Matching Items (151)
Description
In the development of personalized medicine and many other clinical studies, biospecimen integrity serves as the prerequisite for not only the accurate derivation of patient- and disease-specific molecular data from biological specimens but the meaningful downstream validation of biomarkers. However, a large number of preanalytical variables may influence the quality of biospecimens in an undesired way and ultimately render the samples unsuitable for molecular analysis. The limited ability to directly reduce discrepancies caused by preanalytical variables gives rise to the need for development and retrospective application of appropriate tests for assessment of biospecimen integrity. Nevertheless, the most standard approaches to assessing biospecimen integrity involve nontrivial procedures. Thus, the need for quality control tools or tests that are readily applicable and can produce results in a straightforward way becomes critical. As one of the major ex vivo biomolecular degradation mechanisms, oxidation that occurs when blood plasma and serum samples are exposed to thawed states during storage and processing is hard to forestall and detect. In an attempt to easily detect and monitor the degree of oxidation, the technique of Fluorescence Resonance Energy Transfer (FRET) was examined to determine whether this concept could be employed to monitor exposure of samples to thawed conditions when controlled by spontaneous oxidative disulfide bonding. The intended mode of usage was envisioned as a fluorescence liquid being stored in a separate compartment but within the same test tube as archived plasma and serum samples. This would allow the assessment of sample integrity by direct visualization of fluorescence under a hand-held black light. The fluorescent dynamic range as well as kinetic control of the reaction were studied. While the addition of Cu(II) proved to facilitate excellent dynamic range with regard to fluorescence quenching, the kinetics of the reaction were too rapid for practical use. Further investigation revealed that the fluorescence quenching mechanism might have actually occurred via Intramolecular Charge Transfer (ICT) rather than FRET mediated by oxidative disulfide bond formation. Introduction of Cu(II) via copper metal slowed fluorescence quenching to the point of practical utility; facilitating demonstration that storing at room temperature, refrigerating or freezing the samples delayed fluorescence quenching to different extents. To establish better kinetic control, future works will focus on establishing controlled, thoroughly understood kinetic release of Cu(II) from copper metal.
ContributorsZhang, Zihan (Author) / Borges, Chad (Thesis director) / Emady, Heather (Committee member) / Williams, Peter (Committee member) / Chemical Engineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2018-12
Description
As a biology major, many of my classes have included studying the fundamentals of genetics or investigating the way genetics influence heritability of certain diseases. When I began taking upper-division psychology courses, the genetic factors of psychological disorders became an important part of the material. I was exposed to a new idea: that genes were equally important in studying somatic diseases as they were to psychological disorders. As important as genetics are to psychology, they are not part of the required courses for the major; I found many of my peers in psychology courses did not have a grasp on genetic fundamentals in the same way biology majors did. This was a disconnect that I also found in my own life outside the classroom. Growing up, my mother consistently reminded me to limit my carbs and watch my sugars. Diabetes was very prevalent in my family and I was also at risk. I was repeatedly reminded of my own genes and the risk I faced in having this biological disorder. However, my friend whose father was an alcoholic did not warn her in the same way. While she did know of her father's history, she was not warned of the potential for her to become an alcoholic. While my behavior was altered due to my mother's warning and my own knowledge of the genetic risk of diabetes, I wondered if other people at genetic risk of psychological disorders also altered their behavior. Through my thesis, I hope to answer if students have the same perceived genetic knowledge of psychological diseases as they do for biological ones. In my experience, it is not as well known that psychological disorders have genetic factors. For example, alcohol is commonly used by college students. Alcohol use disorder is present in 16.2% of college aged students and "40-60% of the variance of risk explained by genetic influences." (DSM V, 2013) Compare this to diabetes that has "several common genetic variants that account for about 10% of the total genetic effects," but is much more openly discussed even though it is less genetically linked. (McVay, 2015)This stems from the stigma/taboo surrounding many psychological disorders. If students do know that psychological disorder are genetically influenced, I expect their knowledge to be skewed or inaccurate. As part of a survey, I hope to see how strong they believe the genetic risk of certain diseases are as well as where they gained this knowledge. I hypothesize that only students with a background in psychology will be able to correctly assign the genetic risk of the four presented diseases. Completing this thesis will require in-depth study of the genetic factors, an understanding of the way each disease is perceived and understood by the general population, and a statistical analysis of the survey responses. If the survey data turns out as I expect where students do not have a strong grasp of diseases that could potentially influence their own health, I hope to find a way to educate students on biological and psychological diseases, their genetic risk, and how to speak openly about them.
ContributorsParasher, Nisha (Author) / Amdam, Gro (Thesis director) / Toft, Carolyn Cavaugh (Committee member) / Ostwald, Madeleine (Committee member) / Department of Psychology (Contributor) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2018-05
Description
Alzheimer’s disease (AD) is a progressive cognitive and behavior disorder that is characterized by the deposition of extracellular Aβ plaques, intracellular neurofibrillary tangles, and neuroinflammation. Aβ is generated by cleavage of the amyloid precursor protein (APP) by β-secretase (BACE1) and, subsequently, y- secretase. In recent years, there has been an increasing interest in studying and understanding inflammation as a therapeutic target for AD. Inflammation manifests in the brain in the form of activated microglia and astrocytes. These cells are able to release high levels of inflammatory cytokines such as Tumor Necrosis Factor-α (TNF-α). TNF-α is a major cytokine, which is involved in early inflammatory events and plays a role in the progression of AD pathology. There are currently no treatments that target chronic neuroinflammation. However, previous work in our laboratory with transgenic mice modeling AD suggested that the anti-cancer drug lenalidomide could lower neuroinflammation and slow AD progression, though the cellular and molecular mechanisms are yet to be elucidated. Here we hypothesized that lenalidomide can modulate TNF-α production in microglia and decrease amyloidogenesis. Using immortal cell lines mimicking several brain cell types, we discovered that lenalidomide is likely to decrease inflammation by modulating microglia cells rather than neurons or astrocytes. In addition, the drug may prevent the overexpression of BACE1 upon inflammation, thus blocking the overproduction of Aβ. If confirmed, these results could lead to a better understanding of how inflammation regulates Aβ synthesis and provide novel cellular and molecular therapeutic targets to control the progression AD.
ContributorsGujju, Manasa (Author) / DeCourt, Boris (Thesis director) / Olive, M. Foster (Committee member) / Department of Psychology (Contributor) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2018-05
Description
Honeybees (Apis mellifera) are pollinators that face multiple challenges during foraging such as fungicides applied to floral sources. Fungicides are chemicals used to inhibit key fungal mechanisms like metabolism, but their effects remain relatively unknown in bees. In addition, studying the maturing bee can help us identify demographics that are more vulnerable to toxic materials like fungicides. The purpose of this study is test whether maturation and the fungicide Pristine influence the permeability of the blood-brain barrier. Specifically, we use a transportable dye to test how blood brain barrier transporter function responds to toxic insult and how it changes with age. Oral ingestion of Pristine by female workers did not have an effect on blood brain barrier permeability which suggests Pristine may have no or longer term consequences in the bee. However, blood brain barrier permeability changed with the bee's age which could be explained by the regulation of blood brain barrier transporters during natural transitions in hive task or the presence of hemolymph protein filtration
ContributorsPatel, Aamir S. (Author) / Amdam, Gro (Thesis director) / Harrison, Jon (Committee member) / Ozturk, Cahit (Committee member) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2018-05
Description
Dire wolves have recently risen to fame as a result of the popular television program Game of Thrones, and thus many viewers know dire wolves as the sigil and loyal companions of the Stark house. Far fewer recognize dire wolves by their scientific name, Canis dirus, or understand the population history of this ‘fearsome wolf’ species that roamed the Americas until the megafaunal mass extinction event of the Late Pleistocene. Although numerous studies have examined the species using morphological and geographical methods, thus far their results have been either inconclusive or contradictory. Remaining questions include the relationships dire wolves share with other members of the Canis genus and the internal structure of their populations. Advancements in ancient DNA recovery methods may make it possible to study dire wolf specimens at the molecular level for the first time and may therefore prove useful in clarifying the answers to these questions. Eighteen dire wolf specimens were collected from across the United States and subjected to ancient DNA extraction, library preparation, amplification and purification, bait preparation and capture, and next-generation sequencing. There was an average of 76.9 unique reads and 5.73% coverage when mapped to the Canis familiaris reference genome in ultraconserved regions of the mitochondrial genome. The results indicate that endogenous ancient DNA was not successfully recovered and perhaps ancient DNA recovery methods have not advanced to the point of retrieving informative amounts of DNA from particularly old, thermally degraded specimens. Nevertheless, the ever-changing nature of ancient DNA research makes it vital to continually test the limitations of the field and suggests that ancient DNA recovery methods will prove useful in illuminating dire wolf population history at some point in the future.
ContributorsSkerry, Katherine Marie (Author) / Stone, Anne (Thesis director) / Amdam, Gro (Committee member) / Larson, Greger (Committee member) / School of Human Evolution and Social Change (Contributor) / School of Nutrition and Health Promotion (Contributor) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2018-05
Description
Nicotine addiction remains a prevalent public health issue, and the FDA has released a statement outlining the systematic reduction of nicotine to non-zero levels in the coming years. Current research has not yet established the effects of abrupt nicotine dose reduction on vulnerability to relapse, nor has abrupt nicotine dose reduction been evaluated in terms of behavioral economic characteristics of demand and elasticity been evaluated for reduced doses of nicotine. Using a rat model, we first evaluated the comparability of between- and within-session protocols for establishing characteristics of demand and elasticity for nicotine to shorten experimental timelines for this study and future studies. We then tested environmental enrichment and sex as factors of elasticity of demand for nicotine. Using a rat model of relapse to cues, we also examined the effects of nicotine dose-reduction on vulnerability to relapse. We found differences in maximum consumption and demand between the between- and within-session protocols, as well as sex differences in elasticity of demand on the within-session protocol where male demand was more elastic than female demand. Additionally, we found that enrichment significantly increased elasticity of demand for nicotine for both males and females. Finally, preliminary analyses revealed that nicotine dose reduction yields more inelastic demand and higher maximum consumption, and these outcomes predict increased time to extinction of the association between nicotine and contingent cues, and increased rates of relapse. These studies highlight the usefulness and validity of within-session protocols, and also illustrate the necessity for rigorous testing of forced dose reduction on nicotine vulnerability.
ContributorsCabrera-Brown, Gabriella Paula (Author) / Gipson-Reichardt, Cassandra (Thesis director) / Olive, M. Foster (Committee member) / Davis, Mary (Committee member) / Sanford School of Social and Family Dynamics (Contributor) / Department of Psychology (Contributor) / Barrett, The Honors College (Contributor)
Created2017-12
Description
Biomarkers are the cornerstone of modern-day medicine. They are defined as any biological substance in or outside the body that gives insight to the body's condition. Doctors and researchers can measure specific biomarkers to diagnose and treat patients, such as the concentration of hemoglobin Alc and its connection to diabetes. There are a variety of methods, or assays, to detect biomarkers, but the most common assay is enzyme-linked immunosorbent assay (ELISA). A new-generation assay termed mass spectrometric immunoassay (MSIA) can measure proteoforms, the different chemical variations of proteins, and their relative abundance. ELISA on the other hand measures the overall concentration of protein in the sample. Measuring each of the proteoforms of a protein is important because only one or two variations could be biologically significant and/or cause diseases. However, running MSIA is expensive. For this reason, an alternative plate-based MSIA technique was tested for its ability to detect the proteoforms of a protein called apolipoprotein C-III (ApoC-III). This technique combines the protein capturing procedure of ELISA to isolate the protein with detection in a mass spectrometer. A larger amount of ApoC-III present in the body indicates a considerable risk for coronary heart disease. The precision of the assay is determined on the coefficient of variation (CV). A CV value is the ratio of standard deviation in relation to the mean, represented as a percentage. The smaller the percentage, the less variation the assay has, and therefore the more ability it has to detect subtle changes in the biomarker. An accepted CV would be less than 10% for single-day tests (intra-day) and less than 15% for multi-day tests (inter-day). The plate-based MSIA was started by first coating a 96-well round bottom plate with 2.5 micrograms of ApoC-III antibody. Next, a series of steps were conducted: a buffer wash, then the sample incubation, followed by another buffer wash and two consecutive water washes. After the final wash, the wells were filled with a MALDI matrix, then spotted onto a gold plate to dry. The dry gold target was then placed into a MALDI-TOF mass spectrometer to produce mass spectra for each spot. The mass spectra were calibrated and the area underneath each of the four peaks representing the ApoC-III proteoforms was exported as an Excel file. The intra-day CV values were found by dividing the standard deviation by the average relative abundance of each peak. After repeating the same procedure for three more days, the inter-day CVs were found using the same method. After completing the experiment, the CV values were all within the acceptable guidelines. Therefore, the plate-based MSIA is a viable alternative for finding proteoforms than the more expensive MSIA tips. To further validate this, additional tests will need to be conducted with different proteins and number of samples to determine assay flexibility.
ContributorsTieu, Luc (Author) / Borges, Chad (Thesis director) / Nedelkov, Dobrin (Committee member) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2017-12
Description
Brown adipose tissue (BAT) is thought to be important in combating obesity as it can expend energy in the form of heat, e.g. thermogenesis. The goal of this study was to study the effect of injected norepinephrine (NE) on the activation of BAT in rats that were fed a high fat diet (HFD). A dose of 0.25 mg/kg NE was used to elicit a temperature response that was measured using transponders inserted subcutaneously over the BAT and lower back and intraperitoneally to measure the core temperature. The results found that the thermic effect of the BAT increased after the transition from low fat diet to a high fat diet (LFD) yet, after prolonged exposure to the HFD, the effects resembled levels found with the LFD. This suggests that while a HFD may stimulate the effect of BAT, long term exposure may have adverse effects on BAT activity. This may be due to internal factors that will need to be examined further.
ContributorsSion, Paul William (Author) / Herman, Richard (Thesis director) / Borges, Chad (Committee member) / School of Molecular Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2017-05
Description
Vitellogenin (vg) is a precursor protein of egg yolk in honeybees, but it is also known to have immunological functions. The purpose of this experiment was to determine the effect of vg on the viral load of deformed wing virus (DWV) in worker honey bees (Apis mellifera). I hypothesized that a reduction in vg expression would lead to an increase in the viral load. I collected 180 worker bees and split them into four groups: half the bees were subjected to a vg gene knockdown by injections of double stranded vg RNA, and the rest were injected with green fluorescent protein (gfp) double stranded RNA. Half of each group was thereafter injected with DWV, and half given a sham injection. The rate of mortality in all four groups was higher than expected, leaving only 17 bees total. I dissected these bees' fat bodies and extracted their RNA to test for vg and DWV. PCR results showed that, out of the small group of remaining bees, the levels of vg were not statistically different. Furthermore, both groups of virus-injected bees showed similar viral loads. Because of the high mortality rate bees and the lack of differing levels of vg transcript between experimental and control groups, I could not draw conclusions from these results. The high mortality could be caused by several factors: temperature-induced stress, repeated stress from the two injections, and stress from viral infection. In addition, it is possible that the vg dsRNA batch I used was faulty. This thesis exemplifies that information cannot safely be extracted when loss of sampling units result in a small datasets that do not represent the original sampling population.
ContributorsCrable, Emma Lewis (Author) / Amdam, Gro (Thesis director) / Wang, Ying (Committee member) / Dahan, Romain (Committee member) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2017-12
Description
Almost every form of cancer deregulates the expression and activity of anabolic glycosyltransferase (GT) enzymes, which incorporate particular monosaccharides in a donor acceptor as well as linkage- and anomer-specific manner to assemble complex and diverse glycans that significantly affect numerous cellular events, including tumorigenesis and metastasis. Because glycosylation is not template-driven, GT deregulation yields heterogeneous arrays of aberrant intact glycan products, some in undetectable quantities in clinical bio-fluids (e.g., blood plasma). Numerous glycan features (e.g., 6 sialylation, β-1,6-branching, and core fucosylation) stem from approximately 25 glycan “nodes:” unique linkage specific monosaccharides at particular glycan branch points that collectively confer distinguishing features upon glycan products. For each node, changes in normalized abundance (Figure 1) may serve as nearly 1:1 surrogate measure of activity for culpable GTs and may correlate with particular stages of carcinogenesis. Complementary to traditional top down glycomics, the novel bottom-up technique applied herein condenses each glycan node and feature into a single analytical signal, quantified by two GC-MS instruments: GCT (time-of-flight analyzer) and GCMSD (transmission quadrupole analyzers). Bottom-up analysis of stage 3 and 4 breast cancer cases revealed better overall precision for GCMSD yet comparable clinical performance of both GC MS instruments and identified two downregulated glycan nodes as excellent breast cancer biomarker candidates: t-Gal and 4,6-GlcNAc (ROC AUC ≈ 0.80, p < 0.05). Resulting from the activity of multiple GTs, t-Gal had the highest ROC AUC (0.88) and lowest ROC p‑value (0.001) among all analyzed nodes. Representing core-fucosylation, glycan node 4,6-GlcNAc is a nearly 1:1 molecular surrogate for the activity of α-(1,6)-fucosyltransferase—a potential target for cancer therapy. To validate these results, future projects can analyze larger sample sets, find correlations between breast cancer stage and changes in t-Gal and 4,6-GlcNAc levels, gauge the specificity of these nodes for breast cancer and their potential role in other cancer types, and develop clinical tests for reliable breast cancer diagnosis and treatment monitoring based on t-Gal and 4,6-GlcNAc.
ContributorsZaare, Sahba (Author) / Borges, Chad (Thesis director) / LaBaer, Joshua (Committee member) / School of Molecular Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2016-05