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  • Upstream Regulators of TORC1 Signaling Pathway in Saccharomyces cerevisiae

    Vaishampayan, Prajakta (The University of Arizona., 2019)
    Many nutrients including glucose, phosphate and amino acids regulate TORC1 activity and when the cells are in stress or starvation conditions, TORC1 activity is inhibited. It remains unclear how this happens. So, we are interested in mapping the signaling system that talks to TORC1 to ultimately understand how the complex integrates signals to control growth in stress conditions. In this project, the response of TORC1 under nitrogen, glucose and phosphate starvation conditions was checked in strains lacking one or more stress signaling pathway proteins to identify the mechanism which leads to inhibition of TORC1 in stress conditions.
  • Matrix Matters: Biomarker Potentials of Phagocytes, Exosomes, and Cytokines

    Buckley, Maverick J. (The University of Arizona., 2019)
    Overview: The Applied Biosciences Professional Science Masters (ABS-PSM) program at the University of Arizona prepares students in the fields of biological science to enter areas of business and scientific competition. This interdisciplinary course of study involves the completion of an internship wherein students demonstrate scientific inquiry in the context of the goals and economic pursuits of the hosting agency. Students are required to convey how their projects contribute to the ambitions of the company or academic institution as well as to the larger scientific field. In this report, two internship projects completed to fulfill this requirement for the ABS-PSM degree will be described. Biomedical research and diagnostics rely heavily on the use of biomarkers for drug discovery and disease management. Biomarkers are “characteristics that are objectively measured and evaluated as indicators of normal biological processes, pathogenic processes, or pharmacologic responses to therapeutic interventions,” as defined by the National Institutes of Health Biomarkers Definitions Working Group (2001). Drug development and many facets of clinical diagnostics involve the measurement of a combination of biomarkers to evaluate the status of disease in an individual or their physiological changes following some treatment. In contrast to symptoms, biomarkers are not perceived by the patient but rather are observed from outside the patient (Strimbu & Tavel, 2010). The most well-studied biomarkers, such as troponin for the assessment of cardiac injury (Babuin & Jaffe, 2005), are applied regularly in predicting the incidence or outcome of a disease. This use of biomarkers as clinically meaningful surrogate endpoints is entirely justifiable, but only when such a characteristic has extensively and repeatedly proven predictive of outcome (Strimbu & Tavel, 2010). Additionally, the most logical biomarkers are those directly involved in the pathophysiology of a certain pathway, enabling more accurate interpretations of an individual’s disease status to be made. The evaluation of a biomarker’s potential is challenged by the way in which it is measured. Biological samples are wide in variety, and, aside from ensuring the marker is present in the medium at all, determining what sample type is most compatible with existing instruments and what is most associated with a disease or anatomic site is a complex task. Cardiac troponin, for example, is measured in a peripheral blood sample to assess heart damage. The correlation of this enzyme’s concentration in the blood with heart muscle damage makes it a valuable disease indicator because the enzyme is produced in cardiac tissue and immediately released into the circulation (Antman et al., 1996). Neurodegenerative disease markers are markedly complicated because their presence in conventional fluid samples may not be accurately representative of concentrations in the brain. MSDx, Inc. (Tucson, AZ), a company that develops diagnostic solutions for neurodegenerative diseases, has identified phagocytes as a suitable source of biomarkers for this purpose. These cells naturally concentrate and carry remnants of disease pathology through phagocytosis, thereby preserving intact markers of neurodegeneration from the potentially degradative extracellular environment that can be quantified in a blood sample. In comparing this approach to conventional fluid analyses, the first chapter in this report describes a literature review performed on various potential neurodegenerative disease biomarkers and their alterations relative to controls by sample matrix. A second aspect of this project that will be discussed was an exploration of entities involved in the research of exosomes, a type of extracellular microvesicle capable of carrying proteins between cells. Cytokine concentrations in the blood also change during various disease processes such as inflammation, making them attractive potential biomarkers. Their use, though, is hindered by variation in normal levels from person to person and an uncertainty as to the optimal sample matrix in which to quantify them. For biomarkers to be identified among cytokines, it is essential to first determine what constitutes “normal” levels of a selection of pathologically relevant cytokines, whether serum or plasma samples should be used, and what collection and processing practices such as anticoagulant should be employed. This is the overarching goal of a group at the University of Arizona BIO5 Institute whose small pilot study will be the focus of the second chapter of this report. The study involved recruiting a cohort of self-identified healthy adult volunteers at specific time intervals and obtaining blood samples for quantifying their cytokines using a commercial multiplex Luminex-based assay. An abbreviated preliminary analysis of the results was then conducted, which concerned the levels of cytokines obtained at different time points and in different sample matrices, supporting a brief evaluation of the benefits and drawbacks of using certain anticoagulants and preparation strategies over others. Both projects produced exciting findings related to biomarker discovery and measurement. Based on my review, phagocytes seem to be largely ignored in neurodegeneration research, yet represent a promising medium in that they avoid many of the drawbacks associated with measuring central nervous system components outside the brain; as well, they did not appear to be studied elsewhere beyond MSDx nor their potential to carry important biomarkers dismissed. On the other hand, exosomes have been shown to be increasingly studied in neurology with broad potential, but important challenges mostly related to their small size remain. Cytokines, in contrast to such biomarker cargo carriers, are information couriers that may also have biomarker potential. These intercellular messengers mediate inflammatory processes but are in constant production at different degrees depending on one’s immunologic and overall health status. Determining what is normal must first be established and is a major challenge since cytokine levels vary by individual, even in an uninfected state. To advance this process, plasma collected with EDTA as an anticoagulant may be optimal, though serum is attractive due to its widespread use and would help make comparisons with the majority of other cytokine studies possible. Despite its invaluable capability to simultaneously measure multiple cytokines in a single sample, the bead-based assay also has its weaknesses that may be related to the matrix type or antibodies being used. Thus, determining the most appropriate method for sample retrieval, reliable measurement of the marker of interest, and establishment of reference values for novel biomarkers appear to be the most pressing challenges associated with the goals described here.
  • Participatory Democracy, Pluralism, and the Rational Model in Natural Resource Planning: A Case Study of the San Pedro River Initiative Process, Arizona, U.S.A.

    Evans, Luke T. (The University of Arizona., 2002)
    An analysis and critique is conducted of pluralism, the rational model, and participatory democracy in relation to public participation in natural resource planning and policy development. Each theory is evaluated in terms of efficacy, representation and access, information exchange and learning, continuity of participation, and decision-making authority. A case study is used to assess elements of each theory in an actual public participation process, utilizing the above criteria. The study indicates that the need for efficiency precludes the use of practices that would more thoroughly involve the public in decision making processes, and that policy making tends to revert to more traditional, expert-dominated, and exclusive public processes despite efforts to the contrary. Finally, the analysis questions the utility of comprehensive public involvement in natural resource policy making and planning given the constraints of existing legal mandates, polarized public opinions, and the need for decisions made in the larger public interest.
  • Genetic Analysis of Mountain Lion (Puma concolor) Feces from Kofa National Wildlife Refuge, Arizona

    Naidu, Ashwin (The University of Arizona., 2009)
    Investigations on recent records of mountain lions (Puma concolor) and concurrent declines in desert bighorn sheep (Ovis canadensis mexicana) on Kofa National Wildlife Refuge have necessitated the determination of the number of mountain lions and their diet on the refuge. Using genetic analysis, we identified mountain lion feces/scats (n=53) from the Kofa and Castle Dome Mountains in southwestern Arizona. We identified 11 individual mountain lions that included at least 6 males and 2 females. We also identified prey species from bone and connective tissue remains inside the mountain lion scats. Our data suggest that a majority of mountain lion diet (62 %) on the refuge is mule deer (Odocoileus hemionus). These estimates greatly enhance our knowledge of mountain lions in an area where, historically, their presence was considered transient. Additionally, recognizing the need for reliable species identification and to improve species identification from non-invasive samples, we developed a Polymerase Chain Reaction (PCR) primer set that would enable the amplification of the complete cytochrome b gene from a large number of mammalian species. DNA sequence information obtained from the use of this primer set can be used for the development of mammalian species’ databases and referencing. Overall, this project demonstrates the efficacy of genetic techniques and their potential to provide reliable and necessary information on elusive species to wildlife managers.
  • I. Morphological and Nanomechanical Studies of Lipid Bilayers Composed of Polymerizable and Non-Polymerizable Lipids II. Fluidity Studies of Platelet Plasma Membranes

    Fonseka, Nelusha Malithi (The University of Arizona., 2019)
    Two main projects are discussed in this dissertation. The first project: Morphological and nanomechanical studies of planar supported lipid bilayers (PSLB) composed of polymerizable and non-polymerizable lipids as potential platforms for biosensors, is discussed in Chapters 2 through 4. Chapters 2 and 3 focus on PSLBs composed of the polymerizable lipid, bis-SorbPC and Chapter 4 focuses on bis-DenPC16,16. These studies are important because PSLBs are widely studied as platforms for receptor-based biosensors. PSLBs composed of fluid lipids lack the stability necessary for many technological applications due to the relatively weak non-covalent interactions between lipid molecules. Lipid polymerization enhances bilayer stability, but greatly reduces lipid diffusion and membrane fluidity. In an effort to enhance bilayer stability while maintaining fluidity, PSLBs composed of mixtures of polymerizable lipids and fluid lipids were prepared and characterized. Fluidity studies of these bilayers showed that considerable fluidity is retained even when the polymer fraction is substantial, which suggests that these bilayers are phase segregated, composed of polymerized and fluid domains. However, domains had not been observed previously. Chapter 2 of this dissertation describes the work done with atomic force microscopy (AFM) to study the phase segregation of mixed PSLBs composed of the polymerizable lipid bis-SorbPC and the fluid lipid DPhPC. This work provided direct evidence for polymerization-induced phase segregation of these mixed PSLBs, forming membranes composed of fluid and poly(lipid) domains. In these mixed bilayers, DPhPC formed a semi-continuous phase of greater height surrounding island-like domains of poly(bis-SorbPC) of lesser height. Numerous studies demonstrate that retention of bioactivity upon reconstitution of transmembrane proteins typically requires both membrane fluidity and elasticity. Thus, AFM force mapping was employed to study the nanomechanical properties of lipid bilayers, which is described in Chapter 3. This is the first study done to quantify the nanoscale mechanical properties of bis-SorbPC before and after polymerization, and mixed bilayer composed of bis-SorbPC and DPhPC. The results showed that the resistance to rupture and elastic modulus of bis-SorbPC increased upon polymerization. In addition, the results showed that the breakthrough force and the elastic modulus of DPhPC in mixed bilayers were different to pure bilayers due to the size (interface/edge effects) and the purity of the domains in mixed PSLBs. Findings similar to Chapters 2 and 3 are discussed in Chapter 4, with a different polymerizable lipid; bis-DenPC16,16. Comparing the results of bis-SorbPC (Chapter 3) and bis-DenPC (Chapter 4) showed that the position of the polymerizable moiety significantly changed the nanomechanical properties of PSLBs. In addition, no direct evidence of phase segregation was observed in mixed PSLBs composed of bis-DenPC and DPhPC during sub-micron scale morphological and nanomechanical studies. The second project: Fluidity studies of platelet plasma membranes, is the focus of Chapter 5. Therein, the feasibility of employing fluorescence recovery after photobleaching (FRAP) to determine the diffusion coefficient of platelet plasma membranes in response to lipophilic molecules is investigated. Mechanical circulatory devices, used in patients with heart failure to restore blood flow, cause thrombosis due to the abnormal flow of blood and supra-physiologic shear on blood platelets when blood passes through these devices, known as shear-mediated platelet activation (SMPA). It has been hypothesized that the membrane fluidity plays a role in treating SMPA and that the fluidity can be modulated with lipophilic molecules. Accordingly, the possibility of employing FRAP to study the lateral diffusion coefficient of platelet plasma membranes was investigated and a protocol was developed. The results showed that FRAP of platelet membranes is a suitable technique to determine the lateral fluidity of platelets before and after treating with exogenous lipophilic molecules. Therefore, the protocol established here will be helpful to study the fluidity of shear-activated platelets allowing to test the hypothesis. Further, this protocol will be beneficial to investigate the fluidity of platelet plasma membranes in the development of treatment methodologies targeting the material properties of platelets to reduce SMPA.
  • Development of Selective Peptide Ligands Targeting Melanocortin Receptors for Melanoma Prevention and Therapy

    Zhou, Yang (The University of Arizona., 2019)
    Melanocortin receptors are a family of G-protein-coupled receptors (GPCRs) that regulate many important physiological functions. Specifically, melanocortin-1-receptor (MC1R) on skin melanocytes regulates skin pigmentation, which is a natural protection against UV-induced DNA damage, the major risk for melanoma. However, the lack of selectivity to MC1R and the requirements of needle injections of currently available peptide drugs greatly hinder the development of MC1R agonists as melanoma prevention medicines. This thesis mainly describes efforts to develop MC1R selective peptide agonists. Inspired by previous structure-activity studies and receptor mutagenesis studies, the peptide drug design enhances MC1R selectivity through removing strong ionic interactions between ligand and melanocortin receptors other than MC1R, introducing local structural constrains on the backbone (N-methylations) and sidechain (φ and ψ angle constrains) as well as global conformational constrains on cyclic peptides. The other emphasis of this thesis is to develop peptide drugs that are accessible to the target receptors and can be taken in a non-invasive way. To achieve this goal, different peptide drug design templates were exploited. Lead compounds with potentials for different applications include: 1. a long linear peptide suitable to be directly applied to the skin surface and induce skin pigmentation with less concerns on side effects; 2. tetrapeptides with great potential to be developed into oral administration drugs to induce skin pigmentation; 3. cyclic peptides suitable to target the MC3R or the MC4R in the brain; 4. an orally available cyclotide to activate the MC1R. This thesis also describes the opportunity to target MC1R-overexpressing melanoma cells with MC1R agonist for targeted drug delivery. Such approach can bring versatility to the cytotoxic drug selection, which is the key to solve the serious drug resistance problem of melanoma. Future opportunities exist to develop peptide drugs targeting different signaling pathways downstream of melanocortin receptors to regulate physiological functions governed by the same melanocortin receptor subtype.
  • Effects of Climate and Ecological Processes on Engineered Uranium Disposal Cell Performance with Respect to Nearby Subsistence-Based Indigenous Communities

    Joseph, Carrie Nuva (The University of Arizona., 2019)
    Near-surface earthen-engineered disposal covers are used to consolidate uranium mill tailings waste generated from defense-related uranium mines located across the United States. Disposal covers made from compacted soil layers stabilize waste consisting of a mixture of chemical and radiological constituents to limit air, soil, and water contamination above and below ground. However, 20 to 30 years post-construction, vegetative succession, dust deposition, and soil development are changing the as-built engineering design, clearly notwithstanding the longevity standard of 200-1,000 years. Little is known about how natural ecological succession will impact the performance of earthen-engineered covers. Furthermore, there is a dearth of information about how subsistence-dependent Indigenous communities located nearby inactive uranium disposal facilities are impacted by past, present, and future operations of legacy sites. The Department of Energy – Legacy Management (DOE-LM) is evaluating whether natural ecological processes occurring above disposal covers are sustainable and alternate long-term remedies that could reduce maintenance costs and exposures to humans and the environment. Using quantitative and qualitative methodologies, three interdisciplinary studies were completed to address key knowledge gaps for uranium legacy sites. The overall research objectives are: (1) to investigate how natural ecological succession above engineered covers, specifically plant establishment and root intrusion, may be a potential benefit or potential detriment to disposal cover performance; (2) to provide climatological data and future climate trajectories to understand whether engineered cell covers are vulnerable to climate impacts; and (3) to understand community risk perceptions from the unheard voices of indigenous people and the impacts to their livelihoods from uranium legacy sites. In the first study, 144 samples of various vegetation types were analyzed for metal and radiological uptake collected from a broad range of climates and disposal cover designs, where soil development, dust deposition, and moisture provided a favorable habitat for plant growth. The goal was to determine if vegetation was compromising the performance of disposal cell covers by creating an exposure pathway for uranium tailing constituents. It was also important to determine exposure levels because twelve Indigenous tribes living near the study sites have ethnobotanic uses of plants. The results of this research indicate that plant concentrations are not accumulating to toxic levels with the exception of two site locations, where exceedances can be attributed to background soil characteristics. The second study addresses the vulnerability of uranium disposal cell covers to climate change in the U.S. Southwest. We extract monthly precipitation, daily minimum, and daily maximum temperatures to determine climate trends (mean annual temperatures, extreme conditions, and seasonal variation) of the recent past from high-resolution data sets. We also determine future climate by documenting projections from CMIP5 models under two representative concentration pathways (RCP). It was found that there are yearly and seasonal differences in climate outputs compared to historical data. While one site in the southwest experiences a trend towards wet/hot conditions, another site will experience hot/drier conditions under RCP 8.5 projections. From this study, it is hypothesized that allowing plants to grow in regions (Tuba City) that will experience wetter conditions, in normally arid regions, could be an alternative to controlling the water-balance. The third study used indigenous research methodologies to determine the community risk perceptions of two Hopi villages located 7 kilometers (km) downstream from an inactive uranium mill tailings site. Five focus groups were held using the conversational method in which open-ended, broad overview descriptive questions were used as a guide. The results from this study can address broader questions about a needs assessment, exposure and risk assessment, and risk communication, that are unique to the Hopi population but also useful to other tribes. Given the scope of the problem, this dissertation research confirms how natural ecological processes via climate-plant interactions on disposal covers may pose risks to performance, and in other cases it may not. The results will help to prioritize sites that may be good candidates for disposal cell renovations that embraces a sustainable design. Further, the qualitative study provides vital information, not documented in the literature than can be prioritized to bridge partnerships and reduce environmental and health risks.
  • Examining Thresholds for Diabetes, Renin-Angiotensin System Antagonist, and Statin Medication Adherence Quality Measures: The Application of the Law Of Diminishing Returns in Administrative Claims

    Campbell, Patrick James (The University of Arizona., 2019)
    Objective: Despite the limited evidence, a threshold of 80% proportion of days covered (PDC) is used to categorize medication adherence. The objective of this study was to assess the association of 1) antidiabetic, 2) renin-angiotensin system antagonists (RASA), and 3) statin medication adherence, at deciles of PDC, with disease-specific and all-cause economic outcomes (inpatient utilization and total healthcare costs) to identify optimal medication adherence thresholds using the law of diminishing returns. Methods: This retrospective cohort study included individuals from the Truven Health MarketScan® Commercial Claims and Encounters Research Databases (2010-2012) eligible for inclusion in the Pharmacy Quality Alliance diabetes, RASA, and statin medication adherence measures with non-capitated health plans. Generalized linear models (GLMs) with log link and gamma (costs) or negative binomial (utilization) distributions were used to assess the relationship of adherence with economic outcomes while adjusting for covariables (e.g., age, gender, Charlson comorbidity index). An alpha level of 0.01 was set a priori. Beta coefficients were used to compute use ratios and cost ratios and plotted to generate use and cost reduction functions. Marginal use and cost reduction curves were estimated and points of diminishing marginal returns and maximum returns were identified. Results: A total of 404,108 (diabetes), 1,329,576 (RASA), and 1,266,066 (statin) individuals were included in the study cohorts. Of the 120 GLMs that assessed the relationship between adherence and economic outcomes, 116 significant associations were identified (all p<0.0001). Of these, 98 models identified that adherence was associated with lower cost and utilization compared to nonadherence. Eighteen models showed adherence was associated with higher healthcare costs than nonadherence. The following adherence thresholds were identified as the optimal range of medication adherence thresholds (i.e., points of diminishing marginal and maximum returns): between 86% and 91% PDC for diabetes, 83% and 89% PDC for RASA, and 90% and 96% PDC for statin medications. Conclusions: The law of diminishing returns can be successfully applied to medication-taking behavior to derive optimal adherence thresholds in a commercially-insured patient population. Reliance on the 80% PDC adherence threshold should be re-evaluated to optimize benefits of diabetes, RASA, and statin medication adherence. The application of the law of diminishing returns in other patient populations and medication classes is warranted.
  • Synthesis and Application of Tunable Mo2 and W2 Tetraguanidinate Paddlewheel Complexes

    Humphries, Matthew (The University of Arizona., 2019)
    Super-electron-donor dimolybdenum and ditungsten tetraguanidinate paddlewheel complexes have proven to be the strongest reducing agents known. Strong single and double electron donors can be used for applications from H2 production to difficult organic transformations like C–Cl bond cleavage and crosscoupling reactions. The goal of this research was to develop accessible syntheses of the ligands and complexes, to investigate high impact applications of the Mo2 and W2 analogues, and to explore new complexes in hopes to tune their reactivity and solubility. Preparation and handling of the super-base, bicyclic guanidinate ligands and complexes have prevented extensive investigations of their applications. New reproducible syntheses for HTEhpp and W2(TEhpp)4Cl2 were developed. Interaction of the paddlewheel complex Mo2(TEhpp)4 and low concentrations of acetic acid were studied using cyclic voltammetry. Experiments show the electron deficient vacant dimetal axial site and the electron rich guanidinate core work together similar to frustrated pairs. Acetic acid protonates the guanidinate while simultaneously coordinating to the metal center. This newly discovered synergistic bonding decreases the electron donor ability of the complex preventing catalysis for the production of H2. By tuning the dimetal center to a more electron rich, 3rd row transition metal, W2(TEhpp)4, the reduction of H+ to H2 is now favored and is catalytic. Computations were used to explore the nature of these interactions. In addition to H2 production, a mechanistic study of C–Cl bond cleavage in dichloromethane by Mo2(TEhpp)4 shows a novel singlet-to-triplet crossover at the transition state. This work shows the potential for this class of complexes to 19 perform electron transfers via multiple mechanisms (e.g. atom transfer and electron transfer). Upwards of 25 new dimetal tetraguanidinate paddlewheel complexes were explored computationally for their electron donor ability. Some of the ligands have synthetic precedent. These and many others will hopefully be part of the next generation of super-electron-donor complexes.
  • Combined Model Approach to the Problem of Ranking

    Lee, Alexander S. (The University of Arizona., 2019)
    Ranking can be defined in many ways and has been applied in many areas. Over the past years, many research studies have been conducted on ranking methods for decision-making. The issue is that results pertaining to ranking entities are based only on one method, which can be subjective and prone to biases. As a result, results vary from method to method. To resolve this issue, a combined model approach is proposed, in which multiple methods are taken into account. The goals of the combined model are to rank entities more objectively and obtain more reliable results. The combined model has both qualitative and quantitative elements, where the qualitative element is the ranking and clustering, while the quantitative element is comprised of the scores. Since ranking is based on the scores of the entities, it takes into account the distribution of scores. In some scenarios, closely ranked entities can have similar scores, while in other scenarios, their scores can be relatively different even though they are ranked close to one another. The score distribution leads to clustering analysis, where entities are divided into clusters based on the spread of the scores. Hence, the combined model takes into account not only the ranks but also the scores of the entities. The proposed combined model is applied to three areas for this dissertation research. The first is identifying and ranking road hotspots and predicting the number of traffic crashes in road segments using the Empirical Bayesian (EB) enhanced by the Proportion Discordance Ratio (PDR) metric. The effectiveness of the Enhanced EB method is tested and demonstrated through a case study that is conducted in one of the major highways in Phoenix, Arizona. The second is ranking major US metropolitan areas in traffic congestion using unsupervised learning based on the Normalized Scoring Method (NSM), Principal Component Analysis (PCA), and the PDR similarity matrix. In 2015, TomTom ranked Tucson as the 21st most congested metropolitan area in the US, and the unsupervised learning combined model is applied to assess TomTom’s traffic congestion ranking of the metropolitan areas in order to determine if Tucson is highly congested based on the proposed model. The third is ranking and assessing the Hall of Fame (HOF) worthiness of retired Major League Baseball players based on their performance statistics through supervised learning based on Support Vector Machines (SVM) and Neural Networks (NN). Players are considered for the HOF through a voting procedure, where voters are comprised of members of the media, but there is a possibility of voting bias that can favor or go against certain players. Results from all three scenarios show that the proposed combined models are more reliable and can more objectively rank entities in order to correct biases based on previous methodologies.
  • I. Causes of Multiple Diffusing Populations of Fluorescently Labeled Probes in Lipid Membranes II. Evaluation of Phospholipid Membranes Incorporating the Polymerizable Lipid Bis-Denpc (16, 16) and Suitability as Ultra-Stable Platforms for Ion Channel Based Sensors

    Smith, Christopher M. (The University of Arizona., 2019)
    This dissertation is composed of two major projects, though some capabilities and findings from the first project were applied to the second. Project I focuses on advancements made in the understanding of the chemical interactions of a number of commonly used fluorescently labeled phospholipid probes. These probes are used for a variety of studies, including labeling of cellular or artificial membranes, examining transport and communication between different membranes, and determining membrane fluidity. Understanding the chemical behavior and interactions of these probes in membranes can be key for the proper interpretation of experimental data. Utilizing fluorescent recovery after photobleaching (FRAP), in combination with other spectroscopic techniques, multiple diffusing populations of commonly used probes in various artificial lipid membrane formats were identified, as were the causes for these populations. This allows for a fuller description of the fluidity of lipid membranes. These findings are the focus of Chapters 3 and 4 while the hardware developed that enabled critical measurements is the focus of Chapter 2. Project II focuses on addressing key limitations in developing ion channel (IC) based biosensors utilizing artificial lipid membranes. Among these limitations are the weak mechanical, chemical, and electrical stabilities of artificial lipid bilayers due to the weak noncovalent interactions involved in the membrane. To address these limitations, the polymerizable lipid bis-dienoyl phosphatidylcholine (bis-DenPC(16, 16)) was characterized for its ability to form ultra-stable membranes suitable for IC based sensors using the model IC gramicidin A (gA). Special attention was given to determining the membrane fluidity given the requirement of gA that two subunits must laterally diffuse to converge and dimerize to form a conductive pore. These studies are the focus of Chapters 5 and 6.
  • A Society of Individuals: Worker Differentiation and Collective Behavior in Social Insects

    Leitner, Nicole Elise (The University of Arizona., 2019)
    Individuals are not identical. Organisms of the same species usually exhibit diverse phenotypes, a phenomenon that extends not only to physical characteristics but to behavior as well. Indeed, consistent differences in behavior across individuals are now well cited across the animal kingdom and have been shown to be an integral part of the ecology and evolution of an organism. Yet, the proximate, physiological mechanisms upon which natural selection might act to either favor or eliminate variation is often not known. Here, I use social insect division of labor – essentially an exaggerated form of behavioral variation in which nestmate workers specialize in different tasks – as a study system for exploring potential neural mechanisms of inter-individual behavioral variation. This investigation also led me to test and review one of the most widely accepted hypotheses for division of labor, the results of which provide new and important insights into how and why social insect workers choose different tasks. In my first chapter I explored variation among Temnothorax rugatulus ant workers in their peripheral sensory organs, a potentially important, but often overlooked, source of behavioral variation (or, in the context of social insects, task specialization). The idea for this study was based on the popular “response threshold” mechanism for division of labor, whereby workers become specialized in tasks because they vary in their response (often assumed to be ‘sensitivity’) to task-associated stimuli. Potentially, variation in worker sensory organs might influence a worker’s ability to detect such work-associated stimuli. Though workers showed considerable variation in the number of antennal sensory structures, this variation was not related to variation in worker behavior. This led to my second chapter, in which I directly tested the response threshold hypothesis in T. rugatulus ants. Here, I found no evidence for individual variation in response thresholds to three different putative task stimuli, calling into question the ubiquity of this mechanism as a general driver of social insect division of labor. To follow up on this idea, in my third chapter I performed a comprehensive review of the response threshold hypothesis in the social insect literature and found that the empirical evidence for it is not as strong as is commonly assumed. This chapter also provides guidelines for how to test response thresholds, with the goal of becoming a valuable reference work for those studying both social insects and self-organization in complex systems in general. My last chapter scaled up to the colony level. I explored plasticity and constraint in task reallocation by challenging colonies of T. rugatulus with different types of perturbations. By considering multiple types of tasks, this study provided novel insight into how the variety of tasks a colony performs are regulated quite differently. Here, I found that colonies were able to achieve considerable flexibility through the use of two different strategies. Which strategy the colony used was task-dependent. In addition, colonies appeared to have two types of reserve workers upon which to draw when task needs increased, dependent on task type.
  • Modern Contraceptive Use among Women in Sub-Saharan Africa: Individual, Interpersonal, and Healthcare System Indicators

    Asaolu, Ibitola (The University of Arizona., 2019)
    Background: Contraceptives promote maternal and child health by reducing the prevalence of: unintended pregnancies, unsafe abortions, maternal deaths, low birth weight infants, preterm birth, and infant mortality. Despite its benefits, the uptake of contraceptives among women of childbearing age remains low in sub-Saharan Africa. In sub-Saharan Africa, the prevalence of contraceptive use among women in a union is 28%. To advance maternal and child health in this region, it is imperative to identify various individual, interpersonal, cultural, and healthcare system factors that affect the adoption of modern contraceptives. Objective: This project assesses individual, interpersonal, and healthcare systems indicators of modern contraceptive use among women in sub-Saharan Africa. Methods: Quantitative insight into the determinants of modern contraceptive use was obtained through analyses of multiple Demographic and Health Surveys (DHS) and Performance Monitoring and Accountability (PMA) 2020 data. The quantitative research studies were limited to women with a need for contraception, i.e. respondents who want to limit or space childbirth. Multivariable logistic regression models assessed the relationship between modern contraceptive use and: a) individual; b) interpersonal; and c) healthcare system factors. To further explore the barriers and facilitators of modern contraceptive use, this project employed focus group discussions and semi-structured interviews of healthcare providers in Calabar, Nigeria. Data from focus group discussions and interviews were analyzed using the thematic analysis approach. Results: The prevalence of modern contraceptives ranged from 7.8% in Gambia to 68.9% in Kenya among women. Several factors were positively associated with modern contraceptives use. Evidence from the DHS revealed that two domains of women’s empowerment—labor force participation and education—were most consistently associated with increased rates of modern contraceptive use. Results from analysis of the PMA2020 data identified four healthcare factors associated with women’s use of modern contraceptives. Health worker home-visits, adolescent reproductive health services, and polyclinic/hospitals were associated with modern contraceptive use in Ghana, Kenya, and Nigeria. Evidence from focus group discussions showed that women mostly used condoms, emergency contraceptives pills, fertility awareness (rhythm), and folkloric methods of contraception. In addition, myths about contraceptives prevented participants from using certain forms of modern contraception. For instance, participants cited that pills, injectables, and implants caused infertility and diseases among women who use these methods. Conclusion: Modern contraceptive use among sub-Saharan African women can be increased by promoting the health-worker visits, empowering women through education and labor force participation, and correcting myths about the side effects of modern contraceptives. By addressing individual, interpersonal, and healthcare correlates of contraceptive use, findings from this research can inform the design of comprehensive and more effective contraceptive interventions. Therefore, findings from this research can inform the design and implementation of interventions that acknowledge multiple correlates of contraceptive use.
  • HCMV Manipulation of Host Cholesteryl Ester Metabolism

    Dahlmann, Elizabeth Alan (The University of Arizona., 2019)
    Human cytomegalovirus (HCMV) is a β-herpesvirus that infects over 50% of people above the age of 40. Once infected, HCMV establishes a lifelong latent infection with periodic reactivation. Most infections are asymptomatic. However, infection in immunocompromised patients may result in fatal HCMV-related complications. Further, congenitally-acquired HCMV infection is the leading cause of birth defects in the United States. The HCMV virion contains a large double-stranded DNA genome encapsidated by a protein shell that is surrounded by a lipid membrane. Like all enveloped viruses, HCMV steals host lipids to generate its envelope membrane. While previous studies demonstrate that HCMV replication requires lipid metabolism, the details of virally-induced lipid changes remain poorly defined. We performed an untargeted lipidomic screen using liquid chromatography high resolution tandem mass spectrometry to identify and quantitatively measure how infection alters the lipidome of cells. We found that HCMV increases cholesteryl esters (CE) by 24 hours post infection. CE lipids are synthesized by sterol O-acyltransferase 1 (SOAT1) attaching a fatty acyl-CoA to a cholesterol molecule. I hypothesized that early stages of HCMV replication induce CE biosynthesis and that CE are required for viral replication. In support of our hypothesis, we found HCMV induces SOAT1 gene expression. Further, HCMV immediate early pUL37x1 is partially responsible for virally-induced CE accumulation. We found that treating infected cells with a SOAT1 inhibitor blocked CE production and infection. Overall, our findings suggest that HCMV induces CE synthesis that can be targeted to block infection.
  • The Effect of Sentence Contexts on Second Language Word Recognition

    Batel, Essa (The University of Arizona., 2019)
    This dissertation consists of three experiments testing various aspects of sentence processing in non-native English speakers. Experiment 1 tested the effect of constraining sentence context on word recognition time (RT) in the first and second language. English language learners (L2) and English native speakers (L1) performed self-paced reading and listening tasks to see whether a semantically-rich preceding context would lead to the activation of a probable upcoming word prior to encountering it. The pre-access prediction model (e.g., Altmann & Kamide, 1999; McClelland & Rumelhart, 1981) posits that when the preceding context is semantically high-constraining, the perceptual system anticipates a probable upcoming word prior to encountering it. In contrast, the post-access model (e.g., Fodor, 1983; Forster, 1981; Traxler et al., 2000; Van Petten & Luka, 2012) suggests that it is only after a word is encountered that a subsequent process integrates it into the preceding context. The integration process is easier and faster when the word is more congruent with the preceding context. In line with these two models of visual word recognition, auditory word recognition is modeled by the Trace Model (McClelland & Elman, 1986) which suggests that spoken word is influenced by the preceding sentence context whereas models such as the Cohort Model (Marslen-Wilson, 1984) supports post-access integration process. Regarding L2 processing, some studies observed a facilitative effect of sentence context on L2 word recognition (e.g., Kamide, 2003) while others found no effect of sentence context in L2 (e.g., Ito et al., 2017; Martin et al., 2013). In the present experiment, the RTs of English native speakers (L1) and English non-native speakers (L2) were collected in both visual vs. auditory word recognition in semantically high-constraint sentences and semantically low-constraint sentences. A linear mixed effects model showed that both groups of participants were faster on a word when it was proceeded by a semantically high-constraining context. This result was observed in both the visual and the auditory modalities, lending some support for a mechanism facilitating access of target words based on sentence context in both L1 and L2. Experiment 2 also investigated the effect of a constraining sentence context on word recognition, but the target words have multiple meanings (e.g., bank: [1] a financial institution, [2] an edge of a river/lake). A self-paced reading task was performed by both English L1 and L2 participants, in which all sentences were biased for the secondary meaning (i.e., bank as an edge of a river/lake). The results showed that L1 participants were able to use the constraining context to activate the secondary meaning, but L2 participants had some difficulty in activating the secondary meaning even when the preceding context was biased for it. The results of the L1 participants are compatible with the Reordered Access Model (Duffy, Kambe, & Rayner, 2001; Duffy, Morris, & Rayner, 1988; Sheridan, Reingold, & Daneman, 2009) in that a preceding biasing context leads to an activation of the secondary meaning equal to that of the primary meaning. However, the results of the L2 participants are not compatible with this model’s explanation. Experiment 3 tested the recognition process of L2 words in formulaic sequences whose co-occurrence happens more often than by chance (e.g., The new job requires a teacher who is highly qualified) compared to a combination that sounds less natural (e.g., The new job requires a teacher who is hugely qualified) in a sentence context. Based on the Associative Relatedness Model (Postman & Keppel, 1970) that underlies the Spreading Activation theory (Collins & Loftus, 1975), a facilitative recognition of a target word occurs as a result of activation from an associated word. Thus, this experiment tested whether L2 speakers are sensitive to the less natural co-occurrence formulaic sequence in the same way as L1 speakers. Both language groups performed a self-paced reading task. The results showed that the RTs of L1 participants on the target word (e.g., qualified in the above example) is significantly longer in the less natural combination compared to the natural one. However, the RTs of L2 participants on both combination types were not significantly different from one another, although the L2 participants were able to detect the less natural combination in the post-experiment test. This result indicates a difference in processing between the declarative knowledge (i.e., the stored facts) and the procedural processing (i.e., putting the stored knowledge into action), which is more compatible with the declarative/procedural model proposed by Ullman (2001). The final part of this dissertation discusses some general aspects of L2 word processing when embedded in sentence contexts and some points to consider when conducting such tasks with non-native speakers.
  • Evaluating the Psychometric Properties and Determining Clinical Meaningfulness of the Psoriasis Symptom Inventory

    Patel, Mira Janak (The University of Arizona., 2019)
    Plaque psoriasis is a skin condition that affects two to three percent of individuals around the world by causing not only signs (e.g., redness) and symptoms (e.g., flaking), but also, impairments in health-related quality of life (e.g., work function). Evaluation of psoriasis in research studies is important to provide the occurrence and magnitude of treatment benefit, especially from the patient’s perspective. Therefore, the Psoriasis Symptom Inventory (PSI) was developed. The PSI is a patient-reported outcome (PRO) measures that assesses the signs and symptoms of psoriasis. While initial research has been conducted on the assessing the psychometric properties of the measure, more work is still needed to assist in validating the measure. Therefore, this research was conducted to further the quantitative evaluation of the PSI by conducting a secondary analysis that utilized data from two separate studies. The first was a longitudinal, 16-week study that used the 24-hour recall, electronic version of the PSI, while the second study was a prospective, cross-sectional study that used the 7-day recall, paper version of the PSI. Given the differences in the design of the two studies, focused research aims were created to best utilize the given data from each source to evaluate the measurement properties and assess the clinical meaningfulness of the PSI. The aims were targeted more towards research that had not been done on the PSI as of yet. To provide a comprehensive report, eight chapters were developed to detail the research that was conducted. Chapter 1 provided initial background information on psoriasis and the use of clinical measures, including the PSI, to evaluate the skin condition. In addition, the chapter addressed some of the research that has been done on the measurement properties of the PSI, as well as, the unmet research objectives and hypothesis that were conducted for this study. In Chapter 2, a literature review was conducted to primarily provide an in-depth review of the current use of PRO measures in psoriasis, in addition to, previous research that has been conducted when using the PSI. Chapter 3 focused on the data sources for conducting this research including information on data collection and the variables that were available in each dataset. The remaining chapters focused on the results that met the objectives of this research. In Chapter 4, a summary of the psychometric properties (e.g., classical test theory [CTT], item response theory [IRT]) for the PSI (24-hour) were evaluated at Baseline, Week 8, Week 12, and Week 16. The primary findings of this measurement properties analyses indicated good item and measure performance at Baseline. However, due to low sample size, analysis for the remaining timepoints had to be interpreted with caution as treatment benefit affected the choosing of response options by participants. Chapter 5 evaluated the categorization of the PSI responder against other clinical measure responders. Key findings from this analysis showed that the PSI responder had moderate association (kappa= 0.40-0.60) with some of the previously developed clinical measure responders, and therefore, the PSI can provide clinically meaningful interpretation of psoriasis improvement. In Chapter 6, responsiveness was evaluated, as well as, the clinically important difference (CID) and responder (CIR) were determined for the PSI. The PSI total score averaged (mean + SD) at 6.76+5.93 at Week 16, which resulted in a change score of 6.88+7.72 (improvement) from Baseline. The CID for the PSI was 3.39 (95% CI: 3.08, 3.17) while the CIR was 27.16% (95% CI: 20.74, 33.59). In Chapter 7, the psychometric properties of the PSI (7-day) were assessed. With the use of CTT, CFA, Rasch, and IRT, the PSI showed good item and measure performance. Key findings from the analyses showed that the PSI had high internal consistency (CTT; Cronbach’s alpha=0.95), good overall fit (CFA; comparative fit index/Tucker-Lewis index=0.99/0.98), and no misfitting items (Rasch) and overlapping of response options (IRT). Chapter 8 observed the differential item functioning (DIF) of the PSI (7-day) between males and females. From the analysis, one item (Item 2: Redness) was flagged for non-uniform DIF (X223=0.007). All items, including the redness item, showed that the magnitude for DIF was negligible (R2<0.13). When evaluating the test characteristic curve, sex was not associated with difference in response for the PSI. Overall, this research assessed various key quantitative analyses for evaluating the psychometric properties, as well as, clinical meaningfulness of the measure. The findings showed the PSI had good item and measure performance. In addition, the responder status of the PSI associated well with other clinical measure thresholds. Furthermore, the PSI did not show differences in item functioning between sexes. And finally, clinically important difference and responder were established for the PSI to be assessed between treatment groups in future studies. Further research is warranted to assess the PSI in a larger sample and in a real-world setting for both versions for quantitative analyses, such as determining measurement invariance across time and cross-cultural performance of the PSI.
  • Development and Evaluation of Habitat Suitability Criteria for Native Fishes and Assessment of the Relationship Among Riparian Areas and Stream Macrohabitats Type and Fish Presence in Four Central Arizona Streams

    Nemec, Zach (The University of Arizona., 2018)
    Habitat loss is an important reason for fish fauna declines in the southwestern U.S. Several studies have defined habitat conditions for selected native fish species in Arizona, yet habitat use can vary across streams due to a variety of biological and physical factors. In addition, previous studies have focused on effects of instream habitat characteristics and less on how riparian areas structure aquatic communities of the Southwest. Riparian areas affect aquatic communities in a variety of ways, including structuring instream habitat. Macrohabitat (riffle, run, pool) is an important determinant of fish use, and little is known about the effect of riparian vegetation and associated land use activities on the formation of macrohabitat. Therefore, the objectives of my study were to 1) evaluate suitable habitat for native Arizona species, and 2) to investigate the relationships among riparian vegetation and stream macrohabitats type and fish presence in four central Arizona streams. Fish and habitat data were collected in four streams along the Mogollon Rim in Arizona during the 2017 summer field season at base flow conditions. I used the National Hydrography Dataset (NHD) and National Agricultural Imagery Program (NAIP) aerial imagery to estimate the amount of vegetation cover within the riparian areas of each stream. I developed habitat suitability criteria for four native species in three streams. Most generalized criteria did not transfer among study streams, similar to finding from past studies suggesting that stream-specific criteria were more accurate. I found that Smallmouth Bass and Red Shiner had a negative relationship to canopy cover, possibly explained by high temperature tolerances of both species. Desert Sucker and Speckled Dace presence were positively related to presence of riffle habitat, as has been noted in previous habitat studies. Riffle habitat was positively related to increases in riparian vegetation cover. These results can inform researchers, agencies and stakeholders who study and manage Arizona’s riparian areas and instream habitat.
  • Examination of Latinx Bullying Victimization and Depressive Symptoms through a Social-Ecological Framework

    Lutrick, Karen (The University of Arizona., 2019)
    BACKGROUND: Bullying victimization is correlated with depressive symptoms in adolescents. While the literature is extensive, there has been little focus on racial/ethnic minorities, specifically Latinx youth. In the United States, there is some evidence that Latinx adolescents experience bullying victimization and depressive symptoms at higher rates than their non-Hispanic white (NHW) peers. OBJECTIVES: This dissertation is composed of three studies that work together to identify factors of influence in the development of depressive symptoms within Latinx adolescents that experience peer violence: 1) a synthesis of the bullying/depression literature to evaluate Latinx representation and Latinx-specific factors; 2) identification of the interaction between racial/ethnic discrimination and bullying victimization on depressive symptoms; and 3) examination of the role of family and social support as a protective factor in the relationship between bullying victimization and depressive symptoms three years after victimization. METHODS: A systematic review was conducted for research aim one following the Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA) guidelines. For research aims two and three, secondary data analysis was conducted utilizing the Healthy Passages national and longitudinal dataset with permission from the publication committee. Hierarchal regression analyses were conducted for research aims two and three. RESULTS: For research aim one, of 957 studies identified, 17 included a Latinx population of 25% or more. They all identified a relationship between bullying and depression, with nine examining factors related to race/ethnicity or unique to the Latinx population. For aims two and three, a sample of 1,666 Latinx adolescents (grade 7) reported bullying victimization rates of 60% within the previous year. For aim two specifically, 15.7% reported racial/ethnic discrimination and 14.4% reported bullying and discrimination victimization in the previous year. All forms of victimization were found to be significantly related to depressive symptoms, including the interaction between bullying and discrimination victimization (p<.001 for immediate effect, p<.05 three years later). For research aim three, parent/child connectedness had a moderation effect (p<.001, b=-.061; p=.011, b=.006), reducing the likelihood of depressive symptoms three years after victimization. Social support reduced the relationship to depressive symptoms (p<.001, b=.025) but was not a moderator, and global parental monitoring had no significant effect. CONCLUSIONS: The Latinx community is the fastest growing racial/ethnic minority population in the United States, but they are underrepresented in the bullying literature. Studies that included variables unique to Latinx communities such as acculturation and unique family factors found a stronger relationship between bullying victimization and depression. In a Latinx sample, this dissertation identified an increased likelihood of depressive symptoms when bullying and racial/ethnic discrimination were experienced. Additionally, this work found that strong family relationships and strong social support reduced the likelihood of depressive symptoms of bullying victimization even three years after victimization occurred. These two findings identify potential future directions for bullying research and practice. At a minimum, they illustrate a need for an expanded social-ecological lens when measuring victimization as well as the inclusion of family and the development of strong family relationships in bullying interventions.
  • Evaluations of pancreatic cancer with acidoCEST MRI

    High, Rachel (The University of Arizona., 2019)
    Pancreatic ductal adenocarcinoma (PDAC) is a virulent disease which readily develops resistances to prevailing chemotherapies. PDAC is often not diagnosed until the disease presents in an inoperable late stage. Early detection of PDAC while there is still hope for effective treatment is critical to survival. Cancer cells experience a metabolic shift towards aerobic glycolysis as they develop. This atypical metabolism results in the production and export of lactic acid, which results in acidification of the extracellular tumor microenvironment. Thus, acidosis is a biomarker of cancer development. Our research program uses a technique called chemical exchange saturation transfer magnetic resonance imaging (acidoCEST MRI) to measure acidosis in the extracellular microenvironment. The work presented in this dissertation will focus on characterization of acidosis in PDAC using acidoCEST MRI. Chapter 1 reviews components of the tumor microenvironment that are affected by acidosis as well as briefly overviews aspects of the microenvironment of pancreatic cancer. Chapter 2 presents work analyzing the role of the glucose transporter GLUT3 in chemoresistance and acidosis of two pancreatic cancer cell lines. Chapter 3 explores the use of acidosis as a prognostic biomarker of pancreatic cancer development. Finally, chapter 4 highlights future studies which would be a natural continuation of the work in this dissertation or would contribute to the advancement of acidoCEST MRI as a research technique.
  • Novel Host Targets in Respiratory Viral Illnesses

    Zhou, Xu (The University of Arizona., 2019)
    Rhinovirus (RV) and influenza virus infections are the leading causes of airway tract problems that lead to cold and flu and exacerbated asthma, chronic obstructive pulmonary disease (COPD), and other respiratory illnesses. This dissertational study investigated the effects of host factors in these viral infections. Chapter 2 and chapter 3 are related research about rhinovirus, and chapter 4 is research about influenza virus. Human rhinovirus (RV) is the major cause of common cold and it also plays a significant role in asthma and asthma exacerbation. Airway epithelium is the primary site of RV infection and production. In contrast, monocytic cells (e.g., monocytes and macrophages) are believed to be non-permissive for RV replication. Instead, RV has been shown to modulate inflammatory gene expressions in these cells via a replication-independent mechanism. However, Johnston et al. firstly reported a low-grade productive replication of RV9 in monocytes and also showed the RV16 replication in monocytes in following studies. In the present study, we generated the epithelial cell- monocyte coculture system. RV16 (a major-group RV) replication was found to be significantly enhanced in monocytes when co-cultivated with airway epithelial cells. This effect appeared to be mediated by secretory components from epithelial cells, which stimulated RV16 replication and significantly elevated the expression of a number of proinflammatory cytokines. The lack of such effect with RV1A, a minor-group RV that enters the cell by a different receptor, suggests that ICAM1, the receptor for major-group RVs, may be involved. Conditioned media from epithelial cells significantly increased ICAM1 expression in monocytes. Consistently, ICAM1 overexpression and ICAM1 knockdown enhanced and blocked RV production, confirming the role of ICAM1 in this process. In addition, we isolated secretory components from the epithelial conditioned medium and further determined their function. Alpha-Heremans Schmid Glycoprotein (AHSG) is the top matched secretory protein to induce the ICAM1 expression. High AHSG expression is closely related to asthma in mice models and human patients. Thus, we demonstrated that airway epithelial cells direct significant RV16 replication in monocytic cells via an ICAM1-dependent mechanism. This is the first time we demonstrated that AHSG expression is related to asthma. Influenza virus is the major cause of influenza (or the flu). Pandemic flu killed tens of millions of people and seasonal flu yearly outbreak also caused severe illness and hundreds of thousands of deaths. Current options for preventing or treating influenza are either limited (e.g., vaccine) or becoming ineffective due to the emergence of drug resistant strains (e.g., M2 blockers). Thus, new treatments for influenza viral infection are urgently needed. Host targets are relatively stable, so the drugs have high barrier when they are targeting on the host factors. To develop drugs targeting the host, it is important to understand the influenza related pathway in the host cells. In the present study, we generated an influenza infectious model on human airway epithelial cells, which are primary sites for the virus’ infections. We serendipitously found that a small-molecule inhibitor (AG1478), previously used for epidermal growth factor receptor (EGFR) inhibition, demonstrated a potent and broad-spectrum activity against influenza. Surprisingly, the antiviral effect of AG1478 was not mediated by its EGFR inhibitory activity, as influenza was insensitive to EGFR blockade by other EGFR inhibitors or by the knockdown using a small interference RNA against EGFR. Additionally, interferons are the major anti-viral proteins in the cells, its effect has to be considered when we research on the anti-viral activity. A knockout approach using Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) further demonstrated that this antiviral activity was also interferon independent. AG1478 was found to target on Golgi-specific brefeldin A-resistance guanine nucleotide exchange factor 1- ADP ribosylation factor 1 (GBF1-ARF1). AG1478 reversibly inhibit GBF1 activity and disrupted its Golgi-cytoplasmic trafficking. Compared to the two existing GBF1 inhibitors, AG1478 demonstrated lower cellular toxicity and better preservation of Golgi structure, suggesting its antiviral activity may not originate from a non-specific disruption of intracellular vesicle trafficking regulated by GBF1. GBF1 was found to interact with a specific set of viral proteins including M1, NP and PA. AG1478-elicited spatial alternation of GBF1 distribution disrupted these interactions. Because host factors are more genetically stable than viral proteins, host-targeting antivirals might have a higher genetic barrier to drug resistance than direct-acting antivirals. In conclusion, the host factors are crucial for these respiratory viral illnesses, and drugs targeting host factors are a new direction for treating infectious illnesses. 1) For rhinovirus, we demonstrate that epithelial secretions, especially AHSG, direct robust RV replication in monocytes via significantly increased ICAM1. This new information will advance our understanding of the interaction between airway epithelium and inflammatory cells in the context of RV infection and RV-induced disease exacerbation. Blocking ICAM1 or decreasing AHSG expression will be a new direction in treating rhinovirus infections. This finding will open a new venue for the study of RV infection in airway disease and its exacerbation. 2) For influenza virus, through a serendipitous finding, we have discovered a potent and broad-spectrum anti-influenza drug candidate-AG1478. Its antiviral activity is mediated by targeting GBF1.The treatment of AG1478 disrupted this interaction and potentially impaired vRNP transport leading to markedly decreased IAV production. It is a new direction to develop the drugs targeting GBF1. Further development on this candidate target will lead to novel anti-influenza therapy.

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