Life Sciences Commons^™

The Role Of Superantigens During Staphylococcus Aureus Nasal Colonization And Infection, Stacey Xu

Electronic Thesis and Dissertation Repository

Superantigens (SAgs) are potent toxins produced by bacteria such as Staphylococcus aureus that function to overactivate T cells resulting in massive cytokine production and immune activation. Despite decades of research on the structure and function of these proteins, as well as their role in severe diseases such as toxic shock syndrome, the question as to why strains of S. aureus produce SAgs and the role that they play in the life cycle of these bacteria remains unanswered. The contribution of SAgs towards pathogenicity and bacterial survival in vivo were assessed using isogenic SAg deletion knockouts in conjunction with SAg-sensitive humanized …

Identification And Characterization Of Cysteine Protease Genes In Tobacco For Use In Recombinant Protein Production, Kishor Duwadi

Electronic Thesis and Dissertation Repository

Plants are an attractive host system for pharmaceutical protein production. Many therapeutic proteins have been produced and scaled up in plants at a low cost compared to the conventional microbial and animal based systems. The main technical challenge during this process is to produce sufficient level of proteins in plants. Low yield is generally caused by proteolytic degradation during expression and downstream processing of recombinant proteins. The yield of a human therapeutic protein interleukin (IL) -10 produced in transgenic tobacco leaves was found to be below the critical level, and is potentially due to degradation by tobacco cysteine proteases (CysPs). …

Elucidating The Signalling Pathway Of Mer Tyrosine Kinase Receptor In Efferocytosis, Ekenedelichukwu Azu

Electronic Thesis and Dissertation Repository

Efferocytosis is the clearance of apoptotic cells and is necessary for homeostasis. Mer Tyrosine Kinase (MerTK) is a crucial efferocytic receptor whose loss is associated with chronic inflammatory diseases and autoimmunity. While previous studies have shown that MerTK mediates efferocytosis through a unique mechanism that requires integrins, MerTK signalling pathway remains unknown. Given this unusual internalization mechanism, I hypothesized that MerTK signals and engages integrins through a novel signalling pathway different from that used by other phagocytic receptors. Therefore, this study aimed to identify the signalling pathways activated by MerTK, utilizing conventional cell biology and pharmacological approaches.

I found that …

Understanding Recurrent Disease: A Dynamical Systems Approach, Wenjing Zhang

Electronic Thesis and Dissertation Repository

Recurrent disease, characterized by repeated alternations between acute relapse and long re- mission, can be a feature of both common diseases, like ear infections, and serious chronic diseases, such as HIV infection or multiple sclerosis. Due to their poorly understood etiology and the resultant challenge for medical treatment and patient management, recurrent diseases attract much attention in clinical research and biomathematics. Previous studies of recurrence by biomathematicians mainly focus on in-host models and generate recurrent patterns by in- corporating forcing functions or stochastic elements. In this study, we investigate deterministic in-host models through the qualitative analysis of dynamical systems, to …

Characterization Of Efferosome Maturation And The Processing Of Apoptotic Bodies, Yohan Kim

Electronic Thesis and Dissertation Repository

Every day billions of cells in our bodies undergo apoptosis and are cleared through efferocytosis – a phagocytosis-like process in which phagocytes engulf and degrade apoptotic cells. Proper processing of efferosomes prevents inflammation and immunogenic presentation of antigens. In this thesis I determined that the early stages of efferosome maturation parallel that of the pro-immunogenic phagocytosis of pathogens. Mass spectrometry analysis of later maturation stages identified unique regulatory proteins on efferosomes and phagosomes. Keys among these were Rab17 and Rab45 on efferosomes versus Rab6b and PI-4-Kinase on phagosomes. The later would allow for antigen presentation from phagosomes, while the former …

Evolutionary And In Silico Analysis Of The Antiviral Trim22 Gene, Jenna Kelly

Electronic Thesis and Dissertation Repository

Tripartite motif protein 22 (TRIM22) is an evolutionarily ancient interferon-induced protein that been shown to potently inhibit human immunodeficiency virus (HIV), hepatitis B virus (HBV), and influenza A virus (IAV) replication. Altered TRIM22 expression levels have also been linked to autoimmune disease, cancer, and cellular proliferation. Despite its important role in a number of biological processes, the factors that influence TRIM22 expression and/or antiviral activity remain largely unknown. To identify key functional sites in TRIM22, we performed extensive evolutionary and in silico analyses on the TRIM22 coding region. These tools allowed us to pinpoint multiple sites in TRIM22 that have …

Transcriptional Regulation Of Peptidylarginine Deiminase Type Iv: Implications For Rheumatoid Arthritis, Ali Abbas

Electronic Thesis and Dissertation Repository

High titers of anti-citrullinated protein antibodies have been detected in sera of rheumatoid arthritis (RA) patients, implicating citrullinating enzymes in the pathogenesis of RA. Peptidylarginine deiminase type IV (PAD4) is a member of the PAD family of enzymes that catalyze the post- translational modification of arginine to citrulline and has been linked with RA. However, little is known about its transcriptional regulation. Therefore, our aim was to determine how transcription of PAD4 is activated in the myeloid lineage. Using bioinformatics, a potential nuclear factor kappa B (NF-kB) binding site was identified on the PAD4 promoter. Luciferase assays were used to …

Sub-Inhibitory Antibiotics Enhance Virulence, Persistence, And Pathogenesis Of Uropathogens, Lee W. Goneau

Electronic Thesis and Dissertation Repository

In addition to their bactericidal effects, antibiotics are potent signal mediators at sub-inhibitory levels in the environment. The ability to modulate community structure in this niche raises concerns over their capacity to influence pathogenesis in patients during antibiotic therapy. This concept forms the basis of this thesis, and is explored using models of prophylactic therapy for recurrent urinary tract infection (UTI) management.

Sub-inhibitory ciprofloxacin, ampicillin, and gentamicin were found to augment virulence in vitro, increasing adherence and urothelial cell invasion in uropathogenic Escherichia coli (UPEC) and Staphylococcus saprophyticus. In addition, biofilm formation was increased, and swarming motility decreased. …

Study Of Virus Dynamics By Mathematical Models, Xiulan Lai

Electronic Thesis and Dissertation Repository

This thesis studies virus dynamics within host by mathematical models, and topics discussed include viral release strategies, viral spreading mechanism, and interaction of virus with the immune system.

Firstly, we propose a delay differential equation model with distributed delay to investigate the evolutionary competition between budding and lytic viral release strategies. We find that when antibody is not established, the dynamics of competition depends on the respective basic reproduction numbers of the two viruses. If the basic reproductive ratio of budding virus is greater than that of lytic virus and one, budding virus can survive. When antibody is established for …

Characterization Of Staphylococcus Aureus Lipase, Vithooshan Vijayakumaran

Electronic Thesis and Dissertation Repository

USA300, a strain of community-associated methicillin resistant Staphylococcus aureus (CA-MRSA), has become prevalent in the community. Colonization of human skin requires mechanisms that allow this bacterium to overcome the innate immune defenses on the skin, including secretion of antimicrobial lipids. Antimicrobial lipids inhibit S. aureus growth and induce the staphylococcal proteolytic cascade, producing aureolysin (Aur) which processes the lipase glycerol ester hydrolase (Geh). Nearly all S. aureus strains secrete Geh, yet little information exists concerning its function. Using purified Aur and Geh we confirm that aureolysin processes proGeh to Geh. We then confirmed that geh was required for lipase activity …

Role Of Branched-Chain Amino Acid Transporters In Staphylococcus Aureus Virulence, Sameha Omer

Electronic Thesis and Dissertation Repository

Branched-chain amino acids (BCAAs) act as effector molecules that signal a global transcriptional regulator, CodY, to regulate virulence factors in nutrient depleted environments. Staphylococcus aureus contains three putative BCAA transporters (BrnQ1, BrnQ2, BrnQ3) whose role in BCAA uptake is unknown. We hypothesize that BrnQ transporters are involved in BCAA uptake and contribute to virulence in S. aureus by modulating CodY activity. Results from radioactive uptake assays indicate that BrnQ1 is the predominant BrnQ transporter of isoleucine, valine and leucine. Meanwhile, BrnQ2 is more specific for isoleucine. Furthermore, only the lack of BrnQ1 hinders growth of S. aureus in chemically-defined media …

Gene Repression And Cell Cycle Regulation By Pu.1 In Acute Myeloid Leukemia, Rachel Gh Ziliotto

Electronic Thesis and Dissertation Repository

Acute myeloid leukemia (AML) is associated with mutations or chromosomal translocations in genes encoding transcription factors. PU.1 is a transcription factor that is required for the development of nearly all white blood cell types of the immune system, including B cells, granulocytes, and monocytes. Mutation of the gene encoding PU.1, SPI1 in humans and Sfpi1 in mice, is associated with AML. We hypothesized that reduced expression of PU.1 in Sfpi1^BN/BNmyeloid cells will result in the development of AML in transplanted mice due to reduced repression of E2F1, leading to deregulation of the cell cycle. Results indicate that NOD/SCID/γc …

Elucidating The Role Of Cathepsin B In The Lifecycle Of Influenza A Virus, Macon D. Coleman

Electronic Thesis and Dissertation Repository

Influenza virus type A (IVA) is the etiologic agent responsible for the febrile respiratory illness referred to as the flu. Seasonal and occasionally pandemic IVA-associated illness is a significant cause of morbidity and mortality worldwide, and presents a significant burden to the healthcare system. Our previous work showed that the propagation of IVA required the lysosomal protease cathepsin B (CTSB), though the mechanism behind this dependency was not elucidated. This study further examined the role of CTSB by using CTSB-deficient (CTSB^-/-) macrophages and the CTSB-specific chemical inhibitor CA-074 Me (CaMe) in human lung epithelial cells. CTSB^-/- and …

Modeling Leafhopper Populations And Their Role In Transmitting Plant Diseases., Ji Ruan

Electronic Thesis and Dissertation Repository

This M.Sc. thesis focuses on the interactions between crops and leafhoppers.

Firstly, a general delay differential equations system is proposed, based on the infection age structure, to investigate disease dynamics when disease latencies are considered. To further the understanding on the subject, a specific model is then introduced. The basic reproduction numbers $\cR_0$ and $\cR_1$ are identified and their threshold properties are discussed. When $\cR_0 < 1$, the insect-free equilibrium is globally asymptotically stable. When $\cR_0 > 1$ and $\cR_1 < 1$, the disease-free equilibrium exists and is locally asymptotically stable. When $\cR_1>1$, the disease will persist.

Secondly, we derive another general delay differential equations system to examine how different life stages of leafhoppers affect crops. The basic reproduction numbers $\cR_0$ is determined: when …

The Role Of Α5gaba(A) Receptors In Brain Inflammation, Jason K. Wong

Electronic Thesis and Dissertation Repository

Microglia critically regulate brain inflammation. GABA_A receptors that contain the α5-subunit (α5GABA_A) exhibit high sensitivity to GABA and confer tonic activity. Moreover, α5GABA­_A receptors have been associated with brain inflammation. This study investigates the role of α5GABA_A receptors in microglial activation. Immunohistochemistry revealed that in response to intraperitoneal lipopolysaccharide (LPS), α5-subunit null mice exhibited significantly higher expression of IL-1β in hippocampal microglia. Neuronal-glial co-cultures treated with α5GABA­_A receptor inverse agonist L655,708 drastically increased microglial IL-1β expression. Surprisingly, ELISA of media from L655,708-treated co-cultures revealed a considerably lower concentration of IL-1β. Treating cultured primary astrocytes …

Human Adenovirus E1a Binds And Retasks Cellular Hbre1, Blocking Interferon Signalling And Activating Virus Early Gene Transcription, Gregory J. Fonseca

Electronic Thesis and Dissertation Repository

Upon infection, human adenovirus (HAdV) must block interferon signaling and activate the expression of its early genes to reprogram the cellular environment to support virus replication. During the initial phase of infection, these processes are orchestrated by the first HAdV gene expressed during infection, early region 1A (E1A). E1A binds and appropriates components of the cellular transcriptional machinery to modulate cellular gene transcription and activate viral early genes transcription. We have identified hBre1/RNF20 as a novel target of E1A. hBre1 is an E3 ubiquitin ligase which acts with the Ube2b E2 conjugase and accessory factors RNF40 and WAC1 to monoubiquitinate …

Cellular Adaptation Of Macrophages To Anthrax Lethal Toxin-Induced Pyroptosis Via Epigenetic Mechanisms, Chae Young Han

Electronic Thesis and Dissertation Repository

Cellular adaptation to microbial stresses has been demonstrated in several cell types. Macrophages (MФ) are sentinel immune cells fending off invading microbes. Anthrax lethal toxin (LeTx) is a key virulence factor released by Bacillus anthracis that causes rapid cell death, pyroptosis. A small number of RAW246.7 macrophages (~4%) exposed to a non-lethal dose of LeTx become resistant to LeTx-induced pyroptosis for ~ 4 weeks, termed “toxin-induced resistance (TIR)”. Here, I showed that high levels of DNA methyl transferase1 (DNMT1) expression were maintained although global genomic methylation levels were not high in TIR. TIR cells treated with the DNMT inhibitor 5-azacitidine …

The Effect Of Dendritic Cell Mobilization On Cd8+ T Cell Responses To Influenza A Virus, Adil N. Shivji

Electronic Thesis and Dissertation Repository

Influenza A viruses (IAV) cause respiratory infections with potentially catastrophic consequences. Neutralizing antibodies towards surface proteins of IAV prevent reinfection. However, mutations in these proteins allow the virus to evade these antibodies. Enhancing cytotoxic T cell-mediated immunity has been proposed as an attractive strategy to combat flu. Recombinant human FMS-like tyrosine kinase 3 ligand (FL) is known to mobilize dendritic cells (DCs) in mice. Given that DCs are the most potent antigen-presenting cells, I hypothesized that their mobilization by FL will improve the CD8+ T cell response to IAV. Quantification of CD8+ T cell responses to IAV epitopes by intracellular …

Involvement Of Interleukin-33/St2 In Myocardial Dysfunction In Murine Model Of Sepsis, Yoonmi Choe

Electronic Thesis and Dissertation Repository

The disruption of myocardial extracellular matrix (ECM) has been implicated in myocardial dysfunction during sepsis. However, the underlying mechanism(s) are not clear. Interleukin-33 (IL-33) is a cytokine which regulates collagen synthesis in various cardiac pathologies. The purpose of the present study is to test whether IL-33 contributes to sepsis-induced myocardial dysfunction through regulation of matrix metalloproteinase-9 (MMP-9). The in vivo, feces-induced peritonitis (FIP) in mice and in vitro lipopolysaccharide (LPS) treatments to isolated cardiomyocytes were used. In FIP mice, myocardial IL-33 and MMP-9 expression were increased and myocardial contractility was decreased. Myocardial function in FIP mice was improved when treated …

Role Of Il-10 In The Immune Response To Staphylococcus Aureus Nasal Carriage, Teresa Fernandez Plaza

Electronic Thesis and Dissertation Repository

Staphylococcus aureus is a human pathogen with the potential to cause life-threatening diseases, but it can also be found as a commensal in up to 50% of the population. Its main reservoir is located in the nostrils, where macrophages are the principal mucosal APC and elicit a predominantly IL-10 response. Induction of IL-10 requires phosohoinositide 3-kinase activity, upon stimulation with S. aureus ligands through toll-like receptor 2 binding. It is hypothesized that IL-10 production by macrophages in the nasal mucosa allows S. aureus to persist in this niche as a commensal. To test this hypothesis, a cohort of S. aureus …

Restriction Of Hiv-1 Replication By Unique Trim22 Isoforms., Clayton Hattlmann

Electronic Thesis and Dissertation Repository

Understanding how the immune system reacts to HIV infection and why normal antiviral defenses are insufficient to fight infection is a key step towards creating better therapies. Several interferon-induced proteins, such as the tripartite motif protein TRIM22, are capable of restricting HIV-1 replication; however single nucleotide polymorphisms (SNPs) can dramatically impact the actions of these proteins. While the trim22 gene contains numerous SNPs, no study has addressed how these may affect TRIM22 functions. Here we provide the first direct comparison of two TRIM22 unique isoforms. Through confocal microscopy we observed these isoforms exhibit different patterns of localization. In vitro studies …

Serum Iron Uptake And Virulence In Staphylococcus Aureus, Federico C. Beasley

Electronic Thesis and Dissertation Repository

The high affinity iron scavenging glycoprotein transferrin sequesters trace amounts of serum Fe³⁺ to concentrations below what is required to sustain microbial life. Iron may be liberated from this important innate immune factor after interaction with molecules that chelate or reduce Fe³⁺. Organisms with cognate transport systems for these iron coordinating molecules can survive in the bloodstream using transferrin iron.

Staphylococcus aureus is an opportunistic bacterial pathogen. S. aureus executes numerous strategies for overcoming the innate immune barrier of iron deprivation in the bloodstream. In addition to specialized mechanisms for hemoglobin iron extraction, S. aureus can proliferate …

The Role Of Granulocyte Colony-Stimulating Factor In The Intestinal Immune System And The Response To Probiotics, Andrew J. Martins

Electronic Thesis and Dissertation Repository

Loss of immunoregulation in the intestine results in inflammation, such as is observed in the chronic, relapsing, inflammatory bowel diseases (IBD), Crohn's disease and ulcerative colitis. However, our understanding of this process remains incomplete. The cytokine granulocyte-colony stimulating factor (G-CSF) is an important stimulator of granulopoiesis in steady-state and during inflammation, but has also been shown to have anti-inflammatory effects and is a potential therapeutic for IBD. In addition, certain strains of bacteria, termed probiotics, have been postulated to have anti-inflammatory effects in the intestine. The mechanisms by which these strains of bacteria elicit anti-inflammatory effects are not fully understood. …

The Hiv-1 Tat Protein And Adverse Drug Reactions: A Model System Utilizing Jurkat T Cells And Sulphamethoxazole-Hydroxylamine, Kaothara Adeyanju

Electronic Thesis and Dissertation Repository

In 2009 approximately 2.6 million people became infected with the Human Immunodeficiency Virus (HIV). In addition to the estimated 33.3 million people currently living with the virus, this makes HIV/ AIDS an epidemic of unprecedented scale in modern times. Treatment of HIV infection requires antiretroviral agents as well as a number of other drugs such as antimicrobials. Hypersensitivity adverse drug reactions (ADRs) to a variety of drugs are common in HIV-infected individuals, but the antimicrobial Sulphamethoxazole remains a major culprit. Hypersensitivity ADRs cause significant morbidity, with the skin and liver most commonly affected and are among the top causes of …