22689 research outputs found
Sort by
NUCLEIC ACID-BASED ELECTROCHEMICAL SENSORS: DISCOVERING THEIR SIGNAL DECAY MECHANISMS AND EXPANDING THE LIBRARY OF MATERIALS FOR THEIR FABRICATION
Get PDFNucleic acid-based electrochemical (NBE) sensors are a rapidly advancing technology in both academic and commercial spheres of interest. These sensors, which rely on affinity of nucleic acids for specific targets, are capable of detecting, and often quantifying, nearly any molecule of interest in unprocessed biological fluids. As such, they are ideal candidates for deployment in human health applications. For example, NBEs could be interfaced with bandages or prosthetic implants to monitor inflammation and healing. They could also be placed in veins to continuously track and modulate the concentration of therapeutic drugs. However, NBEs are currently limited in lifespan to about 6-12 hours when deployed in the bodies of living animals. If the technology is to be translated for human use – where drug excretion and protein expression occur over days, not hours – then this lifespan must be greatly extended. Additionally, NBEs are most often fabricated via self-assembly of alkanethiols and alkanethiol-modified nucleic acids onto a gold electrode surface. However, this attachment chemistry and electrode material are not compatible with all possible NBE sensor applications. Therefore, there is a need to discover the mechanisms of sensor signal loss over time, design solutions to circumvent those mechanisms, and expand the number of materials that can support NBE sensors. In this work, through modifying the surface functionality of alkanethiol monolayers and the chirality of attached nucleic acids, I discover that an underlying mechanism for signal loss of thiol-on-gold NBEs is electrochemically induced desorption of alkanethiols. Specifically, by increasing the hydrophobicity of the alkanethiols, they remain on the electrode surface for longer, thereby extending the lifetime in buffered solutions. By inverting the chirality of attached nucleic acids, they become resistant to nuclease cleavage, thereby extending the lifetime in biological fluids. Equipped with this knowledge of the limitations of thiol monolayers on gold, I further explore two alternative options for the fabrication of NBE sensors: alkylphosphonic acid monolayers on indium tin oxide and conductive polymers on platinum electrodes. By providing extensive detail on the modification of such materials, I generate an experimental framework to guide future development of NBEs for a variety of potential applications
RECONSTRUCTING BLOOD-RETINAL BARRIER FUNCTION IN RETINOPATHY
No full textDiabetic retinopathy (DR) is a microvascular complication found in patients with diabetes mellitus. Inner blood retinal barrier (iBRB) breakdown, which leads to a decreased blood supply to the retina, is a hallmark of DR. However, the pathogenesis of DR and the molecular mechanisms that govern iBRB breakdown remain unclear. In this thesis, our emphasis is on unraveling two pivotal events contributing to iBRB breakdown: (1) the disruption in endothelial-pericyte interactions and (2) endothelial cell dysfunction.
First, we utilized human induced pluripotent stem cells (hiPSCs) to generate isogenic endothelial cells (iECs) and pericytes (iPericytes), aiming to investigate the loss of cellular communication between these two cell types in DR. We identified VEGFR2 pY951 as a crucial signaling pathway in pericyte-modulated vascular stabilization. Notably, we found that direct endothelial-pericyte contact is essential for downregulating VEGR2 pY951. The inhibition of VEGFR2 pY951 enhances iPericyte recruitment to iECs and 3D vascular networks. We observed increased pericyte engagement, enhanced vascularization, and tissue growth in the retina of both mouse model of oxygen-induced retinopathy and the developing healthy mouse retina. Next, we demonstrated that by modulating Norrin signaling pathways we could obtain retinal-specific endothelial cells (iRECs) and pericytes from hiPSCs. These hiPSC-derived cells exhibited phenotypic and functional similarities to human retina endothelial cells and pericytes
Establishing the prevalence of last-minute submissions and measuring their impacts on research administrators’ pre-award support and review functions, personal workplace experiences, and workplace relationships and stress
Get PDFPracticing research administrators have consistently voiced concerns about the prevalence and wide-ranging negative impacts of last-minute submissions on their workplace functions, experiences, and relationships; however, there is little empirical evidence to support these practical claims as little more than one-off experiences. The purpose of the current study was to provide statistical evidence to support the claim that last-minute submissions are a profession-wide problem using quantitative data analysis to inform professional practice. The researcher sought to answer the following research questions through completion of the study: 1) How often do research administrators encounter last-minute submissions during internal proposal review?; 2) Do last-minute submissions negatively impact research administrators’ pre-award support functions, pre-award review functions, personal workplace experiences, workplace relationships, and workplace stress more often than they do not?; and 3) Are last-minute submissions a profession-wide problem? The researcher conducted a quantitative, non-interventional research study using a cross-sectional survey research design to measure the impact of six independent variables on 26 dependent variables. The researcher-developed questionnaire was comprised of an information form, multiple choice questions (some with text entry options), Likert scales, open-ended text entry questions, and demographic questions. The survey was estimated to take approximately 15-30 minutes to complete, and participants were asked to complete the survey only once. A total of 5 items (dependent variables) out of 26 were found not to be statistically significantly impacted by last-minute submissions. To the researcher’s surprise, there was significant statistical evidence to conclude that research administrators are more likely to encounter on-time submissions than last-minute submissions; however, when last-minute submissions do occur, there is significant statistical evidence to conclude that last-minute submissions do negatively impact research administrators’ workplace functions, experiences, and relationships more often than they positively or neutrally impact them. The researcher also found significant statistical evidence to conclude that last-minute submissions are very/extremely stressful more often than they are moderately stressful (slightly/somewhat). Future research should compare the types of internal submission deadlines and their effectiveness in reducing or mitigating the negative impacts of last-minute submissions
THE H3K4 METHYLTRANSFERASE SET DOMAIN CONTAINING 1 PROMOTES MAINTENANCE AND DIFFERENTIATION OF ADULT GERMLINE STEM CELLS IN THE DROSOPHILA TESTIS
Get PDFMany short-lived cell types in the body are produced from adult stem cell precursors, either continuously or in response to physiological signals or trauma. Drosophila spermatogenesis is a well characterized model system for the study of mechanisms regulating the maintenance and proliferation of stem cells, as well as proper differentiation of precursor cells. In fly testes, two stem cell lineages derived from germline stem cells (GSCs) and cyst stem cells (CySCs) share the same microenvironment and stay in close communication throughout their cellular differentiation pathways. During these processes, the occurrence of genetic lesions or epigenetic mis-regulation may induce a shift in their cellular homeostasis to become ever-dividing cells and block further differentiation, leading to a number of diseases such as cancers.
Polycomb group (PcG) and Trithorax group (TrxG) complexes play critical roles in the cell decision between maintaining proliferating precursor status and initiating the terminal differentiation program. PcG and TrxG complexes are also known to have antagonizing molecular functions in regulating chromatin structure and gene expression. In my thesis work, I studied the biological functions of components in both PcG and TrxG in the early-stage germline including GSCs. Through a series of cell-type and stage-specific Set1 RNAi knockdown experiments in a time course, my results demonstrated a temporal progression of the germline defects, starting from severe germ cell loss to overpopulated early-stage germ cells. This later appearance of early-stage germline overproliferation phenotype suggests an intriguing possibility that a small subset of “survivor cells” that escape earlier cell death could initiate an uncontrolled and fast over-proliferation phase, mimicking a “cancer stem cell” phenomenon. These germ cell defects also result in defective stem cell niche architecture and overpopulated cyst cells, likely through non-cell-autonomous mechanisms. Further, I determined the inception of these germline phenotypes was dependent on the loss of Set1’s methyltransferase activity in early-stage germ cells. Compromising Set1 in the late-stage germ cells or somatic gonadal cells failed to reproduce these defects. In addition, I conducted experiments where a transgene encoding either the wild-type or catalytic inactive forms of Set1 was driven in the Set1 knockdown background. Only the wild-type Set1 was able to rescue these phenotypes. To understand the molecular mechanisms, I performed both RNA-seq to identify gene expression changes upon knocking down the set1 gene. Through this genomic assay, I identified key signaling pathways components, such as Stat92E and Mad, downstream factors of the JAK-STAT and BMP signaling pathways respectively, have upregulated expression when the set1 gene is knocked down. I then performed genetic interaction analyses to further confirm the functional relationship between set1 and these signaling pathways. I found that strong loss of function mutations of stat92E and mad genes significantly suppress the set1 knockdown early germ cell overproliferation phenotype. Together, my study will enhance our understanding of the decision between proliferation vs. differentiation in stem cell lineages, which will have significant impact on stem cell biology and cancer biology. Finally, my finding of this initial cell loss followed by cell overproliferation when knocking down a key histone methyl-transferase raises the concern using inhibitors of these enzymes in cancer therapy
Electron diffraction-based studies on the effects of complexity and order on the microstructural plastic response of multi-principal element alloys
Get PDFMulti-principal element alloys (MPEAs) have become one of the most intensely researched subjects in the contemporary study of metallic materials. These alloys are generally categorized as being comprised of three to five (sometimes more) primary elemental components at or near equiatomic composition. Certain MPEAs are known as medium and high entropy alloys (MEA or HEA) and exhibit a nominally disordered single-phase microstructure. A combination of chemical complexity and relatively simple and well-understood crystal structures such as FCC make the study of M/HEAs interesting for two reasons. First is that these alloys show promising physical properties such as excellent mechanical behavior. They
also provide researchers to better understand fundamental phenomena of materials science.
The research conducted for this dissertation seeks to assess the mechanical response of alloys of varying complexity at multiple length and time scales. First, the effect of strain rate on microstructural development at the grain scale is assessed in chemically pure copper. This provides us with a baseline before moving into more complex ternary and quinary systems in an effort to determine how chemical complexity leads to variation strain-rate dependent behavior. Dynamic and shock testing on the Cantor alloy (equiatomic CoCrFeMnNi) and CoCrNi highlights the role that chemical disorder can play in modulating the mechanical response compared to conventional alloys. Finally, this research develops a technique for the quantitative mapping of ordering behavior in aged CoCrNi and assess how such ordering can affect the alloy’s microstructural development upon deformation. The experimental work relies heavily on microstructural characterization techniques via electron microscopy, especially advanced electron diffraction techniques. These include well established methods for to measure plastic strain evolution and deformation microstructures at both the micro- and nanoscales, as well as newly developed techniques to probe the reciprocal lattice of ordered and disordered lattices in MPEAs. This dissertation concludes that chemical complexity, in terms of both nominal disorder and measurable localized ordering, strongly modulates the deformation response of metallic alloys. Mechanical behavior varies both as a function of the novel pre-deformation microstructures in MPEAs, as well as the anomalous behavior of dislocations and substructure formation throughout the deformation process
DIABETES, VASCULAR RISK FACTORS AND LONG-TERM RISK OF DEMENTIA
No full textThis dissertation examined the association between diabetes and vascular risk factors with the lifetime risk of dementia, elucidating the nuanced impacts of disease onset and management using the Atherosclerosis Risk in Communities (ARIC) study population. This dissertation delved into the role of clinical progression to diabetes in the association of prediabetes and dementia incidence. We also explored the significance of age at diabetes onset, and the cumulative effect of multiple vascular risk factors on long-term risk of dementia cross the life course, accounting for mortality as a competing outcome.
The findings in chapter 1 reveal that while prediabetes was modestly associated with dementia risk, it is the progression to diabetes that drives this association, with early onset of diabetes (before 60 years) substantially amplifying the risk. This chapter provided a fuller picture of the association and suggest that prediabetes is not a robust risk factor for dementia in the absence of a subsequent diagnosis of diabetes. In chapter 2, we documented that early diabetes onset (before age 60) was strongly associated with an increased lifetime risk of dementia despite of greater mortality risk as well as a ~4 years shorter survival free from dementia, comparing to individual without diabetes. Meanwhile, late onset diabetes results in a lower dementia lifetime risk (unadjusted: 25.9%; adjusted: 22.1%) since it elevates mortality risk far more than its effect on dementia. In chapter 3, we explored the double-edged sword of optimal vascular health, which, although delays dementia onset by ~7 years, paradoxically increases the absolute lifetime risk due to extended life expectancy.
The synthesized evidence from these three studies underscores the importance of early detection and management of prediabetes, delaying diabetes onset, optimization of vascular health as pivotal strategies for dementia risk mitigation and delay. Moreover, the insights into the competing risks highlight an essential public health message: the necessity for a balanced approach that extends healthy life years while also preparing for the implications of an aging population with an increased burden of dementia.
In summary, this dissertation work provides valuable contributions to the understanding of prediabetes progression, diabetes onset and the cumulative effect of vascular risk factors on dementia risk, both from relative risk and life-course absolute risk. Additionally, this work has profound implications for public health policies aimed at prevention strategies on the escalating dementia epidemic
CRISPR-Cas9 as Precision Gene Therapy in Pancreatic Cancer
Get PDFMetastatic cancer affects millions of people worldwide annually and is the leading cause of cancer-related deaths. Most patients with metastatic disease are not eligible for surgical resection, and current therapeutic regimens have varying success rates, some with 5-year survival rates below 5%. Here, using pancreatic ductal adenocarcinoma (PDAC) as a model system, I test the hypothesis that metastatic cancer can be genetically targeted with CRISPR-Cas9.
In order to develop a genetic treatment for metastatic cancer, it is important to identify targets that are present in every cancer cell in the patient. I studied five patients with PDAC who underwent a rapid autopsy to determine areas of evolutionary conservation that were maintained throughout all metastases in each patient’s cancer. Using whole genome sequencing validated by high-depth capture sequencing, I found that 90% of somatic CRISPR targets were maintained between primary carcinomas and metastases. I identified regions of loss of heterozygosity (LOH) surrounding tumor suppressor genes and oncogenes in the primary tumor where targets were 100% conserved in metastases. These regions of truncal LOH represent genetic vulnerabilities in all primary and metastatic lesions of these carcinomas.
Using multitarget sgRNA which cut at a known number of sites in the human genome, I determined a relationship between the number of CRISPR-Cas9
targets and growth inhibition in PDAC cells. I found that cutting at 8-10 target sites in non-coding regions was sufficient to induce >95% cancer cell death. Furthermore, only 2-3 targets were required to achieve a comparable level of cell death in the presence DNA double strand break repair inhibition.
Finally, using cocultures of PDAC cells transduced with sgRNA targeting multiple tumor specific loci, I demonstrated selective cell killing of three different PDAC cell lines. Importantly, deep sequencing of patient-matched normal lymphocytes lacking the tumor-specific targets transduced with cas9 and the same set of sgRNA array exhibited no editing at the targeted loci.
In this thesis I have identified the pattern of CRISPR target maintenance between primary tumors and metastases, established a relationship between DNA double strand breaks and cancer cytotoxicity and demonstrated selective elimination of targeted cancer cells in mixed cell populations. This work establishes a proof of principle for a CRISPR-cas9 based gene therapy for metastatic cancer
A Tale of Two Terrestrial Dynamos
No full textMagnetic fields originating from planetary interiors are pivotal for comprehending the dynamics, extreme conditions, and complex chemistry of these regions and their extensive planetary implications. This research introduces innovative techniques for the enhanced observation and characterization of such magnetic fields, focusing on Earth and Mercury, the only terrestrial planets in our solar system with an active core dynamo. The objective is to deepen our understanding of the structure, properties, and dynamics governing these regions.
Magnetic fields, spanning various spatial and temporal scales, arise from multiple current sources, complicating the isolation and characterization of the dynamo field from other source fields. This complexity is due to the superposition of fields in magnetic observations and our inability to monitor the total field ubiquitously and segregate the contributions accurately. This study leverages magnetic data from the Iridium Communications constellation network of satellites to characterize previously inaccessible spatio-temporal scales in Earth's core field. Despite the lower quality of body-mounted magnetometers, the selected Iridium data, under stringent geomagnetically quiet conditions, can map the global magnetic field with uncertainties as low as 3 nT, aligning well with state-of-the-art models.
The data reveal abrupt geomagnetic changes, or geomagnetic jerks, and a large-scale, wave pattern with a period of ~14 months, potentially originating from a wave in Earth’s core, with implications for Earth's deep interior. Additionally, variations on smaller scales, not of external origin, are detected, exhibiting strong geographic coherence, supporting an internal origin and revealing potential heterogeneities in Earth's mantle.
For Mercury, where external fields are prominent, high-fidelity physics-based, magnetohydrodynamic simulations, specifically GAMERA, are adapted to simulate the solar-wind magnetosphere interaction, including the Birkeland currents. This allows quantification and mapping of Birkeland current signals, enabling the isolation of the planet's core field to better precision from MESSENGER observations.
In conclusion, this study, using commercial satellite constellation systems and advanced simulations, pioneers new approaches in unveiling intricate dynamics of planetary magnetic fields in Earth and Mercury. The methodologies employed open avenues for future research, offering insights into the magnetic behaviors of celestial bodies and their role in planetary phenomena, maintaining the scientific integrity for an expert audience
The Pursuit of Understanding: How Curiosity About Exploration and Innovation Inspires Storytelling
Get PDFFor me, the allure of science is centered around our relationship with the natural world—how we strive to discover, work to coexist with nature, and search for ways to address our medical concerns and those of our animal companions. Through this collection of articles I observe how exploration and innovation are at the heart of a wide breadth of scientific endeavours. Such endeavours include exploring the stars and the depths of the sea, improving human and animal health, and innovating solutions to combat beach erosion, algae blooms, and mitigate the impact of beaver dams
Utility of rapid cycle health facility assessments to improve ANC service provision in Sarlahi, Nepal
No full textHealth facilities are central to antenatal care (ANC) service delivery, but the availability and quality of services provided exhibit substantial variability, particularly in lower- and middle-income countries (LMICs). In Nepal, despite significant improvements in maternal healthcare, the country still grapples with a high maternal mortality ratio. Assessments of the 2015 Nepal Health Facility Survey (NHFS) revealed that many health facilities lack readiness to provide essential maternal health services, highlighting the necessity to better monitor factors influencing health service readiness and service provision for ANC services.
Existing readiness and service provision assessments like the Service Provision Assessments (SPAs) and Demographic and Health Surveys (DHS) provide valuable insights but face limitations such as time gaps between data collection rounds, reducing the timeliness of data for decision-making. Additionally, these cross-sectional assessments are less equipped to address the considerable temporal and facility-level variability observed in service readiness and provision. Traditional ANC coverage estimates based on household surveys also often lack the depth required to measure effective coverage, particularly in settings like Nepal where resources, readiness, and service provision may fluctuate over time and across facilities.
To address these challenges, there has been a growing interest in exploring alternative monitoring approaches such as rapid facility-based surveys that offer more frequent, real-time data, even if less comprehensive. This, coupled with additional methodologies – such as linkage of household and facility data - can offer opportunities for more accurate estimates, extending beyond mere service contact and addressing quality of care offered.
In light of these considerations, this dissertation investigates the temporal dynamics of health facility readiness, service provision and service quality for ANC services. By utilizing several data collection tools including facility-based assessments, direct observations, and recall household data, this dissertation aims to provide a more nuanced understanding of facility readiness and its relationship with service provision, as well as their combined value in calculating service quality estimates, together. More specifically, this research highlights the need for more rapid and more frequent ANC health service assessments in regions like Sarlahi, Nepal, where provision of ANC services and maternal health outcomes continue to face persistent challenges