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THE ABILITY OF SEASONAL INFLUENZA VACCINATION TO INDUCE T CELL CROSS-REACTIVITY WITH PEPTIDES DERIVED FROM AVIAN INFLUENZA STRAINS
Get PDFInfluenza viruses pose a persistent threat to global public health due to their ability to mutate rapidly, leading to the emergence of new strains. Current seasonal flu vaccines primarily target antibody-mediated immunity, leaving gaps in protection against novel variants that easily evade the immune system. This thesis investigated the potential of T cell-mediated cross-reactivity to provide broad-spectrum defense against diverse influenza virus strains including avian influenza strains. Through intracellular flow cytometric analysis of IL-2, IFN-γ, and Granzyme B, T cell responses to influenza peptides (Flu B, H5N1, H3N2, Flu A, H7N9, and influenza A Pepmix) stimulation were investigated in a cohort of flu-vaccinated individuals. Results unveiled donor-specific and peptide-specific variations in immune responses. Results from statistical analyses indicated non-significant differences in expressions of the cytokines tested in both pre- and post-vaccinated samples across CD4+ and CD8+ T cell populations, suggesting a larger sample size is needed to account for inter-individual variability. Nevertheless, a downregulation trend was observed for Granzyme B production between pre- and post-vaccination. Despite limitations such as sample size constraints, the findings underscored the promise of T cell cross-reactivity in broadening our understanding of influenza infection immunity dynamics. This study suggests avenues for future vaccine development strategies leveraging cross-reactivity to combat influenza and other infectious diseases
Elevated TGF-β activity in age-related macular degeneration
No full textAge-related macular degeneration (AMD) disease is one of the major causes of severe vision loss in the world. Current therapies primarily target the late stage of this disease including macular neovascularization (MNV) and geographic atrophy (GA) with diverse side effects. Thus, developing new treatments for the early stages of AMD presents a significant challenge. Transforming growth factor β (TGF-β) plays a role in the regulation of fibrosis, recruitment of circulating cells, and induction of neovascularization. Accumulation of age-related oxidative stress increases the level of activated TGF-β. Active TGF-β can induce fibrosis by causing cell loss through apoptosis, stimulating extracellular matrix (ECM) synthesis, and promoting the transition of various cell types into fibroblast-type cells capable of depositing ECM. Furthermore, TGF-β is a potent chemoattractant for circulating cells. It can recruit immune cells and mesenchymal stem cells to induce different biological responses. The relationship between increasing oxidative stress with aging in the subretinal area and the TGF-β signaling activity, and the role of TGF-β in AMD progression are not well understood. Here, we demonstrated that TGF-β is accumulated with AMD pathological induction in RPE and choroid layers. We created AMD mice by injecting the mild oxidative agent, sodium iodate, in 22-month-old aged wild-type mice to induce AMD-like characteristics including retinal pigment epithelium (RPE) disruption, tight junction loss, and immune system activation. Furthermore,
we found that the recruitment of immune cells and stromal cells in the subretinal area can be regulated by the TGF-β signaling pathway. Overall, our work supports the role of elevated TGF-β activation with aging in the immune cell and stromal cell recruitment in AMD progression. Suppressing the increase in TGF-β activation could possess high therapeutic potential in treating AMD
Study of biochemical and biological effects of N6-(2-deoxy-α,β-D-erythropentofuranosyl)-2,6-diamino-4-hydroxy-5-formamidopyrimidine, an epimerizable DNA lesion
No full textDNA is constantly exposed to various exogenous or endogenous agents that induce DNA damage. During this process, the native nucleotides undergo chemical modifications on bases as well as sugar backbones. 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-Oxo·dG) is one of the most common products of guanine oxidation and stands as one of the most well studied DNA lesions. In contrast, only a few studies were done on N6-(2-Deoxy-α,β-D-erythropentofuranosyl)-2,6-diamino-4-hydroxy-5-formamidopyrimidine (Fapy·dG), an epimerizable DNA damage that is closely related to 8-Oxo·dG. It has been established that Fapy·dG is formed in comparable or sometimes even greater amounts compared to 8-Oxo·dG. Furthermore, there is evidence showing that Fapy·dG is more mutagenic than 8-Oxo·dG. It is possible that Fapy·dG is more biologically significant compared to 8-Oxo·dG, but little is known about this DNA damage, mainly because of the challenging synthesis of Fapy·dG-containing oligonucleotide.
Recently, a more practical synthesis of Fapy·dG-containing oligonucleotide was achieved, allowing further study of this lesion. Kinetic analysis was carried out to investigate how Fapy·dG-containing oligonucleotides interact with various polymerases involved in replication and transcription. When it was formed in the template strand, Fapy·dG results in reduced replication bypass and leads to mutagenesis. Adjacent 5-formyldeoxyuridine further enhances its mutagenic effects. When formed in triphosphate as Fapy·dGTP, its incorporation by DNA polymerase is even less efficient. During transcription, Fapy·dG was found to reduce the bypass of RNA polymerase II or T7 RNA polymerase elongation complexes but does not block the promoter-dependent transcription. Cellular studies indicate that Fapy·dG is more mutagenic than 8-Oxo·dG during not only replication but also transcription. Further mechanistic details were obtained from crystallography studies. An important aspect of our study is how different anomers of Fapy·dG interact differently with polymerases. Our studies suggest that α-Fapy·dG exhibits poorer recognition by RNA polymerases and results in higher mutagenic effects during transcription.
Our research provides further information on how Fapy·dG, either in the DNA template strand or in triphosphate form, affects the replication process in-vitro and in cells. Furthermore, we also provided the first analysis on how Fapy·dG affects the transcription process. Based on our research, we believe that Fapy·dG could potentially be more biologically important than 8-Oxo·dG
FACULTY EXPERIENCE OF ACHIEVEMENT EMOTIONS IN THE COMMUNITY COLLEGE ASYNCHRONOUS ONLINE CONTEXT
Get PDFThis qualitative phenomenological study examined how faculty experience of achievement emotions may be influenced by features of the community college asynchronous context (CCAOC). The following overarching question and sub-questions guided the research:
1. How is faculty experience of achievement emotions influenced by contextual features of the CCAOC?
a. How is faculty experience of achievement emotions in the CCAOC influenced by the asynchronous delivery modality?
b. How is faculty experience of achievement emotions in the CCAOC influenced by the learner population?
c. How is faculty experience of achievement emotions in the CCAOC influenced by administrative policies and practices?
The research questions were answered qualitatively by analyzing data from a sample of five community college faculty using a 1-hour, in-depth, semi-structured interview. The interview was recorded and transcribed via Zoom, then analyzed using Moustakas' (1994) transcendental phenomenology model. The study made four key conclusions. First, contextual features in the CCAOC influence faculty experience of achievement emotions via the mechanism of appraisal of capacity to achieve the teaching philosophy and its related subgoals. Second, the asynchronous modality influences faculty experience of achievement emotions, through perceptions of properties of the modality. Third, the learner population influences faculty experience of achievement emotions through perceptions of learner characteristics, engagement, and classroom conduct. Fourth, administrative policies and practices influence faculty experience of achievement emotions through perceptions of administrative engagement and support. Needs assessment research was recommended to evaluate how interventions informed by the four major findings of the study may enhance teacher experience of enjoyment related to instruction
From Bench to Bedside: The Impairment of Cerebrovascular Autoregulation Following Cardiac Arrest and Potential Interventions
Get PDFAnnually, over 350,000 people experience cardiac arrest, but only a mere 10% of them manage to survive. Cardiac arrest is a medical emergency that greatly reduces blood flow to the brain, causing severe brain damage and neurological impairment. The leading cause of death following resuscitation from cardiac arrest is brain injury. Therefore, tools that can quantify brain injury in real-time are critically important, so they may be used to guide lifesaving therapeutic interventions at the bedside. Cerebrovascular autoregulation is the innate ability of the cerebral blood vessels to regulate their blood flow amidst blood pressure variations. It has been used in the clinic as a metric to quantify the level of injury experienced by the brain at rest following resuscitation. In this thesis, I discuss the methods employed for quantifying cerebrovascular autoregulation, including equipment, data analysis techniques, and potential challenges faced in the clinic. I then use a rat model to interrogate the dynamics of autoregulation in the immediate aftermath of resuscitation from cardiac arrest and quantify both the ‘static’ and ‘dynamic’ aspects of cerebrovascular autoregulation post-resuscitation. Next, I introduce an innovative approach to quantify the disruption of autoregulation that is not detectable by standard techniques via quantifying its changes at ultralow frequencies, i.e., in the scale of 1 mHz. Finally, I conclude this thesis by describing the clinical implications of these findings and discussing potential intervention strategies. I also point out the limitations of the study and suggest directions for future research in cerebrovascular autoregulation following cardiac arrest
INVESTIGATING CRISPR-CAS REGULATION IN THE NATIVE BACTERIAL HOST STREPTOCOCCUS PYOGENES
Get PDFAlthough CRISPR technologies have been developed extensively, less is known about how CRISPR-Cas systems function in nature. In bacteria, CRISPR-Cas systems form memories of bacteriophages (phages) by inserting a short piece of phage DNA (a “spacer”) into the CRISPR array in the bacterial genome. Spacers are transcribed into crRNAs which direct the nuclease Cas9 to bind and cleave complementary DNA during a reinfection. Most of the research investigating these steps has been performed in heterologous expression systems or in vitro. To understand how CRISPR-Cas systems are regulated, and to identify other interacting cellular pathways, we must begin to study them in their native bacterial hosts. To address this knowledge gap, we developed tools to study the CRISPR-Cas system found in Streptococcus pyogenes.
This thesis work describes a novel form of CRISPR-Cas transcriptional regulation. In S. pyogenes, a single-guide RNA, tracr-L, directs Cas9 to bind to its own promoter, transcriptionally repressing cas9 expression. During my thesis work, we discovered that Cas9 binding elevates the rate of mutation within its binding site. Cells in which these mutations occur overexpress Cas9, and are thus more immune to phage infection. However, overexpression comes with a fitness cost, and when the phage threat is cleared, overexpressers are outcompeted by wild-type cells. We propose a model whereby the size of the overexpressing subpopulation expands and contracts in response to phage pressure, allowing cells to mount a quick defense to sudden phage epidemics.
Secondly, we describe the development of a transposon sequencing (Tn-Seq) screen to identify additional factors that interact with CRISPR-Cas in S. pyogenes. By using the tools and assays we optimized in S. pyogenes, we will be able to characterize the hits from the Tn-Seq screen and determine how they interact with the CRISPR-Cas system
Rosetta Energy Approximation Using a Machine Learning Approach
Get PDFThe advent of AlphaFold2 has significantly accelerated advancements in protein structure prediction using deep learning. Despite its monumental success, the AlphaFold2-like learning-based methods lack explainability and generalization, which has limited further understanding and application. Traditionally, the minimizing free energy approach, exemplified by Rosetta, has been cornerstone in protein structure prediction, embedding extensive biophysical insights into its methodology. Notably, when researchers encounter improbable structures, Rosetta is used to refine them, enhancing their physical plausibility. The critical interplay between structure prediction and energy optimization highlights a gap in current deep learning approaches, which overlook the integration of energy information. Addressing this, my project aims to incorporate energy-based metrics into deep learning models, enhancing both their predictive performance, generalization and explainability, alleviating AlphaFold2-like models' heavy reliance on Multiple Sequence Alignments (MSAs) and extensive data sets. By employing equivariant graph neural networks, I have begun to approximate Rosetta’s one-body and two-body energy terms, achieving Pearson correlations with Roseta’s energy metrics above 0.7 for most terms. My work has prepared machinery to integrate the energy model into some deep learning models like IgFold, an antibody structure prediction method developed by our lab. This integration aims to enhance IgFold’s performance and its ability to generalize across diverse antibody structures
Learner Characteristics and Online Course Adoption
No full textHigh education institutions have undergone unprecedented challenges after the pandemic with emergency remote teaching (ERT). Students’ online experiences and attitudes towards the unexpected shift to online varied greatly. With a study population of part-time mid-career finance MBA students in China, this dossier investigates the factors contributing to adult learners’ decision to take online and hybrid courses. The mixed-methods study examined how students’ self-regulated learning (SRL) abilities and academic motivation influenced their adoption of online and hybrid courses, which in turn has an impact on their on-time graduation status. The analysis also explores the moderating effect of gender in these relationships. The research findings reveal that students with lower help-seeking behaviors and stronger goal-setting tendencies were slightly inclined towards online/hybrid courses. Those who took more online courses are more likely to graduate on time. In addition, a significant gender difference was noted in SRL help-seeking behaviors, with female students scoring higher than male students. Besides, female students adopt a statistically lower number of total online/hybrid courses as compared to their male counterparts. Qualitative findings further illustrate various patterns through which individual preferences, strategic concerns, and adaptability shaped the experiences and preferences of different students during pandemic-induced disruptions. Students who graduated on time often described a pragmatic approach, acknowledging the shortcomings of online learning but embracing it as a necessary strategy. In contrast, those who delayed graduation strongly preferred in-person learning, often due to concerns over the quality of interactions and technical difficulties associated with online formats. Additionally, the research highlights the critical role of high-quality online learning environments, teaching support, and meaningful interaction and engagement in enhancing students' online experiences. The study results highlight the importance of students’ self-regulated learning abilities in the online environment. Educational institutions could consider integrating SRL skills development into the curriculum. Besides, they may introduce other support schemes for students' emotional support and SRL skill development with a focus on goal setting and help-seeking abilities. Gender differences in online and hybrid course adoption as well as help-seeking behaviors signify the necessity to consider gender-specific motivational orientations. The findings from this study serve as a foundation for future research on targeted interventions that can support SRL strategies and motivation in online learning environments, with implications for educators, policymakers, and institutions navigating the evolving world of higher education
A REVIEW OF THE FOUNDATIONAL STATUS OF STONY CORAL REEFS IN THE GREATER EASTERN CARIBBEAN
Get PDFThere is unequivocal evidence that a failure to reduce greenhouse gas (GHG) emissions and anthropogenic environmental degradation has effectively altered the dynamics of Earth’s climate. The consequential systematic decline of scleractinian (stony) coral reefs throughout the Greater Eastern Caribbean (GEC) has implied the foundational role that stony corals play in facilitating the life-sustaining ecosystem services regional populations have developed to rely on. This capstone project reviews much of the extant literature surrounding the dependency of human populations on GEC stony coral ecosystem services and applies the findings to promote collaboration among members of the newly defined ecoregion. This project is the product of a lifetime appreciation and fascination with coral reefs and concern for the future of vulnerable ecosystems and communities. The ecosystem services facilitated by stony reefs allow populations to establish basic socioeconomic platforms on which to develop (e.g., food security, household income, etc. ), preventing communities from experiencing significant hardship and helping them maintain a sense of self-sufficiency and sustainability. Leaders of GEC countries and territories should collaboratively implement proven solutions to mitigate the impacts of climate change and coral decline and promote climate stabilization through sustainable resource management practices
INTEGRATED GENOMIC AND METABOLOMIC ANALYSIS TO CHARACTERIZE MOLECULAR SIGNATURES OF AUTISM SPECTRUM DISORDER
No full textAutism spectrum disorder (ASD) is a neurological and developmental condition that represents a significant global public health concern, with approximately 1 in 100 children worldwide affected by autism. The causes of ASD are complex and not fully understood, but it is generally accepted that both genetic and environmental factors play critical roles. Research methodologies such as Genome-Wide Association Studies (GWAS) combined with untargeted metabolomics can link critical genes to the metabolic aspects of ASD and reveal new pathways and mechanisms involved in ASD. We got an ASD GWAS dataset from the GWAS Catalog, metabolomic data from Metabolomics Workbench, and one published paper. Genes related to ASD were mapped, and Reactome and STRING databases were used to show potential pathways and gene networks. Differential expression and differential correlation analyses were performed in the metabolomic. The differentially expressed genes were put into STITCH, and functional enrichment was done on differential correlation pairs. The results showed that DPYD stands out as a significant gene and enzyme identified in both GWAS and metabolomic datasets. The pyrimidine metabolism pathway may be linked to ASD. More differences are shown in correlation relationships than in differential expression analysis. Additionally, metabolomic data showed that there may not be a single pathway consistently altered in ASD; instead, there could be numerous minor variations across candidates involving RNA polymerase, alanine, aspartate and glutamate metabolism, cytoskeletal regulation, HIF-1 signaling pathway, and galactose metabolism may be crucial in ASD