Matching Items (19)
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- All Subjects: Traumatic Brain Injury
- All Subjects: Bioengineering
- Creators: Stabenfeldt, Sarah
- Resource Type: Text
Description
Modern medical conditions, including cancer, traumatic brain injury, and cardiovascular disease, have elicited the need for cell therapies. The ability to non-invasively track cells in vivo in order to evaluate these therapies and explore cell dynamics is necessary. Magnetic Resonance Imaging provides a platform to track cells as a non-invasive modality with superior resolution and soft tissue contrast. A new methodology for cellular labeling and imaging uses Nile Red doped hexamethyldisiloxane (HMDSO) nanoemulsions as dual modality (Magnetic Resonance Imaging/Fluorescence), dual-functional (oximetry/ detection) nanoprobes. While Gadolinium chelates and super paramagnetic iron oxide-based particles have historically provided contrast enhancement in MRI, newer agents offer additional advantages. A technique using 1H MRI in conjunction with an oxygen reporter molecule is one tool capable of providing these benefits, and can be used in neural progenitor cell and cancer cell studies. Proton Imaging of Siloxanes to Map Tissue Oxygenation Levels (PISTOL) provides the ability to track the polydimethylsiloxane (PDMS) labeled cells utilizing the duality of the nanoemulsions. 1H MRI based labeling of neural stem cells and cancer cells was successfully demonstrated. Additionally, fluorescence labeling of the nanoprobes provided validation of the MRI data and could prove useful for quick in vivo verification and ex vivo validation for future studies.
ContributorsCusick, Alex (Author) / Kodibagkar, Vikram D. (Thesis advisor) / Stabenfeldt, Sarah (Committee member) / Kleim, Jeff (Committee member) / Arizona State University (Publisher)
Created2014
Description
Approximately 2.8 million Americans seek medical care for traumatic brain injury (TBI) each year. Of this population, the majority are sufferers of diffuse TBI, or concussion. It is unknown how many more individuals decline to seek medical care following mild TBI. This likely sizeable population of un- or self-treated individuals combined with a lack of definitive biomarkers or objective post-injury diagnostics creates a unique need for practical therapies among diffuse TBI sufferers. Practical therapies stand to decrease the burden of TBI among those who would otherwise not seek treatment or do not meet clinical diagnostic criteria upon examination. For this unique treatment niche, practical therapies for TBI are defined as having one or more of the following qualities: common availability, easy administration, excellent safety profile, and cost-effectiveness. This dissertation identifies and critically examines the efficacy of four classes of practical treatments in improving rodent outcome from experimental diffuse traumatic brain injury.
Over-the-counter (OTC) analgesics, omega-3 fatty acids, specialized pro-resolving mediators (SPMs), and remote ischemic conditioning (RIC) were administered before or following midline fluid percussion injury. Behavioral, histological, and molecular analyses were used to assess treatment effects on functional outcome and secondary injury progression. Acute administration of common OTC analgesics had little effect on post-injury outcome in mice. Dietary supplementation with omega-3 fatty acid docosahexaenoic acid (DHA) prior to or following diffuse TBI significantly reduced injury-induced sensory sensitivity and markers of neuroinflammation with no effect on spatial learning. Intraperitoneal administration of omega-3 fatty acid-derived SPM resolvin E1 significantly increased post-injury sleep and suppressed microglial activation. Aspirin-triggered (AT) resolvin D1 administration improved both motor and cognitive outcome following diffuse TBI. RIC treatment in mice demonstrated little effect on functional outcome from diffuse TBI. Untargeted proteomic analysis of plasma samples from RIC-treated mice was used to identify candidate molecular correlates of RIC. Identification of these candidates represents a vital first step in elucidating the neuroprotective mechanisms underlying RIC. The overall findings suggest that omega-3 fatty acid supplementation, SPM administration, and RIC may serve as effective practical therapies to reduce the somatic, cognitive, and neurological burden of diffuse TBI felt by millions of Americans.
Over-the-counter (OTC) analgesics, omega-3 fatty acids, specialized pro-resolving mediators (SPMs), and remote ischemic conditioning (RIC) were administered before or following midline fluid percussion injury. Behavioral, histological, and molecular analyses were used to assess treatment effects on functional outcome and secondary injury progression. Acute administration of common OTC analgesics had little effect on post-injury outcome in mice. Dietary supplementation with omega-3 fatty acid docosahexaenoic acid (DHA) prior to or following diffuse TBI significantly reduced injury-induced sensory sensitivity and markers of neuroinflammation with no effect on spatial learning. Intraperitoneal administration of omega-3 fatty acid-derived SPM resolvin E1 significantly increased post-injury sleep and suppressed microglial activation. Aspirin-triggered (AT) resolvin D1 administration improved both motor and cognitive outcome following diffuse TBI. RIC treatment in mice demonstrated little effect on functional outcome from diffuse TBI. Untargeted proteomic analysis of plasma samples from RIC-treated mice was used to identify candidate molecular correlates of RIC. Identification of these candidates represents a vital first step in elucidating the neuroprotective mechanisms underlying RIC. The overall findings suggest that omega-3 fatty acid supplementation, SPM administration, and RIC may serve as effective practical therapies to reduce the somatic, cognitive, and neurological burden of diffuse TBI felt by millions of Americans.
ContributorsHarrison, Jordan L (Author) / Lifshitz, Jonathan (Thesis advisor) / Neisewander, Janet (Thesis advisor) / Stabenfeldt, Sarah (Committee member) / Willyerd, Frederick A (Committee member) / Pirrotte, Patrick (Committee member) / Arizona State University (Publisher)
Created2017
Description
The research question explored in this thesis is how CRISPR mediated editing is influenced by artificially opened chromatin in cells. Closed chromatin poses a barrier to Cas9 binding and editing at target genes. Synthetic pioneer factors (PFs) are a promising new approach to artificially open condensed heterochromatin allowing greater access of target DNA to Cas9. The Haynes lab has constructed fusions of enzymatic chromatin-modifying domains designed to remodel chromatin and increase Cas9 editing efficiency. With a library of PFs available, this research focuses on analyzing the behavior of Cas9 in chromatin that has been artificially opened by PFs. The types and frequency of INDELs (insertions & deletions) were determined after non-homologous end joining (NHEJ) in PF and Cas9-treated cells using quantitative Sanger sequencing and Synthego’s ICE software. Furthermore, NOME-seq analysis was carried out to map nucleosome position in PF and Cas9 treated cells. Although this experiment was unsuccessful, the heat map generated with data obtained from Synthego ICE predicts a possible presence of nucleosome in the vicinity suggesting that perhaps a fully open chromatin state was not achieved. Linear Regression analysis with certain assumptions confirms that with the increase in distance downstream of cut-site, the editing frequency decreases exponentially. Nevertheless, further experimental work should be carried out to investigate this hypothesis.
ContributorsHamna, Syeda Fatima (Author) / Haynes, Karmella A (Thesis advisor) / Stabenfeldt, Sarah (Thesis advisor) / Tian, Xiaojun (Committee member) / Arizona State University (Publisher)
Created2019
Description
Monitoring complex diseases and their comorbidities requires accurate and convenient measurements of multiple biomarkers. However, many state-of-the-art bioassays not only require complicated and time-consuming procedures, but also measure only one biomarker at a time. This noncomprehensive single-biomarker monitoring, as well as the cost and complexity of these bioassays advocate for a simple, rapid multi-marker sensing platform suitable for point-of-care or self-monitoring settings. To address this need, diabetes mellitus was selected as the example complex disease, with dry eye disease and cardiovascular disease as the example comorbidities. Seven vital biomarkers from these diseases were selected to investigate the platform technology: lactoferrin (Lfn), immunoglobulin E (IgE), insulin, glucose, lactate, low density lipoprotein (LDL), and high density lipoprotein (HDL). Using electrochemical techniques such as amperometry and electrochemical impedance spectroscopy (EIS), various single- and dual-marker sensing prototypes were studied. First, by focusing on the imaginary impedance of EIS, an analytical algorithm for the determination of optimal frequency and signal deconvolution was first developed. This algorithm helped overcome the challenge of signal overlapping in EIS multi-marker sensors, while providing a means to study the optimal frequency of a biomarker. The algorithm was then applied to develop various single- and dual-marker prototypes by exploring different kinds of molecular recognition elements (MRE) while studying the optimal frequencies of various biomarkers with respect to their biological properties. Throughout the exploration, 5 single-marker biosensors (glucose, lactate, insulin, IgE, and Lfn) and one dual-marker (LDL and HDL) biosensor were successfully developed. With the aid of nanoparticles and the engineering design of experiments, the zeta potential, conductivity, and molecular weight of a biomarker were found to be three example factors that contribute to a biomarker’s optimal frequency. The study platforms used in the study did not achieve dual-enzymatic marker biosensors (glucose and lactate) due to signal contamination from localized accumulation of reduced electron mediators on self-assembled monolayer. However, amperometric biosensors for glucose and lactate with disposable test strips and integrated samplers were successfully developed as a back-up solution to the multi-marker sensing platform. This work has resulted in twelve publications, five patents, and one submitted manuscripts at the time of submission.
ContributorsLin, Chi En (Author) / La Belle, Jeffrey T (Thesis advisor) / Caplan, Michael (Committee member) / Cook, Curtiss B (Committee member) / Stabenfeldt, Sarah (Committee member) / Spano, Mark (Committee member) / Arizona State University (Publisher)
Created2018
Description
Traumatic brain injury (TBI) may result in numerous pathologies that cannot currently be mitigated by clinical interventions. Stem cell therapies are widely researched to address TBI-related pathologies with limited success in pre-clinical models due to limitations in transplant survival rates. To address this issue, the use of tissue engineered scaffolds as a delivery mechanism has been explored to improve survival and engraftment rates. Previous work with hyaluronic acid \u2014 laminin (HA-Lm) gels found high viability and engraftment rates of mouse fetal derived neural progenitor/stem cells (NPSCs) cultured on the gel. Furthermore, NPSCs exposed to the HA-Lm gels exhibit increased expression of CXCR4, a critical surface receptor that promotes cell migration. We hypothesized that culturing hNPCs on the HA-Lm gel would increase CXCR4 expression, and thus enhance their ability to migrate into sites of tissue damage. In order to test this hypothesis, we designed gel scaffolds with mechanical properties that were optimized to match that of the natural extracellular matrix. A live/dead assay showed that hNPCs preferred the gel with this optimized formulation, compared to a stiffer gel that was used in the CXCR4 expression experiment. We found that there may be increased CXCR4 expression of hNPCs plated on the HA-Lm gel after 24 hours, indicating that HA-Lm gels may provide a valuable scaffold to support viability and migration of hNPCs to the injury site. Future studies aimed at verifying increased CXCR4 expression of hNPCs cultured on HA-Lm gels are necessary to determine if HA-Lm gels can provide a beneficial scaffold for stem cell engraftment therapy for treating TBI.
ContributorsHemphill, Kathryn Elizabeth (Author) / Stabenfeldt, Sarah (Thesis director) / Brafman, David (Committee member) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2016-05
Description
One of the most prominent biological challenges for the field of drug delivery is the blood-brain barrier. This physiological system blocks the entry of or actively removes almost all small molecules into the central nervous system (CNS), including many drugs that could be used to treat diseases in the CNS. Previous studies have shown that activation of the adenosine receptor signaling pathway through the use of agonists has been demonstrated to increase BBB permeability. For example, regadenoson is an adenosine A2A receptor agonist that has been shown to disrupt the BBB and allow for increased drug uptake in the CNS. The goal of this study was to verify this property of regadenoson. We hypothesized that co-administration of regadenoson with a non-brain penetrant macromolecule would facilitate its entry into the central nervous system. To test this hypothesis, healthy mice were administered regadenoson or saline concomitantly with a fluorescent dextran solution. The brain tissue was either homogenized to measure quantity of fluorescent molecule, or cryosectioned for imaging with confocal fluorescence microscopy. These experiments did not identify any significant difference in the amount of fluorescence detected in the brain after regadenoson treatment. These results contradict those of previous studies and highlight potential differences in injection methodology, time windows, and properties of brain impermeant molecules.
ContributorsWohlleb, Gregory Michael (Author) / Sirianni, Rachael (Thesis director) / Stabenfeldt, Sarah (Committee member) / Barrett, The Honors College (Contributor) / Harrington Bioengineering Program (Contributor)
Created2015-05
Description
The endogenous response of neural stem cell/progenitor (NPSC) recruitment to the brain injury environment following a traumatic brain injury (TBI) is currently under heavy investigation. Mechanisms controlling NPSC proliferation and migration to the brain injury environment remain unclear; however, it is thought that the vascular extracellular matrix proteins (e.g. laminin, fibronectin, and vitronectin) and vascular endothelial growth factor (VEGF) play a role in mediating NPSC behavior through vasophillic interactions. This project attempts to uncover potential VEGF-ECM crosstalk in mediating migration and proliferation. To investigate migration, neurospheres were seeded on ECM-coated wells supplemented with VEGF and without VEGF, and neural outgrowth was measured at days 0, 1, 3, and 8 using differential interference contrast microscopy. Furthermore, single-cell NPSCs were seeded on ECM-coated Transwell membranes with VEGF supplemented media on one side and without VEGF to look at chemotactic migration. Migrated NPSCs were visualized with DAPI nuclear stain and imaged with an inverted fluorescent microscope. To investigate NPSC proliferation, NPSCs were seeded on ECM coated plates as in the radial migration assay and visualized with EdU on day 8. Total proliferation was measured by seeding NPSCs on ECM coated 96-well plates and incubating them with MTT on days 3 and 6. Proliferation was measured using a spectrophotometer at 630nm and 570nm wavelengths. It was found that VEGF-laminin crosstalk synergistically increased radial migration, but may not play a role in chemotactic migration. Understanding the mechanisms behind VEGF-laminin crosstalk in NPSC proliferation and migration may provide crucial information for the design of stem cell transplantation therapies in the future.
ContributorsMillar-Haskell, Catherine Susan (Author) / Stabenfeldt, Sarah (Thesis director) / Addington, Caroline (Committee member) / Barrett, The Honors College (Contributor) / Harrington Bioengineering Program (Contributor)
Created2015-05
Description
The main objective of this research is to develop and characterize a targeted contrast agent that will recognize acute neural injury pathology (i.e. fibrin) after traumatic brain injury (TBI). Single chain fragment variable antibodies (scFv) that bind specifically to fibrin have been produced and purified. DSPE-PEG micelles have been produced and the scFv has been conjugated to the surface of the micelles; this nanoparticle system will be used to overcome limitations in diagnosing TBI. The binding and imaging properties will be analyzed in the future to determine functionality of the nanoparticle system in vivo.
ContributorsRumbo, Kailey Michelle (Author) / Stabenfeldt, Sarah (Thesis director) / Kodibagkar, Vikram (Committee member) / Barrett, The Honors College (Contributor) / Harrington Bioengineering Program (Contributor)
Created2014-05
Description
Traumatic brain injury (TBI) can result in many pathologies, one of which being coagulopathy. TBI can progress to hemorrhagic lesions and increased intercranial pressure leading to coagulopathy. The coagulopathy has been linked to poor clinical outcomes and occurs in 60% of severe TBI cases. To improve hemostasis, synthetic platelets (SPs) have been repurposed. SPs are composed of a poly(N-isopropylacrylamide-co-acrylic-acid) microgel, conjugated with a fibrin-specific antibody and are biomimetic in their ability to deform and collapse within a fibrin matrix. The objective of this study is to diminish coagulopathy with a single, intravenous injection of SPs, and subsequently decrease neuropathologies. TBI was modeled in animal cohorts using the well-established controlled cortical impact and SPs were injected 2-3 hours post-injury. Control cohorts received no injection. Brain tissue was harvested at acute (24h) and delayed (7 days) time points post-TBI, and fluorescently imaged to quantify reactive astrocytes (GFAP+), microglial morphology and presence (Iba1+), and tissue lesion spared. SP-treatment resulted in significant reduction of GFAP expression at 7 days post-TBI. Furthermore, SP-treatment significantly reduced the percent difference from 24h to 7 days in microglia/macrophage per field compared to the control. For microglial morphology, SP-treated cohorts observed a significant percent difference in endpoints per soma from 24h to 7 days compared to untreated cohorts. However, microglial branch length significantly decreased in percent difference from 24h to 7 days when compared to the control. Finally, tissue sparing did not significantly decrease between 24h and 7 day for SP-treated cohorts as was observed in untreated cohorts, implying inhibition of delayed necrosis. Overall, these results suggest decreased neuroinflammation by 7 days, supporting SPs as potentially therapeutic post-TBI.
ContributorsTodd, Jordan Cecile (Author) / Stabenfeldt, Sarah (Thesis director) / Bharadwaj, Vimala (Committee member) / Harrington Bioengineering Program (Contributor, Contributor) / Barrett, The Honors College (Contributor)
Created2019-05
Description
Traumatic brain injury (TBI) is a serious health problem around the world with few available treatments. TBI pathology can be divided into two phases: the primary insult and the secondary injury. The primary insult results from the bump or blow to the head that causes the initial injury. Secondary injury lasts from hours to months after the initial injury and worsens the primary insult, creating a greater area of tissue damage and cell death. Many current treatments focus on lessening the severity of secondary injury. Secondary injury results from the cyclical nature of tissue damage. Inflammatory pathways cause damage to tissue, which in turn reinforces inflammation. Since many inflammatory pathways are interconnected, targeting individual products within these pathways is impractical. A target at the beginning of the pathway, such as a receptor, must be chosen to break the cycle. This project aims to identify novel nanobodies that could temporarily inactivate the CD36 receptor, which is a receptor found on many immune and endothelial cells. CD36 initiates and perpetuates the immune system's inflammatory responses. By inactivating this receptor temporarily, inflammation and immune cell entry could be lessened, and therefore secondary injury could be attenuated. This project utilized phage display as a method of nanobody selection. The specific phage library utilized in this experiment consists of human heavy chain (V_H) segments, also known as domain antibodies (dAbs), displayed on M13 filamentous bacteriophage. Phage display mimics the process of immune selection. The target is bound to a well as a means of displaying it to the phage. The phage library is then incubated with the target to allow antibodies to bind. After, the well is washed thoroughly to detach any phage that are not strongly bound. The remaining phage are then amplified in bacteria and run again through the same assay to select for mutations that resulted in higher affinity binding. This process, called biopanning, was performed three times for this project. After biopanning, the library was sequenced using Next Generation sequencing (NGS). This platform enables the entire library to be sequenced, as opposed to traditional Sanger sequencing, which can only sequence single select clones at a time thereby limiting population sampling. This type of genetic sequencing allows trends in the complementarity determining regions (CDRs) of the domain antibody library to be analyzed, using bioinformatics programs such as RStudio, FastAptamer, and Swiss Model. Ultimately, two nanobody candidates were identified for the CD36 receptor.
ContributorsLundgreen, Kendall (Author) / Stabenfeldt, Sarah (Thesis director) / Ugarova, Tatiana (Committee member) / School of Life Sciences (Contributor) / School of International Letters and Cultures (Contributor) / Barrett, The Honors College (Contributor)
Created2018-05