Hypoxic-Ischemic Encephalopathy (HIE) is a brain injury that commonly causes mortality in neonates. Current treatment consists of therapeutic hypothermia, but close to 50% of affected infants still have a poor outcome (death or severe disability). In order to discover new effective therapies, it is important to compare how different treatments affect the brain in animal studies. The research laboratory has developed a ferret model of HIE because the ferret brain has more complex gyrification compared to rodents. Animals underwent unilateral carotid ligation at postnatal age 17 days (P17), in which one side of the carotid artery was restricted temporarily and the other was restricted permanently. The animals then received periods of hypoxia and hyperoxia. To better quantify the extent of injury, a system involving measurements of the gyri, sulci, and cerebellar exposure was developed. Ex vivo brain measurements were collected from a population of 63 ferret kits at age P42, and adjusted by the weight and sex of the animal. These measurements included the lengths of: the longitudinal fissure (anterior and posterior), lateral sulci, suprasylvian sulci, coronal sulci, pseudosylvian sulci, ansinate sulci, cruciate sulci, presylvian sulci, lateral gyri, suprasylvian gyri, sigmoid gyri (anterior and posterior), coronal gyri, ectosylvian gyri (anterior and posterior), orbital gyri, and the exposure of the cerebellum. In injured animals, significant changes in the longitudinal fissure, ansinate sulci, left coronal sulci, cruciate sulci, presylvian sulci, posterior sigmoid sulci, and exposure of the cerebellum were seen compared to littermate controls. The implications of this measurement system include the ability to accurately characterize the degree of injury in animals with an hypoxic-ischemic brain injury, which will help to show whether potential treatments are neuroprotective.

Tuberculosis is a major infectious disease with approximately 2 billion people infected worldwide. With the emergence of multi-drug resistant tuberculosis (MDR-Tb), efforts to develop efficient treatments with fewer cytotoxic effects are currently under investigation. Dual host and pathogen directed therapeutics hold promise towards more effective treatments against tuberculosis (Tb) through mechanisms that enhance bacterial clearance and reduce tissue damage and immunopathology. We investigate RGFP966, a drug that modulates the host's immune defenses and directly reduces Mtb growth (a histone deacetylase 3 (HDAC3) inhibitor). The aim of my project is to characterize the effect of RGFP966 on Mtb growth. My hypothesis is that the antimicrobial effects of RGFP966 are specific to Mycobacterial species, and the bacterial target of RGFP966 is required for its survival. Within the experiential growth period, I treated Mycobacterium avium and M. Smegmatis in 7H9 broth with a range of RGFP966 concentrations (0.5 uM, 5 uM, and 50 uM) and incubated in culture tubes and 96-well plates for 3-7 days. Within designated time intervals, I analyzed levels of dose dependent inhibition using optical density, luminometry, and fluorometry methods. Current results through optical density and luminometry demonstrate that RGFP966 controls M. Avium growth in 7H9 broth with an MIC50 of approximately 5 uM. Fluorometry and optical density results indivate no restriction of M. smegmatis growth. Comparing these results to previous findings in our laboratory, where RGFP966 lacked effects against common gram-positive and gram-negative bacteria, indicates possible specificity of the antimicrobial effects of RGFP966 to Mycobacterial species. We will further examine the effects of RGFP966 against mycobacterial growth. A comprehensie understanding of its pathogen directed effects combined with its host directed effects hold promise as an adjunctive therapeutic against Tb infection. 

Bloodborne pathogens are wide spread and can pose risks to health care workers and vulnerable patient populations alike. Conventional diagnostic tests for bloodborne pathogens are costly and time intensive, so fast, affordable, and sensitive diagnostic methods are needed that can be performed under low-resource conditions by untrained personnel. Microfluidic self-digitization technology, developed in the Chiu Laboratory, provides the foundation for such low-cost diagnostics. As undergraduates, we work on the fabrication and optimization of cheap, robust devices used to load samples, as well as dPCR of samples required to detect diseases. As part of a larger research endeavor, we are developing a portable instrument that, in combination with our proprietary microfluidic chip technology, will expand rapid diagnostics to low-resource settings around the world. Expanding diagnostics for bloodborne pathogens will both help prevention as well as monitoring treatment of patients already infected.

Magnetic fields (MFs) and cosmic rays (CRs) are decidely important aspects of galactic disk and halo evolution, however, their precise roles are not yet completely understood. While there are many simulations studying galactic evolution, few have deeply explored the exact impact of CRs and MFs in the evolutionary process. The current goal of our research is to learn more about how CRs and MFs contribute to the evolutionary process by looking at how MFs grow and change in the circumgalactic medium (CGM) under the influences of CRs. Using a suite of simulated, isolated disk galaxies, we investigated the role of CRs in MF growth and galaxy evolution by comparing different galactic models, each possessing slightly varied CR physics. We present the role of CR transport on the geometry, strength, and growth rate of MFs in these simulated galactic halos.

During embryonic development of the vertebrate central nervous system, innate immune cells, called microglia, migrate into the retina and brain. Microglia are responsible for phagocytosis, monitoring tissues for pathogens, and inflammatory signaling. Several studies in model organisms show microglia also have roles in neural development, particularly in formation of synaptic circuitry. Defects of the neuronal circuitry can lead to vision loss, therefore it is probative to investigate the role and mechanisms of microglia in wiring the retina. The purpose of this project is to investigate microglia migration and activity in the human retina, where very little is understood. To understand if microglia could have similar effects in human retinal development as in model organisms, we first needed to determine when and where microglia are found at different ages. We collected donated fetal tissues from as early as developmental day 40 through 132 to determine the number and distribution of microglia in tissue sections using Iba1, a microglia-specific antibody, and confocal fluorescence microscopy. To test whether microglia are necessary for retinal development we are using our lab’s fetal retina culturing technique that can be used to manipulate microglia populations. Retinal development can be visualized after deletion of microglia using markers for retinal progenitor cells (EdU) and various synaptic antibodies. So far, we have seen from whole retinal tissues that microglia are present as early as day 58, prior to the majority of synaptic development, and survive in retinal cultures. Earlier stages are being investigated, and we are currently determining methods that will deplete microglia in order to study the effects of their absence on retinal development. In conclusion, we know that microglia are present during synaptic development, and persist during retinal culturing. Therefore, microglia are potentially essential for human retinal development and could be targets for future disease research.

The foraging patterns and populations of predators can provide a better understanding of the nutrient dynamics in ecosystems. Brown bears (Ursus arctos) are a particularly difficult species to study. However, brown bears are a key predator species to study due to their close relationship with Pacific salmon (Oncorhynchus spp.) populations. Noninvasive methods like hair collection and camera traps are emerging as effective ways to study brown bear populations, diet, and behaviors. This study aims to determine whether the use of these methods in the Wood River system (Aleknagik, AK) are unbiased and characteristic of the resident brown bear population. The data used in this study are part of a long-term study (beginning in 2012) taking place in this area involving brown bear predation on sockeye salmon (Oncorhynchus nerka) and other population studies. Using video footage from the 2016 field season, this study analyzes brown bear reactions and avoidance of the barbed wire used to collect hair samples. Expected results are a higher frequency of wire avoidance in adult bears, single bears, during daylight, and equal frequency of avoidance between creeks. The results of this study can establish whether these methods of studying bear populations are accurate, or if they need to be modified to collect more inclusive and representative data. Further study is required to develop a comprehensive appreciation for brown bear populations, habitat use, and other behavioral patterns.

Many environmental factors can influence foraging strategies of predators, such as availability of prey sources. Brown bears feed on many species of animals and plants but Pacific salmon are especially important to their diet. For example, brown bears in the Lake Aleknagik, Alaska ecosystem forage on sockeye salmon as well as other food items. Direct observations and motion-activated cameras have documented bears foraging on salmon but do not reveal variation among individual bears. This study used the distinctive carbon and nitrogen isotope signatures in salmon as a way to assess brown bear diet by processing samples of bear fur collected during the salmon spawning season. As fur sample collection is noninvasive, it provided the opportunity to “resample” bears (individually identified by DNA analysis) multiple times within a season and over the course of multiple seasons. This study demonstrated how various factors, including salmon availability, gender, and location, influence the extent of bear consumption of salmon over time. Understanding the association between brown bear diet and these factors can provide better insight into the importance of both sockeye salmon and alternative food sources to brown bears.

Due to antibacterial resistance and the high recurrence of urinary tract infections (UTIs), studies have shifted to focus on anti-adhesive therapies as alternative to antibiotics. Often treated with antibiotics, UTIs are caused by uropathogenic Escherichia coli (UPEC). The bacterial adhesin, FimH, found on the terminal end of fimbria, hair like structures expressed on the perimeter of UPEC, is the main etiological factor of UTI prevalence and recurrence. FimH increases the virulence factor of E. coli by mediating the initial binding of the bacteria to glycosylated cells in the urinary tract. FimH has two domains. The lectin domain (LD) recognizes and binds the terminal mannose on glycosylated cells lining the urinary tract, whereas the pilin domain acts as an anchor to the fimbria. Previous studies have shown that α-methyl-mannose (αMM) competitively inhibits glycoproteins, such as horseradish peroxidase (HRP), from the FimH active site. We hypothesize that αMM can non-competitively inhibit HRP through a novel mechanism of inhibition. To determine the mechanism of inhibition of HRP in the presence of αMM, we are using Enzyme Linked Immunosorbent Assays to measure the dissociation of HRP in the presence and absence of αMM, after the FimH-HRP complex has formed. We expect to see an increase in the dissociation of HRP in the presence of αMM. HRP in this case, will act as a model to the glycosylated cells lining the urinary tract. This study aims to assist in the design of innovative anti-adhesive therapies that inhibit binding of FimH once bound to glycosylated cells lining the urinary tract.

Retinal diseases tend to effect specific neuron subtypes, ranging from age-related macular degeneration, which is caused by the deterioration of photoreceptors near the central portion of the retina (macula), to glaucoma, which identifies abnormally high intraocular pressure resulting in the death of ganglion cells. Unfortunately, adult mammals are not able to regenerate retinal neurons. Conversely, zebrafish, frogs, and various amphibians are able to completely regenerate their retinal neurons in many different animal models of damage, and restore retinal structure and visual function. The source of regeneration stems from the resident Müller glia cells, which normally provide neuronal support and span all three retinal layers. A critical gene for the initiation of transforming Müller glia into neurons was found to be Ascl1. This led our lab to hypothesize that the introduction and upregulation of Ascl1 in mammalian Müller glia might stimulate them to become retinal neurons after damage, as occurs in these other regenerating species. Indeed, after introducing Ascl1 into the Müller glia of mice, we found newly regenerated retinal interneurons (bipolar cells) that successfully integrated into the retinal circuitry and functionally responded to light stimulus. In addition to Ascl1, we have identified another transcription factor, that when introduced in combination with Ascl1, stimulates the generation of a different retinal interneuron (amacrine cells). Ectopic expression of a proneural transcription factor to stimulate retinal regeneration provides a potential therapeutic intervention for treating blinding diseases, that even now, have few modest treatment options. In contrast to prosthetic devices and stem cell-based therapies, neuronal regeneration via viral injections is advantageous in regards to host tolerance and reducing the invasiveness of a given treatment, since the patient’s own Müller glia are the source cell of the new neurons; and therefore, immunosuppression would likely not be required.

Domoic acid (DA), a common marine neurotoxin found in marine organisms such as shellfish and finfish, is gaining public attention with increasing outbreaks in the USA, Australia, China, and some countries in Europe. Most DA outbreaks are observed along coastal regions, increasing the risk of exposure by seafood consumption. However, little is known about the effects of low level prenatal exposure of DA. To address this growing public health issue, we administered behavioral and cognitive tests on 27 Macaca fascicularis infants prenatally exposed to 0, 0.075, and 0.15 mg/kg/day of DA. Their cognitive capacities were analyzed based on their speed of learning new tests using the Wisconsin General Testing Apparatus (WGTA). The WGTA requires testers to present stimuli to the infant and, depending on the response, reward the infant for a correct response. I was one of several testers that worked with the infants during testing sessions 5 days per week. I also worked with the investigators and data analysis team to analyze the results of the WGTA tests. Results from early cognitive tests of object and spatial discrimination did not indicate a statistically significant detrimental effect of DA on these basic learning tasks. This suggests that DA does not have an impact on basic discrimination learning in the early stages of life (< 1 year). However, due to the potential for delayed toxicity of DA, more complex learning and memory tests are being administered as the infants mature. The results of this study will provide insights into future research and influence policies being implemented regarding seafood consumption and food safety.

The most recent National Vital Statistics Report reports that approximately 9.85% of babies in the United States are born preterm, with 72% of those born late-preterm (at 34-36 weeks of gestation). Using neonatal ferrets at age 17 days old, the Juul lab in the Division of Neonatology at the University of Washington Medical Center has developed a preliminary model of brain injury to mimic late-preterm neonatal injuries. In this species-specific adaptation of the Vannucci Model, the left carotid artery is permanently ligated, along with a temporary (4h) occlusion of the right carotid artery. Ferrets are then exposed to periods of hypoxia and hyperoxia. By looking at data and outcomes from our surgeries, I aim to examine the effects of certain surgical parameters on ferret mortality. These parameters include: time the animal is exposed to isoflurane, the length of surgery, and the amount of time the animal is given to recover between surgery and hypoxia. Aside from mortality, I will also analyze the effects of these parameters on respiratory rate after surgery as well as gross brain injury and data from behavioral testing in an attempt to discern the level of injury in living animals and the most common predictors of death in those that died prior to their determined endpoint.

Giardia lamblia, a microscopic flagellated parasite that causes giardiasis, is a highly divergent eukaryote in which conventional Golgi, endosomes, lysosomes, and mitochondria are absent. Similar to other parasites of medical importance, Giardia lamblia has two life cycle stages - proliferative trophozoite form and water-resistant, nonmotile, infectious cyst form. During encystation when Giardia trophozoites transform into infectious cysts, they secrete cyst wall proteins (CWP1-3) that are trafficked and processed in Encystation Specific Vesicles (ESVs). These vesicles are thought to be stage-induced Golgi in Giardia. Previous work in the lab has shown that the signaling activities of G. lamblia’s single Rho family GTPase, GIRac play an important role in regulating this encystation process. The aim is to characterize proteins in Giardia lamblia that potentially interact with GIRac, currently focusing on homologs of known players in membrane trafficking by examining their order of arrival using morphology of the ESVs based on CWP1 staining. Since this is subjective, there is a need for stage-specific molecular markers. In other eukaryotes, Rab GTPases have been established as markers of membrane identity and directionality of trafficking. Only two out of nine Giardia’s Rab GTPases have been localized and reportedly found at ESVs and based on published images, they appear to be recruited at different stages of ESV maturation. By tagging the N-terminus of all 9 Giardia Rab GTPases with fluorescent tags, we can screen them for their localization to ESVs and perform multi-color imaging to determine the order of arrival of these markers. Ultimately, this finding of stage-specific molecular markers could be a powerful tool to further suggests its potential as a novel target for drug development to treat giardiasis.

Galaxy clusters are collections of galaxies that form the largest gravitationally bound structures in the universe. Because these galaxies are clustered together, they undergo a wide variety of processes and operate under a multitude of mechanics, differing from non-clustered galaxies, like our own Milky Way. One such example is ram pressure stripping. A galaxy that falls through the hot and dense Intracluster Medium (the space between clustered galaxies) is subject to a "wind" force that can strip it of its gas, usually producing a long gaseous tail emanating from the galaxies causing it to be dubbed a Jellyfish Galaxy. This phenomenon can be incredibly impactful as it can "quench" star formation in the galaxy or, in other words, it can cause the galaxy to “die.” A galaxy is considered quenched when it has insufficient gas to form stars, and it is still not well understood why some galaxies become quenched and others do not. Learning more about this process can inform us on the life cycle of both the galaxies themselves and the cluster as a whole. We used the RomulusC simulation data, run on the NSF Blue Waters Supercomputer, which simulated a large galaxy cluster in high resolution. This simulation has allowed astronomers to study ram pressure stripping in a realistic setting for the first time. Comparing the results of this simulation to observations will supplement our understanding of galaxy clusters and the movement of matter in these colossal and highly dynamic systems.