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The GGGGCC (G4C2) hexanucleotide repeat expansion (HRE) in the C9orf72 gene is the most common genetic abnormality associated with both amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), two devastatingly progressive neurodegenerative diseases. The discovery of this genetic link confirmed that ALS and FTD reside along a spectrum with clinical and pathological commonalities. Historically understood as diseases resulting in neuronal death, the role of non-neuronal cells like astrocytes is still wholly unresolved. With evidence of cortical neurodegeneration leading to cognitive impairments in C9orf72-ALS/FTD, there is a need to investigate the role of cortical astrocytes in this disease spectrum. Here, a patient-derived induced pluripotent stem cell (iPSC) cortical astrocyte model was developed to investigate consequences of C9orf72-HRE pathogenic features in this cell type. Although there were no significant C9orf72-HRE pathogenic features in cortical astrocytes, transcriptomic, proteomic and phosphoproteomic profiles elucidated global disease-related phenotypes. Specifically, aberrant expression of astrocytic-synapse proteins and secreted factors were identified. SPARCL1, a pro-synaptogenic secreted astrocyte factor was found to be selectively decreased in C9orf72-ALS/FTD iPSC-cortical astrocytes. This finding was further validated in human tissue analyses, indicating that cortical astrocytes in C9orf72-ALS/FTD exhibit a reactive transformation that is characterized by a decrease in SPARCL1 expression. Considering the evidence for substantial astrogliosis and synaptic failure leading to cognitive impairments in C9orf72-ALS/FTD, these findings represent a novel understanding of how cortical astrocytes may contribute to the cortical neurodegeneration in this disease spectrum.
ContributorsBustos, Lynette (Author) / Sattler, Rita (Thesis advisor) / Newbern, Jason (Committee member) / Zarnescu, Daniela (Committee member) / Brafman, David (Committee member) / Mehta, Shwetal (Committee member) / Arizona State University (Publisher)
Created2023
Description
Understanding how wildlife interact with humans and the built environment is critical as urbanization contributes to habitat change and fragmentation globally. In urban and suburban areas, wildlife and people are often in close quarters, leading to human-wildlife interactions (HWI). In the greater Phoenix Metropolitan Area, Arizona, HWI can involve reptiles such as venomous (family Viperidae, e.g., rattlesnakes) and nonvenomous (family Colubridae, e.g., gophersnakes) snakes. Rattlesnake Solutions, LLC, a local business, removes and relocates snakes from homes and businesses in the Phoenix area and, as a collaborator, has provided records of snake removals. Using these records, I investigated taxa-specific habitat trends at two spatial scales. At the neighborhood scale (n = 60), I found that removals occurred in yards with abundant cover opportunities. At the landscape scale (n = 764), nonvenomous snakes were removed from areas of higher urbanization compared to venomous snakes. Clients of Rattlesnake Solutions, LLC, were asked to answer a short survey, designed by K. Larson and colleagues, regarding the circumstances of their snake removal event and their attitudes, perceptions, and experiences with snakes. I used responses from this survey (n =271) to investigate if prior experience with snakes influences reported attitudes towards snakes. Respondents with prior snake experiences reported more positive attitudes towards snakes and were more consistent across their responses than those without prior snake experiences. Continuing inquiry into the urban ecology of these snakes is important to fostering coexistence between snakes and people that call Phoenix home.
ContributorsEnloe, Annika (Author) / Bateman, Heather L (Thesis advisor) / Lewis, Jesse (Committee member) / Larson, Kelli (Committee member) / Arizona State University (Publisher)
Created2023
Description
The learning journey toward sustainability requires the generation of action-oriented knowledge to understand the progress and potential adaptation of strategies to facilitate collective transformation. One integral area where advancing these strategies is essential is higher education. While efforts in the field have identified relevant learning objectives, pedagogies, and broader contributions for sustainability education to make, the sophistication of approaches to evaluating and enhancing collective learning has remained underdeveloped. This dissertation aimed to address this gap by exploring assessment in sustainability education, with the goal of supporting innovations in transdisciplinary practice by developing distinctive approaches for the field through deliberative processes that articulate design frameworks.The first study, Sustainability-Oriented Assessment, applied a formative intervention within an undergraduate course focused on professional skill development to demonstrate how a student-led rubric co-design process that drew from participative, normative, and integrative approaches contributed to student agency, expansive learning, and self-formation. The study demonstrated how students leveraged challenges and changing perspectives towards development as students and sustainability professionals. The second study, Boundary Crossings and Innovations, applied an exploratory action research approach within a community of practice of program leaders and instructors to identify barriers and possibilities for assessment practices, suggesting conceptual, practical, and logistical innovations. The strategies to enable these innovations focused on defining the field and articulating practice characteristics related to interdisciplinary approaches and transdisciplinary strategies, while meeting demands and facilitating innovations across course, program, and institutional levels. The third study, Assessment-Oriented Sustainability, applied a scoping literature review to develop a framework that integrates problem, solution, and learning orientations to sustainability by infusing educational purposes, processes, and principles to strengthen the efficacy of assessment approaches for transdisciplinary strategies.
Insights from the three studies suggest that the practice of assessment plays multifaceted roles as it facilitates the design, implementation, and evaluation of sustainability endeavors. These roles include evaluative and formative functions as it appraises and advances learning, normative and deliberative functions in encouraging discussion and critical reflection, and adaptive and generative functions by indicating, motivating, and enabling dynamic learning and action for sustainability.
ContributorsKing, Jordan (Author) / Fischer, Daniel (Thesis advisor) / Larson, Kelli (Thesis advisor) / Brundiers, Katja (Committee member) / Arizona State University (Publisher)
Created2024
Description
APOE encodes for a lipid transport protein and has three allelic variants-APOE ε2, ε3 and ε4 each of which differentially modulate the risk for Alzheimer’s disease (AD). The presence of the ε4 allele of APOE greatly increases AD risk compared to the presence of the more prevalent and risk neutral ε3 allele. An imbalance in the generation and clearance of amyloid beta (Aβ) peptides has been hypothesized to play a key role in driving the disease. APOE4 impacts several AD-relevant cellular processes. However, it is unclear whether these effects represent a gain of toxic function or a loss of function, specifically as it relates to modulating amyloid beta (Aβ) levels. Here, a set of APOE knockout (KO) and APOE4 isogenic human induced pluripotent stem cells (hiPSCs) were generated from a parental APOE3 hiPSC line with a highly penetrant familial AD (fAD) mutation to investigate this with respect to Aβ secretion in neural cultures and Aβ uptake in monocultures of microglia-like cells (iMGLs). Conversion of APOE3 to E4 as well as functionally knocking APOE out from the APOE3 parental line, result in elevated Aβ levels in neural cultures, likely through multiple mechanisms including the altered processing of the precursor protein to Aβ called amyloid precursor protein (APP). In pure neuronal cultures, a shift in the processing of APP was observed with the Aβ-generating amyloidogenic pathway being favored in both APOE3 as well as APOE4 neurons compared to APOE KO neurons, with APOE4 neurons exhibiting a greater shift. In iMGLs derived from the isogenic hiPSC lines, expression of APOE, regardless of the isoform, lowered the uptake of Aβ. Overall, APOE4 modulates Aβ levels through distinct loss of protective and gain of function effects. Dissecting these effects would contribute towards a better understanding of the design of potential APOE-targeted therapeutics in the future.
ContributorsRajaram Srinivasan, Gayathri (Author) / Brafman, David (Thesis advisor) / Plaisier, Christopher (Committee member) / Newbern, Jason (Committee member) / Stabenfeldt, Sarah (Committee member) / Wang, Xiao (Committee member) / Arizona State University (Publisher)
Created2024
Description
For the last 10 years, the American Southwest has been experiencing the most persistent drought conditions on record. Based on future climactic predictions, there is a dire need to reduce water usage within Phoenix. An environmentally responsible behavior such as low water use landscaping (xeriscaping), has been shown to reduce household water consumption by 40%-70%. While much is known regarding the relationship between socio-demographics and xeriscaping choices, the influence of other variables remains to be explored. Using data from the 2017 Phoenix Area Social Survey, this study investigates the influence of two additional variables - ecological worldview and place identity on xeriscaping choice. Data was analyzed using two models - Ordinary Least Squares (OLS) and Linear Probability Model (LPM). Ecological worldview and place identity, along with income, ethnicity, and gender, were all found to be positively related to xeriscape preference. Additionally, when compared to the LPM, the traditional OLS was found to still be the most robust and appropriate model when measuring landscape preference. Finally, results suggested that programs to foster identity with the local desert mountain parks may help to increase xeriscaping in the Valley and thus lower residential water use.
ContributorsSampson, Marena (Author) / Budruk, Megha (Thesis advisor) / Larson, Kelli (Committee member) / Gall, Melanie (Committee member) / Arizona State University (Publisher)
Created2018
Description
Urban planning in the neoliberal era is marred by a lack of public engagement with urban inhabitants. Henri Lefebvre’s ‘right to the city’ theory is often treated as a way to empower disenfranchised urban inhabitants who are lacking control over the urban spaces they occupy. Though the right to the city has seen a resurgence in recent literature, we still lack a deep understanding of how right to the city movements work in practice, and what the process looks like through the lens of the everyday urban inhabitant. This dissertation seeks to fill these gaps by examining: 1) how a minority-led grassroots movement activates their right to the city in the face of an incoming light rail extension project in South Phoenix, Arizona, USA, and 2) how their right to the city movement demonstrates the possibility of urban society beyond the current control of neoliberalism. Through the use of participant observation, interviews, and media analysis, this case reveals the methods and tactics used by the group to activate their right to the city, the intra-and inter-group dynamics in the case, and the challenges that ultimately lead to the group’s demise.Tactics used by the group included protesting, organizing against city council, and creating a ballot initiative. Intra-group dynamics were often marred by conflicts over leadership and the acceptance of outside help, while inter-group conflicts erupted between the group, politicians, and pro-light rail supporters. The primary challenge to the group’s right to the city movement included neoliberal appropriation by local politicians and outside political group. By possessing limited experience, knowledge, and resources in conducting a right to the city movement, the grassroots group in this case was left asking for help from neoliberal supporters who used their funding as a way to appropriate the urban inhabitant’s movement. Findings indicate positive possibilities of a future urban society outside of neoliberalism through autogestion, and provide areas where urban planners can improve upon the right to the city. If urban planners seek out and nurture instances of the right to the city, urban inhabitants will have greater control over planning projects that effect their neighborhoods.
ContributorsTziganuk, Ashlee (Author) / Pfeiffer, Deirdre (Thesis advisor) / Larson, Kelli (Thesis advisor) / Ehlenz, Meagan (Committee member) / McHugh, Kevin (Committee member) / Arizona State University (Publisher)
Created2019
Description
Synthetic manipulation of chromatin dynamics has applications for medicine, agriculture, and biotechnology. However, progress in this area requires the identification of design rules for engineering chromatin systems. In this thesis, I discuss research that has elucidated the intrinsic properties of histone binding proteins (HBP), and apply this knowledge to engineer novel chromatin binding effectors. Results from the experiments described herein demonstrate that the histone binding domain from chromobox protein homolog 8 (CBX8) is portable and can be customized to alter its endogenous function. First, I developed an assay to identify engineered fusion proteins that bind histone post translational modifications (PTMs) in vitro and regulate genes near the same histone PTMs in living cells. This assay will be useful for assaying the function of synthetic histone PTM-binding actuators and probes. Next, I investigated the activity of a novel, dual histone PTM binding domain regulator called Pc2TF. I characterized Pc2TF in vitro and in cells and show it has enhanced binding and transcriptional activation compared to a single binding domain fusion called Polycomb Transcription Factor (PcTF). These results indicate that valency can be used to tune the activity of synthetic histone-binding transcriptional regulators. Then, I report the delivery of PcTF fused to a cell penetrating peptide (CPP) TAT, called CP-PcTF. I treated 2D U-2 OS bone cancer cells with CP-PcTF, followed by RNA sequencing to identify genes regulated by CP-PcTF. I also showed that 3D spheroids treated with CP-PcTF show delayed growth. This preliminary work demonstrated that an epigenetic effector fused to a CPP can enable entry and regulation of genes in U-2 OS cells through DNA independent interactions. Finally, I described and validated a new screening method that combines the versatility of in vitro transcription and translation (IVTT) expressed protein coupled with the histone tail microarrays. Using Pc2TF as an example, I demonstrated that this assay is capable of determining binding and specificity of a synthetic HBP. I conclude by outlining future work toward engineering HBPs using techniques such as directed evolution and rational design. In conclusion, this work outlines a foundation to engineer and deliver synthetic chromatin effectors.
ContributorsTekel, Stefan (Author) / Haynes, Karmella (Thesis advisor) / Mills, Jeremy (Committee member) / Caplan, Michael (Committee member) / Brafman, David (Committee member) / Arizona State University (Publisher)
Created2019
Description
Neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease, or amyotrophic lateral sclerosis are defined by the loss of several types of neurons and glial cells within the central nervous system (CNS). Combatting these diseases requires a robust population of relevant cell types that can be employed in cell therapies, drug screening, or patient specific disease modeling. Human induced pluripotent stem cells (hiPSC)-derived neural progenitor cells (hNPCs) have the ability to self-renew indefinitely and differentiate into the various neuronal and glial cell types of the CNS. In order to realize the potential of hNPCs, it is necessary to develop a xeno-free scalable platform for effective expansion and differentiation. Previous work in the Brafman lab led to the engineering of a chemically defined substrate—vitronectin derived peptide (VDP), which allows for the long-term expansion and differentiation of hNPCs. In this work, we use this substrate as the basis for a microcarrier (MC)-based suspension culture system. Several independently derived hNPC lines were cultured on MCs for multiple passages as well as efficiently differentiated to neurons. Finally, this MC-based system was used in conjunction with a low shear rotating wall vessel (RWV) bioreactor for the integrated, large-scale expansion and neuronal differentiation of hNPCs. Finally, VDP was shown to support the differentiation of hNPCs into functional astrocytes. Overall, this fully defined and scalable biomanufacturing system will facilitate the generation of hNPCs and their derivatives in quantities necessary for basic and translational applications.
ContributorsMorgan, Daylin (Author) / Brafman, David (Thesis advisor) / Stabenfeldt, Sarah (Committee member) / Wang, Xiao (Committee member) / Arizona State University (Publisher)
Created2018
Description
Calcium imaging is a well-established, non-invasive or minimally technique designed to study the electrical signaling neurons. Calcium regulates the release of gliotransmitters in astrocytes. Analyzing astrocytic calcium transients can provide significant insights into mechanisms such as neuroplasticity and neural signal modulation.
In the past decade, numerous methods have been developed to analyze in-vivo calcium imaging data that involves complex techniques such as overlapping signals segregation and motion artifact correction. The hypothesis used to detect calcium signal is the spatiotemporal sparsity of calcium signal, and these methods are unable to identify the passive cells that are not actively firing during the time frame in the video. Statistics regarding the percentage of cells in each frame of view can be critical for the analysis of calcium imaging data for human induced pluripotent stem cells derived neurons and astrocytes.
The objective of this research is to develop a simple and efficient semi-automated pipeline for analysis of in-vitro calcium imaging data. The region of interest (ROI) based image segmentation is used to extract the data regarding intensity fluctuation caused by calcium concentration changes in each cell. It is achieved by using two approaches: basic image segmentation approach and a machine learning approach. The intensity data is evaluated using a custom-made MATLAB that generates statistical information and graphical representation of the number of spiking cells in each field of view, the number of spikes per cell and spike height.
In the past decade, numerous methods have been developed to analyze in-vivo calcium imaging data that involves complex techniques such as overlapping signals segregation and motion artifact correction. The hypothesis used to detect calcium signal is the spatiotemporal sparsity of calcium signal, and these methods are unable to identify the passive cells that are not actively firing during the time frame in the video. Statistics regarding the percentage of cells in each frame of view can be critical for the analysis of calcium imaging data for human induced pluripotent stem cells derived neurons and astrocytes.
The objective of this research is to develop a simple and efficient semi-automated pipeline for analysis of in-vitro calcium imaging data. The region of interest (ROI) based image segmentation is used to extract the data regarding intensity fluctuation caused by calcium concentration changes in each cell. It is achieved by using two approaches: basic image segmentation approach and a machine learning approach. The intensity data is evaluated using a custom-made MATLAB that generates statistical information and graphical representation of the number of spiking cells in each field of view, the number of spikes per cell and spike height.
ContributorsBhandarkar, Siddhi Umesh (Author) / Brafman, David (Thesis advisor) / Stabenfeldt, Sarah (Committee member) / Tian, Xiaojun (Committee member) / Arizona State University (Publisher)
Created2019
Description
Several debilitating neurological disorders, such as Alzheimer's disease, stroke, and spinal cord injury, are characterized by the damage or loss of neuronal cell types in the central nervous system (CNS). Human neural progenitor cells (hNPCs) derived from human pluripotent stem cells (hPSCs) can proliferate extensively and differentiate into the various neuronal subtypes and supporting cells that comprise the CNS. As such, hNPCs have tremendous potential for disease modeling, drug screening, and regenerative medicine applications. However, the use hNPCs for the study and treatment of neurological diseases requires the development of defined, robust, and scalable methods for their expansion and neuronal differentiation. To that end a rational design process was used to develop a vitronectin-derived peptide (VDP)-based substrate to support the growth and neuronal differentiation of hNPCs in conventional two-dimensional (2-D) culture and large-scale microcarrier (MC)-based suspension culture. Compared to hNPCs cultured on ECMP-based substrates, hNPCs grown on VDP-coated surfaces displayed similar morphologies, growth rates, and high expression levels of hNPC multipotency markers. Furthermore, VDP surfaces supported the directed differentiation of hNPCs to neurons at similar levels to cells differentiated on ECMP substrates. Here it has been demonstrated that VDP is a robust growth and differentiation matrix, as demonstrated by its ability to support the expansions and neuronal differentiation of hNPCs derived from three hESC (H9, HUES9, and HSF4) and one hiPSC (RiPSC) cell lines. Finally, it has been shown that VDP allows for the expansion or neuronal differentiation of hNPCs to quantities (>1010) necessary for drug screening or regenerative medicine purposes. In the future, the use of VDP as a defined culture substrate will significantly advance the clinical application of hNPCs and their derivatives as it will enable the large-scale expansion and neuronal differentiation of hNPCs in quantities necessary for disease modeling, drug screening, and regenerative medicine applications.
ContributorsVarun, Divya (Author) / Brafman, David (Thesis advisor) / Nikkhah, Mehdi (Committee member) / Stabenfeldt, Sarah (Committee member) / Arizona State University (Publisher)
Created2016