The leading causes of hearing and balance disorders are damage and loss of inner ear hair cells. Noise overexposure and aging can damage the fragile synaptic transmission between presynaptic hair cells (HC) and postsynaptic afferent neurons (AN), leading to “hidden” hearing loss. Hidden hearing loss is a condition where one can show normal auditory sensitivity when tested but still have difficulty in situations such as hearing one person in loud environments. Mitochondria might play an essential role in this connection by regulating transmission and energy supply. Mitochondria are known to adapt their morphologies to meet cellular demands. Studying mitochondrial morphology may reveal solutions to hearing preservation and contribute to our overall knowledge of the auditory system and general biology. Our lab has previously found that presynaptic hair cells harbor unique mitochondrial networks localized at the presynaptic HC release sites (ribbons) regulated by activity. However, the synapse requires postsynaptic neurons to function correctly to maintain healthy auditory transmission, so we examined the postsynaptic ANs. Preliminary data of two postsynaptic ANs revealed mitochondrial networks that extend between different HCs, but substantial evidence was lacking. Using the zebrafish lateral line as a model system, I microinjected fluorescent probes that tag AN mitochondria, Ca2+ uptake, and track depolarization via Channelrhodopsin. I used serial block-face scanning electron microscopy (SBFSEM) to reconstruct zebrafish AN and their mitochondria. SBFSEM cuts thin layers of the fluorescently tagged structures and images each layer to generate high-resolution 3D images. Probes successfully tracked the different functional characteristics. We observed distinctive architecture in postsynaptic mitochondria, confirming our preliminary findings. Further research will assess changes in postsynaptic mitochondria and regulation by synaptic activity. Understanding the connection between mitochondrial architecture at the synapse and its functional mechanisms will contribute to our knowledge of proper synaptic transmission and the pursuit of healthy hearing preservation.

Since 2013 there has been a decline in the harvest of Dungeness crab (Metacarcinus magister) in south Puget Sound. The Pacific Northwest Crab Research Group (PCRG) is a network that focuses on crab population monitoring. PCRG runs a large-scale larval trapping network that uses light traps to attract Dungeness crab megalopae via positive phototaxis. The light traps span the Salish Sea, to recognize migration patterns and better understand Dungeness crab population dynamics. In addition to Dungeness crabs, different crab species are caught in the traps, creating an opportunity to study more species. All crab larvae follow the same larval stages, called zoea and megalopa, which appear similar to the naked eye. This creates a problem because population counts are often done by volunteers who don’t have prior knowledge of megalopa morphology. Gathering reliable population data of the marine organisms caught in the traps requires a clear procedure to identify bycatch. In this research project, I will classify the unknown larvae into morphospecies by carefully looking and documenting individual characteristics of each organism. Documents will note features such as: carapace shape, carapace width, lateral edges of carapace, telson shape and additions (e.g. setae), number of spines, location of spines, antennae, length and width of thoracic legs, coloration, and overall shape. Measurements will be taken through Image-J, which will be documented along with photographs clearly differentiating anatomical characteristics to compare to DNA barcoding to be conducted by Seattle University on these samples. When DNA results come back, I will determine if specimens that had identical characteristics are genetically matched. This information will be used to make an accurate guide down to the species level. My results will be shared with PCRG to improve the accuracy of population counts of different crab species which will increase reliability of future PCRG data.

Dungeness crabs (Metacarcinus magister) are culturally and ecologically essential to Washington; they also constitute the most valuable state and tribal commercial fishery. Recently, there have been sharp declines in adult crab catch, resulting in the closure of the south Puget Sound fishery. Previous research by Alan Shanks has shown larval abundance is a clear driver of adult yield in Oregon, but many gaps in research remain regarding population dynamics in Washington, sustainability, and the impact of shifting environmental conditions. To address these gaps, the Pacific Northwest Crab Research Group (PCRG) was formed by the Swinomish Indian Tribal Community. PCRG is a collective of over 110 state and tribal co-managers, federal agencies, academics, commercial crabbers, and non-profit organizations. As part of PCRG, UWT’s Becker Lab deployed standardized light traps in the south Puget Sound from mid-April to September for two consecutive years, with sampling four days per week. In June 2021, the light trap network experienced a large pulse of Dungeness megalopae succeeding an extreme heat event. With the frequency of heat waves doubling over the last century, understanding temperature and other environmental stress responses in Dungeness crab larvae is imperative. Megalopae (n=55) from six PCRG sites were preserved in the field with 100% ethanol, then returned to the laboratory where I imaged them under a microscope and performed morphometric analysis (carapace width, carapace length, total length) in Image-J. I then compared measurements with recorded temperature, pH, dissolved oxygen, and salinity. Analysis is ongoing, but I expect to see an inverse relationship between environmental factors and Dungeness megalopae size. The aim of my work is to quantify larval growth as it relates to varying environmental conditions in order to better plan restoration and predict adult harvest limits. These data will offer valuable insight for restoration and protection of larval Dungeness crab.

Organic mixed ionic electronic conductors (OMIECs) are a class of semiconducting compounds that have recently sparked major interest due to their unique ability to conduct both ions and electrons when electrochemically doped. This unique property of OMIECs make them great contenders for applications in biosensing and neuromorphic computing, since they are soft organic materials that can change their state of conductivity. The rate at which these polymers can undergo redox reactions is known as the polymer’s doping kinetics, which is an important parameter for understanding these materials. In this work, I measured the doping kinetics of the commercially available polymer, poly(3-hexylthiophene-2,5-diyl) (P3HT), using UV-Vis spectroelectrochemistry to measure the electrochemical oxidation rate in solutions of different anionic species and solution concentrations, as well as varying the film thickness. I predicted that anionic species of larger sizes, at higher concentrations, results in faster doping kinetics of P3HT. I also expect to see faster doping in thinner films of P3HT, when compared to thicker films. My results show that the choice and concentration of the electrolyte plays a large part on the kinetics of electrochemical doping. I showed that using electrolytes that have larger anions were able to generate a faster doping kinetics. Increasing the electrolyte concentrations also increased the kinetics of doping the polymer. I also found that the thickness of the polymer film, when decreased, resulted in a faster doping kinetics. Using P3HT as a model system, I have examined the effects of anion, electrolyte concentration, and polymer film thickness, which are important parameters to understanding the factors that go into making these materials good conductors for a range of applications.

Activins are a class of growth factors belonging to the Transforming Growth Factor-Beta (TGF-β) superfamily that recruit a tetrameric complex of type-I (ACTRI) and type-II (ACTRII) transmembrane serine/threonine kinase receptors to cause downstream signaling in various critical cell-signaling pathways. Inhibition and supplementation of the signaling molecules offer new possibilities in many therapeutic and clinical applications. The overexpression of Activin A has been shown to be a major driver of diseases such as pulmonary hypertension, chronic kidney disease, and cerebrovascular disease. Sotatercept, one of the few existing activin-A-inhibiting therapeutics, is in clinical trials for treating pulmonary arterial hypertension, but, like many ligand traps, it is designed by fusing a large antibody domain to a native activin receptor domain, making it expensive to produce and unstable due to its large size. There is a need for therapeutics that can be produced in bacteria, replicate high efficacy and strong affinity, and minimize side effects. These therapeutics can be developed through de novo protein design, which involves designing proteins from scratch, with a new, unique sequence predicted to fold into its specific corresponding structure or by grafting native protein-protein interfaces onto novel scaffolds. This project aims to use computational de novo protein design methods to develop a therapeutic ligand trap for Activin A. The design pipeline involves scaffolding functional motifs, optimizing protein sequences, predicting protein structures, and filtering designs using metrics calculated by Rosetta, a protein design tool. With this pipeline, we have tested several design strategies, providing knowledge-based guidance and analyzing outputs to inform strategies for future design iterations. Recent designs have passed Rosetta filters, and we have ordered the corresponding genes to express and purify the designed proteins. Once purified, they can be experimentally characterized, testing binding and stability in vitro, and then further optimized to create stable Activin A ligand traps.

Cardiomyopathy is currently the leading cause of death for patients with Duchenne muscular dystrophy (DMD), a severe neuromuscular disease affecting young boys. With no current cure, gene therapy is a promising solution, but supplementation with drug therapies is likely inevitable to fully address the pathology seen in older patients. The use of human-induced pluripotent stem cell (hiPSC) models for drug studies is beneficial due to the direct relevance to human physiology and the potential development of personalized care. Dystrophic hiPSC cardiomyocytes have been shown to exhibit calcium reuptake delays, higher resting calcium levels, and frequent arrhythmias. The Mack Lab previously conducted a preliminary drug screen on healthy and DMD-affected cardiomyocytes and found that certain L-type calcium channel blockers (CCBs) indicated a cardioprotective effect. These drug compounds (namely Nitrendipine and Nimodipine) have been shown to alleviate cardiac fibrosis in patients through vasodilation. In this project, I am validating the beneficial aspects of the drug compounds. I initially hypothesized that treatment of DMD hiPSC cardiomyocytes with L-type CCBs will rescue resting calcium levels and normalize relaxation kinetics. To assess this, I cultured mature hiPSC cardiomyocytes on a Microelectrode Array (MEA) system capable of maintaining physiological conditions while measuring properties of cardiac electrophysiology. The cells were then matured using lactate enrichment to attain further maturity and grown in culture prior to MEA experimentation. Using the MEA, I have found that the QT interval for DMD hiPSC cardiomyocytes was significantly longer than isogenic controls. In current experiments, I am using this platform to validate the effect of L-type CCB compounds of interest in relation to DMD cardiomyopathy. The development of this novel platform may not only have broader implications for DMD drug discoveries and targeted therapies, but it can potentially serve as a powerful preclinical model for other neuromuscular disorders.

A “tough on crime” approach in the late 1900s ushered in more punitive sentencing than ever before in American history. As a result of mandatory minimums, three-strikes laws, and other harsh legislation, the American carceral state boomed. Not only were more people incarcerated, but they were incarcerated for longer periods of time. Since then, federal court cases have challenged the constitutionality of life and lengthy sentences, especially for individuals sentenced when they were juveniles. In the wake of these federal decisions, Washington state has enacted a variety of legal pathways towards post-conviction sentence review and early release. This study explores how learning of the possibility of early release affects incarcerated individuals who previously did not expect to have a chance at release. This research will be conducted through qualitative interviews and a survey with a sample of 12-15 individuals who served lengthy sentences and unexpectedly learned of a chance for early release. The findings of the research are expected to make a variety of contributions. First, the research will fill a gap in the literature on the social-psychological dimensions of imprisonment and on the significance of hope for incarcerated individuals. Second, this research will add qualitative insights to the largely policy-related literature on criminal sentencing. Third, the research may influence early-release policy by demonstrating potential benefits of such programs, which may positively contribute to less stringent sentencing and carceral practices. Fourthly, the research may help to shatter preconceived notions that individuals who have served life/lengthy sentences are “burdens” to society and instead replace such negative views with positive, prosocial, and communal experiences.

Methanotrophs are prokaryotes that naturally consume the potent greenhouse gas methane for energy. Through metabolic engineering at an industrial scale, these microorganisms hold potential to mitigate the contribution of methane emissions to global warming. In particular, Methylotuvimicrobium buryatense can sustain robust growth both in nature and experimental settings; it is a promising engineering candidate. To develop a robust metabolic engineering platform using M. buryatense, biologists require a deeper understanding of the genetic mechanisms by which it functions. Here, I present an open-source software tool designed to interactively explore the transcriptome of M. buryatense. By integrating bulk RNA-seq datasets collected from experiments over the past decade and applying an array of unsupervised machine learning clustering algorithms, we cluster genes by their expression profiles in differing growth conditions. These gene clusters are annotated with gene ontology (GO) terms using statistical enrichment analysis to assist in functional interpretation of the clusters and the genes that comprise them. To enhance domain-expert researchers’ ability to explore and drill-down into specific queries, I unify these cluster-specific analyses in a web-hosted tool using interactive data visualization techniques centered on a ReactJS frontend and Azure Cloud backend. With both exploratory and query-focused use cases, this software tool can support M. buryatense biologist workflows for predicting functions of hypothetical proteins, showcase new or confirming putative regulatory processes, and generate new experimental hypotheses from the presented transcriptomic trends.

Semiconductors are integral to many industries. As electronics trend towards more powerful devices, research efforts now focus on developing tools with the ability to achieve higher feature densities, such as the use of extreme ultraviolet (EUV) photolithography. Current photoresist technologies center around the use of 193 nm wavelength light with photo-sensitive organic thin films. However, these tools cannot produce feature sizes smaller than 20 nm. To overcome this limitation, semiconductor manufacturing is exploring 13.5 nm (EUV) wavelength light. Current commercial polymer photoresists lose sensitivity at this wavelength due to their low atomic absorption and photoemission of EUV light. Transition metal atoms have higher EUV absorption and can be incorporated to improve polymer photoresist sensitivity. To that end, molecular layer deposition (MLD) is a promising, scalable tool capable of creating thin films with angstrom-level precision. Unlike other deposition techniques, MLD does not require the use of solvents for deposition, boasts high compositional control, and is area selective deposition capable. In this work, the synthesis, characterization, and stability of novel thin films deposited using MLD are tested to determine their suitability as EUV photoresists, with the goal of improving semiconductor feature density, reducing the use of hazardous solvents, and decreasing energy and material costs in semiconductor production. Six films were studied using diethyl zinc (DEZ) as an inorganic EUV absorbent with six organic reactants: ethylene glycol; cis-2-butene-1,4-diol; 2-Methylene-1,3-propanediol; 1,5-hexadiene-3,4-diol; 1,4-butyne diol; and 3,4-dihydroxy-1-butene. Film thickness was measured using ellipsometry. Photosensitivity was measured upon exposure to 254 nm wavelength UV-C light. Degradation upon solvent exposure in an inert environment was examined using acetone, chloroform, ethanol, toluene, and deionized water, which proved most effective at removing the films, with thickness reductions up to ~90%. Results of solvent stability and light sensitivity will be used to propose new EUV photoresist processes for production-scale semiconductor manufacturing.

A thoracic aortic aneurysm (TAA) is an expansion of the aorta within the chest. TAA can rupture, often causing sudden death. Mechanisms of TAA formation and growth are incompletely understood, but likely include dysfunction/loss of aortic smooth muscle cells and inflammatory cell accumulation. We hypothesized that quantitative analyses of cell types, transcriptomes, and cell-cell communications in experimental TAA tissue would provide insights into pathogenesis. Because many TAA are caused by heritable mutations, we investigated TAA pathogenesis in a mouse model of TAA caused by a mutation in the type 2 transforming growth factor beta receptor (Tgfbr2^G357W/+ mice). We used single-cell RNA sequencing (scRNA-seq) to quantify cell types and transcriptomes in proximal aortas of Tgfbr2^G357W/+ and wild-type mice. I analyzed transcriptomes with CellChat, a tool that uses scRNA-seq data to quantitatively infer and analyze intercellular communication networks. Tgfbr2^G357W/+ aortas had fewer smooth muscle cells and more macrophages than control aortas. Transcriptome analysis revealed that Tgfbr2^G357W/+ aortas also contained a new subpopulation of fibroblasts (“new fibroblasts”) that was absent in controls. Among all cell types, CellChat identified the new fibroblasts as the strongest source of outgoing cell-cell signals, and smooth muscle cells and macrophages as the major recipients of signals emanating from both the new fibroblasts and from other fibroblast populations. Outgoing signals predicted to emerge from the new fibroblasts are mediated by matrix components (e.g., collagen, laminin), cytokines (e.g., CSF), and other ligands (e.g., angiopoietin). We conclude that TAA in Tgfbr2^G357W/+ mice have fewer smooth muscle cells, more inflammatory cells, and a new population of fibroblasts. These new fibroblasts appear to signal to other aortic cells and may play important roles in inflammation and smooth muscle cell phenotypic alteration/loss. Further characterization of the new fibroblasts and their signaling pathways may reveal new targets for therapies that prevent or stabilize TAA.

Thoracic aortic aneurysms (TAA) are excessive dilations of the aorta, inside the chest. TAA can rupture, causing sudden death. Human TAA are attributed to both decreased and increased TGF-β signaling in aortic smooth muscle cells (SMC). Experimental data from mice clearly shows that decreased SMC TGF-β signaling causes TAA; however, the connection of increased SMC TGF-β signaling with TAA is largely based on correlational data. Some of these data implicate excessive TGF-β signaling in only one of the two SMC embryonic lineages that populate the proximal thoracic aorta: the cardiac neural crest (CNC) lineage. To directly test whether increased SMC TGF-β signaling in either of the two lineages causes TAA, I am generating mice with increased SMC TGF-β signaling in either CNC-derived or second heart field (SHF)-derived SMC. I accomplish this by using a transgene that expresses a constitutively active TGF-β receptor (TBRI-CA) after activation by Cre recombinase. To express TBRI-CA in CNC-derived SMC, I mate mice with a Wnt1-Cre transgene to mice with the TBRI-CA transgene. To express TBRI-CA in SHF-derived SMC, I mate mice with an Mef2c-Cre transgene to mice with the TBRI-CA transgene. I hope to determine whether increased TGF-β signaling in SMC of either lineage causes TAA. However, because SMC TGF-β signaling plays critical roles in embryonic vascular development, I will begin by determining whether activation of TGF-β signaling in either lineage is embryonically lethal. I am analyzing genotypes of pups from each mating, and using Chi Square or Fisher Exact Test to test the null hypothesis that increased TGF-β signaling in embryonic SMC is not lethal. If my hypothesis is supported, I will then be able to determine whether SMC-targeted activation of TGF-β signaling causes TAA. My findings could provide support for development of human therapies that prevent TAA by blocking SMC TGF-β signaling.

Mosasaurs are extinct, enormous lizards that dominated the oceans in the Late Cretaceous, from 90 to 66 million years ago. They were important members of Late Cretaceous marine ecosystems, with some species being the top predators. However, there remain many uncertainties about mosasaur diets, which likely varied considerably among species. Therefore, I aim to investigate mosasaur diets to provide more information on lizard evolution and Cretaceous marine ecosystems. To infer the diets of mosasaurs, I examine the lower jaws and teeth of their closest living relatives, modern lizards, and test for correlations between craniodental morphology and diet. For example, a lizard that eats primarily hard-shelled foods will likely have more robust jaws and teeth than a lizard that primarily consumes insects. To quantify the morphologies of the lower jaw and teeth, I measured the width and height of the jaws at three points, and the height, width, length, curvatures, root length, and cusp numbers of the teeth at the same three points (n = 43 species). I then used the jaw measurements to calculate cross sectional shape values that represent the amount of stress the jaws experience during feeding. Finally, I used phylogenetic regressions and multivariate analyses to test the relationship between jaw/tooth shapes and diets. I find evidence that jawbone heights increase among diets in the following order: insectivores, carnivores, herbivores, and durophagous taxa. Further, bone width is greater in herbivores than in non-herbivorous taxa, and durophagous lizards have the most diverse tooth morphologies. These results provide a foundation for future studies to examine the relationship of jaw/tooth shapes and diet more robustly, with the goal of using modern lizards as analogs for inferring diets of mosasaurs.

Increased awareness of the complexity and importance of soil ecosystems has led to a surge in “regenerative” agricultural practices, which build topsoil and improve soil fertility and nutritional quality of produce. Such practices also sequester carbon in soils, reduce topsoil erosion and reliance on synthetic fertilizers, and increase microbial content and water storage capacity of soils, avoiding many of the negative environmental and ecological impacts caused by more conventional forms of agriculture. While there is substantial anecdotal evidence for the success of regenerative farming, quantitative studies that support farmer experiences are limited. This study aims to help bridge this gap by examining soils in the Puget Sound region to evaluate differences between areas managed regeneratively and conventionally. I visited five local regenerative farms and took two sets of soil samples from each: one from a plot managed regeneratively, and one from a portion of the farm that has not yet transitioned from conventional to regenerative management. Each set of soil samples consisted of soil cores to test for soil organic carbon (SOC), and a soil pit to examine soil horizons. I determined SOC using loss-on-ignition tests, and topsoil depth by measuring the thickness of the A-horizon in the soil profile. The data show that topsoil managed with regenerative practices can be up to 4 inches deeper and contain up to 20% more SOC than when managed conventionally. Within the regeneratively managed plots at the UW Student Farm, there is a strong correlation between the age of the plot and topsoil depth, suggesting growth of topsoil over time. While these findings align with the results of other studies, a more nuanced understanding of how topsoil formation processes and soil ecosystems develop under regenerative management is necessary to support large-scale transitions towards more sustainable agriculture.

Tropospheric reactive halogens are a sink for ozone and influence the oxidizing capacity of the troposphere. Daily measurements of chloride enrichment, bromide enrichment, and iodine in sub- and super-micron aerosol were collected in June 2022 and January-February 2023 in Bermuda during the Bermuda boundary Layer Experiment on the Atmospheric Chemistry of Halogens (BLEACH) campaign. Gas-phase halogen radicals originate from heterogeneous reactions (chemical reactions involving more than one phase of matter) on the surface of halide-containing aerosol. These reactions lead to enrichment or depletion of aerosol halides (e.g., particle phase chloride, bromide, or iodide) relative to their ratios with sodium in sea water. We compare these measurements with simultaneous observations of ozone and gas-phase halogen concentrations in order to understand the relationship between aerosol and gas-phase halogens and their impact on tropospheric ozone abundance. Observations from these campaigns will aid in improving atmospheric model outputs and environmental policy. I extracted the ions from more than 60 quartz air-filter samples and aided in method development with Ion Chromatography (IC). Inductively Coupled Plasma Mass Spectrometry (ICP-MS) was primarily used for iodine measurements. Data analysis and lab work is still ongoing, therefore only preliminary observations are available. We’ve observed that sodium, iodine, chloride, and bromide agree well with previous observations at Tudor Hill (Arimoto et al., 1995; Sander et al., 2013). Higher depletions of chlorine excess and bromine excess and higher iodine and non-sea salt sulfate concentrations are observed during the first half of the summer campaign.

The Hell Creek Formation (HCF) of northeastern Montana is known globally for preserving some of the last non-avian dinosaurs, including Tyrannosaurus rex. In addition to T. rex, several other species of theropod dinosaur lived in the HCF. These theropods differ morphologically from each other in a variety of ways; however, given that many of these taxa are represented primarily from dental remains (i.e., teeth), the best diagnostic feature available is dental morphology, with the exception of toothless theropods such as Anzu. In this project, I am utilizing imaging processing software in order to collect diagnostic linear measurements of key aspects of tooth morphology from tooth-bearing theropods of the HCF and plot this data against time to ascertain if changes occurred in the dentition of individual taxa through the approximately 2-million-year time span recorded in the formation, utilizing specimens from the upper, middle, and lower portions of the HCF. My current dataset of 20 teeth is insufficient for statistical analysis, but already possesses great potential for future use tracking the various observed dental morphologies. The amount and precision of this morphometric data will only increase as I continue to grow the dataset through the imaging and measurement of at least 40 additional theropod specimens. Once complete, the morphological data I produce will assist in testing my current taxonomic identifications through the establishment of ranges of individual variation. Additionally, quantifying observed morphological variation relative to time can provide insight into the continuing evolution of HCF theropods due either to speciation or extinction. Through these measurements, I hope to gain clarity on both the community composition and dental evolution in the HCF theropods, which is vital to understanding the evolutionary history of those dinosaurs that are closest to birds and their ecological standing as a group immediately prior to the Cretaceous/Paleogene mass extinction.

The migratory songbird Gambel's White-Crowned Sparrow (Zonotrichia leucophrys gambelli) experiences drastic seasonal shifts in its song production and stereotypy, driven by changes in distinct song circuit nuclei, namely increased neurogenesis in the forebrain nucleus HVC and increased electrical excitability in nucleus RA. Manipulating the photoperiod and hormone levels of these sparrows in a laboratory setting results in changes very similar to the seasonally-induced behavioral and neurophysiological changes they experience in the wild. We are interested in reducing the number of birds necessary for our studies by developing a protocol to produce seasonal effects in vitro via organotypic slice culture. Organotypic slice cultures preserve the neural connections and functions involved in the seasonal changes we are interested in, so they can be more directly observed and manipulated. We maintained slices of neural tissue from white crowned sparrows, including HVC and RA, on a membrane that allowed for exchange with the media, a technique initially developed by Stoppini et al., in 1991. After the tissue was cultured, it was fixed and resectioned into thinner slices so it could be Nissl stained and imaged to test for the presence of healthy cells. Organotypic culture is an established technique for neural tissue from juvenile animals; we are attempting to use it on tissue from adult, wild caught, white crowned sparrows. There is currently no protocol for organotypic cultures of this tissue, and the nature of the tissue itself poses a challenge. Adult tissue is less plastic than the juvenile or neonatal tissue that is usually used with this technique, so it has more difficulty surviving the shift to culture. Once the protocol is developed, we plan to manipulate the hormonal environment to try to mimic changes that occur during their breeding season.

The alpine zone has been underrepresented in herbarium collections due to its difficulty in access and short growing season. Despite its underrepresentation, the alpine zone presents a unique opportunity to study climate change impacts due to species; limited ability to migrate to more suitable habitats. For this study, we are examining the Cascades Range from the Canadian border to Mount Adams. Our primary objective is to understand the distribution patterns of alpine species richness and how it is influenced by latitude and elevation. Our secondary objective is to see if these patterns in phytogeography correlate to species; life history characteristics of dispersal, pollination mode, and flower color. To research these questions, we created a species list of Washington's alpine plants using 50 Peaks Project data, historical herbarium records, and literature references. To assess latitudinal course patterns of species richness along the Cascades range, we created three relatively equal zones and scored the presence of each species in it. For statistical analysis, the total number of species per zone will be tallied and Chi-square analysis will be performed to test for significant differences in species richness. We will use regression analysis to quantify the relationships between latitude and the number of peaks, and latitude and average elevation. To compare life history traits across the three zones, we will analyze frequency distribution of those traits. Our preliminary results for latitudinal patterns indicate that the North Cascades have the most species while the Southern and Central Cascades are nearly tied. The final results from this study will inform the selection of future collecting locations and future analysis for species richness among peaks for the 50 Peaks Project. Preliminary Run through the Burke Herbarium, the 50 Peaks Project collects plant specimens to document diversity and distribution in Washington's Cascades Range alpine zone.

With the ever-increasing interest in new photovoltaic materials, much attention is being given to the study of nanoparticles and their assembly. One of the primary goals in this field is the self-assembly of particles, such that they can be programmed to form a desired structure given only a template and a solution of particles. In my project, I investigate the effect of proteins (specially designed through de novo synthesis) on the aggregation of gold nanoparticles, with samples prepared in buffers of salt and Tris base. The particles used are nanospheres of sizes 100, 50 and 10 nm, as well as nanorods of different aspect ratios which can offer more information on the directionality of the assembly. To obtain the necessary data on these samples I use a number of spectroscopy techniques (ultraviolet-visible, dynamic light scattering and circular dichroism) and microscopy methods (hyperspectral and scanning electron). A stereospecific response is obtained from the protein-particle mixtures if the materials formed are chiral, that is, if they rotate plane polarized light. I have shown that the proteins stabilize the particles in a salt solution, which is an indication of protein-particle binding - similar results have been correlated in literature to the formation of a chiral organic-inorganic complex. Such complexes would potentially benefit from both the plasmonic properties of the nanomaterial by absorbing light at a particular wavelength in the visible range, as well as the stereospecificity imparted by the protein helix. Being able to achieve such a result is an important step towards understanding the optoelectronic properties of biotemplated nanostructures, which has a diverse array of applications, including materials for solar energy production, photodynamic cancer therapy in which tumor cells can be specifically targeted, and drug delivery systems. It would also be invaluable for the customizable design of catalysts, enzymes, probes, sensors and diagnostic tools.

 Birth control is an important tool to prevent unwanted pregnancies. However, many people who might benefit choose not to use birth control, contributing to a range of negative outcomes, including unwanted pregnancies and sexually transmitted infections. Many factors affect people’s perceptions of birth control options, and ultimately influence their use of contraceptives. Previous research shows racial differences in rates of birth control utilization, but little work has explored why these racial differences exist and if and how culture contributes to birth control attitudes and utilization cross-racially. Furthermore, most previous research in this area focuses exclusively on race, typically reporting only on Black, Hispanic, and White women, with little or no intersectional analysis. The present study investigates how culture influences birth control attitudes, considering both a broader range of racial categories and the impact of multiple intersecting identities on culture. Our goal is to gain insight into the health care decision processes of intersectionally marginalized patients. We accomplish this with a cross-sectional qualitative study. We first pre-screen potential interviewees to recruit participants with diverse cultural backgrounds. Selected participants will participate in structured 1:1 interviews, answering questions about how their cultural background (including race, ethnicity, religion, and family) influences birth control attitudes. We will conduct thematic analysis to determine what aspects of culture impact birth control attitudes. Shedding light on how culture influences the perception and use of birth control provides insight into a broader range of patient populations, allowing for improved contraceptive counseling and education in the medical setting. Recruiting a diverse sample will illuminate the lived experiences of individuals who are typically excluded from research and scholarship, allowing future advancements in birth control to be more representative and sensitive, with the knowledge of all cultural experiences in mind rather than just a select few.

Hexagonal boron nitride (hBN) is essential for nearly all nanoscale two-dimensional (2D) devices, as its flatness and wide bandgap make it an ideal dielectric for applying electrostatic gating. At high electric fields, hBN undergoes electrical breakdown where large currents flow to the sample, damaging the device. Gating fields are therefore limited by hBN’s dielectric strength. While the dielectric properties of hBN have been studied previously, the preparation of those samples differed from that used in practice. I conducted an experiment with the help of my advisors to understand how hBN’s dielectric breakdown characteristics depend on sample thickness and temperature. I began by fabricating three hBN devices, each containing multiple regions of different thicknesses. By measuring the current and varying the voltage on a given region, I was able to locally probe the hBN’s electric breakdown characteristics with thicknesses ranging from 5 to 24 nm. I tested each device multiple times using a cryostat at a range of temperatures from 4 to 300 K. During initial measurements, I observed an increase in the breakdown voltage with temperature in hBN between 15 and 24 nm thick, conflicting with previous reports. I repeated these measurements with a finer resolution which yielded the same result. The thinnest hBN regions showed no temperature dependence, confirming the absence of systematic temperature effects. I am currently fabricating more devices to reproduce this temperature dependence and working with my advisors to find a theoretical basis for this observation. Understanding how thickness and temperature effect hBN’s dielectric strength will allow researchers to construct more resilient devices, facilitating the study of 2D materials at higher electric fields. Moreover, the study of defects in hBN remains an active subject of research for quantum information applications, and a probe of the capacitive properties of hBN may shed light on this topic.

 The diversification of many vertebrate groups was spurred by the use of novel food resources, and jaw functional morphology provides clues about the adaptations associated with dietary diversification. The external dimensions along the mandible reflect strength to resist bite forces, which are in turn associated with physical properties of the diet. Variation in these dimensions along the jaw and between different species therefore may reflect adaptations of the jaw to specific diets. We applied this biomechanical framework to investigate the relationship between jaw robustness and diverse diet types in bats. Using mandibles of more than 60 species, we quantified the external dimensions at interdental gaps to generate mandibular strength profiles. The strength profiles of frugivorous, insectivorous, and omnivorous bats showed similar patterns, with a trend of increasing jaw depth toward posterior teeth. All diet types showed a high level of variation in jaw shape along the toothrow, suggesting differences in the functional roles of different teeth. Insectivores showed the greatest within-guild variation in jaw shape, while nectarivores had noticeably gracile symphyses. Further, insectivorous bats showed relatively deep jaws at the canine and premolars, which may be associated with the use for prey capture, while frugivores have relatively deep jaws at the posterior molars, possibly linked to adaptations for crushing seeds and pulp. These results suggest that mandible strength profiles reflect dietary adaptations in bats.

 Dravet syndrome (DS) is a genetic form of epilepsy characterized by febrile seizures in infancy, developmental delays, and sudden unexpected death in epilepsy (SUDEP) as a result of being drug-resistant. Finding new and innovative treatments is essential to reducing the risk of SUDEP and other symptoms in DS patients. Using a mouse model of DS (SCN1a+/- mouse), I showed that a machine learning-based detection algorithm could be used to detect interictal spikes (IS), which are abnormal neuronal discharges typical of epilepsy. The goal of the current experiment is to see whether the prior findings can be generalized to a larger dataset and whether the detected IS can be used to predict seizures before their onset. Data is collected by implanting two electrocorticographic electrodes and one electromyography electrode, as well as a wireless body temperature sensor in DS mice. Ambient temperature is controlled so that the animal’s core body temperature is initially maintained at 37°C and then is gradually increased by 0.5 °C every 2 min until a seizure is observed or the core body temperature reaches 42.5 °C. A machine learning model previously trained using manually scored data from the de la Iglesia lab is used to autonomously detect IS in the collected data. My results so far showed a moderate yet significant positive correlation between ambient temperature increases and IS frequency and points to a positive correlation between IS frequency and seizure onset. However, these results did not include the continuous recordings of body temperature. In the current experiment, I test if these correlations hold using a larger sample size and including continuously recorded body temperature, which may have more predictive power than ambient temperature. Our long-term plan is to design a closed-loop experiment that uses the algorithm to predict and stop seizures before their onset.

Weight loss (WL) is recommended for people with obesity to mitigate cardiometabolic risk, but its effect becomes limited when reaching a WL-plateau (WL-PL), when the rate of WL becomes minimal despite efforts to continue losing weight. The biological basis for the WL-PL is not fully understood. The goal of this study is 1) to test two diet-regimens in the development of a WL-PL in mice with diet-induced obesity (DIO), and 2) to identify subsequent changes in mitochondrial function. We hypothesized that despite similarities in caloric intake, higher fat content will make high-fat diet (HFD-CR20) mice protect their adiposity and reach a WL-PL sooner than low fat diet (LFD-CR20) mice. We also hypothesized that mitochondrial function will be reduced in mice that have reached a WL-PL. To test hypothesis 1, individually housed DIO mice were divided into two groups, and were provided with 80% of their ad libitum caloric intake with either a high-fat, or low-fat diet daily for ~2.5 weeks. Body weights were recorded daily. A WL-PL was identified by weight stability (<0.5% change BW/day) following weight loss. HFD-CR20 mice reached a WL-PL phase after 10 days of caloric restriction. LFD-CR20 mice did not achieve a plateau within the study time frame. LFD-CR20 mice lost more weight than HFD-CR20 (-10.8% ± 2.2 vs. -5.2% ± 1.8, p<0.05 respectively), which was attributed to a greater loss in adiposity, measured by an EchoMRI, (-23.3 g ± 6.0 vs. -3.3 g ± 3.0, p<0.05). To test hypothesis 2, mitochondrial function was assessed by high resolution respirometry at the study endpoint. We will further analyze this data to identify differences in mitochondrial function attributed to the WL-PL. This work will improve our understanding on the biological mechanisms behind resistance to weight loss to help advance obesity treatments in humans.

Obesity is a condition characterized by excessive fat accumulation, resulting in increased risk for chronic diseases like cardiovascular disease and diabetes. Weight loss can effectively reduce the burden of cardiometabolic risk factors, but weight loss maintenance is difficult to achieve. Mitochondria are key organelles within cells that are responsible for the breakdown of substrates to produce energy. Mitochondrial dysfunction is implicated in obesity, but little is known about the role of mitochondrial dysfunction in weight loss maintenance. Additionally, females are often underrepresented in obesity research, partly attributed to female mice being more resistant to develop obesity compared to males. The aim of this study is to compare mitochondrial function in liver, adipose tissue, and skeletal muscle, following weight loss in female mice with diet-induced obesity (DIO). We hypothesized that obesity would result in a reduction of mitochondrial function across tissues, and weight loss to further reduce it. We provided CB6F1 female mice with a high fat diet, where 87% of them developed DIO. DIO mice were separated into two groups: one underwent 20% caloric restriction for 4 weeks (HFD-CR), and the other group remained on ad libitum high fat diet for the same intervention (HFD-AL). A healthy weight control group was maintained on a regular chow diet. Their weight and food intake were recorded daily. Body composition was assessed twice, before and after the 4-week intervention period. We had an unexpected finding, where mice lost 5-10% of their body weight prior to the intervention period. HFD-AL mice regained lost weight at study endpoint, while the weight of HFD-CR mice remained weight reduced until study endpoint. CR mice had lower adipose tissue mass compared to HFD- AL mice. Future analyses will include comparisons of mitochondrial content and function in different tissues. Findings will provide more insight into the effects of weight maintenance and regain on mitochondrial function.

Heart disease is the leading cause of death in the United States. Echocardiography is used clinically to highlight cardiac structures, wall motion, and contraction abnormalities to diagnose heart failure. Heart failure is classified into diastolic (relaxation) or systolic (contraction) heart failure using left ventricular ejection fraction (LVEF), a measurement of the amount of blood pumped during each contraction. Global Longitudinal Strain (GLS) analysis, which measures the stiffness and deformation of the heart wall during contraction, has recently emerged as a more sensitive metric of systolic function that predicts cardiovascular mortality in patients. Elamipretide (ELAM;SS-31) is a mitochondrial-targeted intervention that improves aging heart mitochondrial and diastolic function. In this study, we used an aging mouse model to compare sex-specific outcomes in systolic function by LVEF and GLS. We hypothesized that GLS would be a better predictor of systolic dysfunction in mice than LVEF, and that ELAM would repair aging systolic dysfunction. We compared young (4-6 mo) and old (25-26 mo) male and female mice using 2D echocardiography to obtain left ventricular parasternal short and long axis images. A cohort of aged male mice was imaged before and after 8-week ELAM treatment. These images were analyzed using Vevo LAB software using conventional echocardiography to measure LVEF and speckle-tracking echocardiography (STE) for GLS and LVEFStrain. Statistical analysis was performed using GraphPad Prism Software. Limited change in LVEF was observed by conventional echocardiography. Using STE, GLS and LVEFStrain declined with age. Treatment with ELAM restored GLS in aging mice to young levels. Here we show that the more sensitive STE analysis reveals that aging mice exhibit both systolic and diastolic dysfunction. The research supports our hypothesis that ELAM treatment improves systolic function in aging. Future treatments to target systolic dysfunction can be assessed in an aging mouse model using STE.

Despite decades of research very little is known about how mitochondria control stress responses. Therefore, new and innovative models are needed to understand the mechanisms of mitochondrial stress response. We developed a new mouse model of skeletal muscle mitochondrial stress to mimic the aging process in young animals to determine if mitochondrial oxidative stress replicates age-related skeletal muscle and mitochondrial dysfunction. We generated a mouse model to induce skeletal muscle mitochondrial redox stress to mimic skeletal muscle aging by knocking down superoxide dismutase 2 (SOD2). My project was to determine whether this model works in vivo. I fed animals a Doxycycline (DOX) chow diet (0.625g/kg) to induce SOD2 knockdown (KD). After 3-week DOX feeding, tissues were collected and processed for western blotting (WB). WB's were run for SOD2 in gastrocnemius, quadriceps, liver, kidney, heart and brain tissue. Normally, SOD2 is expressed in all tissues that contain mitochondria, so comparing SOD2 expression levels across tissues in KD animals validates tissue specificity. HNE adducts, a marker of oxidative stress, were measured by WB to confirm increases in oxidative stress associated with SOD2 KD. Three-week DOX feeding showed significant decreases in SOD2 protein in gastrocnemius (p<0.001) and quadriceps muscles (p<0.0001) compared to unfed littermate controls of the same genotype. There were no differences in SOD2 protein in heart, brain, liver or kidneys between DOX and control groups. HNE protein adducts were also significantly increased in skeletal muscle of DOX compared to controls (p<0.05). SOD2 is knocked down in skeletal muscle in response to DOX feeding. The increase in HNE adducts confirms that the knockdown of SOD2 causes an increase in oxidative stress. This model can now be used to explore the physiological mechanisms of inducing mitochondrial redox stress in young animals to recapitulate the effects of aging in a controlled manner. 

Obesity is associated with mitochondrial dysfunction. A study previously conducted in our laboratory produced preliminary data suggesting a reduction in T-cell mitochondrial function in subjects with obesity relative to healthy weight controls. The purpose of this present study is to determine whether changes in T-cell mitochondrial function (MITO) reflect MITO changes occurring in liver and skeletal muscle, which are known for having greater influence in glucose homeostasis and energy expenditure. We hypothesize that mice with diet-induced obesity (DIO) will exhibit reduced MITO in T-cells that will correlate to a decline in MITO in liver and skeletal muscle. Adult male C57Bl/6J mice were divided into two groups–the control group was fed a standard chow diet whereas the experimental group was fed a high-fat diet for fifteen weeks. Body weight and food intake were measured every week. Body composition was performed at the endpoint. MITO was measured via high-resolution respirometry in permeabilized liver tissue, skeletal muscle fibers, and splenic T-cells. DIO mice had higher body mass than standard CHOW [52.8g±1.8 vs. 35.1g±2.6] that was explained by an increase in fat mass [19.9g±0.9 vs. 6.1g±1.8] and lean mass [31.5g±1.7 vs. 26.1g±1.1]. We found that differences between DIO and CHOW in mitochondrial leak respiration, maximal oxidative capacity, maximal electron transport chain, and ADP sensitivity were not the same across all tissues. We will proceed to determine which aspects of MITO are correlated between different tissues and assess if variations in respiration are associated with differences in mitochondrial content. An improved understanding of how DIO affects different types of cells regarding oxidative capacity might provide key insights into the development of therapeutics and other preventative approaches to improve immunity and cardiovascular fitness in obesity.