Honors Program Theses: Spring 2023

Permanent URI for this collectionhttps://hdl.handle.net/10675.2/625516

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Reversing Alzheimer's Disease Pathology in Zebrafish by Photobiomodulation Treatment
(Augusta University, 2023-05) Velez, Daniela
Zebrafish (Danio rerio) are one of the widely used research models due to their characteristics such as being transparent during development, sharing 70% of genes with humans, and having conserved features of vertebrate aging including deterioration of mitochondrial and cognitive functions. Neurodegenerative diseases, such as Alzheimer’s disease (AD), Parkinson’s disease (PD), and Huntington’s Disease (HD), are associated with aging and characterized by amyloid plaques and tau tangles leading to progressive neuronal loss and cognitive deficits. While affecting approximately 15% of the world population, neurodegenerative diseases are currently not curable which is why this study was conducted with the following two aims: (1) generate a pharmacological zebrafish model of AD by adding small amounts (100 nM) of okadaic acid (OKA) directly to fish tanks for 9 days and (2) reverse the AD pathology and impaired cognitive function by applying a sequential photobiomodulation (PBM) therapy. When examined after OKA treatment, zebrafish brains displayed histological hallmarks of AD including amyloid plaques, neurofibrillary tangles, vacuoles, and accumulation of GSK3β-pSer9. Furthermore, behavioral studies using a T-maze revealed that OKA-treated zebrafish spend significantly less time in the reward arm compared to untreated controls (15.2% vs. 50%). In contrast, a sequential PBM therapy at two different doses (i.e., 3 J/cm2 and 4.5 J/cm2) significantly reduced formation of amyloid plaques, neurofibrillary tangles, and vacuoles, as well as accumulation of GSK3β-pSer9 in brains of OKA-treated zebrafish while also improving their cognitive function as evidenced by being able to recall the reward arm and spending more time there (55 and 57%, respectively) similar to controls. Together these findings suggest that (1) a fast and cost-effective zebrafish model of AD can be established by OKA treatment and (2) PBM therapy is a promising approach to reverse AD pathology.
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Use of Ferric Oxide, Aluminum, and Copper as Green Reduction Catalysts for the Synthesis of Nylon 6-6
(Augusta University, 2023-05) Flint, Abigail; Department of Chemistry and Biochemistry
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Investigation of HphB and an ArAT in a Homologation Pathway Department of Chemistry & Physics
(Augusta University, 2023-05) Parviainen, Anna; Chemistry
Natural products (NPs) are nonessential metabolites produced by organisms that are often used in the production of pharmaceuticals. Their natural purpose is to protect their producing organism from the environment, making them an attractive medium for the starting point of drug discovery. NPs are generally very diverse in structure and therefore difficult to reproduce or alter structurally using organic synthesis. Understanding how NPs are synthesized in Nature aids researchers in developing new tools to produce novel NP-like bioactive compounds. The enzymatic pathway this project investigates is a homologation pathway found in the cyanobacterium Nostoc punctiforme PCC 73102 that serves to add a methylene (-CH2-) group to the side chain of an amino acid. Characterization of enzymes in this pathway could lead to the ability to artificially homologate other natural products, a tool that would be valuable because the addition of a methylene group has been shown to improve the biological activity of compounds. The two enzymes involved in this pathway that are being investigated in this project are dehydrogenase/decarboxylase HphB and an aromatic aminotransferase (ArAT). The goal of this project is to clone, express, and purify these enzymes and then test their enzymatic activity to characterize their detailed functions. The two enzymes of interest have been successfully purified by the biochemical technique affinity column chromatography, and the activity of ArAT is currently being investigated by applying the analytical method high performance liquid chromatography (HPLC) to experimental assays. The knowledge gained from this project will shed light on an unknown enzymatic pathway, which could ultimately lead to the development of a new enzymatic tool.
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The Role of Rbx1 in Cardiac Development
(Augusta University, 2023-05) Varghese, Priyanka; Biology
The human heart is one of the first organs to form and function during the development of a human embryo. Heart development is a highly complicated and tightly regulated process that culminates in the formation of the four-chamber heart. During the 6 to 7 weeks of human embryo development, the bottom of the heart tube forms the two atria, and the middle of the tube forms the two ventricles. The ventricular walls then begin to divide into all 4 chambers. Each chamber includes an entrance and exit for blood flow. Abnormal heart structure can impair heart contractility and can lead to Congestive Heart Disease (CHD). CHD is very prevalent in many children in where the heart does not pump a sufficient amount of blood into one’s body. According to the Center for Disease Control and Prevention (2008), “CHDs affect nearly 1% of?or about 40,000?births per year in the United States”. CHD can be caused by genetic or environmental cues that disrupt heart development (Lage et al., 2012). The mechanisms that lead to CHD are unfortunately not fully understood.
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Design and Synthesis of Potential Antiviral Drug Candidates
(Augusta University, 2023-05) Moore, Jade; Chemistry
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Investigation of Vitamin B12 for the treatment of traumatic brain injury
(Augusta University, 2023-05) Onyekachi, Joy; Biology
This study investigates the potential of vitamin B12 as a treatment for traumatic brain injury (TBI). TBIs are complex injuries that can result in a wide range of symptoms, and current treatments include medications, surgery, and therapies. The pathology of TBIs includes excitotoxicity, mitochondrial dysfunction, and apoptotic cell death. Vitamin B12 has been shown to have neuroprotective effects in various neurological disorders, but its potential as a treatment for TBI has not been extensively studied. This study uses a rodent model of TBI to investigate the effects of vitamin B12. The study aims to contribute to the understanding of the mechanisms underlying TBI and to identify potential therapeutic targets for this complex injury. The rats underwent CCI surgery and were divided into three groups: control group, injured non-treated group, and experimental group. Each group consisted of 10 adult rats, and cognitive and behavioral testing was conducted using the Foot Fault Task (FFT). The FFT allowed for the quantitative assessment of the motor deficits that commonly occurred following brain injury. The assigned task for the project was to score and analyze the foot fault data, providing important insights into the motor deficits experienced by the rats and the potential impact of vitamin B12 on their recovery. Figure 1 shows a linear decline in the number of foot faults made by the rats, indicating improved motor coordination and balance. However, statistically B12 did not have any significant effect on the treatment of TBI.
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The role of bHLH transcription factor Bmal2 in arterial endothelialcircadian rhythms and remodeling: Sex dependent effects in mice
(Augusta University, 2023-05) Moratin, Jordan; Cell and Molecular Biology
Cardiovascular disease remains the number one cause of mortality in humans. An important influence in the progression of artery disease is the long-term effect of disruptions in daily patterns or circadian rhythms. Not sleeping well at night, and daytime sleepiness associate with cardiovascular disease. In addition, a cardiovascular system that does not rest well at night is also bad. A blood pressure reading that does not decrease at night called non-dipping hypertension worsens cardiovascular disease. One important impact of broken rhythms is to cause disease in arteries. Thus, understanding the mechanisms that control 24-hour daily patterns is important. One key component gene of circadian rhythm is the transcription factor Bmal1. Vascular disease is a progression that begins as an adaptation to hypertension, diabetes, and hypercholesterolemia, whereby blood vessels change their structure in response to these changes in the bloodstream through a process called vascular remodeling. Remodeling is a process whereby arteries, arterioles, and even veins change their size and cellular structure (muscularity). Using mouse models of genetic disruption, our lab previously found that Bmal1 has an important role to control vascular remodeling, and when Bmal1 is disrupted, a vascular disease phenotype occurred. The lab also found that the endothelial cell layer of arteries contributes to the disease in Bmal1 knockout (KO) mice. These observations seemed the same in both males and females, thus, were sex independent. Bmal2, is a paralog of Bmal1, and interacts with Bmal1, and is more selectively expressed in the endothelium. However, the role of Bmal2 in remodeling is not clear. To understand the role of Bmal2 in vascular disease, I have implemented a widely used experimental animal model of arterial ligation to induce vascular remodeling. I have ligated the left common carotid artery (LC) in two groups of mice, control wild-type mice (no genetic mutation) and the experimental Bmal2-KO (global knockout) mice. After two weeks, I isolated the LC and fixed the arteries in optimal cutting temperature (O.C.T.) compound and conducted histological processing (cut cross sections with a cryotome and staining with hematoxylin and eosin). I then quantified the changes in structure in the artery using the ImageJ program on digitized microscope images. My findings show that inward remodeling and wall-hypertrophy in male Bmal2-KO mice are similar to wild-type mice. The inward remodeling observed in the male WT and male Bmal2-KO mice is consistent with the normal response of what has been observed in this ligation model, inward accompanied by wall hypertrophy. However, I saw something different in the female Bmal2 KO mice undergoing the ligation for two weeks. Female Bmal2-KO mice exhibited robust inward remodeling that was accompanied by intimal hyperplasia. My data suggest that there are sex-specific differences in remodeling controlled by Bmal2.
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The Role of KCTD 17 in the Striatal G-Protein Coupled Receptor Signaling
(Augusta University, 2023-05) Momin, Saniya; Cell and Molecular Biology
Many neurological disorders cause a wide array of detrimental effects, such as movement disorders. Despite an extensive amount of research on the underlying structure and signaling of neurodevelopmental disorders, such diseases remain largely uncured. After decades of unclear etiology, we now know of numerous genetic marker sessential for disease progression. Unfortunately, this revelation has not yet enhanced patient outcomes, suggesting precedence to investigate novel genetic players. Research is currently rising upon the potassium channel tetramerization domain (KCTD) family due to neurological diseases from patients with mutations in various KCTD genes. Yet,our understanding on the roles of KCTD proteins is in its infancy. However, several KCTDs (KCTD2, 5, 8, 12, and 16) have been shown to interact with components of the protein network that interrogate neuromodulatory signals through G Protein Coupled Receptors (GPCRs). Abundant evidence indicates an essential role in neuromodulatory GPCR signaling across a spectrum of movement disorders. Intriguingly clinical variations in KCTD17, which has a high level of homology to KCTD2/5, is causal of movement disorders. Therefore, we have examined KCTD17’s role in the GPCR signaling pathway. In addition, previous reports have demonstrated a relationship between primary cilia and KCTD17. Intriguingly, components of the GPCR signaling pathway have been localized to primary cilia, however the role of primary cilia in neuromodulatory signaling has not yet been explored. We tested the significance of this pathway by observing the effects of cilia on KCTD17 to help us better understand the function of primary cilia to KCTD17 regarding neuromodulatory signaling. The long-term goal was to analyze how the loss of KCTD17 can affect motor control. We used the CRISPR/Cas9 approach to see how KCTD17 can affect the neuronal activity via GPCR signaling and primary cilia disruption. We found that disruption of primary ciliaindeed influences GPCR signaling. We are hopeful the results will shed light to find further treatment in the neurodevelopmental disorders associated with the protein.
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COVID-19 in Nursing Homes : A Comprehensive Survey of Healthcare Management Practices
(Augusta University, 2023-04) O'Keefe, Anabelle; Health Services and Music
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Investigation of Fear Memory Induced by Contextual Fear Conditioning
(Augusta University, 2023-05) Pokharel, Kritika; Cell and Molecular Biology
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The Function of ELAVL Genes During Neuronal Reprogramming
(Augusta University, 2023-04) Williams, Christine; Chemistry
Spinal cord injuries often impair a person’s daily life by limiting mobility. While potential treatments such as stem cell transplantation are being investigated in animal models, these treatments have not translated in clinical trials and entail risks including triggering an immune response and tumorigenesis. Neuronal reprogramming has emerged as an alternative method of treating spinal cord injuries to restore the mobility lost by patients. When an injury to the spinal cord occurs, reactive astrocytes surround the site of injury where neurons have died. Neuronal reprogramming involves the reprogramming of surrounding glial cells into functional neurons. While neuronal reprogramming has great potential in treating neurological diseases, the underlying molecular mechanisms of this unique biological process are still not fully elucidated. Previous research has shown that when overexpressed, the transcription factor, NeuroD1 can convert reactive astrocytes into functional neurons at a high conversion efficiency. The overexpression of NeuroD1 leads to upregulation of other genes including the two genes, ELAVL2 and ELAVL4. Here, I aim to determine if these two genes have reprogramming ability on their own. Upon cloning into overexpression vectors, I examined the function of these genes in reprogramming of glial cells (U251 glioblastoma cell line). Using techniques like western blot and immunocytochemistry, cells overexpressing these genes were analyzed. Overexpression of ELAVL2 and ELAVL4 resulted in no significant change in morphology and no expression of the young neuronal marker, DCX. In conclusion, this study does not support the hypothesis that the overexpression of ELAVL2 and ELAVL4 could result in neuronal reprogramming.
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Christ and the Criminal: a Literary Case Study of Fyodor Dostoevsky's The Idiot and Brothers Karamazov
(Augusta University, 2023-05) Polhill, Eleanor; English
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Southeastern Fiddler Crabs : Distribut io ns & Characteristics of Burrow Ornamentation
(Augusta University, 2023-05) Patel, Staumi; Biology
An experimental design has been chosen to exhibit the distributions and characteristics of fiddler crab burrow ornaments , specifically present on the southern coast of the United States. This thesis will demonstrate the growing evidence of non-biological ornamentation found in the burrows constructed by fiddler crabs. To further this research, two sampling sites have been pinpointed along the southeastern coast, Edisto Island and Hunting Island, to survey the existing features of fiddler crab burrow ornaments. The distribution of burrow structures was compared to local regional differences between South Carolina versus Georgia and the Southeast versus the Northeast coast. Furthering this research, it was also postulated that burrowing behavior of fiddler crabs is affected by the sediment size and the construction of the burrow ornaments/hoods. To identify the presence of fiddler crab semi domes and ornaments of the type, a continuation of observations of the ecological features, such as mating, predation, and environmental heterogeneity will be factored.
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Replacement of o-xylene with Avocado Oil as the Solvent System in a Diels-Alder Reaction
(Augusta University, 2023-05) Bangiyev, Sandra; Biology
This study is investigating environmentally friendly alternatives to o-xylene as a solvent in the Diels-Alder synthesis using butadiene sulfone as a precursor to 1,3-butadiene. This reaction is used as a learning laboratory experiment in the Augusta University introductory Organic Chemistry class as well as in a multitude of other universities. O-xylene has a variety of detrimental health impacts ranging in severity due to the length and duration of exposure to the solvent. Not only is it a health hazard, but it is also an environmental pollutant. One of the major concerns of o-xylene is that due to its high volatility, several environmental pollutants can be released into our atmosphere. With these health and environmental impacts in mind, the aim of this green chemistry project is to find a solvent that could replace o-xylene without affecting the type and quality of the product produced. Due to the availability, affordability, compatibility, simple extraction method, and easy access to third-world countries, it is hypothesized that avocado oil would be an excellent green replacement for this reaction.
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The effect of exercise mode on time perception during exercise
(Augusta University, 2023-05) Johnson, Kade; Kinesiology
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The Effect of Human Immunodeficiency Virus Antiretroviral TherapyDolutegravir on the Metabolic Health of Mice
(Augusta University, 2023-05) Ajala, Priscilla; Chemistry
Human immunodeficiency virus (HIV) and its more advanced form acquired immunodeficiency syndrome (AIDS) has been around for decades and affected millions worldwide. Over the years, scientists have developed novel antiretroviral therapies (ART) to prevent the progression of HIV to AIDS and increase life expectancy in people living with HIV (PLWH). Although this treatment is beneficial in blocking viral replication, it also has adverse side effects on the metabolic and cardiovascular health of PWH. Specifically, issues like obesity that were previously written off as nutritional rehabilitation have now become a more common occurrence in people living with HIV (PLWH). Fundamentally, this project focused on the cardiometabolic impact of the ART, dolutegravir, an integrase inhibitor used to treat PLWH by preventing the integration of viral DNA into the hosts. The mice were used as a model for how the drug dolutegravir affects humans. In the study, we placed the mice on a 3-month diet of dolutegravir and had a control set of mice that are fed with normal chow. From there we kept track of the weight gain by weighing the mice manually and then used nuclear magnetic resonance (NMR) to measure body composition including the percentage of body fat, lean mass, and fluid content. At the end of the three-month period, the mice were placed in calorimetric cages (CLAMS cages) to monitor the metabolic function of the mice; specifically heat production, oxygen consumption and carbon dioxide respiration. After all the live data was collected the mice were euthanized and different parts of their bodies were used in a series of analytical experiments such as wire myography, and RNA/DNA PCR analysis, comprehensive lab animal monitoring system cages (CLAMS). We hypothesized that the Integrase inhibitor Dolutegravir utilized in ART contributes to the weight gain seen in patients with HIV using ART by decreasing heat production by decreasing the non-shivering thermogenesis brown adipose levels and uncoupling protein (UCP) levels. The experiment showed signs of increased body weight in the mice treated with dolutegravir as well as decreased heat production and decreased brown adipose tissue and UCP-1. The decreased brown adipose tissue and UCP-1 could explain the decrease in heat production which could lead to weight gain in humans despite the lack of significant evidence in mice. This study provides insight into the effects of dolutegravir on mice, but more importantly shows how it could potentially be affecting patients that are treated with this drug. It also gives room for improvement on the different classes of drugs used in combination with antiretroviral therapy and which classes of drugs are resulting in adverse side effects and which classes are not. It opens the space to find new drugs that still help in blocking viral replication, but also decrease the number of adverse side effects on people with HIV.
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Expression and Purification of HphB and ArAT: Enzymes in the L-Phenylalanine Homologation Pathway
(Augusta University, 2023-05) Reynes, Juan-Paolo; Cell and Molecular Biology
Natural products (NPs) are organic molecules produced by microorganisms and plants that are often used in drugs and drug leads. Organisms use biological catalysts called enzymes to perform the biosynthesis of NPs. Homo-L-phenylalanine (L-Hph) is an uncommon amino acid derived from L-phenylalanine (L-Phe) by a chemical process called homologation. L-Hph is used as a building block of some peptide NPs to enhance their biostability. HphB and aromatic amino acid aminotransferase (ArAT) are two out of four enzymes involved in the biosynthesis of L-Hph. In the proposed biosynthesis of L-Hph, HphB catalyzes dehydrogenation/decarboxylation to convert 2-benzyl-3-hydroxybutanedioic acid (B3HB) to 2-oxo-4-phenylbutyric acid (OPB). ArAT is responsible for the first and last step of the homologation pathway: the conversion of L-Phe to phenylpyruvic acid (PPA) and 2-oxo-4-phenylbutyric acid (OPB) to L-Hph. This project focused on cloning, expression, and purification of HphB and ArAT. After cloning of genes encoding HphB and ArAT into the expression plasmid pET-28a, HphB was overexpressed in E. coli and purified by an affinity column chromatography. It is now ready for the enzymatic assays for its characterization. However, purification of ArAT was unsuccessful as the protein was insoluble. It was cloned to be expressed with different tags that can enhance the protein solubility, such as 6×histidine tag at N- and/or C-terminus as well as maltose binding protein tag, all of which did not make the protein soluble in a large scale. Further expression experiments using different constructs and conditions will be performed in the future. Characterizing enzymes involved in the homologation of L-Phe to L-Hph could allow for future engineering of the pathway to produce L-Hph analogs and other homologated amino acids. These compounds can be used to increase the potency and stability of existing drugs, as L-Hph is used as a chiral building block in some pharmaceuticals, such as angiotensin-converting enzyme (ACE) inhibitors.
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Synthesis of Novel NSAID Hybrid Conjugates as Potential Anti-inflammatory and Anti-Cancer Agents
(Augusta University, 2023-05) Chavez, Julio; Chemistry
A series of piperidone-ibuprofen conjugates were synthesized through the reaction of 3E,5E-4-piperidones with the appropriate ibuprofen-amino acid conjugates in DCM in the presence of EDC and DMAP. All the synthesized conjugates were well characterized by spectroscopic techniques. Most of the conjugates reveal antiproliferative properties against A431 (squamous skin) cell lines with potency higher than that of reference drugs 5-fluorouracil and sunitinib. Many of the synthesized agents also reveal remarkable antiproliferative properties against a HCT116 (colon) cancer cell line.
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Dietary and Caffeine Intakes of Firefighters Located in the Southeast
(Augusta University, 2023-05) Hachem, Garrett; Kinesiology
Firefighters are prone to coronary heart disease, cancer, obesity, and high blood pressure possibly due to the working conditions they face including irregular sleeping/eating habits, high- stress levels, and exposure to carcinogens. This study focused on the nutritional aspect and observed nutritional habits including caffeine consumption in local firefighters. I hypothesize firefighters will have a high caloric intake (Over 3,000 kcal/day) and caffeine consumption (Over 400 mg/day) due to the aforementioned health factors. A total of 15 local firefighters participated in this study. Dietary logs of the firefighters were collected for five days over a seven-day period to get a baseline of the participants’ regular eating habits. The data analysis will be completed using MyFitnessPal and will consist of protein, carbohydrates, fats, fiber, sugar, cholesterol, sodium, and caloric intake. In addition, they are asked to log their daily caffeine consumption.
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Inflammatory responses to chorda tympani nerve sectioning in TNFR1 Knockout miceand their role in taste bud degeneration
(Augusta University, 2023-03) Boothe, Khaylie; Neuroscience and Regenerative Medicine
Cytokines such as tumor necrosis factor (TNF) play a key role in the development and maintenance of pain after peripheral nerve injury or infection. These are pro-inflammatory cytokines- small proteins that are crucial in controlling the proliferation, recruitment and activity of immune cells. Mechanisms responsible for taste bud regeneration are not fully understood, sothis hinders strategies towards restoring natural taste sensation after trauma, cancer treatment,and even viral infection. A striking example of this is a loss of taste, commonly attributed to long-term SARS-CoV-2. Previous research conveys that TNF receptor signaling is required for taste bud regeneration after experimental nerve sectioning. Interleukin-1 (IL-1) receptor also showed that neural responses to taste were also affected after nerve sectioning. Based on this, we proposed that neutrophil recruitment due to CT nerve injury would be dampened in Tnfr1/2 knockout (KO) compared to the wild-type mice receiving injury. To test the hypothesis, CT nerve axotomy was performed in anesthetized C57BL/6J (wild-type mice) and Tnfr1/2 KO mice. Mice were euthanized 6 hours post-injury, and tongues were harvested, frozen, and cryosectioned. Hour 6 C57BL/6J (wild type mice) and Tnfr1 KO mice cryosections were stained with MPO+ neutrophil marker, and DAPI nuclei marker then analyzed for MPO+ neutrophil pixels using MetaMorph imaging software. Significant neutrophil responses were observed at a greater rate in TNFR KO mice than in the wild-type mice, which does not support the hypothesis. These findings suggest that other cytokines, like IL-1, can compensate for neutrophil recruitment. Further studies will focus on different time points after injury as well as observing the mucosa as a region of interest to observe the migration of neutrophils from the vasculature to the papillae.