ASU Electronic Theses and Dissertations
This collection includes most of the ASU Theses and Dissertations from 2011 to present. ASU Theses and Dissertations are available in downloadable PDF format; however, a small percentage of items are under embargo. Information about the dissertations/theses includes degree information, committee members, an abstract, supporting data or media.
In addition to the electronic theses found in the ASU Digital Repository, ASU Theses and Dissertations can be found in the ASU Library Catalog.
Dissertations and Theses granted by Arizona State University are archived and made available through a joint effort of the ASU Graduate College and the ASU Libraries. For more information or questions about this collection contact or visit the Digital Repository ETD Library Guide or contact the ASU Graduate College at gradformat@asu.edu.
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- All Subjects: Biology
- Creators: LaBaer, Joshua
- Creators: Kusumi, Kenro
Description
Type 1 diabetes (T1D) is a chronic autoimmune disease characterized by progressive autoimmune destruction of insulin-producing pancreatic β-cells. Genetic, immunological and environmental factors contribute to T1D development. The focus of this dissertation is to track the humoral immune response in T1D by profiling autoantibodies (AAbs) and anti-viral antibodies using an innovative protein array platform called Nucleic Acid Programmable Protein Array (NAPPA).
AAbs provide value in identifying individuals at risk, stratifying patients with different clinical courses, improving our understanding of autoimmune destructions, identifying antigens for cellular immune response and providing candidates for prevention trials in T1D. A two-stage serological AAb screening against 6,000 human proteins was performed. A dual specificity tyrosine-phosphorylation-regulated kinase 2 (DYRK2) was validated with 36% sensitivity at 98% specificity by an orthogonal immunoassay. This is the first systematic screening for novel AAbs against large number of human proteins by protein arrays in T1D. A more comprehensive search for novel AAbs was performed using a knowledge-based approach by ELISA and a screening-based approach against 10,000 human proteins by NAPPA. Six AAbs were identified and validated with sensitivities ranged from 16% to 27% at 95% specificity. These two studies enriched the T1D “autoantigenome” and provided insights into T1D pathophysiology in an unprecedented breadth and width.
The rapid rise of T1D incidence suggests the potential involvement of environmental factors including viral infections. Sero-reactivity to 646 viral antigens was assessed in new-onset T1D patients. Antibody positive rate of EBV was significantly higher in cases than controls that suggested a potential role of EBV in T1D development. A high density-NAPPA platform was demonstrated with high reproducibility and sensitivity in profiling anti-viral antibodies.
This dissertation shows the power of a protein-array based immunoproteomics approach to characterize humoral immunoprofile against human and viral proteomes. The identification of novel T1D-specific AAbs and T1D-associated viruses will help to connect the nodes in T1D etiology and provide better understanding of T1D pathophysiology.
AAbs provide value in identifying individuals at risk, stratifying patients with different clinical courses, improving our understanding of autoimmune destructions, identifying antigens for cellular immune response and providing candidates for prevention trials in T1D. A two-stage serological AAb screening against 6,000 human proteins was performed. A dual specificity tyrosine-phosphorylation-regulated kinase 2 (DYRK2) was validated with 36% sensitivity at 98% specificity by an orthogonal immunoassay. This is the first systematic screening for novel AAbs against large number of human proteins by protein arrays in T1D. A more comprehensive search for novel AAbs was performed using a knowledge-based approach by ELISA and a screening-based approach against 10,000 human proteins by NAPPA. Six AAbs were identified and validated with sensitivities ranged from 16% to 27% at 95% specificity. These two studies enriched the T1D “autoantigenome” and provided insights into T1D pathophysiology in an unprecedented breadth and width.
The rapid rise of T1D incidence suggests the potential involvement of environmental factors including viral infections. Sero-reactivity to 646 viral antigens was assessed in new-onset T1D patients. Antibody positive rate of EBV was significantly higher in cases than controls that suggested a potential role of EBV in T1D development. A high density-NAPPA platform was demonstrated with high reproducibility and sensitivity in profiling anti-viral antibodies.
This dissertation shows the power of a protein-array based immunoproteomics approach to characterize humoral immunoprofile against human and viral proteomes. The identification of novel T1D-specific AAbs and T1D-associated viruses will help to connect the nodes in T1D etiology and provide better understanding of T1D pathophysiology.
ContributorsBian, Xiaofang (Author) / LaBaer, Joshua (Thesis advisor) / Mandarino, Lawrence (Committee member) / Chang, Yung (Committee member) / Arizona State University (Publisher)
Created2015
Description
Skeletal muscles arise from the myotome compartment of the somites that form during vertebrate embryonic development. Somites are transient structures serve as the anlagen for the axial skeleton, skeletal muscle, tendons, and dermis, as well as imposing the metameric patterning of the axial musculoskeletal system, peripheral nerves, and vasculature. Classic studies have described the role of Notch, Wnt, and FGF signaling pathways in controlling somite formation and muscle formation. However, little is known about the transformation of myotome compartments into identifiable post-natal muscle groups. Using a mouse model, I have undertaken an evaluation of morphological events, including hypertrophy and hyperplasia, related to the formation of several muscles positioned along the dorsal surface of the vertebrae and ribs. Lunatic fringe (Lfng) deficient embryos and neonates were also examined to further understand the role of the Notch pathway in these processes as it is a modulator of the Notch receptor and plays an important role in defining somite borders and anterior-posterior patterning in many vertebrates. Lunatic fringe deficient embryos showed defects in muscle fiber hyperplasia and hypertrophy in the iliocostalis and longissimus muscles of the erector spinae group. This novel data suggests an additional role for Lfng and the Notch signaling pathway in embryonic and fetal muscle development.
ContributorsDe Ruiter, Corinne (Author) / Rawls, J. Alan (Thesis advisor) / Wilson-Rawls, Jeanne (Committee member) / Kusumi, Kenro (Committee member) / Fisher, Rebecca E. (Committee member) / Arizona State University (Publisher)
Created2012
Description
The most abundantly studied societies, with the exception of humans, are those of the eusocial insects, which include all ants. Eusocial insect societies are typically composed of many dozens to millions of individuals, referred to as nestmates, which require some form of communication to maintain colony cohesion and coordinate the activities within them. Nestmate recognition is the process of distinguishing between nestmates and non-nestmates, and embodies the first line of defense for social insect colonies. In ants, nestmate recognition is widely thought to occur through olfactory cues found on the exterior surfaces of individuals. These cues, called cuticular hydrocarbons (CHCs), comprise the overwhelming majority of ant nestmate profiles and help maintain colony identity. In this dissertation, I investigate how nestmate recognition is influenced by evolutionary, ontogenetic, and environmental factors. First, I contributed to the sequencing and description of three ant genomes including the red harvester ant, Pogonomyrmex barbatus, presented in detail here. Next, I studied how variation in nestmate cues may be shaped through evolution by comparatively studying a family of genes involved in fatty acid and hydrocarbon biosynthesis, i.e., the acyl-CoA desaturases, across seven ant species in comparison with other social and solitary insects. Then, I tested how genetic, developmental, and social factors influence CHC profile variation in P. barbatus, through a three-part study. (1) I conducted a descriptive, correlative study of desaturase gene expression and CHC variation in P. barbatus workers and queens; (2) I explored how larger-scale genetic variation in the P. barbatus species complex influences CHC variation across two genetically isolated lineages (J1/J2 genetic caste determining lineages); and (3) I experimentally examined how CHC development is influenced by an individual’s social environment. In the final part of my work, I resolved discrepancies between previous findings of nestmate recognition behavior in P. barbatus by studying how factors of territorial experience, i.e., spatiotemporal relationships, affect aggressive behaviors among red harvester ant colonies. Through this research, I was able to identify promising methodological approaches and candidate genes, which both broadens our understanding of P. barbatus nestmate recognition systems and supports future functional genetic studies of CHCs in ants.
ContributorsCash, Elizabeth I (Author) / Gadau, Jürgen (Thesis advisor) / Liebig, Jürgen (Thesis advisor) / Fewell, Jennifer (Committee member) / Hölldobler, Berthold (Committee member) / Kusumi, Kenro (Committee member) / Arizona State University (Publisher)
Created2016
Description
Emerging pathogens present several challenges to medical diagnostics. Primarily, the exponential spread of a novel pathogen through naïve populations require a rapid and overwhelming diagnostic response at the site of outbreak. While point-of-care (PoC) platforms have been developed for detection of antigens, serologic responses, and pathogenic genomes, only nucleic acid diagnostics currently have the potential to be developed and manufactured within weeks of an outbreak owing to the speed of next-generation sequencing and custom DNA synthesis. Among nucleic acid diagnostics, isothermal amplification strategies are uniquely suited for PoC implementation due to their simple instrumentation and lack of thermocycling requirement. Unfortunately, isothermal strategies are currently prone to spurious nonspecific amplification, hindering their specificity and necessitating extensive empirical design pipelines that are both time and resource intensive. In this work, isothermal amplification strategies are extensively compared for their feasibility of implementation in outbreak response scenarios. One such technology, Loop-mediated Amplification (LAMP), is identified as having high-potential for rapid development and PoC deployment. Various approaches to abrogating nonspecific amplification are described including a novel in silico design tool based on coarse-grained simulation of interactions between thermophilic DNA polymerase and DNA strands in isothermal reaction conditions. Nonspecific amplification is shown to be due to stabilization of primer secondary structures by high concentrations of Bst DNA polymerase and a mechanism of micro-complement-mediated cross-priming is demonstrated as causal via nanopore sequencing of nonspecific reaction products. The resulting computational model predicts primer set background in 64% of 67 test assays and its usefulness is illustrated further by determining problematic primers in a West Nile Virus-specific LAMP primer set and optimizing primer 3’ nucleotides to eliminate micro-complements within the reaction, resulting in inhibition of background accumulation. Finally, the emergence of Orthopox monkeypox (MPXV) as a recurring threat is discussed and SimCycle is utilized to develop a novel technique for clade-specific discrimination of MPXV based on bridging viral genomic rearrangements (Bridging LAMP). Bridging LAMP is implemented in a 4-plex microfluidic format and demonstrates 100% sensitivity in detection of 100 copies of viral lysates and 45 crude MPXV-positive patient samples collected during the 2022 Clade IIb outbreak.
ContributorsKnappenberger, Mark Daniel (Author) / Anderson, Karen S (Thesis advisor) / LaBaer, Joshua (Committee member) / Roberson, Robert (Committee member) / Lindsay, Stuart (Committee member) / Arizona State University (Publisher)
Created2023
Description
According to the World Health Organization, cancer is one of the leading causes of death around the world. Although early diagnostics using biomarkers and improved treatments with targeted therapy have reduced the rate of cancer related mortalities, there remain many unknowns regarding the contributions of the tumor microenvironment to cancer progression and therapeutic resistance. The tumor microenvironment plays a significant role by manipulating the progression of cancer cells through biochemical and biophysical signals from the surrounding stromal cells along with the extracellular matrix. As such, there is a critical need to understand how the tumor microenvironment influences the molecular mechanisms underlying cancer metastasis to facilitate the discovery of better therapies. This thesis described the development of microfluidic technologies to study the interplay of cancer cells with their surrounding microenvironment. The microfluidic model was used to assess how exposure to chemoattractant, epidermal growth factor (EGF), impacted 3D breast cancer cell invasion and enhanced cell motility speed was noted in the presence of EGF validating physiological cell behavior. Additionally, breast cancer and patient-derived cancer-associated fibroblast (CAF) cells were co-cultured to study cell-cell crosstalk and how it affected cancer invasion. GPNMB was identified as a novel gene of interest and it was shown that CAFs enhanced breast cancer invasion by up-regulating the expression of GPNMB on breast cancer cells resulting in increased migration speed. Lastly, this thesis described the design, biological validation, and use of this microfluidic platform as a new in vitro 3D organotypic model to study mechanisms of glioma stem cell (GSC) invasion in the context of a vascular niche. It was confirmed that CXCL12-CXCR4 signaling is involved in promoting GSC invasion in a 3D vascular microenvironment, while also demonstrating the effectiveness of the microfluidic as a drug screening assay. Taken together, the broader impacts of the microfluidic model developed in this dissertation include, a possible alternative platform to animal testing that is focused on mimicking human physiology, a potential ex vivo platform using patient-derived cells for studying the interplay of cancer cells with its surrounding microenvironment, and development of future therapeutic strategies tailored toward disrupting key molecular pathways involved in regulatory mechanisms of cancer invasion.
ContributorsTruong, Danh, Ph.D (Author) / Nikkhah, Mehdi (Thesis advisor) / LaBaer, Joshua (Committee member) / Smith, Barbara (Committee member) / Mouneimne, Ghassan (Committee member) / Vernon, Brent (Committee member) / Arizona State University (Publisher)
Created2018
Description
Next-generation sequencing is a powerful tool for detecting genetic variation. How-ever, it is also error-prone, with error rates that are much larger than mutation rates.
This can make mutation detection difficult; and while increasing sequencing depth
can often help, sequence-specific errors and other non-random biases cannot be de-
tected by increased depth. The problem of accurate genotyping is exacerbated when
there is not a reference genome or other auxiliary information available.
I explore several methods for sensitively detecting mutations in non-model or-
ganisms using an example Eucalyptus melliodora individual. I use the structure of
the tree to find bounds on its somatic mutation rate and evaluate several algorithms
for variant calling. I find that conventional methods are suitable if the genome of a
close relative can be adapted to the study organism. However, with structured data,
a likelihood framework that is aware of this structure is more accurate. I use the
techniques developed here to evaluate a reference-free variant calling algorithm.
I also use this data to evaluate a k-mer based base quality score recalibrator
(KBBQ), a tool I developed to recalibrate base quality scores attached to sequencing
data. Base quality scores can help detect errors in sequencing reads, but are often
inaccurate. The most popular method for correcting this issue requires a known
set of variant sites, which is unavailable in most cases. I simulate data and show
that errors in this set of variant sites can cause calibration errors. I then show that
KBBQ accurately recalibrates base quality scores while requiring no reference or other
information and performs as well as other methods.
Finally, I use the Eucalyptus data to investigate the impact of quality score calibra-
tion on the quality of output variant calls and show that improved base quality score
calibration increases the sensitivity and reduces the false positive rate of a variant
calling algorithm.
This can make mutation detection difficult; and while increasing sequencing depth
can often help, sequence-specific errors and other non-random biases cannot be de-
tected by increased depth. The problem of accurate genotyping is exacerbated when
there is not a reference genome or other auxiliary information available.
I explore several methods for sensitively detecting mutations in non-model or-
ganisms using an example Eucalyptus melliodora individual. I use the structure of
the tree to find bounds on its somatic mutation rate and evaluate several algorithms
for variant calling. I find that conventional methods are suitable if the genome of a
close relative can be adapted to the study organism. However, with structured data,
a likelihood framework that is aware of this structure is more accurate. I use the
techniques developed here to evaluate a reference-free variant calling algorithm.
I also use this data to evaluate a k-mer based base quality score recalibrator
(KBBQ), a tool I developed to recalibrate base quality scores attached to sequencing
data. Base quality scores can help detect errors in sequencing reads, but are often
inaccurate. The most popular method for correcting this issue requires a known
set of variant sites, which is unavailable in most cases. I simulate data and show
that errors in this set of variant sites can cause calibration errors. I then show that
KBBQ accurately recalibrates base quality scores while requiring no reference or other
information and performs as well as other methods.
Finally, I use the Eucalyptus data to investigate the impact of quality score calibra-
tion on the quality of output variant calls and show that improved base quality score
calibration increases the sensitivity and reduces the false positive rate of a variant
calling algorithm.
ContributorsOrr, Adam James (Author) / Cartwright, Reed (Thesis advisor) / Wilson, Melissa (Committee member) / Kusumi, Kenro (Committee member) / Taylor, Jesse (Committee member) / Pfeifer, Susanne (Committee member) / Arizona State University (Publisher)
Created2020
Description
Satellite cells are adult muscle stem cells that activate, proliferate, and differentiate into myofibers upon muscle damage. Satellite cells can be cultured and manipulated in vitro, and thus represent an accessible model for studying skeletal muscle biology, and a potential source of autologous stem cells for regenerative medicine. This work summarizes efforts to further understanding of satellite cell biology, using novel model organisms, bioengineering, and molecular and cellular approaches. Lizards are evolutionarily the closest vertebrates to humans that regenerate entire appendages. An analysis of lizard myoprogenitor cell transcriptome determined they were most transcriptionally similar to mammalian satellite cells. Further examination showed that among genes with the highest level of expression in lizard satellite cells were an increased number of regulators of chondrogenesis. In micromass culture, lizard satellite cells formed nodules that expressed chondrogenic regulatory genes, thus demonstrating increased musculoskeletal plasticity. However, to exploit satellite cells for therapeutics, development of an ex vivo culture is necessary. This work investigates whether substrates composed of extracellular matrix (ECM) proteins, as either coatings or hydrogels, can support expansion of this population whilst maintaining their myogenic potency. Stiffer substrates are necessary for in vitro proliferation and differentiation of satellite cells, while the ECM composition was not significantly important. Additionally, satellite cells on hydrogels entered a quiescent state that could be reversed when the cells were subsequently cultured on Matrigel. Proliferation and gene expression data further indicated that C2C12 cells are not a good proxy for satellite cells. To further understand how different signaling pathways control satellite cell behavior, an investigation of the Notch inhibitor protein Numb was carried out. Numb deficient satellite cells fail to activate, proliferate and participate in muscle repair. Examination of Numb isoform expression in satellite cells and embryonic tissues revealed that while developing limb bud, neural tube, and heart express the long and short isoforms of NUMB, satellite cells predominantly express the short isoforms. A preliminary immunoprecipitation- proteomics experiment suggested that the roles of NUMB in satellite cells are related to cell cycle modulation, cytoskeleton dynamics, and regulation of transcription factors necessary for satellite cell function.
ContributorsPalade, Joanna (Author) / Wilson-Rawls, Norma (Thesis advisor) / Rawls, Jeffrey (Committee member) / Kusumi, Kenro (Committee member) / Newbern, Jason (Committee member) / Stabenfeldt, Sarah (Committee member) / Arizona State University (Publisher)
Created2020
Cancer autoantibody biomarker discovery and validation using nucleic acid programmable protein array
Description
Currently in the US, many patients with cancer do not benefit from the population-based screening, due to challenges associated with the existing cancer screening scheme. Blood-based diagnostic assays have the potential to detect diseases in a non-invasive way. Proteins released from small early tumors may only be present intermittently and get diluted to tiny concentrations in the blood, making them difficult to use as biomarkers. However, they can induce autoantibody (AAb) responses, which can amplify the signal and persist in the blood even if the antigen is gone. Circulating autoantibodies is a promising class of molecules that have potential to serve as early detection biomarkers for cancers. This Ph.D thesis aims to screen for autoantibody biomarkers for the early detection of two deadly cancer, basal-like breast cancer and lung adenocarcinoma. First, a method was developed to display proteins in both native and denatured conformation on protein array. This method adopted a novel protein tag technology, called HaloTag, to covalently immobilize proteins on glass slide surface. The covalent attachment allowed these proteins to endure harsh treatment without getting dissociated from slide surface, which enabled the profiling of antibody responses against both conformational and linear epitopes. Next, a plasma screening protocol was optimized to significantly increase signal to noise ratio of protein array based AAb detection. Following this, the AAb responses in basal-like breast cancer were explored using nucleic acid programmable protein arrays (NAPPA) containing 10,000 full-length human proteins in 45 cases and 45 controls. After verification in a large sample set (145 basal-like breast cancer cases / 145 controls / 70 non-basal breast cancer) by ELISA, a 13-AAb classifier was developed to differentiate patients from controls with a sensitivity of 33% at 98% specificity. Similar approach was also applied to the lung cancer study to identify AAbs that distinguished lung cancer patients from computed-tomography positive benign pulmonary nodules (137 lung cancer cases, 127 smoker controls, 170 benign controls). In this study, two panels of AAbs were discovered that showed promising sensitivity and specificity. Six out of eight AAb targets were also found to have elevated mRNA level in lung adenocarcinoma patients using TCGA data. These projects as a whole provide novel insights on the association between AAbs and cancer, as well as general B cell antigenicity against self-proteins.
ContributorsWang, Jie (Author) / LaBaer, Joshua (Thesis advisor) / Anderson, Karen S (Committee member) / Lake, Douglas F (Committee member) / Chang, Yung (Committee member) / Arizona State University (Publisher)
Created2015
Description
Glioblastoma (GBM), the most common and aggressive primary brain tumor affecting adults, is characterized by an aberrant yet druggable epigenetic landscape. The Histone Deacetylases (HDACs), a major family of epigenetic regulators, favor transcriptional repression by mediating chromatin compaction and are frequently overexpressed in human cancers, including GBM. Hence, over the last decade there has been considerable interest in using HDAC inhibitors (HDACi) for the treatment of malignant primary brain tumors. However, to date most HDACi tested in clinical trials have failed to provide significant therapeutic benefit to patients with GBM. This is because current HDACi have poor or unknown pharmacokinetic profiles, lack selectivity towards the different HDAC isoforms, and have narrow therapeutic windows. Isoform selectivity for HDACi is important given that broad inhibition of all HDACs results in widespread toxicity across different organs. Moreover, the functional roles of individual HDAC isoforms in GBM are still not well understood. Here, I demonstrate that HDAC1 expression increases with brain tumor grade and is correlated with decreased survival in GBM. I find that HDAC1 is the essential HDAC isoform in glioma stem cells and its loss is not compensated for by its paralogue HDAC2 or other members of the HDAC family. Loss of HDAC1 alone has profound effects on the glioma stem cell phenotype in a p53-dependent manner and leads to significant suppression of tumor growth in vivo. While no HDAC isoform-selective inhibitors are currently available, the second-generation HDACi quisinostat harbors high specificity for HDAC1. I show that quisinostat exhibits potent growth inhibition in multiple patient-derived glioma stem cells. Using a pharmacokinetics- and pharmacodynamics-driven approach, I demonstrate that quisinostat is a brain-penetrant molecule that reduces tumor burden in flank and orthotopic models of GBM and significantly extends survival both alone and in combination with radiotherapy. The work presented in this thesis thereby unveils the non-redundant functions of HDAC1 in therapy- resistant glioma stem cells and identifies a brain-penetrant HDACi with higher selectivity towards HDAC1 as a potent radiosensitizer in preclinical models of GBM. Together, these results provide a rationale for developing quisinostat as a potential adjuvant therapy for the treatment of GBM.
ContributorsLo Cascio, Costanza (Author) / LaBaer, Joshua (Thesis advisor) / Mehta, Shwetal (Committee member) / Mirzadeh, Zaman (Committee member) / Mangone, Marco (Committee member) / Paek, Andrew (Committee member) / Arizona State University (Publisher)
Created2022
Description
Structural Equation Modeling (SEM) is a multivariate analysis methodology that could potentially be utilized to examine the barrier effect that river systems have on genetic differentiation. In this project, river systems are split into the variables of Daily Average Discharge, Average River Width, and Seasonality measurements and regressed onto the genetic differentiation, measured as Fst. This data was collected from the USGS database (U.S. Geological Survey, 2020), sequencing files from differing literature, or Google Earth measurements. Different Structural Equation Modeling models are used to model different system structures as well as compare it to more traditional methodologies like Generalized Linear Modeling and Generalized Linear Mixed Modeling. Ultimately results were limited by the small sample size, however, interesting patterns still emerged from the models. The SE models indicate that Discharge plays a primary role in the genetic differentiation of adjacent river populations. In addition to this, the results demonstrate how quantification of indirect effects, particularly those relating to discharge, give more informative interpretations than traditional multivariate statistics alone. These findings prompt further investigations into this potential methodology.
ContributorsMaag, Garett (Author) / Dolby, Greer A. (Thesis advisor) / Kusumi, Kenro (Thesis advisor) / Stokes, Maya F. (Committee member) / Barly, Anthony (Committee member) / Arizona State University (Publisher)
Created2023