Individuals from stressful environments, such as those who grew up with low socioeconomic status (SES), exhibit an inflammatory response to stress which reflects a physiological present focus. Despite the negative long-term effects of elevated inflammation, the bodies of people with low childhood SES favor immediate survival under stressful conditions, at the expense of long-term health. However, less is known about whether people from low SES childhood environments also exhibit a psychological present focus in response to stress. The current research was designed to experimentally examine the impact of stress exposure on the impulsive decision making of undergraduates from various economic backgrounds. Participants completed baseline measurements of impulsivity, including their self-reported ability to delay gratification and sense of self-control. Next, participants were randomly assigned to either the stress or control condition of the Trier Social Stress Test, before completing the impulsivity measurements again and providing information about their childhood environment and demographics Psychological present focus was measured by comparing baseline and post-stress levels of impulsivity. Preliminary results indicate that stress exposure does not impact the psychological present-focus of people with relatively high childhood SES. However, results trended such that people with relatively low childhood SES reported a reduced ability to delay gratification and lower self-control after experiencing stress, compared to control. These patterns suggest a psychological present-focus in response to stress among those from stressful early life environments. However, results are preliminary and data collection is still ongoing.

Social connection, and the protection it provides, was a determining factor for our ancient ancestors’ ability to achieve their evolutionary goals of survival and reproduction. In turn, social exclusion, the loss of this connection, posed a serious threat to these goals, spurring the development of several cognitive and behavioral recovery strategies to combat its harmful effects. One potential strategy for women following experiences of social exclusion is increased motivation to engage in short-term sexual relationships that may quickly alleviate the safety and affiliative concerns associated with social exclusion. However, the relationship between social exclusion and women’s short-term mating (STM) motives remains relatively unexamined. The present research investigated the influence of social exclusion on women’s STM motives, and how individual differences in chronic concerns about exclusion influence this relationship. I predicted that being socially excluded, compared to included, would lead women to have increased STM motives. Furthermore, I predicted that individuals’ differences in chronic concerns about exclusion would moderate this relationship. To test these hypotheses, I primed feelings of social exclusion and inclusion using the future alone paradigm, and then measured several dimensions of unpartnered women’s self-reported STM motives (including sexual unrestrictedness, openness to sexual intercourse, and desired mate investment). Results did not support the hypothesized relationship. Instead, they indicated that excluded women exhibited lower STM motives, specifically less sexual unrestrictedness, than included women. However, this relationship was moderated by chronic exclusion concerns, such that, for socially excluded women, the more chronically concerned with social exclusion they were, the greater their expressed sexual unrestrictedness. For social included women, their trait exclusion concerns were unrelated to their expressed sexual unrestrictedness. Thus, for women that have chronically high exclusion concerns, increased sexual unrestrictedness following social exclusion may be a compensatory mechanism to mitigate the negative effects of being excluded. The implications of these findings for women’s interpersonal and intimate relationships will be discussed.

Terror management theory suggests that the potential for anxiety from the awareness of death can be buffered by a cultural worldview. Mind-body dualism, the belief that the mind and the body are separate, might affect people’s mortality concerns. Given that the body is threatening given its vulnerability to death, individuals who perceive the mind and body as being connected (vs. separate) should experience higher mortality-related thoughts and defense of their cultural beliefs. Past research found that mind-body dualism was related to afterlife belief, which was able to buffer existential concerns (Heflick et al., 2015). Based on these findings, the current research investigated how mind-body dualism moderated the effect of the creaturely body on death-related concerns. The result showed that people who perceived the mind-body relationship as more separate showed significantly fewer death concerns after reading an essay emphasizing the creatureliness of the body, whereas people who held beliefs in a more interrelated mind-body relationship showed heightened death concerns after the creaturely body prime.

Alzheimer’s Disease (AD) is the most common form of dementia that mainly impacts the brain, specifically inducing neuronal cell death in the central nervous system. AD is characterized by the secretion of the protein Tau, and the formation of plaques made up of Beta-amyloid protein. Tau and Beta-amyloid plaques activate the secretion of inflammatory cytokines by microglial cell. The resulting inflammation triggers neuronal cell death, which leads to damage and cognitive decline over time. The cytokines secreted by microglial cells activate the Nf-kB signaling pathway. Activation of Nf-kB results in gene expression and secretion of TNF-a, a cytokine known to be associated with inflammation. This leads to a feedback mechanism that results in greater inflammation.
Our lab has demonstrated that a variety of anti-inflammatory compounds derived from IDT (iso-indolin dithione), targets the Nf-kB pathway by reducing the levels of TNF-a at the protein/translational level. BV-2 cells, a mouse microglial cell line were used in this study. Inflammation was stimulated by exposing these cells to LPS to trigger the activation of the Nf-kB signaling pathway. We hypothesize that the drugs tested reduce levels of TNF-a secreted by BV-2 mouse microglial cells, and therefore, block the development of disease-associated CNS inflammation seen in Alzheimer’s disease

Bacillus anthracis is a gram-positive bacterial pathogen that causes the deadly infectious disease anthrax. Bacillus anthracis contains two plasmids, pX01, and pX02. These plasmids were found to be necessary for the virulence of B. anthracis. However, Bacillus anthracis contains over 5,000 chromosomal genes and we believe that there are additional virulence genes that have yet to be discovered. Our lab constructed a transposon mutant library with random disruptions in the B. anthracis Sterne genome to screen for novel virulence factors. This library has been successfully used to identify the chromosomal genes clpX and yceGH and show their importance for B. anthracis virulence. To find additional novel virulence genes, we used the same transposon library and screened around 1,000 mutants using hydrogen peroxide, a reactive oxygen species (ROS). ROS are involved in the immune defense and the mutants that are attenuated in its presence may have a disrupted gene that contributes to the pathogenicity of B. anthracis. We obtained two mutants that were repeatedly susceptible to hydrogen peroxide in vitro. To determine the virulence of these mutants in an animal model, we will be performing an in vivo assay using the waxworm, Galleria Mellonella. Mutants that have reduced virulence in G. mellonella will be further tested to determine the location of the transposon in the genome to find out which genes are disrupted. The findings of this research could be used as potential therapeutic drug targets and could offer insight into the mechanisms that B. anthracis uses for its pathogenesis.

Reintroductions have become increasingly common to help restore populations of Texas horned lizards (Phrynosoma cornutum). Reintroduction success of any species can be shaped by many factors including genetics, selection of suitable reintroduction sites, etc. Our primary goal has been to determine whether release techniques- specifically site selection and release method- contribute to the reintroduction success of captive-bred hatchling Texas horned lizards. In 2020 and 2021, we reintroduced over 500 captive-bred hatchling Texas horned lizards from the Ft. Worth, Dallas, and Caldwell Zoos to Mason Mountain WMA (Mason County, TX). Lizards were randomly assigned to one of two release sites and were placed either in clumps of 20+ lizards (Site 2 2020 & Site 1 2021) or were dispersed 5 m from one another (Site 1 2020 & Site 2 2021) at release. We used harmonic radar to track lizards and monitor survivorship outcomes and growth rates from release (September or October) until most lizards began brumating in early December. We found that survival outcomes were associated with both release site (χ22, 509 = 34.5, p<0.0001) and release method (χ22, 509 =15.09, p=0.005). We achieved the highest survivorship (26.4%) when lizards were dispersed at Site 1. Preliminary dietary and prey availability assessments suggest that survivorship differences between locations may be related to differences in food availability. Our findings suggest that future reintroduction attempts may have higher success rates if 1) sites are selected that meet the specific resource requirements of hatchlings, and 2) lizards are dispersed from one another at release.

Mercury (Hg) is released by coal-burning power plants and artisanal gold mines into the atmosphere. Mercury deposited from the atmosphere into aquatic ecosystems can be converted into a neurotoxic form, methyl mercury (MeHg). In aquatic ecosystems, Hg biomagnifies which can affect organisms feeding at higher trophic positions. However, monitoring Hg in the environment to assess the risk to biota is not straightforward. For example, measuring Hg in sediment may not reflect the amount of Hg bioaccumulating in organisms. This difference in the Hg in the sediment and the amount bioaccumulating is partly determined by the bioavailability of Hg. Measuring bioavailability is very complex so sentinels can be used to detect Hg in food chains. Sentinels are defined as organisms that accumulate Hg within their tissues without significant adverse effects. Riparian spiders have been proposed as sentinels. Riparian spiders eat emerging aquatic insects which transport Hg from aquatic ecosystems and because of this, riparian spiders are sentinels of aquatic Hg contamination. The objective of this study was to measure Hg levels in wolf spiders (family: Lycosidae). From May 2021- June 2021, spiders were collected at four sites, two on the Clear Fork and two on the West Fork of the Trinity River, and the spiders were preserved in 95% ethanol. The spiders were sorted based on sex and their body size was measured. Hg will be analyzed using direct mercury analysis. This study attempts to determine the effect of sex and body size on the amount of Hg in wolf spiders, factors that have not been assessed previously. It also attempts to detect differences in the concentration of Hg between the Clear Fork and the West Fork of the Trinity River.

In the pathogenesis of neurodegenerative inflammatory diseases, such as Alzheimer’s disease, there is an abnormal buildup of redox metal ions that associate with β-amyloid plaques and convert oxygen into oxygen radicals. These radicals are highly reactive with cellular components and lead to oxidative stress that induces damage and death of neuronal cells which is associated with the cognitive decline of Alzheimer’s disease. Bifunctional macrocyclic compounds with antioxidant properties are a promising potential therapeutic to reduce levels of reactive oxygen species (ROS) and increase neuronal cell survival via the ability to chelate dysregulated metal ions and radical scavenging. In this project, novel macrocyclic compounds were tested for their efficacy in reducing intracellular levels of H2O2-induced ROS and H2O2-induced cytotoxicity. Intracellular ROS levels and cell survival were quantified in FRDA and BV-2 cells using the DCFH-DA and MTT cytotoxicity assays.

Alzheimer’s disease (AD) is often associated with chronic inflammation and cognitive dysfunction. In studying how AD-like pathologies change and affect learning and memory, our lab aims to optimize an object location memory (OLM) testing paradigm in mice. Briefly, a mouse is placed into an arena with two identical objects for a training session. Four hours later, one of the objects is moved to a novel location, and the mouse is placed back into the arena for the testing session. Because mice exhibit a preference for novelty, memory is assessed as the amount of time the mouse spends exploring the moved object divided by the total time spent exploring both objects. Our goal is to identify testing parameters that make this task both accurate and efficient for our lab’s use, as we will add this learning paradigm to a battery of behavioral tests to be used in future experiments. In the current study, the OLM protocol will be performed twice according to two different experimental timelines that test the effects of adding an additional training session to the original protocol.

BRCA1 is a gene found in humans that, when mutated, has been linked to breast and ovarian cancer. A homolog version of this gene, known as brc-1, exists in an organism called the Caenorhabditis elegans. This is a species of nematode worm that has the potential to be used as a model organism to study this homolog gene that is associated with human breast cancer. Previous studies with C. elegans have shown links between the brc-1 gene and DNA damage responses, cytochrome p450, or cyp, transcription levels, and ratios of male phenotype worms. This project focused on studying whether these brc-1 functions are dictated by the enzymatic activity of the protein made by this gene. To measure these phenotypes, we used a strain of C. elegans with a brc-1 mutation engineered to lack enzymatic activity of the BRCA1 protein toward nucleosomes. In order to determine how this lack of enzymatic activity affects brc-1 functions, we measured levels of reactive oxygen species (serving as a proxy for DNA damage), numbers of male offspring, and cyp levels in the mutant and wild-type C. elegans. Our initial results indicate the effects of enzymatic activity towards nucleosomes on the aforementioned phenotypes.

Anthrax is an infectious disease caused by Bacillus anthracis, which is a spore forming bacterium. Even though the anthrax toxins and capsule, encoded on 2 plasmids pXO1 and pXO2, play crucial role in the pathogenesis of anthrax infection, evidence suggests that chromosomal genes also play a role. The ClpX ATPase was discovered to be crucial for B. anthracis virulence via protection against host antimicrobial peptides. In this study, we want to investigate the role of clpX in regulation of other stressors including acidic stress, temperature stress, salt stress, and non-cell envelope active antibiotics. We found that clpX is necessary for survival in an acidic environment and growth under heat stress. We demonstrate that acidic stress resistance is mediated by the formation of the ClpXP protease using a ClpX complementation plasmid that is incapable of interacting with ClpP. There is no association between clpX with other stressors. We conclude that the ClpX is required for B. anthracis pathogenicity via defenses against host antimicrobial peptides and for survival in an acidic environment. Understanding the role ClpX in the regulation of stress responses will ultimately infer us with new target for either directly combating infection or improving the efficacy of already available medicines.

Impacts of Pollen-Donor Distance and Nutrient Availability on
Reproductive Success in a Carnivorous Plant

Halia Eastburn and John Horner

The maintenance of genetic diversity has important consequences for the survival of plant populations. Because plants are sessile, the distance between plants is often inversely correlated with relatedness. Therefore, the distance between pollen-donor and recipient can determine the level of inbreeding or outbreeding. Both pollen-donor distance and nutrient availability can affect reproductive success in populations of flowering plants. Populations of the carnivorous plant Sarracenia alata have dwindled and become extremely fragmented due to human development and agriculture. The purpose of this study was to examine the effects of pollen-donor distance and prey capture on reproductive success in S. alata. We hand-pollinated flowers with pollen from varying distances [0 m (self-pollinated) and 35, 60, 90, 125, and 190 m], and we prevented prey capture in half of our study plants. We measured seed production and germination to estimate reproductive success. Pollen-donors from greater distances sired a greater number of seeds but pollen-donor distance did not affect germinability. There was no effect of prey capture alone nor an interaction of pollen-donor and prey capture on seed production or germination. More research is needed to understand nutrient allocation for reproduction over multiple years and natural variance in prey capture which might affect reproductive output in subsequent seasons.

Our project focused on the conservation of activity of the protein BRD1 in C. elegans. C. elegans is a strong model organism for our study because BRD-1 is the worm ortholog to BARD1 in humans. Specifically, our focus is on its function as an enzyme to attach ubiquitin to the H2A tail of nucleosomes. We studied a structural mutation of BRD-1 that we predicted would interfere with its ability to bind its substrate, the nucleosome. We hypothesized that BRD-1 is bound to the nucleosome at this mutation site based on prior research in the human protein. Therefore, we integrated mutations found in humans into the DNA that codes for C. elegans BRD-1. A typical mutagenesis protocol was used to implement the mutations and then we expressed the proteins in E. coli cells. After that, nucleosomes were reconstituted by dialysis, and enzyme activity was assessed using a ubiquitination assay. These assays showed that BRD-1 in C. elegans does bind the nucleosome demonstrating conservation of the BARD1 function. Determining that function is conserved allowed us to determine that C. elegans is an appropriate organism to test mutations found in humans. This research has future clinical potential due to the ability to test mutations encountered in humans using a model organism and can aid with clinical treatment plans to help avoid the development of cancer.

Many anti-inflammatory drugs are currently in use to treat neuroinflammation in the brain which can result from Alzheimer's disease, Parkinson's disease, traumatic brain injury, and more. In collaboration with a company, P2Dbiosciences, we are testing drugs that can modulate the function of inflammatory cytokines such as TNF-alpha, with the goal of reducing neuroinflammation and thus benefiting people suffering from the neurodegeneration and cognitive decline associated with neuroinflammation. We hypothesize these drugs work by inhibiting the signaling associated with inflammatory cytokines.
Two different assays were developed to identify the mechanism of action of these cytokine modulating anti-inflammatory drugs. BV2 cells in culture were used for these assays to model how the drug affects mouse microglial cells (immune cells resident in the brain). The first assay uses a luciferase reporter gene to determine if NF-kB promoter activity is disrupted when cells are treated with drug. The second assay uses quantitative RT-PCR (qPCR) to measure changes in TNF-alpha mRNA levels when cells are treated with drug. Levels of TNF-alpha mRNA were also quantified over a period of time following drug treatment to determine whether the degradation time of the TNF-alpha mRNA was affected by treatment.

Invasive species, such as the Red Imported Fire Ant (Solenopsis invicta, hereafter, RIFA), can negatively impact native species via predation and modifying prey behavior. RIFA exist in two colony types, monogyne (single queen) and polygyne (multiple queens), and polygyne colonies are known to contain higher densities of fire ants than monogyne colonies. Texas horned lizard (Phrynosoma cornutum) eggs and hatchlings are suspected prey of RIFA’s foraging and aggressive behaviors. In this study, we collected fire ants from Karnes City and Kenedy to determine if Texas horned lizard density is lower around polygyne colonies. We collected and sequenced 30 ants, of which 20 were RIFA. Counter to our expectations, there seemed to be no correlation between RIFA colony type and Texas horned lizard density. Furthermore, we found evidence that monogyne and polgyne colonies were coexisting. In future studies, we think larger sampling sizes and determining ratios of polygyne to monogyne colonies within the same area would be useful for further testing the hypothesis that colony type may affect horned lizard density.

Emerging evidence suggests that the immune system is vulnerable to disruption in response to a wide variety of chemical contaminants; thus, there is a need to test chemicals for immunotoxicity. To understand how chemicals impact the ability of the immune system to ward off infection, a model system featuring fathead minnows (FHM, a common toxicological model) infected with Yersinia ruckeri (a bacteria) has been used. Using this model system, the impacts of chemicals on immune system function can be evaluated via pathogen resistance challenges, where a fish is infected with a pathogen and their ability to defend against the pathogen and survive infection is determined. However, the use of Y. ruckeri is unfavorable given that fish must be injected with Y. ruckeri to develop an infection, a process that is time consuming and inconsistent with natural routes of exposure. Thus, the goal of this project was to develop a new host-pathogen system for FHMs by identifying a pathogen that induces infection via immersion. To do this, the ability of three bacterial pathogens, Flavobacterium columnare, Aeromonas sobria and Aeromonas allosaccharophila, to infect FHMs via immersion was evaluated. Results of this study revealed that of the three bacterium evaluated, only F. columnare was capable of inducing an infection via immersion in FHMs and infection was only successful following a fin-clip procedure, in which a small portion of the caudal fin is removed. Overall, this result establishes the potential for a FHM-F. columnare model system for future use in immunotoxicity testing.

Nickel is the most prominent heavy metal in the effluents associated with crude oil extraction and production. Given that these effluents are released into the ocean, investigating the toxicity of nickel on marine life is pertinent. A current method for evaluating the toxicity of oil effluents is the sheepshead minnow larval growth and survival (LGS) test, which exposes larval fish to varying concentrations effluents or associated single chemicals over a 7-day period. However, current legislation, like the Frank E. Lautenberg Act, requires that animal testing be refined whenever possible to enhance animal welfare. The fish embryo toxicity (FET) test, which investigates chemical toxicity using fish embryos over a 7-day exposure period, is a potential alternative method capable of meeting legislative needs related to animal welfare. The objective of this study was to determine if a sheepshead minnow FET test is a viable replacement for the sheepshead minnow LGS test. To accomplish this, the results of sheepshead minnow LGS and FET tests using nickel were compared. The results of this study show that the LGS test is more sensitive than the FET test. In addition, evidence suggests that it may be possible to improve the sensitivity of the FET test by including sublethal metrics as FET test endpoints.

Bacillus anthracis is a bacterial pathogen that causes the often lethal disease anthrax. This research aims to characterize the role of potential virulence genes in Bacillus anthracis. Virulence is a pathogen’s ability to damage the host. Studying virulence allows us to understand infection mechanisms and develop novel ways to target pathogens. Previous work identified a collection of potential virulence mutants (Franks et al, 2014) each containing a genetic disruption that renders a gene non-functional. These mutants were pulled out in initial screenings but were never characterized further. We confirmed that one mutant, TN2, also exhibits decreased virulence in a Galleria mellonella survival assay. We know that TN2 has a disruption in a promoter region that we hypothesize controls two genes: a putative BNR repeat domain protein (TN2A) and a glycosyl-like 2 transferase family protein (TN2B). For my project, I attempted insertional mutagenesis to inactivate these genes with the goal of confirming that the genes are linked to virulence, rather than unintended mutations elsewhere in the genome. After successfully creating insertional mutant 2B, through the disruption of the TN2B gene, I am working to further characterize the mutant to determine its role in immune evasion. Specifically, I will compare the ability of the wild-type and mutants to survive exposure to various antimicrobial defenses conserved in humans and waxworms. This research could help identify a novel bacterial virulence factor and its potential mechanisms of action thus expanding our understanding of bacterial pathogenesis.

The gram-positive bacterium, Bacillus anthracis, is responsible for the deadly disease Anthrax. B. anthracis is dangerous due to virulence factors, or defenses the bacteria uses to infect a host. We hope to better understand how this bacterium interacts with its hosts by studying the genes necessary for virulence. Bacterial mutants, which have a change in their genetic sequence, sometimes show reduced ability to cause disease in a host. Studying these mutants helps us understand the bacteria’s infection method. Previously our lab created a library of mutants using a technique called transposon mutagenesis and then screened these transposon mutants for phenotypes linked to decreased virulence. This resulted in the identification of 11 transposon mutants that were less effective at causing disease in the nematode Caenorhabditis elegans (Franks et al.). While all 11 mutants could be interesting for further characterization, it is necessary to prioritize them as this is still too many to study. In this project, we tested these mutants using a second infection model, the caterpillar Galleria mellonella. G. mellonella is an ideal model due to its optimal size for injection, conserved innate immune defenses, and previous success as an infection model for B. anthracis (Malmquist et al.). We found that only one of these 11 mutants, TN2, had reduced virulence in both C. elegans and G. mellonella. Future research will focus on confirming the genetic change in this mutant and determining the mechanism by which it contributes to infection. This could lead to new antibiotic targets in the future.

Mercury (Hg) is released into the environment by coal-burning powerplants and artisanal gold mines. Aquatic bacteria then convert the inorganic mercury into highly toxic methyl mercury. Turtles acquire mercury through their diet, and it bioaccumulates throughout their long lifetime. Toenail clippings can be used to determine Hg concentrations in turtles. Toenail samples were collected from Trachemys scripta elegans (red-eared sliders) in the Brazos River near Granbury and the Clear Fork of the Trinity River as it flows through Fort Worth. All toenails were dried in a 60℃ oven and put into a direct Hg analyzer that uses thermal decomposition, gold amalgamation, and atomic absorption spectrometry to determine total Hg. Toenails from the Brazos river had significantly more Hg on average than those in the Clear Fork, 658.302µg/kg and 400.146µg/kg respectively. The results were unexpected as the Brazos river near Granbury is considered less polluted than the Clear Fork of the Trinity, which is supported by observations of insect larvae such as hellgrammites which were common in the Brazos but absent in the Clear Fork of the Trinity. Our hypothesis is that red-eared sliders in the Brazos are feeding at a higher trophic level than those in the Clear Fork. Fecal samples and a lack of invertebrate prey in the Clear Fork suggest red-eared sliders primarily feed on algae. In the Brazos river we observed several species of insect larvae underneath rocks and hypothesize that the red-eared sliders are feeding on this abundant food source. Mercury is known to biomagnify and therefore red-eared sliders in the Brazos are likely ingesting more mercury than those in the Clear Fork.

Alzheimer’s Disease (AD) and Traumatic Brain Injuries (TBI) are global societal problems affecting millions of people and costing billions of dollars per year.1,6 Hallmarks of AD include memory loss, cognitive decline, depression, and confusion due to unchecked inflammation in the brain caused by the overproduction of pro-inflammatory cytokines by the immune system.1,9,11 TBI occurs when a sudden trauma damages brain cells, which activate the immune response potentially leading to chronic inflammation and a multitude of symptoms affecting cognitive, somatic, and emotional processes.3,4,12 There is currently no cure for AD, nor is there an effective treatment for chronic inflammation caused by TBI. P2D Bioscience® has manufactured a series of drugs successfully targeting inflammation in a 3XTgAD mouse model.21 To understand the cellular mechanism of the novel drugs, we used SDS-PAGE electrophoresis and Western Blot analysis to investigate protein levels within the NFB pathway, which modulates inflammation. We monitored the inhibitor of NFb, IB, to determine whether the drugs were blocking the phosphorylation and degradation of IkBa and subsequently blocking the pro-inflammatory effects of activated NFB. We show that the drug is blocking the degradation of IB, and therefore the pro-inflammatory genes associated with the NFB pathway are not being transcribed. Increasing our understanding of the cellular mechanism of action is imperative for the progression of drug development because it can be used to evaluate potential side effects.

Alzheimer’s Disease (AD) is a neurodegenerative disease that primarily affects elderly populations. AD engenders memory loss and cognitive decline, and its prevalence is rapidly growing. It is estimated that 14 million Americans will have AD by the year 2050. Therefore, it is imperative for researchers to examine the underlying biological mechanisms responsible for AD. Previous research has demonstrated that chronic inflammation is linked to the hallmark AD pathology, amyloid beta (Aβ). Aβ is a protein that disrupts neuronal communication and increases the production of effector proteins called pro-inflammatory cytokines. Microglia function like immune cells in the brain, and when they are activated by inflammatory triggers, such as Aβ, they secrete pro-inflammatory cytokines. Although cytokine release is initially a healthy response, excess cytokine production is harmful to the brain and exacerbates AD pathologies. Prior research has demonstrated that pro-inflammatory cytokines are upregulated in the serum of AD patients. Therefore, cytokines are a crucial target for AD therapeutics.

The current experiment will examine the temporal inflammatory response of microglial cells following lipopolysaccharide (LPS) insult. LPS is a component of common bacteria and can induce inflammation in microglial cells. We will treat cells with several different concentrations of LPS and assess cytokine production at several different timepoints. To do this, we will collect cell supernatant (secretions) and measure multiple cytokines using an ultrasensitive electrochemiluminscent assay. Data collected from these experiments will be used in many future studies of potential therapeutics and dietary supplements. In fact, data from these experiments will be used by current and future departmental honors students to determine the optimal treatments and times for their experiments. This project is incredibly relevant because AD is currently the 6th leading cause of death in the United States. Data collected will help us pinpoint proper testing procedures for therapeutic compounds that are developed.

Cells use diverse mechanisms to prevent DNA damage and tumor formation. Two tumor suppressors employed in this effort are the focus of our study: breast cancer type 1 susceptibility protein (BRCA1) and BRCA-1-associated RING Domain protein 1 (BARD1). These two proteins form a complex that suppresses the generation of estrogen-derived free radicals. Inherited mutations in BRCA1 or BARD1 are associated with an increased risk of developing breast or ovarian cancer in humans. The model organism Caenorhabditis elegans possesses the orthologs BRC-1 and BRD-1 which can be readily mutated, offering an attractive model to study biochemical functions. However, it is unknown if BRC-1/BRD-1 also regulates the transcription of estrogen metabolism (cyp) genes to control the production of free radicals as noted for the human homologs. Utilizing gene expression analysis and estrogen exposure assays, this study demonstrates that BRC-1/BRD-1 has a conserved function of regulating cyp genes in C. elegans. However, our data also shows that BRC-1 and BRD-1 do not necessarily protect DNA from free radical damage upon estrogen exposure, despite its proven inhibition of cyp genes expression. Further investigation is required to determine the function of these cyp gene homologs in C. elegans. Our findings of this additional conserved function of the BRCA1/BARD1 homologs in C. elegans further validate its use as a model organism to better understand the myriad ways BRCA1/BARD1 protects the genome.

BRCA1 and p53 have been proven to interact in tumor suppressor pathways for hereditary breast and ovarian cancer. Finding the physical binding location associated with this interplay is important in assessing cancer risk and determining molecular details of the interaction. This project aimed to identify the protein binding region of p53 with the intrinsically disordered region of BRCA1. We cloned select regions of human BRCA1 and p53 protein into E. coli bacteria, then harvested and purified the proteins. A pull-down assay was performed to test binding affinity between a segment of p53 and two different length BRCA1 constructs. The assay showed that neither the construct that contained BRCA1 amino acids between 772-1126 nor the construct with amino acids between 896-1190 interacted with p53. This indicates that these amino acids alone are not sufficient for binding of p53 and BRCA1. Our results could also indicate that a third-party binding mediator is required in vivo. This information expands upon our knowledge of the p53 and BRCA1 binding interaction and can be used in a clinical setting to evaluate risk associated with mutations in the experimental regions.

Alzheimer’s Disease (AD) is a progressive neurodegenerative disease associated with old age and marked by deficits in memory and learning skills. AD pathology is characterized by amyloid-beta (AB) accumulation, which leads to plaque formation and ultimately neuronal death. Additionally, AB activates microglial cells, which function as an immune cell in the brain. Microglial cells secrete proteins that induce inflammation, known as pro-inflammatory cytokines. The chronic activation of pro-inflammatory cytokines engenders neuroinflammation and oxidative stress, which then further exacerbates AD pathologies. This project aims to study the effectiveness of cannabidiol (CBD) as a potential treatment for AD, due to its known anti-inflammatory properties. We will measure the inflammatory response of cultured BV2 immortalized mouse microglial cells following lipopolysaccharide (LPS) treatment. We will then include a CBD treatment to study its therapeutic capabilities in reducing inflammation. We hypothesize that treatment with CBD will decrease the pro-inflammatory cytokines TNF-alpha and IL-6 induced by LPS stimulation. We will perform enzyme-linked immunosorbent assays (ELISAs) to detect and quantify the cytokine levels.The overall goal of the research is to demonstrate the capacity of CBD to minimize the immunological mechanisms that drive AD pathologies. Our research will contribute to the understanding of the link between the immune system and central nervous system in AD development. AD is the sixth leading cause of death in America, but the availability of therapies is limited. CBD represents a natural and possible effective therapy for those suffering from Alzheimer’s disease, and our research will contribute to determining its efficacy.