFL7), is a protein that is exclusively expressed by endothelial cells during angiogenesis and is necessary for the proper assembly of a sprout. However, EGFL7 is also upregulated in many tumors such as glioma and non-small cell lung cancer, associated with poor patient prognosis. We have previously shown that EGFL7 expression by tumor cells inhibits tumor angiogenesis, which is contrary to its requirement for angiogenesis when expressed by endothelial cells. As a …