Current mobile navigation applications use large amounts of computing power and battery to frequently update the server on the user’s location. This makes sense, considering that the end user is expected to be moving. When the user is not moving however, possibly while sitting at a stop light, or waiting for the bus to arrive, the app continues to send the user’s location to the server, expending energy, and computational power for absolutely no gain. To address this issue, our research team decided to develop a smarter way of sending data to the navigation servers. The algorithm developed quickly analyzes the location data before sending it to the app’s server. The algorithm considers whether the user is stationary, moving with a constant velocity, or changing velocity, and uses this information to determine when to send location data to the server. If the user is stationary, then sending the same coordinates to the server is redundant and can be simplified by only sending the coordinates of the user once the user’s position changes. When the user is moving with a constant velocity, the sampling frequency can be lowered to further reduce data size. To develop and test this algorithm, our research team made an app that tracks the user’s coordinates, and sends them to the server when the algorithm approves. This allows our team to test the algorithm’s filtering, and make sure that the server still knows where the user is always. Upon large scale testing of the application, we expect it to greatly reduce the amount of data sent to navigation servers when tested for a common use case.

Targeted gene modification and correction is a transformative technology that is currently progressing from basic research and development to applications in medicine, industry and agriculture. Gene targeting systems include four types of targeted nucleases: LAGLIDADG homing endonucleases (LHEs), zinc-finger nucleases, TAL effector nucleases, and clustered regularly interspaced short palindromic repeats/CRISPR-associated endonucleases (CRISPR/Cas9). Each of these systems has the ability to make double strand breaks in DNA. LHEs are promising tools for gene targeting due to their small size (~35 kDa) and high specificity (14-20 bp). One specific LAGLIDADG homing endonuclease is I-LtrWI. To fully understand the LHE family, wildtype, and six variants of I-LtrW1 were overexpressed and purified by nickel affinity chromatography. Using site-directed mutagenesis, variants of I-LtrW1 were generated with an amino acid residue change in the catalytic sites. The purified enzymes were assayed for activity using supercoiled DNA containing a target DNA sequence specific to I-LtrW1. The activity was monitored by agarose gel electrophoresis in an effort to understand catalytically important residues. The results will be used to develop a general method for converting LHEs into LHE nickases, enzymes that make single-strand breaks in target DNA sequences.

The goal of this study was to understand more precisely the placement of the Scalesia clade within Helianthinaea– specifically what taxa are sister to Scalesia and what traits diverged between Scalesia and its sister clades. Previous research has shown Scalesia to be monophyletic and placed Scalesia as sister to Pappabolus and basal to Viguiera, but with low support due to homoplasy. We used extracted ITS, ETS, and psbA sequence DNA from seventeen Scalesia samples from the Galapagos and sequence DNA from ninety two species from mainland South America, North America, and Mesoamerica that we obtained from Genbank. Bayesian analyses of concatenated ITS +ETS + psbA sequence data, maximum likelihood analyses of ITS+ETS+psbA/ITS+ETS, and individual gene region data implied that Scalesia is sister to Pappabolus, Heiseria pusilla, Viguiera simsioides, and Syncretocarpus sericeus with high likelihood support and is monophyletic consistently throughout all trees. Our data also implied that all Scalesia taxa is very closely related and very recently diverged. An interesting split between a woody ancestor (Zaluzania grayana) and an herbaceous, derived species (Scalesia baurii) was observed. Further taxonomic work, especially on the reclassification of Aldama and Viguiera is still necessary.

Modern humans carry loads on a daily basis, and this increases energy expenditure, changes gait kinematics, and increases vertical ground reaction forces (GRF) proportional to the burden. Few studies have attempted to understand the effect of load position on mediolateral and anteroposterior GRFs, where changes in body center-of-mass imposed by the loading conditions may affect GRFs disproportionately. This study aims to assess peak GRFs when carrying a load in a sling in five positions. Our null hypothesis is that mediolateral and anteroposterior GRFs will increase proportionally to the extra load. We expect that position will affect GRFs due to accommodation to the changes in body center-of-mass imposed by the loading conditions. GRFs were recorded by a 3D forceplate for seven participants (5 females, 2 males) walking at their normal velocity carrying a 10 kg load in five carrying conditions: (i) No load; (ii) Front; (iii) Left Side; (iv) Back; (v) Right Side. Each participant completed 8 trials for each condition. Maximum GRFs were determined using a custom-written Matlab. After adjusting the unloaded condition for the increase in mass of the burden, two-sample t-tests, conducted in Stata, were used to determine whether the loaded conditions increased proportionately. The preliminary results indicate that increases in GRFs in the mediolateral and anteroposterior directions are not different from our null hypothesis (all p’s> 0.05). Further study is required in order to understand how load position influences, body position, gait biomechanics, and GRFs in humans.

Cell movement during development is crucial as the location of a cell determines what that cell will ultimately become. Cells require both adhesion and motility for movement. Adhesion is cells holding on to each other via proteins called cadherins. Motility is movement through projections called fillopodia and lamelliopodia. We are interested in the protein p120-catenin, which determines how cells move. We purpose that phosphorylation, or the addition of negative PO4- groups at specific amino acid residues changes p120 catenin’s function. Specifically we are interested in tyrosine (Y) residues 228 and 335 which likely effect p120 antagonistically, by mediating adhesion and motility, respectively. In order to mimic the process of phosphorylation we mutated each residue to either glutamate (E) or phenylalanine (F). Glutamate, which is charged, mimics phosphorylation, while phenylalanine cannot be phosphorylated, but is otherwise similar to tyrosine. To examine which pathway is activated, we quantified whether proteins associated with motility or adhesion were interacting with p120 catenin for each mutation. The primary technique we used to explore this is coimmunoprecipitation. Coimmunoprecipitation works by using antibodies to link p120 catenin to magnetic beads, as well as any proteins interacting with it. The beads can then be extracted and we can determine the amount of each protein interacting with p120 catenin through a process called western blot. So far we have verified that our mutations for Y228 and Y335 produce proteins, which guarantees that any later results are due to phosphorylation. Also preliminary coimmunoprecipitations suggest Y228 does mediate adhesion. Additionally injection of Y228E in embryos, results in embryos with shortened tails and poor yolk constriction, which are associated with excessive adhesion. Taken together these results strongly support our hypothesis for Y228 mediating adhesion. Based on these results, in the future we plan on examining Y335 and verifying our Y228 results.

The family of Transient Receptor Potential (TRP) proteins contains ion channels with a wide array of functions, including invertebrate phototransduction, responding to painful stimuli, responding to temperature changes, and many others. Of particular interest is the polymodal receptor TRPV1, which responds to many stimuli, including capsaicin, heat, pH, etc. TRPV1 is known to play a role in the sensation of both pain and heat; however, the structural dynamics that underlie TRPV1’s ability to transduce these signals are still incompletely understood. Therefore, an understanding of the activation, regulation, and structure of TRPV1 are of clear importance. To address these questions, I have sought to use Xenopus laevis oocytes as an expression platform to perform studies on the structure and function of TRPV1. Since TRPV1 is not naturally expressed in Xenopus oocytes, the genetic information needed for the cell to build the protein was provided for the oocyte via microinjection of RNA. After an incubation period of several days, Western blot techniques were applied to analyze the presence and strength of expression of TRPV1. To better understand specific structural changes made when in the activated conformation, the fluorescent, non-canonical amino acid L-ANAP was integrated into TRPV1 using amber stop codon suppression and an engineered tRNA synthetase. Integrating the non-canonical amino acid ANAP enables structural and functional analysis of the membrane protein via fluorescence microscopy. The results of applying such methods to understand the function of TRPV1 will provide insight into the use of TRPV1 for therapeutic purposes, most prominently the reduction of the sensation of pain.

Environmental conditions following the Late Permian mass extinction, one of the largest mass extinctions to happen on our planet, are not well understood. However, studying paleo-environmental conditions necessitates the use of proxies such as magnesium (Mg), as direct observation is not possible. One method to constrain environmental conditions is by using geochemistry, but until the recent introduction of multi-collector inductively coupled plasma mass spectrometry (MC-ICP-MS) it was difficult to precisely measure Mg isotopes. Now, using MC-ICP-MS, fluctuations of ð²⁶Mg in the oceans can be studied in order to understand changes in seawater chemistry, and the extent of continental weathering, at a time in question. Clean dolomite samples from Zal, Iran, from the Griesbachian, Dienerian, Smithian, and Spathian stratas, have been obtained. They span ~5000kya following the Late Permian mass extinction. Mg has been isolated from these samples using resin column chemistry. The Mg has then been analyzed using MC-ICP-MS, and numerical modeling has determined the ð²⁶Mg for each sample. The expected result was that the ð²⁶Mg would differ from current seawater levels. Current seawater ð²⁶Mg is -0.83, but studies using dolomite show a broader spectrum over time, ranging from -3.25 to -0.38. Based on results, this research could be used to constrain the primary water sources that contributed to the paleo-oceans, as well as track changing ocean chemistry due to influx of chemical precipitate from continental weathering. These changes could be quantified over the time range of our four samples. The data could also be used in correlation with a ð⁸⁷Sr study on weathering rates that was performed on the same dolomites. Ultimately, analogies between past conditions and current issues regarding climate change could be drawn.

We have evaluated the viability of a system for detecting auxin with a yeast culture using the red buildup of P-ribosylformyl glycinamidine as a reporter that can be used to reliably identify auxin in water without the need for advanced equipment. To accomplish this we have fused an IAA17 protein to an ADE2 enzyme with a linker that preserves function, then transformed that into a yeast strain that lacks native ADE2 and has an Fbox AFB2. We plated these on adenine deficient agar with various concentrations of auxin to test for sensitivity. As controls, we plated basic strains w303a and w303alpha and found that the ADE2-lacking w303alpha still needs a little bit of adenine or it will not grow at all. We tested two different types of media, YPD and SC with minimal adenine concentration, both of these have just enough adenine to keep the yeast alive but still induce the red color. We conclude that the method of detecting auxin via the degradation of an IAA17 tagged protein is a viable strategy, but that using ADE2 as our target enzyme presents complications when considering the ultimate goal of our project. The reporter is not simple to use as a very specific concentration of supplemental adenine is required to allow growth but still induce the ADE pathway. To mix this media would require high-precision equipment usually available only in a lab. This problem can be solved by changing the reporter to allow the yeast to be functional in any media, which will be our next project.

 Many children have lasting pain problems after surgery that can greatly impact their recovery and health-related quality of life. Emerging research has identified psychosocial factors as important predictors of children’s postoperative outcomes, suggesting that an intervention addressing these factors could positively impact both short-term and long-term recovery from pain and quality of life. However, there are few data from children, families, or healthcare providers assessing psychosocial needs and receptiveness to a behavioral pain management intervention in the perioperative period. Involving healthcare provider stakeholders in the development of an intervention is essential to ensuring feasibility, acceptability, and successful implementation of the intervention. Seventeen semi-structured interviews were conducted with pain clinicians, nurses, surgeons, physical therapists, and child life specialists who provide perioperative care at a children’s hospital. Interviews probed the following areas: 1) current experience with perioperative preparation and care, 2) recommendations on content and format of an ideal psychosocial intervention, and 3) potential barriers to implementation and participation in a perioperative program. After transcription, we coded and analyzed interviews using inductive thematic analysis to identify consistent and relevant themes across provider groups. Independent members of the research team reviewed codes at each stage for credibility and representativeness. Providers said that a pain management program would address current gaps in surgery preparation, and that children and their parents need to learn effective coping strategies to positively impact their recovery. Providers also suggested necessary content, delivery methods, and ways to enhance program effectiveness. Provider themes, as well as themes from patient and family interviews, are being used to develop a family-based psychosocial intervention targeting children at risk for poor postsurgical outcomes. Next steps for this research include design, implementation, and evaluation of the perioperative intervention aimed at reducing the likelihood that children will develop lasting pain problems after surgery.

Listeria monocytogenes is the causative agent of listeriosis, an infection which causes flu-like symptoms in most individuals after ingesting contaminated food, but can severely harm immunocompromised individuals, newborn children, and the fetus of an infected pregnant woman. L. monocytogenes is a facultative intracellular pathogen which escapes a phagocytic vacuole early in its life cycle. Once in the cytosol of the enterocyte, the master virulence regulator PrfA activates virulence factor production. However, what initiates PrfA and allows L. monocytogenes to sense that it is in the cytosolic environment is still not understood. Recent discoveries have indicated that the translation of at least one of the nine virulence genes, actA, requires several genes involved in bacterial redox regulation, including the protein YjbH. In the Bacillus subtilis model, a homologous YjbH interacts with the protein Spx, an important regulator of thiol-specific oxidative stress. Under normal conditions, YjbH binds to Spx and facilitates its degradation via the ClpXP protease. During disulfide stress, YjbH aggregates, freeing Spx to regulate oxidative stress response genes. Although the B. subtilis and L. monocytogenes YjbH are homologs, the function of the L. monocytogenes YjbH is uncharacterized. My research project seeks to extract and purify the protein. I first generated two strains of E. coli that express a tagged version of either the wild type YjbH or a mutant YjbH which lacks two cysteines that may be involved in its interaction with Spx. The second half of my project will focus on limiting the aggregation of recombinant YjbH. After successful purification of the proteins, these two strains will be a useful tool in characterizing the function of L. monocytogenes YjbH, and its role in the virulence pathway. Understanding the role of YjbH in the virulence pathway could ultimately lead to novel ways to inhibit the pathogenicity of L. monocytogenes.

The Olympia oyster (Ostrea lurida) is the only native oyster along the west coast of the U.S. and has been commercially extinct in Puget Sound since the 1940s. Olympia oysters are a beneficial part of the ecosystem by increasing intertidal species diversity through construction of oyster reefs that provide filtration, habitat, and food. Within the last two decades, they have been the subject of restoration efforts in Washington State. It is important to track the oyster in its larval planktonic stage to aid in restoration of the species because, during this stage, bivalves may travel vast distances of coastline as veliger larvae. To better understand larval transport, plankton samples were collected from two bays in Western Washington during the summer of 2015, Fidalgo Bay to the north, and Dyes Inlet in central Puget Sound. Plankton samples were collected weekly for 5 weeks during ebb and flood tides from two different depths in the water column, one meter from the surface and 0.25m from the bottom of the bay at both locations. O. lurida larval abundance in plankton samples were hand counted and identified using microscopy. The results indicate a high larval abundance in the ebb tide at Fidalgo bay between June 23 and June 30. In Dyes Inlet there is an abundance peak of larvae in the ebb and flood tide on July 9 and larvae were found in both surface and bottom samples at each site. Looking at the densities of oyster larvae in two bays and different tidal conditions can suggest where larvae are dispersing within their natal bays.

Archaeologists have explored the use of emerging technologies that utilize photogrammetric data in order to facilitate the preservation of archaeological sites and material culture in digital form. While conducting survey, mapping, and excavation at two sites on the Grande Ronde Reservation, a nineteenth-century schoolhouse and an early encampment, the 2016 Field Methods in Indigenous Archaeology field school collected consistently scaled photographs of the excavation units and their stratigraphic profiles, as well as aerial drone photography. Importing this data into Agisoft PhotoScan, a software package design to construct three-dimensional models based on photographs, I generated a series of point clouds, or visuals consistenting of data-points acquired from the overlapping photographs; these were given digital textures to mimic the various real-world materials and surface present at each site, and then assembled into scale models, or models that preserve the proportions of the sites in miniature. A similar procedure was used to construct models of artifacts found at both sites, as well as digital elevation models of both the schoolhouse and the encampment based on drone photography. The results demonstrate the utility of photogrammetry in ongoing efforts to preserve material culture and the context in which it is uncovered. These models will be shared with the Tribal Historic Preservation Office, which oversees cultural heritage management for the Confederated Tribes of Grande Ronde, and with the broader public. Making this data accessible to those outside the discipline is an important step towards outreach and education. Ultimately, this and other photogrammetry and digitization projects will contribute to efforts to expedite the return of material cultural heritage to tribal communities as well as encourage the development of collaborative, community-based partnerships between tribes and archaeologists.

According to the World Health Organization, an estimated, 780 million people do not have access to clean water sources. Disasters are another potential cause of disruption to the supply of clean water. When the safety of a water source is unknown, a cost effective method to determine water safety is necessary to reduce unsafe water related disease and death. Prior synthetic biology research has been conducted utilizing transgenic yeast as a biosensor for heavy metal detection, such as using plasmids expressing Green Fluorescent Protein (GFP) which are promoted by heavy metal binding transcription factors. While able to detect copper ions in a dose dependent fashion, the relative instability of the plasmid, reporters which are difficult to detect in situ, and limited targeting to copper and silver ions make the efficacy of the designs impractical. Using modern methods of plasmid integration and the modulation of the copper sensitive pCUP1 promoter expression we sought to produce a stable and more sensitive biosensor for the detection of copper ions in water. We successfully integrated the genetic cassette pCUP1-yeGFP into the yeast genome and produced a viable biosensor for the detection of aqueous copper. We tested this yeast strain via a cytometer after exposure to copper of varying concentrations at varied time intervals. The results showed a dose response curve that positively correlated with copper concentration in a semi-linear fashion, which indicated the pCUP1-yeGFP transgene was a viable sensitive reporter for the detection of copper. Currently, variable constitutive promoters are being altered to increase copper detection sensitivity by modulation of the expression of the metal binding transcription factor Ace1. The team is currently exploring the effect of Ace1 upregulation on the sensitivity of copper sensing. After investigating sensitivity modulation, an exploration of practical field detectable reporters, such as violacein, will be undertaken.

Targeted therapies designed to inhibit hyperactive oncogenic signaling have demonstrated some encouraging clinical responses. However, many of these responses are not durable as tumors may develop new mutations to “work around” pathway inhibition leading to drug resistance. Technologies that enable more efficient discovery of “suites” of drugs that inhibit multiple pathway nodes are needed. The SABRE (Splice Acceptor Brilliant Reporter) platform was developed to provide an unbiased method to screen large compound libraries and dramatically improve the speed and efficiency for which novel targeted therapeutics can be identified. SABRE is built on the premise that oncogenic changes in gene transcription can be harnessed as powerful reporters of pathway activation status. SABRE utilizes gene trap technology coupled with a drug selection process to isolate cells that generate a robust “off to on” luciferase signal in response to pathway inhibition. Via massively parallel comparative analysis, multiple traps let nature provide the best reporter for further analysis and drug discovery. We employed the SABRE technology to identify insertion sites specific to the MAPK (mitogen-activated protein kinase) pathway, an oncogenic pathway that is implicated in a plethora of cancers. Experiments were performed using the human melanoma cell line A375, containing BRAFV600E mutation. SABRE lentiviral transduced A375 cells were treated with trametinib, an inhibitor of MAPK pathway, and clones were isolated that emitted a positive luciferase signal upon drug treatment. To determine if these reporters were MAPK pathway specific, the platform was used to screen a 6000+ compound library. Results from the screen found that 70% (28/40) of the top drug hits were known to directly modulate the MAPK pathway revealing the power of the SABRE gene trap technology to generate a panel of reporters specific to an unlimited number of cancer signaling pathways. SABRE holds the promise for discovering untapped drug therapies for cancer.

The loss of mechanosensory hair cells in the inner ear gives rise to many hearing and balance-related disorders. The most common causes of acquired hair cell loss are normal aging, ototoxic medications and frequent exposure to unusually loud environments, noise exposure. Two main drug classes, aminoglycoside antibiotics and cisplatin-based chemotherapeutic drugs, are known to be ototoxic, with usage resulting in varying levels of temporary or permanent hearing loss through hair cell death. Our goals are to understand and eventually prevent hearing loss due to treatments with these drugs. Because of the inaccessibility and complexity of the mammalian inner ear, hair cells in the zebrafish lateral line have emerged as an outstanding system for studying hair cell death. The external location of the hair cells allows for direct access and visualization of the hair cells, in order to study the cellular pathways underlying hair cell death, and for large drug library screens to identify compounds that protect against ototoxic effects. As a result of a drug screen and subsequent compound analog analysis, a compound named ORC-13661 has been found to be robustly protective against several aminoglycosides in a dose-dependent manner. We have found that there are distinct mechanisms that underlie antibiotic-induced toxicity, distinguished by different temporal events. Some aminoglycosides, including neomycin, kill hair cells rapidly, within one hour of exposure. Other aminoglycosides, including gentamicin, have a much slower time course, taking up to 24 hours to kill hair cells after an initial one-hour exposure. We find that ORC-13661 provides maximal protection to both classes of aminoglycosides if it is present at the same time as the toxic compound. Additionally, the delayed aminoglycoside exposure paradigm gives us an opportunity to use compounds with known effects on cell death pathways to shed light on new mechanisms underlying hair cell death.

Joubert syndrome (JS), a rare recessive genetic disorder, is characterized by abnormalities in the cerebellum and brainstem. Notably, JS is a ciliopathy because the known causative genes have products that localize to the primary cilium, a vital organelle acting as the nexus between many signalling pathways. Understanding more about JS will provide insights into this emergent group of disorders revolving around the cilium. Causal mutations for JS need to fulfill three criteria: They need to be rare, deleterious, and present in both alleles (copies of a gene) in the patient. Patients in our cohort are sequenced for rare deleterious variants (RDVs) to identify variants that satisfy the first two criteria. To determine whether the segregation pattern of the variants satisfies the third criterion, we sequence both parents, each of whom should carry one of the variants. My research uses next-generation sequencing to determine the segregation pattern for families in whom RDVs have already been identified. Specifically, I am using molecular inversion probes (MIPs) to target-sequence putative causal mutations in the parents and then using the sequencing data to determine the segregation pattern of these variants. If the MIPs data show RDVs for a causative gene are on the same allele, then the variants cannot be the cause of JS in the family and we will have to consider other variants or perform additional sequencing to identify the causal mutations. The data will likely show that the mutations are in both alleles because the mutations targeted with MIPs are mutations that have passed the filters for being rare and deleterious and are robust candidates for being causal. Ultimately, the goal of my research is to elucidate the genetic basis of JS in hopes of improving risk recurrence analysis, prenatal diagnosis, and help discover biological mechanisms of JS.

The advancements of microelectronic structures for high performance electronics are now turning to 3D chip integration technology. The use of vertical connections with high aspect ratios, called Through-Silicon Vias (TSVs), is becoming a popular method for fabricating electronic multi-chip systems and devices. Since these vias do not run along the surface of the wafer, the high density of the vertical vias allows for more interconnections to be made between chips in a stacked system. The goal of this research was to establish the procedure in which TSVs are produced to maximize the highest quantity of connections (maximum packing of number of number of usable vias per unit area). The experiments that were performed involved the fabrication of TSVs while addressing possible reasons for error in each step. The steps that were improved upon and are discussed include best photoresist spinning techniques, minimizing time between exposure and development, and best development techniques. These improvements offer a higher repeatability of the procedure to result in the same yield and quality of TSVs for future production.

Little is understood about how supermassive black holes affect their host galaxies in terms of growth and shape. Using a large simulation called Romulus25, we are able to study how galaxy properties correlate with black hole properties throughout time. From this simulation we draw a sample of 40 massive galaxies and analyze their properties to understand how they are affected by the growth of supermassive black holes. We examine galaxies with a variety of star formation histories and investigate how the black hole is involved in determining these histories. We seperate the galaxies into three categories: 1) Galaxies with on going star formation 2) Galaxies that stop forming stars 3.) Galaxies that stop and then restart star formation. We show how the black hole activity changes between categories. These results may lead to solving the mystery: what causes the star formation in the galaxies to quench, (and in some cases) un-quench? 

Alzheimer’s disease (AD) and cancer are two of the most devastating public health challenges in modern society costing 361 billion dollars and 685,240 lives each year. As these two classes of pathologies progress they both generate protein stress inside cells. Tumor cells are often characterized by greatly increased protein production while most neurodegenerative diseases including Alzheimer’s disease, Parkinson’s disease, and Huntington’s disease are classified as a “proteinopathies”; characterized by the accumulation of the misfolded proteins. Some recent reports suggest that the Parkinson’s disease related protein α- synuclein influences cancer growth in mice and cell culture models. The goal of our project is to uncover the cellular pathways that are shared between neurodegeneration and oncogenesis. We will study this using the C. elegans genetic model system. I have generated a transgenic line that ubiquitously expresses a human form of the AD-related protein, Amyloid-βeta, and observed that it develops similar tumors to cancer-model worm lines. We are also generating transgenic worm lines that ubiquitously express the other canonical neurodegenerative proteins α-synuclein and Tau to see if they also form tumors. These transgenic lines will allow me to knock down different cellular pathways using RNA interference (RNAi) which silences or inhibits the gene expression using double stranded RNA. First we will look chaperone functioning because this class of proteins is necessary for the cells to deal with protein stress. Research in our lab has already shown that RNAi of some chaperones decreases the occurrence of tumors in a worm cancer line. Next, we will generate worm lines with fluorescent reporters of the tumor altering genes and reporters of cancer-related cell-signaling pathways. Thus, we will begin to uncover the ways in which AD-related protein stress impacts cell processes that promote cancer. This could lead to new gene targets for future therapies.

In the US, someone needs blood every two seconds, making blood transfusion a frequently performed procedure in hospitals. Transfusion of incompatible blood due to clerical error in the clinical blood typing procedure can induce a multitude of life-threatening immune responses in the recipient’s body. The most prevalent typing procedures are based on the overly simplified characterization of surface antigens on an individual’s red blood cells (RBCs) into the commonly known ABO blood type system. However, studies have shown that the antigens embedded in the RBC membrane, comprising of glycolipids and glycoproteins, are structural determinants of variations in the blood type outside of the ABO system, elucidating the limitations of current typing methods. This project follows the hypothesis that gas-phase hydrogen/deuterium exchange (HDX) MS techniques can be employed to characterize and further elaborate the structure of these RBC antigen constituents. To test this hypothesis, genotypically and phenotypically defined donor blood samples are processed to obtain RBC ghosts, containing only the membrane components with the hemoglobin removed through an osmotic lysis procedure. Glycoprotein and lipid fractionation, followed by digestion are employed to isolate carbohydrate components in preparation for HDX-MS analysis, which provides insight into structural composition of the carbohydrate antigens. Through this study, we expect to develop a deeper understanding of the variations in the carbohydrate RBC antigens which dictates the blood type of individuals. This knowledge will be crucial in developing glycobiological sensors that can be functionalized onto silicon chips used by Ratner Lab, facilitating novel blood typing techniques.

My project focuses on macroscopic analysis of indigenous pottery from the Iron Age in Mallorca, Spain. My first goal is to develop an efficient method of analysis that can be used with large samples of pottery collected in the Landscape, Encounters and Identity Archaeology Project. I am recording a number of traits for individual ceramic pieces, such as color, texture, and temper. For each of these traits, I am evaluating the replicability, speed, and amount and types of tools needed to record them. My second goal is to use this information to understand the variability within indigenous pottery. Traditionally, this pottery has been divided into two types based off of fairly rigid criterion, which masks the potential variability within each type. This method of analysis, along with multivariate statistical analysis of the data collected, will help to characterize the variation within each type in order to better understand indigenous pottery production techniques.

Joubert syndrome (JS) is an autosomal recessive neurodevelopmental disorder, characterized by the "molar tooth sign" on axial brain MRI. JS is caused by mutations in 35 known genes, all of which encode proteins that localize to the primary cilium, a microtubule-based organelle that projects from the surface of most cells. Our current understanding of the underlying molecular mechanism causing JS involves lack of ciliary localization of ARL13B and INPP5E, two JS-associated proteins. ARL13B is required for the ciliary localization of INPP5E. Normal INPP5E localization maintains a phosphatidylinositol subtype distribution in the ciliary membrane. My research investigates ciliary phenotypes in cell lines harboring causal mutations in C2CD3, which encodes a protein that localizes to the primary cilium’s basal body, using existing images of patient cell lines. Loss of C2CD3 function in mouse models has presented lower ciliation rates and shorter cilia lengths. Previous research in our lab on two other JS-associated basal body proteins, OFD1 and KIAA0586, identified normal ciliary ARL13B-INPP5E levels. To determine ARL13B and INPP5E localization, cells from two control and two different C2CD3 patient cell lines were grown and serum-starved, inducing ciliation. Cells were stained with antibodies to acetylated tubulin (marking cilia), and INPP5E or ARL13B (marking protein of interest). Using Fiji, an image analysis software, I “painted” each cilium by outlining the signal in the acetylated tubulin reference channel to define a mask. Using that mask, I quantified ciliary ARL13B and INPP5E. I measured ciliation rates using the number of masks divided by the number of nuclei, and cilium lengths using the “skeleton length” function. Based on data from mouse models, we predict that C2CD3 patient lines will have fewer and shorter cilia, but normal ARL13B-INPP5E localization. These expected data would exclude the hypothesis that INPP5E mislocalization is an obligate part of the biological mechanism underlying JS.

What impacts did colonial bias create regarding the representations of the Tahitian people in discourse surrounding the 1995 reinstitution of French nuclear testing in French Polynesia? French nuclear testing in French Polynesia occurred from 1966-1996 at Mururoa and Fangataufa Atolls. Nuclear testing was suspended for 3 years from 1992 until 1995, and was met with great resistance and riots in Tahiti by the Tahitian people as well as worldwide when it resumed. I used critical discourse analysis (CDA), drawing off ideas from Machin and Mayr, to critically analyze different forms of discourse surrounding the representations of the Tahitian people upon the reinstitution of nuclear testing. CDA is a useful tool to expose power imbalances within discourse and to challenge and disrupt the power and status quo within discourse. This creates a space to actively challenge and work against the discourse at hand, while providing an opportunity for new forms to be produced. My research shows whose voices are missing from discourse, thus allowing discourse from multiple voices to be produced in the future. Furthermore, this research serves as a basis for undergoing CDA on discourse surrounding other nuclear testing sites, allowing the discourse to be challenged and changed to provide accurate and well-rounded representations surrounding nuclear testing.

Both content knowledge and problem-solving strategies are important components for academic success in biology. All problem-solving strategies begin with the ability to understand the problem being presented. In the context of biology, this means identifying the core concept being asked in complex questions. Identifying keywords, words that are important or significant, can help students identify these core concepts. The purpose of this study was to explore how well students can identify keywords and if this ability is correlated to exam performance. Students in an Introductory Biology course took weekly practice exams (PE), after each PE they either graded their own or a peer’s PE. In addition, students were asked to “identify the keywords in the question that should have caused you to provide the complete and correct answer given in the key”. This experimental design also allowed us to explore how grading an anonymous PE affects quality of students’ keyword selections. Students’ keyword selections were compared to a keyword rubric created by the research team. Data was coded by two researchers to 90% inter-rater reliability. On average, students only correctly identified 39% of keywords each week. Multiple regression models were used to explore correlations between correct identification of keywords and performance on both PEs and course exams. Interestingly, students who self-graded were more consistently engaged with the keyword activity as compared to peer-graders. In both cases, percent keywords correct could predict overall exam performance. These results indicate that if instructors want to help students improve their problem-solving skills of complex biological problems, helping students recognize keywords may be one solution. Furthermore, identifying which students have trouble simplifying complex exam problems and helping these students improve their ability to identify keywords may be beneficial in addressing learning disparities within large introductory classes.

Chronic pain affects millions of people worldwide, and current treatments are often ineffective and come with unwanted side effects. Modern research focuses on studying the mechanisms of pain sensation in the nervous system, with the goal of using our improved knowledge to develop more efficient analgesics. Our research focuses on neurons located in the Trigeminal and Dorsal Root Ganglia (TRG and DRG), as they are known to play an important role in the sensation of touch and pain, and on genes regulating the development and function of these neurons. Recent advances in gene editing methods, particularly the CRISPR/Cas9 system, allow us to mutate specific genes of interest a living animal and monitor the effect of such mutations within a matter of days. To this end, we have developed a behavioral assay which determines the effect of CRISPR induced mutations on pain sensation in zebrafish larvae. CRISPR injected larvae are exposed to noxious stimuli such as AITC (mustard oil) and harmful temperatures. The CRISPR treated larvae’s response to the painful stimuli determines the impact of the specific gene we targeted on nociception. Through this assay we have identified two genes, zmat4b and zbtb7b, as potentially important to the sensation of noxious stimuli. Further experiments are now required to determine the precise mechanisms through which these genes play a role in the sensation of pain.

Nearly 94% of adolescents report daily online smartphone use; thus, understanding specifics regarding time spent on smartphone apps is important. Self-reported time spent on apps has been shown to be unreliable. New methodology is needed to better understand app use among highly connected adolescents. The purpose of this study was to test a novel methodology to investigate smartphone application use among older adolescents. A sample of freshman at a large university was recruited through freshman interest groups and university forums. For 9 sequential weeks participants submitted weekly screenshots of their mobile application use showing their most-used apps including social media and messaging tracked by battery usage. Incentives were given at both week 5 and week 9. Analysis included paired t-tests. We evaluated a total of 134 screenshots (82.7% response rate) submitted by 18 participants who were age = 18 (SD=1.1), 72% female. The average number of apps listed on battery usage was 16.1 (SD=1.6). Average battery use of social media apps was 16% (SD = 6.2) and average battery use of messaging apps was 11.8% (SD =6.8). Among social media, Snapchat and Facebook were among the most popular apps used with all participants having both/one consistently in their top 5 battery usage for at least 8 weeks. A total of 9 participants had “home/lock screen” among their top 5 battery usage for 3 or more weeks. These 9 participants had an average of 30.2% of battery usage for their most used app on weeks when home/lock screen was among top 5 apps, compared to 18.2% on weeks without home/lock screen in their top 5 apps (p=0.01). Phone tracking and screenshot data collection offers a novel and feasible methodology to understanding app use among highly connected populations. Additionally, frequent home/lock screen may be an indicator of compulsive phone checking.

American Crows (Corvus brachyrhynchos) form predictable groups called pre-roost aggregations shortly before moving to their communal roost each night. Tens to hundreds of crows reliably congregate on these highly vocal aggregations, and, thus, they provide an excellent opportunity in which to investigate social communication. We predict that there are patterns between call characteristics (caw quality, number of syllables, etc.) and behavioral responses of listening crows on pre-roost aggregations. We tested this by placing remotely activated field cameras in strategic locations on the Discovery Hall building rooftop at the University of Washington Bothell campus, a known pre-roost aggregation site. These cameras took video every five minutes throughout the pre-roosting period on multiple evenings. We used charcoal to mark a grid on the roof to keep track of crow movements during each recording. We then analyzed the video recordings taken each night, noting all discernable vocalizations and movements. Each approach, withdrawal, vocalization, and dispersal behavior exhibited within five seconds of a call was categorized as a potential “response” to that call. We have begun to correlate the acoustic characteristics of crow calls to the observed behavioral responses. Preliminary qualitative analysis suggests listening crows are more likely to withdraw from a caller giving one versus multi-syllable calls, and crows typically only respond with a vocalization to calls with more than one syllable. We will use data gathered in this study to create audio files of calls to use in future playback studies to experimentally determine if certain crow call characteristics elicit specific behavioral responses.

Supernovae enrich the interstellar medium, allowing it to create fascinating and exciting objects such as stars, planets, and people. However, supernovae are difficult to study because, most of the time, we do not observe these supernovae directly, since the whole event can occur in a very short time. Fortunately, they leave behind bright remnants that can give us information about the supernova and even the stars that existed before. We have observed one such supernova remnant in the nearby spiral galaxy NGC 300, which is approximately three million light years away from Earth. We have acquired multi-wavelength data of this source from Chandra X-ray Observatory, XMM-Newton X-ray Observatory, and Hubble Space Telescope. Using this variety of telescopes, we can understand how this supernova remnant behaves in both the x-ray and optical energy bands which gives us a more complete picture of our source. From our Hubble data, we gain detailed measurements of the size and shape of the source. From our Chandra and XMM-Newton data, we can shed light on the properties of this supernova remnant by performing a spectral analysis on the x-ray data. This spectral analysis gives us information about the temperature and metallicity of the source, as well as the density of the surrounding gas. With these quantities, we can better understand this supernova remnant and supernova remnants like it which, in turn, let us better understand the life of the star that created this source.

Climate change has lowered the reproductive success of the Magellanic penguin (Spheniscus magellanicus) population at Punta Tombo, Argentina contributing to the population’s 37% decline since 1987. While increased rainfall and storm intensity directly affect reproductive success through chick mortality, the indirect effects of weather can also impact fitness. We predicted that temperature would alter penguin behavior in two ways: 1) fewer penguins would leave the colony on hot days since traveling in heat requires more energy, and 2) penguins leave earlier on hot days to escape the highest temperatures of the day. Automatic scales that record weight, time, and direction were placed where penguins frequently exit and enter the colony. The scales also recorded the Radio Frequency Identification (RFID) number of the penguins for 5,000 crossings, allowing us to calculate their trip duration. Days were considered hot if the maximum temperature was greater than two standard deviations above mean maximum temperature (>28℃, n=11 days) and cold days were less than two standard deviations below (<20℃, n=11 days). Temperature did not affect when penguins exited or entered the colony, however, more penguins exited and entered the colony on hot days than on cold. A greater number of penguins took short trips (<= 12 hours) on hot days than on cold days, suggesting they only go to the beach to cool down, rather than to forage. Increasing temperatures could lead to parents leaving their chicks alone at the nest for longer durations of time without foraging which could increase chick mortality.

It is widely recognized that there is limited understanding of many animal- plant interactions and the complex networks they form with other species. I conducted an observational study, in the Coshu Cashu field station in the Manu National Park in Peru, to determine the plant-pollinator relationship that Zygia coccinea, a flowering tree in the Fabaceae family, has with the insects that visit it. Very few studies have been conducted on Z. coccinea and the few that have been conducted focused on classifying the characteristics of the plant and did not look at the plant’s ecological role in the environment nor at other species associated to it. It is important to understand the dynamic interaction between vegetation and their insect population to gain a more thorough understanding of the complexity of the ecosystem dynamic and how it may be impacted by alterations in the composition of species present. I used two different observational techniques, scan and focal, in order determine the diversity of the insect network associated with Z. coccinea. I observed 101 different individuals, classified 19 distinct morphospecies, and identified individuals in seven different Orders. In particular, we observed an insect of the Order Homoptera, a small cicada-like pink pigmented insect that was found being herded by ants (similar to other ant-aphid interactions) which seems to be a new species for science, and further research needs to be conducted. This study is important because it contributes to the knowledge of specific insect-plant interactions. In addition, it was the first time an insect visitation network was identified for Z.Coccinea and upon the establishment of this network, a potential new species to science may have been found.

The ubiquitous presence of technology in daily life has made for a multifaceted discourse on rethinking how to approach teaching, learning and studying in the modern age of distraction. Across the globe, individuals from various walks of life are worried about children, their lack of engagement with nature and the hazardous ramifications to come as a result. Richard Louv gave this growing disconnect an identity when he termed it “Nature-Deficit Disorder”; which looks at the human costs of alienation from nature. The increased dominance of screens in childhood upbringing not only enables a series of rather unsettling childhood trends such as obesity, depression, and attention disorders, but it also strengthens the societal narrative that technology is the future, and nature is a thing of the past. In this research, I explore the inner workings of the outdoor and nature-based preschool movement in Seattle, Washington. How can preschools be more effective in promoting a reverence for nature to the upcoming generations? In particular, I look at how preschools promote a reverence for nature, how children experience and define nature when afforded such exposure at a pivotal time developmentally and how that differs from past generations, and how outdoor and nature-based education models help to expand and redefine “quality standards” that contribute to the achievement gap. In order to answer these questions, I will partake in critical observation of two classrooms from separate outdoor nature-based preschools. Analysis of current curriculum and education-models highlight ways in which these institutions aim to foster an innate sense of appreciation of the natural world for their students. A series of series of interviews with teachers and administrative staff will provide insight into the progression and perceived trajectory of the outdoor and nature-based preschool movement and a better understanding alternative educational pedagogies.

Many infectious diseases are caused by bacteria growing in biofilms, a group of cells that stick together and adhere to surfaces, such as the human lung or implanted medical devices. Biofilms affect millions of people worldwide each year, often contributing to death. When a biofilm is successfully established, bacteria are up to 1,000-times more resistant to antimicrobial agents and extremely resistant to clearance by the immune system. These molecular fortifications make bacterial infections difficult to treat, with the possibility of relapse weeks or months after the use of antibiotics. Many infectious diseases caused by biofilm-forming bacteria disproportionately affect the elderly and immunocompromised, like Legionella and Bordetella, which cause pulmonary infections. My research focuses on preventing biofilm formation, and forcing bacteria that have already formed a biofilm to disperse. Molecular engineering was used to design a protein to activate LapG, a protein effector responsible for balancing whether the bacteria are free floating versus anchored. When LapG is active, bacteria shed their adhesive proteins anchoring them to surfaces, and become free floating. I am trying to develop a protein-based drug to fight biofilm-based infections by permanently switching LapG to the “on position.” Because this drug will be made of protein, there will be no toxic breakdown products, and the highly specific binding inherent to protein-based drugs will minimize the risk of side effects. Furthermore, protein-based drugs are genetically encodable, allowing us to genetically modify self-replicating probiotics, such as yogurt cultures or baker’s yeast, to produce this medicine. This will empower low cost drug manufacturing and distribution because local communities in developing countries can produce these therapeutics using readily available cooking equipment and ingredients, and administer them without fear of overdose or toxicity. The goal is to test the LapG binder in animal models of chronic infection, eventually moving to human clinical trials.

Type II diabetes mellitus (T2DM) is a prevalent disease that relates to pancreatic islet beta-cell failure. With no cure and numerous secondary complications - blindness, kidney failure, heart attack, and stroke - T2DM places an enormous burden on our society and healthcare system today; the disease is projected to be the seventh leading cause of death by 2030. Within approximately 90% of T2DM cases, the majority of the amyloid deposits is made up of amylin, also known as islet amyloid polypeptide (IAPP). Unlike other organisms, human IAPP is capable of self-aggregating and forming amyloid fibers or beta-sheet plaques through its unique chemical properties. Our group’s previous findings suggest that T2DM and other amyloid diseases contain misfolded protein aggregates and form toxic oligomers through a non-standard alpha-sheet secondary structure. We propose a peptide-based therapeutic to combat T2DM via utilizing synthetic alpha-sheet compounds complementary to the amyloid-associated alpha-sheet structure to target the toxic oligomer form of IAPP. In this study, the efficacy of this peptide-based therapeutic is assessed through inhibitory and toxicity reduction effects of designed peptides on IAPP aggregation and oligomer cytotoxicity. The designed peptides are co-incubated with IAPP and their aggregation is monitored via a Thioflavin-T assay. The preliminary results on IAPP suggest that our peptides are effective in reducing IAPP aggregation and allow us to conduct cytotoxicity experiments. Effects of our compounds on cell viability are currently being conducted on a human-derived neuroblastoma cell line (SH-SY5Y). The cytotoxicity of oligomer determined using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) cell viability assay. MTT is a yellow compound that will be reduced to purple formazan only in living cells. The cells are plated in a 96-well plate with 15x10⁴ cells/mL.The IAPP is co-incubated with designed peptides and applied to cells. The results of the MTT assay provide the relationship between inhibition of oligomer formation and reduction of oligomer toxicity. The alpha-sheet platform provides a novel potential therapeutic for treating T2DM and other amyloid diseases.

Since DNA replication and transcription are concurrent in most bacteria, collisions (or conflicts) occur between these two processes. Conflicts have deleterious effects on cells, including double strand breaks and mutagenesis, especially when a gene is transcribed head-on to replication. Because conflicts increase mutagenesis in specifically genes that are oriented head-on to replication, this process may be a conserved strategy for targeted evolution. Consistent with this, a bioinformatics analysis determined that many genes in the head-on orientation are involved in stress response. We previously showed in B. subtilis that the error prone translesion synthesis (TLS) polymerase YqjH is required for the asymmetric mutagenesis of genes in the two orientations and that transcription-coupled nucleotide excision repair (TC-NER) is also involved in this repair pathway. Mutation rate (or reversion assays) and survival assay experiments performed under DNA damaging conditions, however, indicate that a different TLS polymerase, YqjW may be responsible for mutational asymmetry, and that YqjW functions in the same pathway as TC-NER. Thus, YqjW may facilitate targeted evolution under DNA damage conditions. 

As part of my Cultural Anthropology class I conducted an oral history interview of Tom Ficca (Colville) with my group to help document the history of Edmonds Community College’s involvement with Indigenous communities for its fiftieth anniversary. I learned that sharing culture and spirituality has played a significant role in Tom’s life and helped connect him with the college. He described his childhood as “phenomenal” and valued his connections to his community on the Colville Reservation over the past 55 years. He is a spiritual leader and a master carver. He has helped build canoes for different tribes and connected with Edmonds CC about five years ago through canoe journeys and carving projects with the Snoqualmie Tribe, where he worked as a carver and skipper. As a carver he collaborated with students and faculty in the construction of q'wəld'ali (Place of the Cooking Fire) or Cultural Kitchen on the college campus. As a skipper he has welcomed students to support and paddle on Tribal Canoe Journeys. Over the last three years Tom has been working with families that are in the streets in Anchorage Alaska where he helped set up a church and non-profit organization to provide food, water, gloves, socks, dry clothes, etc. throughout the year. Tom Ficca wishes to start a strip cedar canoe as well as moving forward on cultural projects such as canoe building, story pole carving, and plank house construction. Tom would also like to see more future work and involvement in the community and in different garden areas. All these accomplishments and other works to be done in the future are driven by Tom’s passion and commitment to culture and his spiritual side.

Student-graded practice exams (PEs) have been used to facilitate learning in large classrooms. In a large introductory biology class, we investigated the impact of self vs peer-grading of PEs. We hypothesized that students who graded their own PEs would grade more accurately than peer graders because they are more motivated and reflective in their grading. Over time this more accurate grading would lead to increased performance on course exams. One question from each week’s PE was graded by a team of experts. We calculated PE grading accuracy by comparing the expert grade to the grade assigned by the student. We analyzed the correlation of PE grading accuracy with exam performance. For each metric we investigated the impact of treatment, incoming GPA, and student demographics (e.g., gender, socioeconomic status, first generation, race/ethnicity) using general linear models. There was no difference in average expert PE grade between the two treatments. Incoming GPA was the only significant predictor of expert PE grade with GPA positively correlated with expert PE grade. There was a significant interaction between incoming GPA and treatment on grading accuracy. Students with high GPAs who graded peers consistently underestimated their peers’ PE performance. Students with lower GPAs who graded peers either graded similarly to experts or slightly overestimated their peers’ PE performance. Students who graded their own PEs more accurately predicted their PE performance regardless of incoming GPA. There was an interaction between GPA and treatment on course exam performance, as high GPA self-graders earned more course exam points than high GPA peer-graders. Average GPA students earned the same amount of course exam points regardless of treatment. Given these findings and the logistical complications of peer-grading in large classes, instructors can use the more easily administered self-grading method of providing feedback on low-stakes practice exams.

A growing number of parents are opting out of vaccinations for their children, which presents a prominent public health risk. Studies have shown that parents are now more likely to look to Web 2.0 communities, such as blogs, than rely on medical expertise for health advice, especially on vaccination. However, there is scant literature outlining how parents contribute to online discussion of vaccination. The purpose of this study was to explore parent involvement in online vaccine communication, by analyzing content in a setting that is informal, unmediated, easily accessible, and intended to express vaccination opinions to the non-descript, lay, parent population. Content analysis was conducted to evaluate a sample of online comments on popular, parenting blog posts on vaccination. Investigators entered the search terms “top parenting blog vaccination posts” into the Google search engine, and included posts that met specified inclusion criteria, such as parent-focused and parent-written. This sample was analyzed using an adapted codebook, vetted through inter-rater reliability testing. The codebook included variables for participant discussion role, examples include providing information or asking questions; stance: pro, con, or neutral; and argument rhetoric, including emotionalization or fact-based. The comment dates were also mapped over a timeline of relevant, vaccine-related events (i.e. Measles outbreak at Disneyland). Investigator analysis compared types of rhetoric and content by stance of the commenter. Statistical and qualitative analysis of this data was performed to reveal trends towards better understanding of anti-vaccination sentiments, and inform efforts to improve vaccination coverage. A divergence is anticipated in the rhetoric and discussion contribution between the two sides of the conversation. Preliminary results favor a tendency of pro-vaccination comments to rely on scientific evidence, and anti-vaccination comments to rely of emotionalization.

Effective drug delivery to solid tumors is made difficult by high cell density and poor blood flow to certain areas within the tumor. Previous studies have shown that pulsed high-intensity focused ultrasound (pHIFU), using highly focused pulses of ultrasound with a slow repetition frequency, can increase drug delivery to tissue through bubble formation. For this technology to become viable in a clinical setting, it is necessary to develop an image reconstruction technique to detect and characterize bubble activity. Passive cavitation detection (PCD) is the gold standard in the field to characterize bubble activity, but it provides minimal information on where the activity occurred. Our work focused on developing and refining previous Doppler-based imaging techniques that were shown to be sensitive to bubble formation, size and location. pHIFU pulses were delivered to optically transparent tissue-mimicking gel phantoms using either a HIFU or diagnostic ultrasound probe. The ensuing cavitation bubble activity at the focus was evaluated with three different methods: Doppler-based ultrasound imaging, PCD and direct observation using a high-speed camera. Ultrasound imaging was performed by an ultrasound probe connected to the Verasonics programmable ultrasound system, and the imaging data was post-processed in Matlab. The Doppler-based cavitation maps generated under varying pHIFU exposures and Doppler sequence parameters were then compared against PCD measurements and high-speed video recordings to select the most sensitive regime and the best quantitative metrics to characterize the cavitation-induced damage from these maps. Preliminary analysis indicates that integrated Doppler power in the region of interest correlates to the cavitation dose measured by PCD, which, in turn, was shown to correlate to the degree of tissue permeabilization.

Mutator phenotypes due to mutations in genes encoding DNA polymerases or mismatch repair proteins lead to increased error rates during DNA replication that accelerate the evolution of cancer cells and contribute to chemotherapy resistance. Work in the yeast Saccharomyces cerevisae indicates that excessive DNA replication errors can lead to error-induced extinction (EEX), where every cell within the population dies due to a random lethal mutation. Thus, a possible direction for cancer therapy may be to modulate mutation rates of mutator cells – to either increase mutation rates to a lethal level or suppress mutation rates to their baseline levels in order to slow the rate of tumor evolution. Genetic pathways that may be targeted to modulate mutator phenotypes have yet to be fully elucidated. To identify such pathways, we isolated spontaneous haploid yeast eex mutants that escaped from error-induced extinction caused by combining a mismatch repair defect (msh2Δ) with the pol2-L439V allele, which models a human cancer mutation affecting DNA polymerase epsilon. We crossed the eex mutants to wild type yeast, generating diploid strains that were heterozygous for msh2Δ, pol2-L439V, and the eex mutation. Following sporulation, we measured the mutation rates of haploid MMR-proficient pol2-L439V progeny in order to assess the presence or absence of the antimutator mutation. For each mapping experiment, cells showing evidence of suppressed mutation rates were pooled separately from those exhibiting the normal pol2-L439V mutator phenotype. Unique mutations present only in the antimutator pool were identified by Illumina Next Generation Sequencing. Candidate eex mutations were engineered into fresh strains to assess the antimutator phenotype. Our compiled list of antimutator mutations and their functions in yeast may allow us to identify analogous mutations in the human genome and explore new methods of cancer therapy by which specific genes can be targeted to lower mutation rates to normal levels.

Teaching undergraduate engineering students about the design process can be a difficult task yet is a critical part of an engineering education. A common method for design teaching involves the use of prescriptive design models, processes, and diagrams that indicate how one “should” do design. In this study we looked at student insights about design through the use of descriptive models, rather than prescriptive models. We question how engaging students in research data about design and descriptive models can be used as a teaching methodology that allow students to draw conclusions and further inform their design learning. In this study, 90 undergraduate engineering students from two different courses participate in a design activity. This activity engages students with descriptive visual data of first year and senior student design processes. Throughout the activity, students fill out a paper worksheet with 6 open response questions. A subset of 30 worksheets were randomly selected for preliminary analysis. Three researchers qualitatively coded the 6 questions for student insights in a grounded bottom up manner and a code book of student insights were developed. These codes were generated through iterative individual coding and group discussions between the three researchers. These qualitative codes have been applied in future work (outside the scope of this project) to the remainder of the student worksheets. The poster presentation displays the core results of student insight codes, preliminary conclusions, and our research process. However, this research is still an ongoing project. From this study we better understand how students can learn design from an activity that actively engages them in research data about design.

Over the past few years’ silicon photonics has proven to be a viable bio-sensing platform for various diagnostic applications. The small size, label free detection and ability to be multiplexed with various assays make it a very attractive technology and potential replacement for many clinical tests. Silicon photonics has proven to be a viable technology for ABO blood typing. This technology has the potential to achieve a higher level of automation than the current clinical method along with a lower run time and lower costs. Two large limitations in clinical translation of this technology are reducing the time it takes to run the assays and the amount of processing that must be done before running them. Indirect blood typing using this technology currently requires centrifuging whole blood to isolate plasma or serum to use in the assay. This sample processing can take anywhere from 20-40 minutes and is not ideal for a clinical setting. This project seeks to increase silicon photonics clinical viability by designing an inline filtration system to isolate the plasma or serum from whole blood as it is running through the assay. This would mean that whole blood would not have to be centrifuged for indirect blood typing. Instead, the blood will be filtered in the assay and then travel through a paper network for reagent delivery recently developed in the Ratner Lab. Additionally, this assay will be developed for the commercial Genalyte silicon photonic test setup. This requires the development of a custom mount for the system along with determining positioning for the paper network. This research may help to replace the agglutination diagnostic test that has been in use for over 100 years and help save medical and monetary resources as well as patient lives.

Expansion Microscopy (ExM) is a revolutionary fluorescence microscopy technique that enables super-resolution imaging of fixed biological specimens on ordinary microscopes. By chemically incorporating the specimens into a hydrogel polymer matrix, enzymatically digesting the proteins that hold the structures together, and physically expanding the hydrogel complex, this technique yields expansion of fourfold along each x/y/z axis and a spatial resolution of ~70nm. The original ExM approach utilized special tri-functional antibodies containing a DNA-dye-linker complex to attach the fluorescent dyes to the expandable polymer. Previously, our group has developed an improved and simplified ExM protocol, which is compatible with conventional antibodies and fluorescent proteins, by using only commonly available reagents, yielding increased fluorophore retention for cultured cells and tissue specimens. In this project, we extend this new approach of ExM to enable ultrahigh resolution imaging of Drosophila melanogaster tissues which had previously been incompatible with expansion. To do so, we adapted our existing ExM protocol to Drosophila embryos, larval brains, and larval and adult body walls by utilizing an enzyme system that can efficiently digest the tough outer cuticle of the organism. We assessed the uniformity of expansion achieved by our new procedure using correlative pre-expansion and post-expansion imaging and found the distortions to be minimal (<2%). With our new ExM approach, we were able to resolve features of presynaptic active zone structure and study the interactions between the epidermis and dendrites of somatosensory neurons in Drosophila. This work should open up this powerful super-resolution microscopy method to a large community of researchers who utilize Drosophila as an important model system in biology.

Aneuploidy is an irregular number of chromosomes caused by errors in cell division. Down syndrome is a common disease resulting from aneuploidy, where there are three copies of chromosome 21. I am in interested in why these errors occur and the mechanism that corrects them. Prior to anaphase, an attachment of microtubule fibers to chromosomes must occur to allow chromosomes to segregate properly. The attachments form by trial-and-error. Microtubule fibers will continue to attach and detach to chromosomes until stability is achieved. In the 1960s, Dr. Bruce Nicklas manipulated individual chromosomes using a microneedle to physically misalign chromosomes and pull on chromosome attachments to create artificial tension. The Asbury Lab is interested in recreating his experiments using tools that were not available at the time, such as small molecule inhibitors, to determine whether certain molecules such as Aurora B kinase are necessary for chromosomes to correct themselves. I use drugs such as ZM, an Aurora B kinase inhibitor, and Taxol, a microtubule polymerizer and common chemotherapy agent. I treat live cricket cells with drugs such as these, and record the treatment’s effect on meiosis. One common phenotype we are expecting is a delayed transition from metaphase to anaphase due to inhibition of a protein required to allow the cell to continue into anaphase. Once my research determines whether the drugs delay cell division and the effective concentrations, the lab can test the importance of the drug’s molecular targets in the error correction mechanism. Further research into the mechanisms of chromosome segregation has implications for understanding what causes aneuploidy, the mechanistic and biophysical mysteries of cell division, and the rising of cancerous cells.

The forefront of modern medical research involves decoding the biological pathways that make disease transmission and perpetuation possible. Protein-based site-specific affinity reagents could allow us to antagonize harmful molecular interactions employed by pathogens, disrupt their toxic effects, and develop cutting-edge treatments. Consequently, the ability to express and purify rationally-designed proteins is the key to assessing their structure and function. To increase solubility and stability during recombinant expression, a genetic fusion to a small-ubiquitin-like-modifier (SUMO) is commonly used. The highly-specific SUMO protease is then used to cleave the SUMO fusion, leaving a completely intact and unmodified protein-of-interest. The current protease used originates from Saccharomyces cerevisiae. Known as Ulp1, the instability of this enzyme due to thermosensitivity and solvent-exposed hydrophobicity presents considerable limitations and can result in incomplete digestion of SUMO-tagged constructs. We present two approaches to mitigate these complications. First, we have discovered a homologous protease with a conserved catalytic site in the thermophilic eukaryote Chaetomium thermophilium. Second, using the Baker Lab’s pioneering computational design program, Rosetta, we have exchanged the hydrophobic surface residues of Ulp1 with hydrophilic side chains with the aim of improving stability in aqueous solution. We have characterized the kinetic function of these protease variants by monitoring digest efficacy through gel-based assays. Effectively, the improved proteases can equip the scientific community with a powerful new tool to facilitate protein-based drug design research.

How are female athletes, specifically gymnasts, being portrayed in the media? We as a society look at their bodies versus their ability to do the sport. I used critical discourse analysis by analyzing the ways in which athletes are portrayed in gymnastics. I used critical discourse analysis because it allows me to show the bias in reporting of the media and negative portrayals of female gymnasts. Using articles and social media, I showed the way female bodies are being represented in certain words and phrases as well as pictures. This work heavily draws on the theories of David Machin and Andrea Mayr. I found it is important because as a society we need to change the way we talk about athletes and their bodies. It’s not right or valid to judge an athlete on what their body looks like and disregard their athletic ability. Through this research, I can begin to open up the discussion about their athletic ability instead of their bodies.  

Nucleic acid (NA) extraction is a critical step in molecular diagnostic assays, since it can impact the sensitivity and specificity of the subsequent amplification step such as the Polymerase Chain Reaction (PCR). Most commercially-available NA extraction approaches capture both disease target NA and other non-target NAs. However, these non-target NAs compete with the target NA for resources and as a result can reduce the efficiency of PCR to detect a disease target of interest. Here, we present a strategy to specifically capture NA sequence of HIV from lysed infected human CD4 cells. After cell lysis, restriction enzymes are used to excise the HIV pol sequence from proviral DNA integrated in human genome. After a heat-denaturing step, biotinylated DNA probes hybridize to the HIV pol sequence. This solution containing the duplexes of pol gene/biotinylated probes is transferred to a microcentrifuge filter tube containing streptavidin agarose bead resin. Subsequent centrifugation retains the duplexes of biotinylated probes and HIV pol while removing other undesirable non-target NAs and possible contaminants in cell lysate. Finally, the target sequence is eluted from the biotinylated probes and with sodium hydroxide and subsequently neutralized prior to addition to PCR. For initial optimization of this capture system, synthetic HIV pol at a concentration ranged from of 50 – 100,000 copies were used as mock specimens. The method presented achieved 60% ~ 100% HIV DNA target recovery. In the next stage of this work, the effectiveness of capture will be tested with the upstream sample preparation procedures. We are currently investigating capture efficiency and restriction enzyme activity in different sample solute conditions in an effort to streamline the two steps. The successful development of this sequence-specific capture will potentially improve the sensitivity of the PCR assay and allow for early HIV diagnostics.

We are studying the regulation of p120 catenin in zebrafish embryonic development. P120 catenin plays an essential role in the pathway for cell migration and normal embryonic development. Our research on cell migration and the extension of the dorsal axis can be applied to all vertebrates. These functions are also relevant to many clinical issues such as Spina Bifida in humans, wound healing, and cancer metastasis. The particular protein we're studying (p120 catenin) facilitates the activation of Rac1 GTPase when it is bound to both Vav2 and Rac1 GTPase. Vav2 is a guanine nucleotide exchange factor (GEF) that activates Rac1 by exchanging GTP for GDP. Rac1 GTPase activates the actin polymerization and lamellipodia. There has been research suggesting that the dephosphorylation of certain tyrosine residues in p120 catenin is necessary for this binding and subsequent Rac activation. Rac activation promotes cell migration in zebrafish embryonic development. This research has prompted us to ask how phosphorylation at Y217 and co-phosphorylation at Y228-Y217 of p120 catenin affect gastrulation of zebrafish embryos. Based on previous research, we expect embryonic injection of mRNA containing Y217E and Y228E-Y217E mutations to result in abnormal gastrulation. Abnormal phenotypes exhibit delays with cell migration and yoke constriction. In contrast, mutations Y217F and Y228F-Y217F should result in a rescue phenotype similar to the wild type, since the phenyalannine residues are not negatively charged. In order to test this, we designed primers for each mutation, and inserted them using PCR. We used gel electrophoresis to confirm the effectiveness of the PCR by comparing the size of the PCR product with its expected length. We amplify the DNA in E. coli, followed by DNA purification, and sequencing. With confirmed mutations, the next step is linearization of the DNA followed by production of mRNA. We are now in the process of synthesizing mRNA for the multiple DNA mutations we have created. We will then inject them into zebrafish embryos with knocked down wild type p120 catenin and look for resulting phenotypes.

The modern human foot, with its pronounced longitudinal arch (the ratio of difference between mid-foot and total foot area) and suite of derived morphologies, evolved in an unshod context on natural substrates. Today, unshod (barefoot) populations are uncommon, and the majority of modern humans walk shod (wearing shoes) on artificial substrates. The unshod populations that do exist today typically have wider feet than do western populations, but western populations have a more variable longitudinal arch index and therefore exhibit more variation, from “flat” to “arched” feet. The reason for this difference remains obscure. The purpose of this study was to investigate the effect of childhood footwear choices on arch index variation in adults. Participants (56 women, 19 men, mean age: 24±7.2 years) were asked about their footwear, both in childhood (<18 years) and adulthood (>18 years), to classify those who wore rigid/supportive shoes throughout development and those who were frequently unshod or wore unsupportive shoes. Participant footprints were captured using a RSscan pressure plate in three stances: (i) static double stance; (ii) static single stance; and (iii) dynamic. Arch indices were then calculated for each participant and multiple regression with Bonferroni correction used to determine if childhood footwear predicted adult arch index. The results indicate that childhood footwear does not predict adult arch index (all p-values > 0.05). The implications of these results are that longitudinal arch index variability in western populations is not determined by childhood footwear choice. Further research is required to shed light on this phenomenon.

Anti-retroviral therapies (ART) are an effective strategy to treat HIV patients by suppressing viral replication and reducing transmission. However, error-prone HIV reverse transcriptase (RT) can cause single nucleotide polymorphisms (SNPs) associated with drug-resistant HIV. To ensure effective treatment, HIV patients must be tested for drug-resistant SNPs prior to initiating ART. The Oligonucleotide Ligation Assay (OLA) is a lab-based sensitive method for detecting drug-resistant SNPs in HIV, developed and validated by the Frenkel lab at Seattle Children’s Research Institute (SCRI)- however complex workflow has limited implementation in low-resource labs. To address the need for an HIV drug resistance test in these regions, the UW Bioengineering Lutz lab, in collaboration with the Frenkel lab and the UW Bioengineering Lai lab, are attempting to re-engineer the lab-based traditional OLA into an equipment-free, point-of-care format. We are currently exploiting innovative chemistry and autonomous wicking properties of paper-based microfluidics to integrate the OLA into a credit-card-sized cartridge device. Multichannel capillary-based sequential delivery platform is used to supply the reagents required in all main steps of the OLA: nucleic acid extraction, amplification, ligation, and detection, to the different regions of the paper-based device. To enable incubation during amplification and ligation a pull-tab technology is used to physically disconnect the circuit. After these reactions are completed, the tab is pulled to re-activate the next sequential delivery steps. Currently, a proof-of-concept integrated device has been designed. To initially test the microfluidic physics, five fluid sources were colored to permit visualization of the flow. A successful controlled sequential delivery to mimic all steps in the OLA has been demonstrated. Next, the chemistry of each step will be incorporated into this cartridge device. The success of this work will enable non-instrumented, sensitive detection of HIV drug resistance and viral load that requires only two user steps: addition of a rehydration buffer and activation of pull-tabs.

In this paper, we studied the terrestrial mammalian populations through surveying both the northeastern and southwestern sides of Cocha Cashu within Manu National Park, Peru. The northeastern side of Cocha Cashu, which is an oxbow lake formed by the river meandering, is dominated by primary Amazonia rainforest while the southwestern side is mainly an early successional forest. Our study goal was to document the mammalian species diversity in Cocha Cashu by focusing on large-sized mammals. Previous studies have focused on rodents, marsupials, and bats, but the ecology of the larger mammals remains unknown. We conducted our surveying using five camera traps placed on each side of the lake at corresponding locations of projected terrestrial mammalian hotspots. We monitored these sites by checking them once daily and therefore had four nights and three full days of surveying. Through the identification of species from the photos on our camera traps we were able to find the species richness, total number of sightings at each site, and total number of sightings per species as well as comparing the number of sightings and species between night and day. The primary forest side had a total of five species (Leopardus pardalis, Dasyprocta, Cuniculus paca, Mazama americana, and Tapirus) and ten sightings, while the early successional side had a total number of four species (Dasyprocta, Tayassu pecari, Panthera onca, and Tapirus) and six sightings. Ultimately, our results show that primary forests in a tropical climate support a higher diversity of terrestrial mammals, as well as a higher number of individual mammals when compared to early successional forests. These data may assist future researchers in placing camera traps in areas of high mammalian diversity and density. Large mammals are keystone species in ecosystems and their existence affects prey populations and the composition of the surrounding flora. This research also confirmed the existence of many important keystone species such as the Tayassu pecari, Leopardus pardalis, and Panthera onca and provided records of occurrence in different types of habitat, which may help future conservation of different species and which type of habitat they prefer.

Small Ubiquitin-like Modifier, SUMO for short, is a protein involved in cellular regulation which can also be used as a biochemistry tool to express difficult-to-express proteins in the lab. I am investigating several SUMO proteases in order to learn more about the regulatory pathways it is involved in. We are focused on the traditionally used protease from Saccharomyces cerevisiae, a thermophilic version from Chaetomium thermophilium, as well as several redesigns of a homologous protein from the Saccharomyces SUMO protease to improve solubility by mutating solvent-exposed hydrophobic amino acids to polar amino acids using a computational protein design program called Rosetta. I am using thermal melting techniques to find melting points and gel comparison techniques to analyze the efficiency of each protease. Understanding of the kinetics and properties of these proteases and comparing them to each other will yield a greater understanding of how the system works, which versions are more efficient for in vitro use, and how the system can be manipulated in vivo.The goal of my research is to create a set of biochemical tools associated with SUMO proteins to be used in cancer research, as the SUMO system is known to be dysregulated in cancer. Understanding of the role SUMO plays in cancer will provide context for the development of a new generation of therapeutics, and understanding of the SUMO system as a whole will improve a current biochemical technique.

In modern medicine little has been done on non-western populations related to pregnancy sickness symptoms. Questions about why certain foods cause pregnancy sickness have never been looked at in rural parts of the world. The purpose of this study is to find certain association among pregnancy-related sickness and women of color , which could in fact be a cultural or population specific behavior. Our hypothesis was that certain foods would be correlated with pregnancy-related sickness in pregnant mothers in Bangladesh, India. The data set that was used was from a study, “Hormones and Pregnancy-Related sickness in Bangladesh”. This study was conducted in a rural part of Bangladesh in order to investigate the association between reproductive hormones and nausea, vomiting and dizziness in pregnancy. The aim of this study was to be able to correlate the effect of hormones on the severity of the pregnancy related symptom by using repeated-measures logistic regression analysis of 1,232 observations form 115 women in the first twenty weeks of pregnancy. Urine samples were taken form these women along with survey data that asked them about food adversions and also the severity of their pregnancy-related symptoms. The methodology of this study will primarily use a cross-sectional analysis of survey data taken and the urine samples taken as well during the initial study to see possible correlations. The results help understand whether there are correlations to specific food adversions and also higher levels of hormones in the urine samples which would indicate some biological mechanism to food adversion in pregnant women. 

The characterization of a genetically modified strain of yeast is a tedious and laborious process, which in standard cases, may require upwards of one hundred twenty hours of regular attendance to a single characterization experiment. This is common when developing a novel genetic circuit with several distinct parameters and states. The state-of-the-art method involves loading the yeast strain into a proprietary lab-on-a-chip to perform the strain characterization. Our method, however, improves upon the state-of-the-art method by being cheaper, more flexible, and more integrated into existing lab equipment. We employ a CNC-milled polycarbonate microfluidic chamber with mix, temperature, and inducer and sampling controls. This chamber connects directly to a flow cytometer while running, affording real-time, fully automated strain characterization. We have found this apparatus, the affectionately-titled Cytopendix, to be suitable for sustaining yeast cultures and permitting real-time chamber sampling with the flow cytometer. The nature of its simple design and the speed of CNC milling has granted us significantly shorter design iteration periods than current lab-on-a-chip methods. Furthermore, its mixing, temperature, and sampling controls are robust and easy to reprogram. We encountered particular difficulty in accurately dispensing discrete sample volumes, but we have discussed viable and actionable solutions to this problem. Through a multitude of design iterations, we have vastly improved the functionality, usability, and reliability of the system and expect it to develop into a significant improvement over the state-of-the-art methods in this field. The Cytopendix has, overall, proved to bear significant potential as a flexible and extensible instrument for fully automated yeast strain characterization.

Aging can be defined as the time-related deterioration of physiological functions that are essential to the fertility and survival of a living organism. In the Kaeberlein Lab, we study how cells change with age and how these changes contribute to age-associated diseases in order to help people live healthier lives. The eukaryote Saccharomyces cerevisiae (yeast) is used as a model for cellular aging. Due to its conservation of cellular functions through evolution, modeling aging using yeast can provide insights into how mammalian cells age. Through replicative lifespan (RLS) analysis, we are able to determine the effects of interventions on aging by measuring the number of divisions that a single yeast cell goes through before it stops dividing. We are testing various nutraceuticals in order to identify novel pharmacological interventions that increase lifespan. In a screen of twenty nutraceuticals, we found that Pterocarpus marsupium (PME) extract significantly extended lifespan. P. marsupium is a deciduous tree native to the Indian regions. It is well-known to Ayurvedic medicine thanks to its purported curative and lenitive properties. The flowers are said to be effective in lowering fever; its heartwood may possess purifying and detoxifying effects; its wood is used to treat chest and body pain as well as indigestion. Additional studies will test whether the RLS-extending effects of PME can be attributed to pterostilbene, a compound found in PME that is structurally related to resveratrol. We tested concentrations of pterostilbene ranging from 1-50 µM to identify its effect on replicative lifespan. These studies will allow for further development of PME and pterostilbene for use as an intervention to promote healthy longevity.

Today, volatile gas anesthetics are commonly used in medicine for various procedures. While they are effective and safe for most adults, multiple or long exposures of anesthesia to young children and infants is thought to carry the risks of neurodevelopmental defects due to the well-established neurotoxic effects of anesthetics on the developing nervous system, referred to as anesthesia induced neurotoxicity (AIN). If the mechanisms of anesthesia and AIN can be studied and better understood, strategies to prevent defects associated with AIN can potentially be developed. At the Seattle Children’s Research Institute, I am assisting as an undergraduate researcher in a project to better understand AIN. My project specifically involves using C. elegans, a laboratory invertebrate species, to study the effects of anesthesia on neurotoxicity in specific developmental stages, including programmed cell death (apoptosis). The short life span and rapid reproductive cycle of these nematodes make them great models for this study because isoflurane exposure has been observed to induce AIN in these organisms. I am working to research the specific developmental stages of C. elegans that isoflurane affects the most. In lab, I am culturing worms to known stages of development, using green flourescent protein (GFP) reporter strains as markers to study apoptosis (specifically in the apoptoic proteins ced-3, ced-1, and ced-10), and exposing worms to isoflurane during these stages. Using PCR, western-blotting, and imaging techniques, I am analyzing the proteins, neurons, and developmental stage(s) impacted by AIN. This should provide new insight into the proteins and specific neurons that anesthesia induced neurotoxicity targets. By carefully analyzing the mechanisms of AIN, I hope to gain knowledge about how it can damage normal neuronal function in C. Elegans that can eventually be applied to humans.

Mussels, Family Mytilidae, are vital to intertidal ecosystems and to a burgeoning aquaculture industry. They act as ecosystem engineers within their environment by controlling resource availability to other organisms and by shaping the structure of their habitat. Unfortunately, with the rise of anthropogenic climate change, mussel populations are seeing growing threats to their ability to survive and reproduce. This includes lower recruitment, increased susceptibility to disease, and weakening of the byssal thread that adheres mussels to their substrate. Understanding the life history of these organisms, in addition to what food sources and conditions are most suitable for their growth, is essential for preserving this economically and ecologically vital species group. Mytilus trossulus, a common species of saltwater mussel ubiquitous in the aquaculture industry was used in this study. The mussels were grown at two different depths on aquaculture lines for comparison of the environmental effects at those depths. The first layer at 1 meters depth consisted of a mixed surface layer rich in phytoplankton growth and represented a warmer, more variable ocean environment. The second layer was found below the surface mixed layer at 7 meters depth and represented a colder, more constant environmental regime. By analyzing the food sources of these mussels through fatty acid extraction, we will draw conclusions not only about what nutrients are most beneficial to mussel growth, but also what effects the environmental differences between the two depth strata has on the mussels. The purpose of this study is to both provide data for the care of mussels in aquaculture farms and to make inferences about what a changing climate will mean for these organisms.

Didelphodon vorax is a large-bodied, North American metatherian, closely related to today's marsupials. D. vorax is posited to have been durophagous (i.e. feeding on hard-shelled organisms or bone), and is considered one of the largest mammalian forms of the Late Cretaceous, living among the dinosaurs. Fossil finds for this taxon are rare and geographically constrained to the North American Western Interior, which often times impedes comparative intraspecific studies. Through quantitative and qualitative measures, including both linear measurements and comparative description, we compiled an extensive review of multiple specimens. We analyzed each specimen, comprised of various mandibular elements, in the context of ontogeny (i.e. the development of an organism from early stage to maturation) utilizing a principal components analysis. We seek to investigate the morphological modifications which characterize and differentiate juvenile and adult forms of D. vorax. Our results shed light on the relationship between ontogeny and morphology in this taxon. 

Historical forest landscape patterns are necessary to generate landscape level restoration treatments. Though landscape and disturbance patterns due to fires are well understood and recorded on public forest lands east of the Cascade Mountain Range, factors like fire suppression, long fire return intervals of west-side forests, lack of documentation, climatological differences, and extensive timber harvest complicate implementing similar strategies on landscapes west of the Cascades. By examining the distribution of smaller forest patch sizes, in addition to patches that were subjected to low-severity burning as a part of the 2015 Goodell burn for mortality and survival, we can derive and quantify factors necessary for early seral habitat creation (the critical establishment phase in ecological succession; historic proportions of early seral habitat have been approximated, and current levels are comparably lower). By interpreting stereo imagery and burn severity maps of the Goodell burn, and by employing GIS and remote sensing techniques, historically compatible models and landscape-level restoration strategies for both terrestrial and aquatic habitat can be utilized in guiding and monitoring restoration efforts.

Galanin is a small protein found in the gastrointestinal tract, and several places within the central nervous system. My project is to test the function of galanin-expressing neurons in the hindbrain of mice, specifically the projection from the Nucleus of the Solitary Tract to the Parabrachial Nucleus. Contrary to effects shown in other brain areas, my preliminary data suggest that the activation of galanin expressing neurons in in the hindbrain decreases feeding, in mice. These neurons are activated artificially in genetically modified mice, allowing us to then measure food intake and body weight. Their activation decreases food intake under normal conditions, but not with prior food restriction. I predict that these neurons are activated by visceral signals from afferents, or neurons that carry input from sensory organs, which are then responsible for the ensuing lack of appetite. By expressing a fluorescent protein in these neurons, I can also trace this projection to see where other connections are. Then, by examining Fos, a marker of neuronal excitation, I will investigate the activation of downstream projections. This is relevant to many current health issues, such as obesity and anorexia, as the circuits involved in hunger and appetite are not entirely understood yet.

Identifying the bacteria that inhabit a stream is important to determine the overall health of the stream and its inhabitants. Previous studies have stated Proteobacteria, Bacteriodetes and Firmicutes to be the dominant phyla found in water microbiomes, while Flavobacterium, a microbe responsible for many salmon diseases, is the dominant genus. For this project, we performed a microbial analysis of Alder and Fry Creek through biochemical assays. 8 water samples, each from different sites, were plated onto Mannitol and MacConkey plates to determine the relative proportions of fecal coliforms within the aerobic bacterial population. Additionally, the Alder Creek samples were characterized through 16S sequencing. Our findings are in agreement with previous studies, with the exception of two sites, which were located at the intersection of residential areas and natural habitat. These sites were consistent with the major phyla expectation, except their major genus were Arcobacter and Crenothrix. Arcobacter species are typically found in sewage and some are human pathogens. Infections from some Arcobacter species are similar to a Campylobacter jejuni infection. The Crenothrix genus contains iron precipitating species that have been known to form deposits that clog or stain pipes and make water look unappealing. These findings contribute to our understanding of the health of the water, specifically related to fish bearing streams running through residential and industrial regions. This project will continue as a three-year study to monitor changes in the microbiome during the course of stream restoration.

Müller glia (MG) are the predominant glia of the neural retina. In fish and birds, retinal injury causes these glial cells to de-differentiate into retinal progenitor cells (RPCs), and these RPCs repair the damaged tissue by generating new neurons. This process does not happen in mammals, and so retinal damage is permanent. microRNAs (miRNAs, miRs) are small RNA molecules that regulate gene expression. Several miRNAs are expressed in the RPCs, but not in the Müller glia, and we call these RPC-miRs. This study focused on if artificially expressing the RPC-miRs would stimulate mammalian MG to regenerate new neurons in postnatal mouse retinal explants. Two leading questions were explored: Will the Müller glia treated with RPC miRNAs regenerate new neurons? What is the best time window to convert Müller glia into retinal progenitors with high efficiency? Retinas from postnatal (P) 4 up to P12 wild type or transgenic reporter mice were dissected from surrounding eye tissue and electroporated to introduce RPC artificial miRNAs (such as miR-15a, miR-17, miR-19, and miR-20a mimics) or control miRNA mimics, along with a reporter mRNA.  Explants were cultured for 3/4 or 6/7 days ex vitro (DEV), fixed, cross sectioned, stained (immunofluorescent labeling) and analyzed using confocal microscopy. The proliferation marker EdU was added to track cell proliferation. Our data showed that RPC-miRs led to an increased proliferative response in the Müller glia, indicating that the cells were adopting an RPC-like state. Moreover, the age of the explants was an important factor; the areas that were successfully electroporated became smaller in older explants and therefore fewer Müller glia can be targeted in older tissue. Our data shows that miRNAs from RPCs can promote Müller glia to adopt some RPC-like characteristics and therefore could be used to stimulate regeneration in the retina.

During embryogenesis, phosphorylation of serine and threonine residues on p120 catenin causes disassociation from cadherin transmembrane proteins and activation of downstream pathways leading to cell motility. We have mutated a single amino acid on p120 catenin, T310, to determine its involvement in cell migration of the dorsal axis during zebrafish embryogenesis in vivo. T310 is a target of glycogen synthase kinase 3 (GSK3) and phosphorylation of this site facilitates disassociation from membrane-bound N-cadherin in astrocytes, decreasing cell-cell adhesion and activating the cell motility pathway. We created two mutations of p120 catenin: T310E that mimics the negatively charged phosphate and T310A containing a neutral residue that cannot be phosphorylated. Either mutated or wild type mRNA was co-injected with morpholino, which knocks down endogenous p120 catenin, to determine the injected protein’s ability to rescue the short-tailed knockdown phenotype. As expected, co-injection of wild type p120 catenin mRNA and morpholino rescued the damaged phenotype resulting in embryos with normal tail extension. Co-injecting either T310A or E with morpholino resulted in partial rescue, suggesting that phosphorylation at this site may be variable during development or that the effects cannot be measured by phenotypic quantification methods. We are currently studying the effects of phosphorylation at other sites, S268 and S269, which may give insight on the downstream effects p120 catenin in cell motility, including morphological bone abnormalities and metastasizing cancer cells.

Research has shown there are potential benefits to social media use in reaching underserved populations such as runaway homeless youth. Social media use in professional social service practice has often been seen as a taboo subject, and anecdotal reports suggest it is something to refrain from because of the potential for ethical dilemmas. However, studies have shown that many social work students feel the National Association of Social Workers Code of Ethics is vague in addressing correct professional social media use. The purpose of this study is to gain practitioner perspectives on professional social media use. The study recruited 20-30 practitioners working in youth-serving social service agencies who regularly come into contact with clients, using snowball sampling as the primary source of recruitment. A 10-20 minute phone or in-person interview was conducted with each participant gauging perspectives on social media use, agency policies on social media, whether ethical codes of the profession adequately address social media use, and how a practitioner would address an online ethical dilemma. The data included qualitative and quantitative components and was analyzed for differences of opinion regarding education level, years in the field, and familiarity with social media. The researchers anticipate that a majority of the participants will report inadaquate training on social media use and that a majority of participants will have encountered clients on social media networking sites before. Furthermore, the researchers anticipate a majority of participants will feel the ethical codes of the profession do not adequately address social media use. This study could inform further social work research into the efficacy of professional social media use as a benefit to the helping relationship.

Empathy is the capacity to understand or feel what another person is experiencing from within the other person’s frame of reference. Empathy has been shown to be steadily declining in college students. Use of various types of media has been shown to increase empathy if the media is prosocial, relating to a positive behavior, while violent or antisocial media is shown to decrease empathy. College students are highly connected through social media. There has been little research on the impact of mobile app usage on empathy. The purpose of this study is to compare smartphone app usage to the Interpersonal Reactivity index, a measure of empathy. A sample of approximately 30 college students with iPhones from two universities were recruited through class Facebook pages and email listservs. The study was conducted through an online survey that uses an established measure (Interpersonal Reactivity Index) to determine perspective taking, empathetic concern, fantasy, and personal distress of each participant. Additionally, participants were asked to submit a screenshot of their weekly app usage that shows time spent on apps. A codebook was used to identify these apps as prosocial or antisocial. Analysis included comparing scores from the Interpersonal Reactivity Index to the amount of time spent on apps according to the screenshot. From these sets of data it will become more clear what kind of smartphone app usage is prosocial, what kind is antisocial, and the effects of app usage on perspective taking, fantasy, empathetic concern, and personal distress amongst college students.

Our laboratory has identified a group of neurons that expresses calcitonin gene-related peptide (CGRP) in the parabrachial nucleus (PBN), which are involved in appetite suppression and responses to fear. For example, CGRPPBN neurons mediate the suppression of food intake during a conditioned taste aversion (CTA) and are activated following conditioned fear responses [1,2,3]. Here, we investigate whether CGRPPBN neurons mediate anorexia associated with a diet deficient in essential amino acids (EAA-def anorexia). We inactivated CGRPPBN neurons by injecting an associated adenovirus (AAV) carrying the tetanus toxin light chain (Tet tox) in the PBN of CalcaCre/+ mice (Calca is the CGRP gene) and showed increased food intake and decreased body weight loss compared to control animals on a prolonged diet deficient in EAAs. In a pair fed group of mice injected with the Tet tox virus, there was not a significant difference in body weight loss. These results suggest that CGRPPBN neurons modulate appetite during EAA-def anorexia, but are not responsible for a change in energy homeostasis during EAA-def anorexia.

Frailty is an age-related, clinically recognizable state of heightened vulnerability that is defined by having 3 of the following criteria: muscle weakness, slow walking speed, exhaustion, unintentional weight loss, and low physical activity. Frailty is associated with increased risk of disability, hospitalization, and mortality among older adults and therefore there is a critical need to identify potentially modifiable risk factors. It is conceivable that chronic pain increases the risk of frailty through multiple biopsychosocial mechanisms, including physical deconditioning. Accordingly, we sought to determine whether pain and the number of pain sites increases risk of frailty in a nationally representative, prospective cohort study of community-dwelling older adults. We analyzed data on 7,601 Medicare beneficiaries aged ≥65 years who participated in the National Health and Aging Trends Study. Participants were interviewed annually in their homes from 2011 through 2015. Over the 4-year follow-up period, the cumulative incidence of frailty was 17.6% (1,327 new cases). Incidence increased with advancing age and was higher in women than in men. Importantly, frailty was significantly more likely to develop among older adults with pain (23.0%) than in those without pain (12.5%). When adjusted for demographic and health characteristics, the risk of frailty was 54% higher in those with pain than without pain (Incidence Ratio [IR]=1.54; 95% confidence interval [CI]: 1.32-1.80). Frailty incidence also increased significantly with the total number of pain sites. Compared to those without pain, the adjusted IRs for frailty were 1.14 (95% CI: 0.89-1.47), 1.34 (95% CI: 1.10-1.64), 1.52 (95% CI: 1.23-1.88), and 2.08 (95% CI: 1.66-2.61) for older adults with 1, 2, 3, and ≥4 pain sites, respectively. In summary, pain was independently associated with increased risk of frailty among community-dwelling older adults. This finding suggests that multicomponent intervention programs should integrate pain management to more effectively prevent frailty among older adults.

We investigated how grading treatment, self-evaluation versus peer-evaluation, on weekly practice exams (PE) in an Introductory Biology course affects the advice students give to improve academic performance. We hypothesize that students who suggest more resources and study strategies perform better on course exams because they are aware of how improvements need to be made and what resources to use. Students in the class (approximately 550) take a PE on Thursday and then grade the PE responses on Friday. The grading groups were split up into two treatments: half of the class self-evaluated whereas the other half evaluated their peers’ responses. In addition, they were prompted to answer two questions: 1) Pick a single thing that you/this student should work on. 2) What resource should they consult before the next exam? We used this information to explore how study strategies and resources that students suggest relate to their test scores. To analyze the responses, we created specific codes corresponding to student answers. Results of the codes indicate that the students’ responses fall into four categories for question 1 and two sets of categories for question 2 (with three and two codes in each set). When asked to provide ways to improve PE performance, the four specific codes were being descriptive, increasing knowledge of content, interpreting the question, and time management. When providing resources, the two sets of categories were active versus passive resources and expert, peer, and self resources. Over the course of ten weeks, students who suggested more strategies and resources tended to perform better on exams. As well, both self-evaluators and women in the course suggested more strategies and resources than peer-evaluators and men respectively. Given this study, instructors can better understand what resources students are utilizing but also which study strategies tend to be more successful for students.

Photolithography is a process used in nanofabrication to create patterns on different types of substrates. These patterns are used to define structures that are either built up (additive) or etched (subtractive) to form functional layers. These functional layers allow for the creation of different kinds of electronics, such as resistors and semiconductors on an incredibly small scale. The process of photolithography involves the use of light to expose a pattern on a substrate. In order to expose a pattern on a substrate or wafer, the use of photoresist is essential. Photoresist is a viscous solution, which is deposited on wafers. The wafer then goes through a process called spin coating which rapidly spins the wafer to create an even surface. This even surface is crucial in order to properly expose a pattern on the wafer. In order to have an even coating and minimize the discrepancies across a wafer’s surface, there are many processes that a wafer must go through in order to ensure the best quality for exposure. This poster discusses techniques including the storage of the wafer, the handling of the wafer and the depositing of photoresist. Analyses of these techniques are discussed as well as critical parameters for the environment in which they are kept.

 The percentage of obese children in the United States has tripled since the 1970’s, with one in five school-aged children now categorized as obese. Obesity development and response to intervention has been found to depend on factors such as impulsivity, though its influence as a major risk factor is unknown. This project seeks to determine if impulsivity affects weight reduction outcomes in family-based behavioral treatment. I hypothesize that increased impulsivity will be associated with increased food intake, increased food reward value and subsequently poorer weight reduction outcomes. Forty-nine obese males and females aged nine to eleven years old were evaluated before and after they participated in a 6-month family-based behavioral treatment intervention to determine the relationship between impulsivity and treatment outcomes. Impulsivity was assessed using a Developmental Neuropsychological Assessment. Each subject’s food reward value was measured using interviewer-administered tasks in which participants were questioned on their choice between preferred snack foods vs. an appealing alternative. Calorie intake was measured by providing study participants with a buffet consisting of a wide variety of foods, with which they had access to for thirty minutes. Calorie intake and composition was measured by weighing back uneaten food. Success of obesity treatment was measured in the comparison of adiposity using BMI z-scores, given that a z-score decrease of at least 0.25 improves blood pressure, lipids and insulin sensitivity. Obese children categorized as impulsive are predicted to consume more calories and rate a food reward value compared to non-impulsive obese children, resulting in poorer weight reduction outcomes, indicating that childhood obesity is correlated with impulsivity on the basis of an inability to engage cognitive control of food intake. This research will provide new insights into the neurobiological basis of child obesity, potentially improving the cost-effectively and long-term results of obesity interventions.

MicroRNAs (miRNA) are non-coding RNA (ncRNA) which post-transcriptionally regulate gene expression. In cancer, many miRNAs are overexpressed leading to the down-regulation of tumor suppressing genes. While the general processing steps of miRNA are known, the specific reason certain miRNAs are overexpressed in cancer remains unclear. Our particular miRNA of interest is miR-21, which inhibits genes encoding tumor suppressing proteins, and upregulation is associated with cancer. Specifically, we are interested in understanding how the pre-miRNA-21 processing step is regulated to generate functionally mature miRNA, and what proteins are involved in this process. Many RNA binding proteins are involved in miRNA biogenesis and influence transcription levels. This project explores the possible role of the RNA binding protein NOVA. NOVA is a neuron-specific RNA-binding protein exclusively expressed in the brain and neuronal tissues except for patients with Paraneoplastic opsoclonus-myoclonus ataxia, where it is found site of malignancy. NOVA is composed of 3 K-homology RNA binding domains selective for the sequence 5’ -YCAY-3’ (Y = pyrimidine). This KH domain recognition sequence is present in the hairpin loop in pre-miR-21. Additionally, in-vitro DICER processing of pre-miR21 is very slow relative to other miRNAs, however, if NOVA interacts with the pre-miR-21, this could stimulate its processing. We are trying to determine a possible mechanism of NOVA stimulated miR-21 expression through binding a conserved YCAY sequence. MiR-21 and NOVA have been isolated and purified, and their binding affinities are being evaluated using electro mobility shift assays. If binding is indicated, gene knockdowns will be preformed in cells to assess how NOVA affects miR-21 levels. It is difficult to design microRNA specific inhibitors, however, finding inhibitors for proteins is possible. If NOVA is found to lead to miR-21 up-regulation, a possible cancer treatment could be designed by targeting NOVA to lower levels of miR-21. 

Thioester reactions are essential for performing all types of biochemical and organic processes. The reactive C-S bond serves as a gateway to other acyl groups in a multitude of organic chemistry reactions, and thioesters are necessary for processes such as thioester exchange, Claisen condensations, and hydrolysis. These reactions are typically enzyme-catalyzed and are vital for controlling levels of acyl CoAs, S-palmitoylation, and for fatty acid biosynthesis. Due to the strong reactive center of a thioester, the mechanism for thioester hydrolysis is not as extensively studied or completely known as the mechanism for the oxoester. There are two types of mechanisms that are focused on when it comes to thioester hydrolysis; a step-wise mechanism with formation of a tetrahedral intermediate or a concerted mechanism where the nucleophile and leaving group leave simultaneously. This research concentrates on comparing the hydrolysis mechanism of the thioester formylthiocholine with the oxoester analog, formyl choline, which is synthesized in the lab. In addition to synthesizing formyl choline, the isotopic derivated deutero-formyl choline was made. To study the transition-state structure, deutero-formyl choline (d-Fch) and hydrogen-formyl choline (h-Fch) are mixed together and reacted in acidic, basic, and neutral conditions. Kinetic isotope effect values are obtained by measuring the rates of hydrolysis by NMR spectroscopy. To date, the KIEs for the formyl-H have been determined in basic conditions and yield an inverse KIE. This suggests that the rate determining step is during the formation of the tetrahedral intermediate, where the carbonyl-C undergoes an sp2-sp3 conversion. This knowledge provides initial data for the comparison with the thioester to understand the mechanism of hydrolysis.

Rivers are conduits for transporting sediments and organic material through the watershed and to the ocean. The Elwha River recently was involved in the largest dam removal in history within the USA. Studies of the dam removal impacts have never been done as extensively as they have for the Elwha River Restoration project. Now, the river is no longer blocked, and 21 million m³ of sediments have been made available for erosion, and it is not fully known where the sediments are depositing and how they are affecting the ecosystem. In order to quantify where organics are being stored in the river system, sediment samples were collected along the river and shoreline, and grain size and loss on ignition (LOI) analysis performed on them. Most of the samples with more than 2% organic content were at sites in the reservoir and side channels, and organics less than 1% were retained on the beach and riverbank. Therefore, finding organic matter in the river and shoreline systems can inform us how the ecosystem is recovering, and whether overtime organics are fully transported to the ocean, or whether organics will be consumed, transformed, or altered along the way.

Adolescents without college attending parents and from minority communities have lower post-secondary enrollment rates. Non-profit college preparedness organizations targeting these adolescents can provide support to help increase these enrollment rates. An important aspect of reaching adolescents is outreach through social media, given its prevalence among this population. However, pilot evaluations show infrequent social media use by college preparedness organizations, and it remains unclear which online outreach approaches yield the highest user engagement. The purpose of this study is to understand approaches used by college preparedness organizations in social media outreach, and which factors are associated with user engagement. In this content analysis study, seven college preparedness organizations listed as partner organizations on the Seattle government website were randomly selected. A codebook was developed to evaluate outreach approaches using components derived from the Facebook Influence and non-profit organization models, which will use variables for how organizations communicate information, build communities, create immersion, and facilitate action in their social media posts. The codebook was used to evaluate each organization’s posts from their Facebook and Twitter profiles for a period of a year. Likes, Shares, and Retweet totals were also collected for each post. Associations between social media outreach approaches and user engagement were examined. Organizations may use more community and immersive orientated posts to generate more likes and followers. The results of this study could potentially be used to develop strategies that will allow other college preparedness organizations or organizations in other outreach based fields to increase their user engagement.

Duplex ultrasound scanning with both color Doppler and Doppler spectral waveforms is relied upon for diagnosis of vascular disease and selection of patients for further evaluation and treatment. In most duplex ultrasound applications, classification of disease severity is based primarily on alterations in blood flow velocities. Our research group in the Applied Physics Laboratory is collaborating on the development of a Doppler Ultrasound Simulator for training and assessment of scanning skills. The ultrasound simulator allows learners to repeat examinations as many times as necessary to acquire and improve skills. With the addition of clinical cases covering a range of disease severity, this duplex ultrasound simulator will be a useful tool for training health-care providers in vascular ultrasound applications and for assessing their skills in an objective and quantitative manner. The Doppler Ultrasound Simulator extracts virtual blood flow measurements from computational flow models, with real-time scanning linked to a physical mannequin. The Simulator reproduces the key components of a clinical Doppler ultrasound examination including steering the Doppler beam, selecting the size and location of the Doppler sample volume, specifying the Doppler angle relative to the vessel axis, and adjusting the pulse repetition frequency (scale control) and baseline to avoid aliasing. My research role in this larger-scale project is to use a data processing program, MATLAB, for image reconstruction, processing and analysis. MATALB is used for the creation of 3D image volumes, which are used as the foundation for vessel reconstruction as 3D computer models. MATLAB is also a useful tool for correction of image motion artifacts. My effort in this project involves applying image correcting algorithms to create 3D volumes with enhanced vessel alignment and minimized interference from real-time shifting of 2D ultrasound images.

The Olympic Mountains Experiment (OLYMPEX) was a ground validation field campaign centered on the Olympic Peninsula designed to provide physical validation and verification of satellite precipitation measurements from the constellation of satellites known as the Global Precipitation Measurement (GPM). The goal of OLYMPEX was to validate and document precipitation measurements in midlatitude frontal systems moving from the Pacific Ocean to the coast and then to the high terrain and leeside. The assets featured in OLYMPEX included extensive ground instrumentation, soundings and dropsondes, several radars, and aircraft all helping to monitoring storm systems as they approached and then traversed the Olympic Peninsula during the intensive field operations from November 2015 through March 2016. Ground instrumentation was located at a number of different elevations mainly focusing on the windward southwest side of the Olympic Mountains and scattered in other areas of the Olympic Peninsula in locations where precipitation has not been previously measured. This presentation examines patterns of orographic precipitation enhancement using this extensive gauge network and ground radar data to describe and quantify patterns of enhancement and how they vary with synoptic conditions, specifically during the December 8-10th atmospheric river event. The key synoptic parameters that affected the patterns of enhancement included the stability, the strength and direction of the low level flow, the intensity of synoptic scale forcing and the transport of water vapor. These synoptic parameters varied with storm sector. Deep stratiform precipitation with high melting level and low level jet produce strong orographic enhancement along initial windward slopes. Post-frontal conditions accompanied by a low melting level and convective cells are associated with a greater degree of enhancements over interior locations. Pre-frontal and blocked flow conditions show no significant level of enhancement between coastal and inland sites. These findings provide never-before-seen detail of the relationships between storm sectors and orographic precipitation enhancement and can be applied wherever midlatitude cyclones cross coastal mountain ranges around the world.

Aging remains as the biggest risk factor for developing the most prevalent diseases found in developing countries: cancer, cardiovascular disease, and neurodegeneration. Reduced insulin/IGF signaling (IIS) extends lifespan in diverse organisms, but the underlying molecular mechanisms that regulate this remain unclear. Recently, non-coding RNAs, i.e. those RNAs not translated into proteins, have become a hot topic, being identified as regulators of gene expression during aging. Circular RNAs (circRNAs) are one such form of non-coding RNAs. Although they are identified in a diverse number of species, the function of most circRNAs is unknown. CircRNAs are distinct from their linear counterparts because they are devoid of the terminal structures (e.g. 5’cap or a poly A tail) that frequently determine the fate of RNA transcripts thereby escaping normal RNA turnover. It has been established that circRNAs are abundant and differentially expressed; they can serve as either transcriptional regulators, and/or microRNA sponges. Furthermore, some circRNAs are produced at the expense of linear RNA, thereby affecting gene expression and thus tissue or organismal function. CircRNAs are also evolutionary conserved, which begs the question why they are so important. Recently, the Partridge lab conducted gene expression profiling of long-lived IIS mutants and identified differentially expressed circRNAs, which may influence IIS-mediated longevity and thus aging. The Partridge lab have generated in vivo Drosophila models, mutating individual circRNAs through genomic engineering (CRISPR/Cas9) and the Gal4/UAS system, to study their function. Using these novel circRNA mutants, I will contribute to discovering the in vivo function of circRNAs in aging by performing quantitative experiments. I will analyze circRNAs mutant lifespan and phenotypes associated to longevity e.g. development, fecundity, stress resistance, neurodegeneration. Genetic epistasis experiments will determine the role of the given circRNAs in insulin signaling. Finally, I will use molecular biology techniques to unravel the mechanism underlying any observed phenotype.

Atherosclerosis is a disease in which cholesterol accumulates in the walls of arteries, limits blood flow, and causes heart attacks and strokes. We aim to prevent and reverse atherosclerosis with a gene therapy that increases levels of apolipoprotein A-I (apoA-I), a protein that removes cholesterol from the artery wall. This therapy increases expression of apoA-I specifically in endothelial cells (EC), cells that line the artery wall. Our initial vectors for expressing apoA-I contained the murine endothelin-1 (mET-1) promoter (an EC-specific promoter) both without (1XETE) and with (4XETE) three additional copies of an endothelin-1 enhancer (ETE). We used two approaches to increase apoA-I expression: 1) literature searches for regulatory sequences that increase EC gene expression in other contexts; 2) novel bioinformatics approaches that identify EC-specific cis-regulatory sequences (CRMs) throughout the genome based on ENCODE data, relative local abundance of EC-related transcription factor binding sites, and interspecies conservation. We ligate regulatory sequences into the apoA-I vectors mentioned above. We transfect these constructs into cultured EC and measure apoA-I expression. Using the first approach, we found that addition of a shear-stress-response element into the 4XETE vector did not alter apoA-I expression. Addition of a single MEF2C enhancer sequence into the 4XETE vector decreased expression; however, after this decline, addition of more copies of the MEF2C enhancer increased apoA-I expression. Lower expression may be due to unfavorable interactions between multiple enhancer elements. Using the second approach, we identified eleven CRMs (55-352 bp) clustered around the promoters of genes highly expressed in EC: CDH5, EFEMP1, THBS1, and VWF. In conclusion, trial-and-error addition of enhancers has not reliably improved EC apoA-I expression. A bioinformatics-based approach identified CRMs associated with high transcriptional activity in EC. We inserted the eleven CRMs into both the 1XETE and 4XETE apoA-I vectors and will test whether they increase EC apoA-I expression.

Peptoids are a novel polymer with a molecular structure similar to proteins. They can be used for numerous applications such as drug delivery agents, scaffold material, and nucleation agents. Certain peptoids can also self-assemble into functional and tunable structures such as nanosheets and nanofibers. Recent experimental research uncovered that chains of N-ethylcarboxylate – N-ethylphenylchloryl (Nce-Npe) peptoids can aggregate to form clusters in a calcium chloride solution and form nanofibers in the presence of a mica surface. It was also found that calcium ions facilitate the aggregation of these peptoids. In order to characterize the solution-state behavior of these peptoids on the atomic scale, we used molecular dynamics simulations. Molecular dynamics simulations utilize Newton’s laws and fitted empirical equations to produce the trajectory of the peptoids over time. We setup several systems of varying size and ion concentration and ran these simulations using a three-site water model and an AMBER99sb-ildn forcefield with parameters optimized to describe peptoid systems more accurately. We ran these simulations on Hyak, a high-performance computing cluster at UW, using the GROMACS software package. Through analysis of the peptoid trajectory, we found that aromatic pi-stacking drives clustering and ion-carboxylate interactions dictate the structure and stability of the peptoid clusters. We also observed an increase in both the rate of aggregation and number of ion interactions as the concentration of calcium ions increased. These simulations are essential to understanding the self-assembly of peptoids and similar polymers in complex environments.

Hypertension is a life-threatening issue affecting 31% of American adults and demands close monitoring. All conventional blood pressure (BP) devices are cuff based and are inconvenient to wear and fail to provide continuous BP. There is a need for a convenient, miniaturized, and cuff-less blood pressure (BP) monitoring device. During the past several years, studies have shown that BP is correlated to pulse transit time (PTT) obtained from electrocardiogram (ECG) and photoplethysmogram (PPG) or radial pulse. This project has successfully demonstrated that same side ECG and PPG can be used to obtain PTT that correlates inversely with BP, hence paving the way for a continuous device. We have shown two possible configurations for our device which differs in placement of sensors. The first device is designed with contact-electrode ECG and PPG sensors, located on the bicep and the ear; while the second one, which is a wrist-worn device, consists of a noncontact ECG circuit and a piezoelectric pulse sensor. The measured PTT is regressively correlated with BP and a regression coefficient between 0.3-0.9 is obtained. To further improve the accuracy of the BP device, the research work also investigates different algorithms that can be used to estimate BP. Online data available in Physionet database has been used to verify accuracy of popular regressive algorithms and effect of inclusion of other physiological factors like heart rate (HR) and artery thickness in the relationship. Our results indicate that inclusion of HR improves the correlation between PTT and BP. Results of device configuration and algorithm improvement obtained can enable next-generation devices, providing essential continuous BP monitoring off-the-clinic for hypertension patients as well as healthy people.

Bees and flowers trade nectar for reproduction. Rhinos provide food to birds in exchange for removal of parasites. Mutually beneficial interactions like these pervade the natural world, but their impact on the process of evolution is not well understood. Here, we test whether each species in a microbial mutualism repeatedly adapts to the changes in their partners, or coevolves. In our model system 22 communities of the Methanococcus maripaludis (archaea) and Desulfovibrio vulgaris (sulfate reducing bacteria) evolved for over 3000 generations in the laboratory. To test whether coevolution occurred, we separated the D. vulgaris and the M. maripuludis in the mutualistic community at 1000 generations. We then paired these 1000 generation D. vulgaris populations with M. maripaludis at different evolutionary time points and measured the growth rate and yield of each population. If M. maripuludis coevolved with D. vulgaris then we would expect the 1000 generation D. vulgaris to perform better with future partners (0-1000 generations) than past partners (1000-2000 generations) because mutualists are expected to have positive effects on each other's fitness. A preliminary experiment with 6 communities indicated the opposite, that D. vulgaris grew faster with past generations compared to future generations. The yield of communities was similar across pairings. These results could indicate D. vulgaris has coevolved with M. maripaludis, and that there is an underlying antagonism between these species that has not been recognized. We present the results of a more extensive experiment, testing more mutualistic communities at more time points, and looking for patterns of coevolution in both species.  
 

Brain computer interfaces (BCIs) hold an amazing potential to increase our knowledge of the brain and to help improve the lives of countless people. However, almost all work currently being done with BCIs is performed in a strictly controlled laboratory setting. In order to create BCIs that can function robustly in the real world we must examine brain signals during naturalistic tasks like speech. Through our partnership with Harborview Neurology we are able to get week long continuous audio/visual/and electrocorigraph (ECoG) data from long term monitoring of epilepsy patients. A significant problem with this data set is that there is so much data that manual annotation of the subjects' speech would be incredibly time consuming. Our work focuses on using high throughput analysis techniques to automatically annotate our data set in order to efficiently study the brain. We present a pipeline to perform automatic speaker tracking of the implanted patients in our clinical recordings. Speaker tracking is the task of figuring out all the times in which a particular speaker spoke during a recording or across several recordings. We have developed an automatic pipeline that extracts all human speech from a recording and then uses a support vector machine (a machine learning algorithm) to differentiate the subject's speech from anyone else that might speak inside the hospital room. Accuracy for every step is above 80%. The methods developed in the study will allow us to automatically generate timestamps of subject speech across our entire data set. This will allow us to perform useful analysis on the ECoG signal in order to better understand the underlying neural basis of natural speech and develop more robust BCIs to improve the lives of countless people.

Malaria, a disease caused by the Plasmodium parasite, affects half of the world’s population and kills about 500,000 people each year. Infection begins when the infectious form of the parasite, the sporozoite, is deposited into the skin by a Plasmodium-infected Anopheles mosquito. Each sporozoite infects one hepatocyte, and develops into thousands of red blood cell infecting merozoites. Merozoite infection of red blood cells leads to a cyclic expansion of parasites in the blood stream that results in malaria associated morbidity and mortality. No fully protective malaria vaccine exists, but immunization with genetically attenuated parasites (GAPs) that halt in the liver confers long-lasting immunity in mice. Understanding the innate and adaptive immune response to liver stage Plasmodium infection is critical to the enhancement of GAP vaccine efficacy and the generation of human malaria GAP vaccine. Our lab has identified the type I Interferon response (IFN-1) as the prototypical innate immune response to liver stage GAP infection. Yet the host sensors or pattern recognition receptors (PRRs) necessary for the initiation of this IFN-1 response remain unknown. To identify possible inducers of IFN-1 signaling, we injected mice with either mosquito salivary gland debris or GAPs, isolated their hepatocytes, and examined the transcript levels of different PRRs. Transcripts for PRRs critical for the intracellular sensing of pathogen nucleic acid were upregulated upon GAP infection. We will examine the ability of mice lacking these PRRs to induce an IFN response in infected livers. We will also determine whether parasite DNA and/or RNA is sensed by these PRRs. Since Plasmodium is a complex pathogen, we predict that multiple candidate PRRs might be necessary to drive the IFN-1 response to liver stage infection.

Despite their inability to express proteins, long intervening noncoding RNAs (lincRNAs) have demonstrated significant biological function; they regulate both cancer and physiological epidermal development and target chromatin-remodeling genes. Their repertoire of activity is constantly growing with the discovery of each new lincRNA. The lincRNA OIP5-AS1 (Cyrano) is a 4.5kb transcript required for proper embryogenesis in zebrafish, and unlike most lincRNAs, possesses multiple stretches of conservation found in at least 52 vertebrates. While the mechanism for how Cyrano regulates embryogenesis is unknown, it is believed that Cyrano functions primarily through a 26-nucleotide region that is perfectly conserved in every tetrapod; perturbation of this region resulted in a lack of proper development of zebrafish embryos. The secondary structure of the 26-nucleotide conserved region was revealed to contain an RNA binding domain, 5’–CCAU-3’, for RBMX, an RNA binding protein that is highly conserved in mammalian species. I hypothesize that Cyrano interacts with RBMX to aid in the process of embryogenesis, not only in zebrafish, but in other species as well. By using in vitro gel shift assays and in cellulo pull-down experiments, I aim to determine if the lincRNA Cyrano binds to the RBMX protein. Understanding this protein-lincRNA interaction will give more insight to the process of embryogenesis, and how lincRNAs function overall.

Prolonged activation of cardiac fibroblasts after acute wound healing post myocardial infarction results in excessive extracellular matrix deposition in the myocardium and formation of fibrotic scar at the site of injury. This adverse fibrotic remodeling disrupts physiological function in the heart and leads to increased risk for heart failure which is detrimental for patient outcomes. The reasons why activated fibroblasts, known as myofibroblasts, persist after wound stabilization are poorly understood due to the lack of a myofibroblast reporter system capable of direct longitudinal study. Current reporting modalities are invasive, resource and time intensive, wasteful, and indirect. The present project aims to design and develop an in vivo, repeatable, and real time reporter system for myofibroblast cell fate as a surrogate for cardiac fibrosis. To achieve this, we have designed a chimeric promoter to drive firefly luciferase vector expression specifically in myofibroblasts. We have built 9x tandem repeats of serum response factor (SRF) and nucleated factor of activated T-cells (NFAT) binding sequence since they are key mediators in myofibroblast activation. These repeats were ligated onto smooth muscle alpha actin (ACTA2) and collagen type 1 alpha 2 (COL1A2) minimal promoters, and these constructs were cloned into pGL3 luciferase reporter vectors. The specificity of the promoter was evaluated and optimized through in vitro luciferase assays with NIH 3T3 primary mouse fibroblasts.  With the successful creation of this reporter, wasteful use of animal models will be mitigated and direct mechanisms in the pathobiology of chronic cardiac fibrosis can be elucidated. Furthermore, efficacy of novel therapies can be evaluated in vivo.

Through analyzing the properties of the host galaxies to Supermassive Black Hole (SMBH) mergers, we will be able to determine what phase of galaxy evolution these mergers track. We do this by analyzing ROMULUS25, a large cosmological simulation of the universe, from the Big Bang to present day. The data from ROMULUS25 includes the evolution of thousands of galaxies and is able to resolve the internal structure of galaxies. Most galaxies have SMBHs in their core, and when galaxies merge, it is believed that their SMBHs merge as well. Project LISA, as well as other future projects, will be able to detect gravitational waves coming from these mergers. With simulations like ROMULUS25, we will be able to better predict which galaxies are likely to host these SMBH mergers, which will give context to future gravitational wave studies with respect to galaxy evolution theory.

Nearly 1 in 3 children in the U.S are already overweight or obese. Pediatric obesity represents a major health crisis with many psychological and physiological consequences. The American Medical Association states that obese and overweight children should receive moderate-to-high intensity behavioral interventions. Currently, it is estimated that about 17% of children age 2-19 in the U.S. are considered obese. The SHIFT study focuses on examining the short and long term efficacy of peer-versus professionally delivered FBT (family based training) on child weight outcomes and is a model that has been developed with studies done in the past. Previous studies like COMPASS and FOCUS tested the treatment model that SHIFT utilizes and EPICH/PPP were designed to pilot test the treatment model with peers. After conducting those studies, modifications were made to create a model for the SHIFT study. The treatment consists of a 5 month period in which 20 family sessions are conducted that are oriented toward helping them learn and implement behavioral skills for eating and physical activity change. The weight criterion for children is a BMI that is 20 to 100% above the median BMI and they should have at least one overweight parent (BMI>25) because these children are at a greater risk of maintain obesity into adulthood. The sessions include diet, physical activity, and behavior change that are discussed through parent only/child only groups as well as individual family meetings. Families come in every week and then are separated into child/parent groups in which we discuss prevention strategies for obesity and promote a healthy lifestyle and the children learn about ways to include physical activity in their daily lives. SHIFT works as a new model for FBT delivery that decreases costs and increases intervention availability while maintaining the treatment efficacy by using non-professional interventionists to deliver FBT.

Alcohol is used among 45% of college students, and this behavior is associated with up to 1800 deaths per year. Social Media has been shown to influence student's alcohol use. Little is known about the role of alcohol-related Facebook events in promoting safe or problematic alcohol use. The purpose of this study was to investigate the presence of safety messages in alcohol-related Facebook events, as well as students' perceptions of these posts. This mixed methods study involved content analysis and qualitative interviews. Researchers will use the Facebook Discover feature, which enables the user to search for events meeting given criteria, to identify events in the Seattle, Washington area categorized under "Nightlife". The search was be conducted at 4 time points 1-2 days apart, each weel, and the top 4 events will be included, yielding a total of 100 events for analysis. Researchers evaluated events for the following variables: alcohol-related public event, number of people attending, interested, and invited, and inclusion of alcohol safety related messaging. Then researchers identified ten 18-22 year old college students to participate in qualitative interviews by approaching them at popular campus locations. Interviewees viewed a slideshow of common themes present in alcohol-related Facebook events content analysis findings and answered questions about their attitudes about attending such events. These interviews were recorded, transcribed, and undergo thematic qualitative analysis. The results will quantify the proportion of Facebook events with alcohol-related content on Facebook events available under a specified public search. The results from this study can be used to consider ways that safety messaging can be improved on alcohol related Facebook events.

The idea of gender as binary effectively classifies gender into two distinct categories and implicitly emphasizes that the two should be different from each other. Transgender individuals, whose biological sex does not correspond to their current gender identity, expand the traditional binary gender construct. The current study examines how adults attribute gendered characteristics to the author of a written vignette when the author is identified as cisgender or transgender. It is hypothesized that adults will identify different gender related attributes to the author depending on the identified gender of the author. Participants will be randomly assigned to one of five vignettes that each displays the same short story with a different gender identified for the author: a trans-man author, a trans-woman, a man, a woman, or a control (no author specified). Participants will then proceed to the same questionnaire (a Likert-type scale), where they will be asked about their impression of the author. One-way ANOVAS will be conducted to compare the global scores and subscale scores across each of the five authorship conditions. It is expected that there will be a significant difference in the global score of the Gender Attribute Scale, whereby the cisgender female author is attributed more traditionally feminine characteristics and the cisgender male author is attributed more traditionally masculine characteristics. Given the paucity of research in transgender attributions, we do not have a prediction on the direction of difference for the transgender woman and transgender man authors. The new area of inquiry in this study is the gendered attributions adults make about transwomen and transmen. Should we find differences in the ratings of femininity and masculinity between the cisgender and transgender identified authors, we might use the finding to interpret how steadfast the notion of a gender binary is.

Joubert syndrome (JS) is a genetic neurodevelopmental disorder with medical features including a distinctive hindbrain malformation and developmental delay. All 35 JS-associated genes to date encode gene products localizing to the primary cilium but the mechanism behind JS is still unknown. The primary cilium serves as a cellular antenna for signaling, and when dysfunctional, can lead to the above features and more. One JS-associated gene, INPP5E, is an enzyme that controls phosphatidylinositol (PI) subtype distribution in the ciliary membrane. We hypothesize that abnormal distribution of PI subtypes PI(4,5)P₂ and PI(4)P in primary cilia is central to the JS mechanism, and therefore should be present across genetic causes. To test this hypothesis, I used fluorescently-tagged PI biosensors, domains from naturally occurring proteins that bind to different PI subtypes, and a domain from serotonin receptor (5-HT₆) that marks cilia. The PI biosensors were already tagged with GFP (green fluorescent protein) so I replaced GFP for RFP (red fluorescent protein) on the PI(4,5)P₂ biosensor, and inserted BFP (blue fluorescent protein) on the 5-HT₆ domain, allowing me to distinguish all three molecules in the same primary cilium. To do this, I removed the GFP sequence from our PI(4,5)P₂ plasmid using restriction enzymes that cut DNA then inserted RFP (amplified from a template plasmid carrying RFP) using DNA ligase. I confirmed the RFP insert by restriction enzyme digests specific for RFP and by direct DNA sequencing. PI(4,5)P₂-RFP plasmids were transformed into cells with PI(4)P-GFP and our BFP ciliary marker, then live-imaged. These tools will make it possible to determine if abnormal PI subtype distribution in primary cilia is a unifying cellular defect in JS patient cell lines. If so, we can then explore the effects of altered PI distribution and determine whether modulating PI distribution has the potential for mitigating the effects of Joubert syndrome.

Impairments in the DNA mismatch repair system give rise to mutations in microsatellite loci, which are short, repetitive stretches of DNA. Also known as microsatellite instability (MSI), these mutations manifest as new alleles of variable length. MSI is an important marker of cancer outcomes, and moreover, can directly drive oncogenesis. Despite their clinical relevance, microsatellite loci are challenging to reliably sequence using existing high throughput sequencing methods. Here, we have performed exploratory analyses using high throughput next-generation sequencing (NGS) in conjunction with single molecule molecular inversion probes (smMIPS) to enable typing of large numbers of cancer-relevant microsatellite loci. smMIPs integrate multiplex targeted sequencing with molecular-tag-based error correction, resulting in a cost-effective method for large-scale, accurate genotyping of microsatellite alleles. smMIPs are 80bp oligonucleotides, each containing a random, unique molecular tag, that hybridize to flanking regions around microsatellite loci. The targeted sequence is copied into the probe through the action of DNA polymerase and ligase. Capture reactions are then exonuclease treated, PCR amplified, and sequenced using the Illumina NGS platform. Error correction is achieved through consensus mapping of probes associated with a unique molecular tag, eliminating analytic errors otherwise inherent to the process. We have developed a panel of probes for detecting microsatellite instability events in 150 cancer-linked loci. We have compared our data to conventional NGS methods and have demonstrated higher accuracy genotyping compared to existing methods. The combination of high-throughput sequencing and robust error correction suggests that further development of this technology will be well suited for clinical laboratories searching for streamlined and accurate approaches to MSI diagnostics. The intent of this project is to develop a smMIP panel that detects microsatellite instability in heterogeneous tumor samples and that offers other advantages over existing testing methods.

On Twitter, there are over 52,000 tweets containing the word “antibiotic”, many of which portray inaccurate information. Young adults are social media’s most avid users and may view such content. It remains unclear if and how college students decipher information in inaccurate tweets. The purpose of this study is to investigate how youth perceive and assess the accuracy of Twitter posts about antibiotics. College students were recruited from the Seattle area through snowball sampling and participated in focus groups. Four focus groups were conducted with a range of 3-5 students in each group. Researchers developed a mock Twitter feed composed of real tweets discussing antibiotics. The tweets chosen conveyed accurate information (e.g., “Antibiotics don’t help viral infections…”), made inaccurate claims (e.g., “Doctor gave me some antibiotics for my flu…but a least I’ll recover”), or embodied disconnect from health assertions (e.g., “Finished antibiotic Tue...”). The facilitator read a vignette about being a college student, sick during finals weeks, who turned to Twitter, and searched “antibiotics”. Students were asked to imagine that they are this person when answering questions throughout the focus group. The mock Twitter feed was presented. Students discussed initial perceptions of the tweets, followed by being asked to group the tweets in any way they saw fit. Then students were asked to categorize the tweets again, but this time into accurate or inaccurate claims about antibiotics. After, a discussion was held about the categorizing processes. The focus groups were recorded, transcribed, and analyzed for reoccurring themes throughout the discussions and categorizing process. This study helps inform health professionals of a need to improve education about antibiotics particularly around evaluating the accuracy of social media content.

Vitamin D₃ is an essential hormone involved in the regulation of calcium and phosphate homeostasis. The major circulating form of vitamin D₃ is 25-hydroxyvitamin D₃ (25(OH)D₃) which is the biomarker often used as a measure of vitamin D status. In addition, the sulfonated form (25(OH)D₃-sulfate) has also been found to be a major metabolite, often circulating in plasma at concentrations comparable to, and at times exceeding, levels of the unconjugated form. Sulfate conjugation occurs primarily in the liver and is catalyzed by the enzyme sulfotransferase 2A1 (SULT2A1). In this study, we evaluated allele variants of the SULT2A1 gene in an effort to better understand factors that may contribute to the inter-individual variation of plasma vitamin D sulfate concentrations. Using liver samples obtained by St. Jude Children’s Research Hospital and the University of Washington, our analysis revealed the presence of three common single nucleotide polymorphisms (SNPs). Of the three SNPs, only one SNP (rs296361) was found to be significantly associated with SULT2A1 mRNA and protein expression as well as vitamin D sulfonation activity. Specifically, we found that homozygous carriers of the mutant allele had significantly reduced expression of SULT2A1 both at the RNA and protein level as well as significantly reduced sulfonation activity. The results of this study provide valuable insight into the role that genetics plays in the disposition of vitamin D, and allows us to better understand factors that may impact inter-individual differences in plasma vitamin D levels.

The Oligonucleotide Ligation Assay (OLA) specifically detects point mutations in the HIV polymerase gene conferring drug resistance. Screening patients for drug-resistant HIV is a necessary step in determining the proper treatment. However, the traditional OLA is complex and lengthy, making it non-ideal for low-resource settings. We are exploiting a novel approach to adapt the OLA into a point-of-care, paper-based format. To enable downstream isothermal ligation and paper-based detection steps, the amplification step of the OLA must be designed to preferably generate many copies of single-stranded HIV DNA (ssDNA). Typically, this can be achieved using a Linear-After-The-Exponential Polymerase Chain Reaction (LATE-PCR), which uses an uneven ratio of forward and reverse primers. Though the LATE-PCR is a useful tool in producing large amounts of ssDNA, it involves a plethora of parameters whose contributions to the reaction output are not well understood. Thus, the LATE-PCR is often optimized via trial and error and may not yield the most desirable outcome. In order to minimize wasted resources from unnecessary trials and achieve a high-efficiency amplification, we aim to understand the behavior, predict the outcome, and develop a platform for LATE-PCR optimization. To accomplish this, we have observed the behavior of LATE-PCR reactions by measuring ssDNA output when altering key reaction parameters such as the number of cycles run, primer concentration ratios, enzyme concentrations, enzyme rates, salt concentrations, and primer sequences. Analysis of these results will allow us to understand the effects of these parameters on the thermodynamics and kinetics of the LATE-PCR. This will form the basis of a computational model which will accurately predict reaction behavior, and provide specific LATE-PCR reactions given user-defined reaction parameters and desired ssDNA output. Future research will focus on expanding the LATE-PCR model to other instrument-free isothermal amplification approaches for a simplified point-of-care OLA.

In addition to its known roles in regulating cell survival, inflammation, and cardiac hypertrophy, the transcription factor nuclear factor-kB (NFkB) has been implicated as a maladaptive mediator of cardiac ischemic injury, but the underlying mechanisms remain undefined. Our objective was to assess the contribution of NFkB-p65 to myocardial injury, pathological remodeling, and ventricular dysfunction after myocardial infarction using cardiac-specific knockout mice. Intriguingly, ablation of NFkB-p65 in the heart protected against adverse remodeling and heart failure following myocardial infarction for 2 weeks. NFkB-p65 knockout mice showed reduced cardiac hypertrophy, fibrosis, and pulmonary congestion compared to control mice. Better-preserved cardiac function and less ventricular dilation were also observed in NFkB-p65 knockout mice after MI. NFkB-p65 knockout and control mice were also subjected to acute ischemia/reperfusion (I/R, 60 min ischemia & 24 h reperfusion). Loss of NFkB-p65 also protected the heart against acute I/R damage as evidenced by decreased infarct size. Accumulating evidence has demonstrated that oxidative stress participates in several aspects of cardiac remodeling after myocardial infarction, including loss of cardiomyocytes by apoptosis and necrosis, inflammatory/fibrogenic responses, and hypertrophy. Here, we examined the role of NFkB-p65 in regulating cell survival/death induced by oxidative stress. We showed that NFkB-p65 silencing in mouse embryonic fibroblasts significantly reduced necrotic cell death induced by reactive oxygen species (ROS). Specifically, ablation of p65 prevented cellular uptake of propidium iodide induced by H₂O₂ or tBHP (tert-Butyl hydroperoxide), as well as the release of HMGB1 (high mobility group box 1) into culture supernatant (a biomarker of necrosis). No changes in apoptosis markers were detected under these conditions, suggesting that NFkB-p65 mediates ROS-induced necrosis, but not apoptosis. These results identified a novel role for NFkB-p65 in mediating myocardial injury and ROS-induced necrosis, suggesting that NFkB-p65 may serve as a therapeutic target for myocardial damage and remodeling following myocardial infarction.

Exposure to abnormally high levels of estrogenic compounds can trigger dramatic developmental and sexual deformities in fish and wildlife. For instance, bisphenol A (BPA), found in plastics, is a common pollutant that threatens the natural environment and human consumers by acting as an endocrine disrupter. However, testing for these estrogenic chemicals requires samples to be processed at a laboratory with expensive equipment. Cell biosensors that are engineered to detect pollutants and produce a visual response are a low-cost, tunable, and highly sensitive alternative. Here, we show our work to expand access to estrogenic pollution detection by genetically engineering Saccharomyces cerevisiae, brewer’s yeast, to produce green fluorescent protein (GFP) until it detects ß-estradiol, a human estrogen. Our biosensor expresses a CRISPR transcription factor (CTF) which includes an estrogen receptor and mxi1 repression domain, coupled with a guide RNA that targets the promoter for GFP. We observed with a cytometer that upon ß-estradiol induction, the CTF-expressing yeast demonstrated heightened GFP expression and larger, possibly un-dividing cells. We hypothesize that our CTF is reducing cell growth and division rates, thus increasing the concentration of GFP molecules per cell, and show our work to investigate this prediction. In the future, we plan to tune the sensor to repress fluorescence only when reaching a hazardous concentration of  ß-estradiol. In addition, we will measure our biosensor’s ability to detect similar estrogens such as BPA, in the hopes that it can function as a universal estrogenic sensor.

As wolves (Canis lupus) recolonize Washington and other parts of the United States it is important to understand what sort of effect their presence will have on other species. Wolves are known to influence the behavior, distribution and abundance of coyotes (Canis latrans) and many studies have been done on this topic. However, none have studied how human presence may mediate the strength of mesopredator suppression exerted by apex predators. Mesopredator suppression is a phenomenon in which the top predator in a food chain suppresses the behavior and number of mid-level predators. We examined the ecological effectiveness of wolves in a human-dominated, multiuse landscape, an area utilized by ranchers, hunters, and many species of wildlife with few protections. We hypothesized that humans will act as super-predators and reduce mesopredator suppression of coyotes by wolves. We used motion activated cameras to track temporal niche partitioning of humans, wolves, and coyotes in northeastern Washington State. Temporal niche partitioning is a method by which competing species can utilize the same space or resources by using it during different seasons or times of day. The results of this research offer insight into the mesopredator release hypothesis and provide information necessary for implementing more effective conservation efforts that focus on combining top-down and bottom-up trophic cascades and take human presence into account.

Human pluripotent stem cells (hPSCs) are defined by their ability to self-renew, proliferate, and differentiate into the three germ layers. Controversy remains as to the role of canonical Wnt/β-catenin signaling (β-catenin dependent) in hPSC maintenance and mesoderm formation. Wnt/β-catenin signaling has shown to be important in the context of cellular homeostasis and embryonic development, and improper signaling can cause growth of cancerous tumors. By determining the role of this signaling pathway in hPSCs, we aim to improve our overall understanding of the molecular processes behind hPSC directed differentiations and early embryonic development. We assessed the role of Wnt/β-catenin signaling in hPSCs by generating a CRISPR/Cas9 mediated knockout (KO) of the signal transducer β-catenin. Expression levels of protein biomarkers of pluripotency (SSEA4 and OCT4) were evaluated and no difference was noted between wild type (WT) and β-catenin KO hPSCs. Additionally, the relative level of cell proliferation between WT and β-catenin KO cells was analyzed by quantification of EDU incorporation using flow cytometry, and no difference was noted between the two cell types. After confirming β-catenin KO hPSCs can self-renew and proliferate, we assessed the ability of β-catenin KO cells to differentiate to form mesoderm in vitro. In order to further demonstrate the level of pluripotency of the KO cells, we also attempted to differentiate the KO cells towards ectoderm. Analysis of the effectiveness of both the mesoderm and the ectoderm differentiations show that while KO cells cannot effectively form mesoderm, they are still able to form ectoderm. In this study, we demonstrate that hPSCs lacking β-catenin can still proliferate, fail to form mesoderm, but can still form ectoderm. Thus, β-catenin may not be required for maintaining the proliferative characteristics of hPSCs, but is required for formation of the mesoderm germ layer, and is therefore an important component of early embryonic development.

The Hille lab studies the movement of cells during embryogenesis using the model organism of zebrafish. Currently, we are focusing on how the phosphorylation of certain residues on the protein p120 catenin affects both cell movement and cell-cell adhesion during the early stages of embryonic development. Protein p120 catenin is imperative in the motility or adhesion of cell-cell junctions, and knocking it down results in wider somites and abnormal notochord protrusions as discussed in the Hsu et al paper published earlier by the lab. We are focused on how specific mutations in the genetic sequence of this protein affect cell migration and adhesion in the early development of zebrafish. Other members of the lab have mutated a phosphorylated tyrosine residue (Y228) within the DNA encoding for p120 to either phenylalanine and glutamic acid residues. We are then taking the mutated embryos and staining the somites and presumptive slow muscle cells using a myoD probe in order to visualize the resulting phenotype. This process involves a four-day protocol that is repeated weekly throughout the quarter in order to produce the best staining and results. After hybridization is complete, we then dissect the embryos under a dissecting scope and image them using a high-powered microscope. Using software, we analyze the images, measuring somite width as well as the number of somites in order to determine the developmental stage of the embryo. We then compare somite length and mesodermal formation between both mutated and uninjected, control embryos to gain a better understanding of which amino acid residues in the p120 protein are important in its function during embryogenesis.

Understanding cold stress response pathways in plants is key to the development of new agricultural techniques, which help prevent crop loss or degeneration due to cold snaps or early frosts. Epigenetic modifications by histone acetyltransferases appear to regulate these pathways. HAC1 is a histone acetyltransferase identified in Arabidopsis thaliana that is involved in cold stress response, and binds with transcription factors to mediate gene expression. CsWRKY46, a transcription factor recently identified in cucumbers, was found to be deeply involved in cold stress response. We identified the WRKY71 gene as the Arabidopsis homolog most similar in sequence to CsWRKY46. The WRKY71 transcription factor has also been identified as part of the response pathway to general abiotic response. We hypothesized that HAC1 regulates cold response in A. thaliana by localizing with WRKY71 and acetylating histones of nearby cold response genes such as COR47. We predicted that in cold temperature, WRKY71 and COR47 transcripts would be up-regulated only if the HAC1 protein is present. We designed an experiment with 6 A. thaliana plants: 3 wild type and 3 Salk_080380 hac1 mutants. Wild type Colombia ecostrain and hac1 A. thaliana were subjected to a 4°C cold temperature treatment of 0, 2, or 6 hours under constant light cycle conditions. RNA was extracted from the leaf tissue of each plant and RT-PCR and gel electrophoresis were performed to measure gene expression of HAC1, WRKY71, COR47, and e1f4-III (control gene). Results from this pilot study of HAC1/WRKY71 interaction with RT-PCR assays were inconclusive due to nonspecific primers. Without RT-PCR results with which to analyze the relative expression levels of eIF4-III, COR47 and WRKY71 throughout cold stress, we can neither support nor refute our hypothesis. We propose repeating the experiment with larger, more specific primers and examining WRKY71-HAC1 protein interaction through a co-immunoprecipitation assay or similar test.

The increasing demand for clean and non-fossil energy has escalated the integration of renewable power sources into the distribution system. Distributed Generation (DG) units are small-scale electrical power generators that are placed within the distribution network, which typically uses renewable energy as its source. Renewable DGs have the potential to reduce the environmental impact when the integration into the distribution system is carefully optimized. However, serious technical issues will be raised when the integration is not properly implemented. My research focuses on the optimal siting and sizing of renewable DG units within distribution systems. Both deterministic and probabilistic models of renewable based DG units will be discussed in details along with characteristics of well-known renewable generators. The effects of DGs on distribution networks will be examined by defining several objective functions. The optimal DG siting and sizing problem will be introduced and formulated as an optimization problem and several solution methodologies will be discussed. The method that we have developed is a step by step process consist of setting up an optimization problem by linearizing the power flow equation at steady-state operating points and by calculating linear sensitivity coefficients for constraints such as voltage and flow limits of distribution systems. The linear programming problem is then used to find the optimal siting and sizing of the DG units. The method works best for large-scale systems because of its calculation simplicity, which makes it capable of producing an accurate solution while demanding a lower computation power and time, making it superior to artificial intelligence techniques. Finally, a simple case study that simulates the ideal placement of DG units within a distribution network will be provided.

Studies have shown that isoflurane, a common anesthetic agent, can negatively impact lifespan in Caenorhabditis elegans, and repeated or extended isoflurane exposure causes neurotoxicity and neurological defects in mammals, including mice and human. In an effort to counteract the effects of anesthesia, rapamycin, one of the most popularly studied drug in ageing, may provide a good candidate. Its inhibition of the mechanistic target of rapamycin (mTOR) has proven to modulate pathways in ageing and mitochondrial diseases, and recent data from the Morgan and Sedensky laboratory at Seattle Children’s Research Institute suggests it may prevent the detrimental effects of isoflurane on neuronal function. We are studying the impact of isoflurane treatment and rapamycin using C. elegans, a multicellular eukaryote that has proven to be an exceptional research model in the study of lifespan and health during aging. C. elegans has been used to study pathways involved in aging for decades, and the first genes regulating lifespan in mammals were discovered in these microscopic worms. C. elegans can be grown on solid media plates and their lifespan measured through synchronized growth and plate counts. We treated wild-type and various GFP reporter worm strains with isoflurane and/or rapamycin and assessed their effects on lifespan using various techniques to measure age-related changes, including fluorescence microscopy for the accumulation of autofluorescent pigment lipofuscin, western blotting, and lifespan curves. We anticipate that these experiments will open the door toward further studies on the effects of anesthesia on aging.

Complications and mortality from glioblastoma multiforme (GBM) are often the result of tumor metastasis and thus, understanding the underlying mechanisms in glioma cell migration to help minimize tumor diffusion is of vital importance. Previously in our lab, aligned co-polymer blend chitosan-polycaprolactone (C-PCL) nanofibers have been shown to promote a migratory phenotype and an upregulation of invasion-related genes in GBM cells after 24 hours of culture. Here, we aim to further investigate the effects of nanofiber chemistry on the migratory and invasive character of GBM cells by coating C-PCL nanofibers with hyaluronic acid (HA), a major component of the brain extracellular matrix (ECM). Human U-87 MG GBM cells, a model glioma cell line, were used to study the GBM behavior on the nanofibers. Proliferation assays to determine the biocompatibility of the nanofiber platforms with and without HA were evaluated followed by cell morphology analysis to evaluate elongation and alignment. Overnight migration studies were conducted to determine if migration profiles on the nanofiber platforms were similar to those reported in vivo. Drug resistance and invasion-related genes were assessed to determine if culture on HA-coated C-PCL nanofibers could increase malignancy in GBM. Finally, flow cytometry was used to pinpoint the population of cells expressing a more invasive phenotype and thus provide an idea if HA-coated C-PCL nanofibers were a suitable in vitro platform for developing high-throughput anti-invasion cancer therapies.

The hippocampus, a brain area traditionally thought to play a key role in memory consolidation, has been shown to be involved in encoding contextual information during reward-motivated decision-making tasks. Projections to and from the hippocampus connect it with structures known to be involved in decision-making behavior and encoding cost/benefit values, making it well situated to contribute to the circuitry processing choice selection. Inactivation of hippocampal areas in rats have been shown to increase variability of choice preference between large rewards with an associated delay, and small rewards received immediately. To elucidate the role of the hippocampus in encoding economic factors of rewards, we recorded from place cells in a hippocampal subregion known to be necessary for spatial coding of salient objects and events, called CA1, during a delay-discounting maze-based task. The task was designed on an elevated T-maze to have three blocks during each daily recording session. During each block the rat chooses several times between a large reward (4 sugar pellets) with a long delay specific to that block (10, 20, or 40 seconds) and a small reward (1 sugar pellet) with a short delay of 3 seconds across all blocks. The rats were familiarized with the delays at the beginning of each block by undergoing 5 forced trials on each side. Behavior measured over 10 free choice trials then completed the block. As expected, preference for the large reward was maximal during 10-second blocks and declined as the delay increased. Preliminary analysis of place fields shows remapping between different delay conditions indicating spatial encoding of reward costs by hippocampus. Remapping was also observed between free and forced trials. These results imply that the place field representation of maze areas reflects contextual spatial information that includes the expected cost of rewards.

The Food and Drug Administration (FDA) approved drug rapamycin lengthens lifespan and rescues rodents from age-related diseases. Previous studies have shown that a 3-months regimen of rapamycin for old mice of the C57BL/6 strain changes the composition of gut bacteria that includes increased bacterial species such as lactobacillus, bacteroides and segmented filamentous bacteria (SFB). In mouse models with a mitochondrial disease, there was a decreased amount of bacteria and an extremely short lifespan. Injection of rapamycin increased the amount of bacteria and prolonged lifespan, showing an association between the quantity of bacteria and lifespan. We are interested in the effect that this experiment would have on a different strain of mice, Balb/c, which had not been previously tested for the rapamycin effect on lifespan nor the change in the composition of gut bacteria. First, we conducted a 90-day injection study on Balb/c mice, in which the mice were injected with different frequencies of rapamycin via an intraperitoneal (IP) injection. After collecting fecal samples at three different time stages – at the start of the 90-day injection period, immediately after the 90-day period, and 3 months after the 90-day period, we ran a qPCR (quantitative polymerase chain reaction) for each of the fecal samples collected, which quantifies the relative amount of microorganisms in the sample. We also monitored the lifespan of those rapamycin-treated Balb/c mice. This allowed us to see if the lifespan extension and gut microbiome change observed in the original model is reproducible in a different mouse strain. We hope that our findings help to support a future translational research strategy that applies the effect of rapamycin on specific types of bacteria to produce an intervention on gut bacteria to slow aging in pet animals and humans.

The real nonnegative inverse eigenvalue problem (RNIEP), introduced by Suleìmanova in 1949, is a long standing problem which aims to find necessary and sufficient conditions on a set of real numbers σ such that there exists an entry-wise nonnegative matrix whose spectrum is σ. Given that the RNIEP is a longstanding unsolved problem, any and all positive results are welcome. Paparella has previously shown that all sets which satisfy certain conditions, known as a Suleìmanova spectra, are realizable by a special matrix. This special matrix, named permutative matrix, is a square matrix where every row is a permutation of the first row. We seek to use recent results and other well known methods to discover any beneficial properties permutative matrices may hold, and how they may further tackle the RNIEP.

Alcohol consumption among college students has been well documented to increase during school gamedays. While studies have focused on the effects of peer and event specific influences on college drinking, there remains a gap in understanding between behavior and social media content such as Instagram. Instagram’s unique lifestyle and filter driven platform have facilitated the glamorization of alcohol displays. The purpose of this study is to assess whether the use of Instagram and viewing school gameday content amplify the receptivity of alcohol behavior. Undergraduate students ages 18-25 were recruited from a Pacific Northwest University using a convenience sample. The survey was structured using a Likert-scale questioning pattern to assess attitude towards alcohol, and four distinct alcohol scenarios will be shown to participants. Responses were used to assess whether the variations in alcohol attitudes changed while school sports affiliation was present in some scenarios. Further questionnaire regarding past experiences of viewing alcohol displays and Instagram use were also included to support findings. This survey will then aim to capture whether undergraduates favor alcohol depictions on Instagram with greater probability when the school team is made visible. Findings could be used to spread social media messages in proximity with sporting events to prevent problematic alcohol behavior. Future studies could incorporate a multi-university study focusing on school sports polls to determine whether school rankings correlate with alcohol consumption.

Sulfate conjugation is an important metabolic pathway which has been shown to play a vital role in modulating the biological activity and disposition of drugs and endogenous compounds. Studies have shown that sulfotransferases (SULTs) have a broad substrate selectivity that includes hormones, neurotransmitters and xenobiotics. SULT proteins are expressed ubiquitously in human tissue, but is largely expressed in the gastrointestinal tract, liver and bone marrow. Numerous SULT isoforms exist in humans and the most predominate of these isoforms in the liver is SULT1A1. In this study, we evaluate the impact of various single nucleotide polymorphisms (SNPs) present in the SULT1A1 gene using samples obtained by St Jude Children’s Research Hospital and the University of Washington. Of particular interest is the effect these SNPs have on the RNA and protein expression of SULT1A1. In total, 17 SNPs within the SULT1A1 gene was identified. A subsequent analysis using RNA-seq data found that 7 out of the 17 SNPs were common variants with an allele frequency greater than 5%. To assess the effects of these SNPs on the protein expression of SULT1A1, samples of human liver cytosol was analyzed using a mass spectrometry based protein quantification method. While this protein analysis continues to be an ongoing effort, we expect results from this analysis to be consistent with genotypic trends observed in our RNA-seq dataset. By evaluating the relationship between various SNPs in the SULT1A1 gene and their effect on SULT1A1 RNA and protein levels, we hope to expand on our knowledge of the impact SNPs have on the disposition of substrates that undergo sulfonation.

DNA polymerase (Pol) proofreading and mismatch repair (MMR) cooperatively guard against DNA replication errors and cancer. Defects in these activities produce “mutator” phenotypes characterized by elevated levels of base-substitutions and frameshifts. Haploid yeast with combined defects in these activities rapidly go extinct, with some strains failing to even form visible colonies - a process termed error-induced extinction (EEX). Haploid eex mutants escape extinction by acquiring antimutator mutations that suppress the mutator phenotype; however, sixty percent of eex mutants escape extinction through spontaneous polyploidization, which insulates them from mutation accumulation by doubling their number of chromosomes. In evolution experiments with pol2-4 msh2Δ strains, which display a synthetic-sick phenotype due to Pol ε proofreading and MMR defects, polyploids routinely beat out antimutator mutants, which maintain haploidy. Whether strong diploid mutators also escape EEX by polyploidization is unknown. To investigate whether polyploids arise during the evolution of mutator diploid strains, we propagated pol3-01/pol3-01 msh6Δ/msh6Δ diploids (Pol δ proofreading and base-base MMR defective) with a mutation rate an order of magnitude below the diploid error threshold. Mathematical modeling suggests that although these diploid mutators are not immediately inviable, they should be overtaken by an antimutator or tetraploid after 200 generations. We found a single tetraploid culture in 89 independent cultures propagated for 300 generations. We measured mutation rates of isolates from cultures that remained diploid and repeatedly found antimutator phenotypes. Thus, spontaneous antimutator alleles and polyploidization rescues haploid and diploid cells from EEX with markedly different efficiencies. Differences in the relative frequency of each escape mechanism may reflect the nature of the mutator alleles, the starting ploidy of the cells, or the magnitude of the initial mutation rate. Our findings in diploids suggest that mutator cancer cells near the edge of error-induced extinction may similarly be under selection for spontaneous antimutator mutations and polyploidization.

Though the rising popularity in hybrid vehicles comes as a benefit to the environment, it has proven to be a detriment to the tax revenue raised at the gas pump. The state of Washington projects to not meet the fiscal needs for civil infrastructure repair in the foreseeable future because of this shift in the market. In preparation for a future dominated by electric vehicles, Washington State has proposed implementing a tax program that charges according to the number of miles driven on Washington roads. Our team’s mission is to develop a mobile app that can dynamically track the miles driven by a vehicle. This app will be able to extract a baseline mileage count from a picture the taxpayer takes of their car’s odometer reading. It will also be able to track nontaxable miles driven out of state by using the car’s on board diagnostic system and GPS data. These nontaxable miles will qualify for a tax rebate at the end of a billing period. Our app will not only sufficiently support government’s road charge policy and help recover the lost gas tax caused by increasing amount of hybrid vehicles, but will also provide drivers in the state with the convenience to obey the policy and save money while travelling out of the state.

Immunotherapy has yielded exciting results in the clinic, primarily against blood cancers. Moving toward applying immunotherapy to solid cancers, the field has seen success in treating certain cancers, such as lung cancer, with immunotherapy. However, only a fraction of patients respond to treatment, highlighting a need for continued research in this area. The ‘KP’ model is a genetically engineered mouse model of lung cancer, relying on Cre-recombinase inducible activation of oncogenic Kras and inactivation of the tumor suppressor p53, the two most frequently mutated genes in human lung adenocarcinoma. Tumors are initiated by delivery of Cre into lung alveolar cells by the administration of Cre-expressing viruses. Using this virally-induced model, we can control the timing of tumor induction and tumor burden, and introduce additional genetic modifications into tumors. However, the use of viruses can cause inflammation, potentially impacting the immune responses that we are studying. Improving this model will make the study of lung cancer more physiologically relevant. Therefore, we are developing a system for separating viral infection from tumor initiation, dependent on a tamoxifen-inducible Cre-ER allele. By developing viruses that express Cre-ER, mice can be infected and the induction of tumors can be delayed until a timepoint when virus-related inflammation has subsided. Then, Cre activity can be induced by administration of tamoxifen. Additionally, this system allows us to control the number tumors that form, by titrating the doses of tamoxifen, to better recapitulate human disease. To date, we have cloned DNA fragments and performed Gibson Assembly, and are currently validating clones. We have produced lentiviruses and titered the viruses using GreenGO cells, a cell line carrying a Green Fluorescent Protein reporter of Cre activity. To test Cre-ER functionality and sensitivity, we will use GreenGO cells to assay differences in Cre–ER activity, using different concentrations of tamoxifen in vitro.

Plants play a vital role in maintaining a healthy ecosystem such as providing oxygen, food and shelter. Global climate change can create selective pressures that have the potential to affect plants physiologically, and their genes provide the plants with the ability to potentially adapt to climate change. In regards to water, climate change may not just be about drought but also the increase of variable precipitation, where there are longer periods of drought followed by a large amount of rainfall. This is likely to also affect expression of drought tolerance genes in plants. By studying the gene expression response in genetically different plants grown alone and in competition under variable water treatments, we can gain better insight into the genotype-phenotype relationship of drought tolerance in plants, and the molecular changes that initiate these responses. During harvesting, plant biomass was measured and then shoot tissue was frozen with liquid nitrogen and stored at -80C. To correlate the collected phenotype data to gene expression, five genes known to be involved in drought response in Arabidopsis thaliana and expressed in shoots were picked from the Drought Stress Gene Database (DroughtDB) and the Arabidopsis Information Resource (TAIR). RNA will be isolated from the harvested shoot tissue, and reverse transcription and PCR will be used to measure the changes of gene expression of the drought response genes.This research will help us understand how environmental changes affect gene expression, as well as how genetically diverse plants interact with each other under stress conditions and how this affects productivity. In the long term, this understanding can be used to improve plant productivity to maintain a healthy ecosystem especially when the climate is changing drastically. 

Extant mammals derive from three lineages: monotremes, marsupials, and placentals. The divergence of marsupials and placentals is estimated to have occurred 160 million years ago. Marsupials typically have very small, underdeveloped offspring born after a short gestation period, which are then nursed for long periods of time in a pouch. In contrast, placental mammals give birth to large, more developed young, which are nursed for a shorter period of time. Sexual dimorphism, the anatomical differences between males and females, has been demonstrated in the pelves of many species, both marsupial and placental, but no studies to date have compared the degree of sexual dimorphism in the pelves of these two groups. This study compares representative pelves from 30 species, 9 marsupial and 21 placental, across a wide range of taxa. One male and one female pelvis from each species was scanned using either a laser scanner or a microCT scanner to create a 3D surface scan. A set of landmark points were placed on each pelvis and the distances between points were calculated. The pelves were compared to calculate a value representing the amount of sexual dimorphism in each species. These values were then compared between placentals and marsupials using a t-test. Locomotion, adult female size, offspring size, number of offspring per litter, were included in the analysis as possible influencing factors. Placentals are expected to show a greater degree of sexual dimorphism due to the burden on the females for bearing larger, more developed offspring. More specialized modes of locomotion are expected to reduce the amount of sexual dimorphism. This study provides important insight into the evolutionary development of the two dominant mammalian lineages today. To our knowledge, no other study has examined sexual dimorphism of the pelvis in such a broad range of taxa.

The goal of this project is to analyze and compare the carbonyl (C=O) stretch of ethylene carbonate before and after deuteration. This molecule is commonly used as a primary component of electrolytes in batteries, and exhibits a strong C=O stretching mode. The C=O mode is a sensitive reporter to an electric field through a process known as the Stark effect. A shift in peak position is associated with a quadratic Stark effect, while a widening of the lineshape is representative of a linear Stark effect. Many times, a combination of the linear and quadratic Stark effect manifest in analysis of the lineshape. Unfortunately, the absorption peak of C=O stretching is split into a doublet due to the presence of a fermi resonant mode, causing a significant misrepresentation of the C=O lineshape. This would result in a poor estimate of an electric field associated with a linear and/or quadratic Stark effect. Substituting the hydrogen atoms on ethylene carbonate for deuterium results in a reduction of the previously mentioned fermi resonant effect, enabling a better interpretation of C=O lineshape for the analysis of Stark effects. Recently, we have observed a shift in the C=O peak position of carbonate-based electrolytes at the interface silicon nanoparticle-based electrodes. It is our hypothesis that this change in vibrational peak position is due to the previously mentioned Stark effect, a measurable electric field at the nanoparticle interface. This is significant because one could correlate the magnitude of this hypothetical interfacial electric field with the performance of a battery containing Si nanoparticles. To better estimate the linear and quadratic Stark effect contributing to the magnitude of this interfacial electric field in future studies, we have characterized the lineshape of deuterated vs. non-deuterated ethylene carbonate, as well as attempted to synthesize the material in both forms using inorganic catalysts.

The Cretaceous-Paleogene (K-Pg) mass extinction is the most recent of the ‘big five’ mass extinctions and led to the extinction of all dinosaurs except birds. Theropod dinosaurs, which were mostly carnivorous, were important members of latest Cretaceous terrestrial ecosystems. A popular hypothesis for the K-Pg mass extinction is that a meteor impact caused rapid extinction of many taxa 66 million years ago (Ma). The fossil record of dinosaur skeletons is not sufficient to assess the timing and pattern of the K-Pg mass extinction and its potential cause(s). Here, I documented theropod diversity using a sample of 783 fossil teeth from eight localities that span most of the depositional duration of the Hell Creek Formation (HCF; ca. 1.9 million years). To test the hypothesis that the bolide-impact was the sole cause of the mass extinction, I wanted to determine if significant changes in theropod diversity occurred before the impact at the K-Pg boundary. New and updated taxonomic identifications were based on the current literature. Raw, range-thru, and rarefied taxonomic richnesses as well as relative abundances and heterogeneity indices (evenness, Simpson’s) were calculated for each of the fossil localities. In total, eight theropod taxa were identified from the HCF. Richness shows a slight decrease from eight taxa in the lowermost HCF locality to five to seven taxa higher in section. Rarefied richness was lowest in the uppermost HCF. Although the changes observed in richness are minor, relative abundance and heterogeneity values differed significantly from the middle to upper HCF localities implying that major changes in theropod faunas occurred locally within the last few hundred thousand years of the Cretaceous prior to the bolide impact. Similar patterns have been observed in the amphibian and mammalian faunas suggesting other environmental factors (e.g., volcanism, climate change) contributed to the causes of the mass extinction.

Aging is characterized by time-dependent deterioration of cellular functions and increased risk for disease and death. The onset of age-related diseases including cancer, Alzheimer's, and other neurological disorders decrease the healthspan of humans. Healthspan is defined as the period of life without infirmity. My research focuses on understanding the basic biology of aging and determinants of healthspan within the emerging interdisciplinary field of geroscience. I use the model system Saccharomyces cerevisiae, a unicellular fungus, to study aging at the cellular level. To analyze cellular aging, I focus on replicative lifespan (RLS), which quantifies the number of daughter cells produced by individual mother cells before they reach replicative senescence and cease to divide. Our replicative lifespan team is composed of a group of undergraduate, graduate, and post-doctoral scientists at the University of Washington who use light microscopy and manual micromanipulation to quantify the RLS of hundreds of yeast mother cells each week. There are four important steps to analyze RLS in yeast: assay setup, incubation to outgrow cells, dissection of daughter cells from mother cells, and quantification of the number of daughter cells produced prior to replicative senescence. Using the replicative lifespan assay, I investigate genetic, environmental, and pharmacological treatments to determine their impact on cellular aging. Research produced by our RLS dissection team has led to a better understanding of evolutionarily-conserved factors that modulate aging, as well as potential treatments for certain human mitochondrial diseases.

Accumulating evidence supports the idea that stress and emotion are crucial in appraisal of situations and in deciding coping processes. Additionally, negative moods have been shown to increase the likelihood of engaging in self-defeating behavior such as impulsive acts. Despite the breadth of research into psychological stress responses and impulse control as a facet of personality, limited attention has been given to a possible link between the two constructs. As a connection between stress and impulsivity may have adaptational consequences, it has prompted new research questions about the role of stress and emotion and how they implicate coping responses. Evidence for impulsivity as a possible coping response is supported by a ‘priority shift’ theorized by researchers, in which an individual chooses to engage in impulsive behavior to attain immediate gratification, thereby shifting their attention from long-term to short-term goal pursuit. This coping response may be motivated by a demand for mood repair. A 10-day Ecological Momentary Assessment (EMA) style survey is used in conjunction with a baseline questionnaire to assess college students’ responses to stressors and negative mood. Data are from a sample of 100 UW students aged 18-20. Participants answer three surveys a day including items from the Perceived Stress Scale and the UPPS Impulsive Behavior Scale. In analysis, we test the relationship between stress levels and levels of impulsivity, and examine negative affect as a possible mediating variable. We have also hypothesized a threshold relationship in which stress will only predict impulsivity if it is exceeds a sufficient level, and will structure statistical analyses accordingly.

Currently, only photo- or thermochromic filaments exist. We are the first group to synthesize a professional grade mechanochromic filament for extrusion 3D printing. The mechanochromic unit used in this poly(ε-caprolactone) (PCL) polymer-based filament is spiropyran. By shining UV light, or stretching the filament, the spiropyran activates and undergoes a reversible isomerization from the yellow-colored closed spiropyran form to its purple-colored open merocyanine counterpart. This filament could have a huge range of uses. For example, it could be used in safety and sports equipment where high impact, potentially concussive forces would result in activation of the polymer, and notify the wearer to seek medical attention. It could also be used as a coating that allows earthquake assessment of steel beams, as well as a diagnostic tool in orthotics and wearable prosthetics to determine if they need to be exchanged. We made this filament by sending our precipitated polymer to Functionalized Incorporated, who integrated it into a blend with more PCL to create over 25 kilograms of filament. We are now testing the filament on our extrusion 3D printers. Future work will focus on blending the spiropyran-PCL polymer with a softer, more elastomeric filament such as Ninjaflex to achieve lower strain activation that is also reversible, allowing for repeated use of a single printed object. Once we have our first samples, we will analyze the mechanical properties of the blends, their changes in strain of activation, and the potential for multiple cycles of activation. We intend to commercialize this filament, and make the material available to other research groups to study the material mechanics and mechanochemical properties. This is the only mechanochromic filament in existence, and its potential uses within these customizable print materials are numerous and important.

Fundamental understanding of how effector T-cells are transformed into quiescent memory T-cells is vital for developing immunotherapeutic treatments for autoimmune diseases and cancer, and for the development of vaccines against diseases such as AIDS, tuberculosis and malaria. Previous research on conversion of memory precursor effector cells (MPECs) into memory T cells has found evidence of Treg cell involvement in the process. Treg cells are regulatory helper T cells which selectively inhibit effector T cells to prevent and suppress overly strong immune responses and autoimmune responses. Research has also shown that MPECs produce their own IL-2, a cytokine involved in immune tolerance and activation. Based on these data, we hypothesize that direct interaction between MPECs and Treg cells, possibly via MPEC-produced IL-2, is required for conversion of MPECs into memory T-cells. We also addressed colocalization of Treg cells with short-lived effector cells (SLECs). SLECs are IL-2 non-producing effector cell counterparts of MPECs and do not form memory. Potential MPEC or SLEC interaction with Treg cells was visualized through immunofluorescence microscopy of the secondary lymphoid organs (spleen and lymph nodes) of experimental mice injected with lymphocytic choriomeningitis virus (LCMV). To bypass the low precursor physiological frequencies of MPECs we employed the P14 LCMV GP33-specific TCR transgenic mouse system, which is a well-regarded model system for studying memory differentiation. To test the role of IL-2 in cell-cell interactions, we used IL-2 agonist and antagonist antibodies. A detailed understanding of how Treg cells are involved in the process of MPEC to memory T cell conversion is key to developing novel immunotherapeutic treatments.

MSN2 is an environmental stress response transcription factor that is activated in yeast by multiple different stresses—e.g. glucose deprivation, heat, oxidative stress, misfolded proteins among many others—and in turn activates a collection of downstream transcriptional programs. Interestingly, while MSN2 becomes increasingly activated with age, deletion of MSN2 increases the replicative lifespan of the budding yeast. We show that in a glucose-rich environment, MSN2 misreads the environment and drives a pathological glucose starvation response that limits lifespan. Our experiments are facilitated by a novel budget microfluidic/microscopy method rather than the golden standard of microdissection. This innovative system analyzes data through time-lapse images to develop replicative lifespan curves rather than picking off daughter cells with a needle. Microfluidics drastically increases the throughput of experiments and allows for whole-lifespan monitoring of aging yeast cells. The system allows us to efficiently further the research of MSN2 and other transcription factors so that we can potentially translate this to other complex organisms that have similar types of transcription.

Quorum Sensing (QS) is a chemical signal based communication system by which bacteria regulate gene expression according to changes in population density. This communication mechanism is used by the environmental bacterium and opportunistic human pathogen P. aeruginosa, which is a major cause of Cystic Fibrosis (CF) lung infections. P. aeruginosa employs two interconnected QS systems: las and rhl. These systems are regulated by the transcription factors LasR and RhlR, each of which binds a unique chemical signal that further activates transcriptional regulation. LasR and RhlR together control the expression of virulence factors critical for many infection processes. In laboratory strains, the LasR and RhlR QS systems are arranged hierarchically, such that LasR transcriptionally activates RhlR. Although this QS hierarchy is common to laboratory strains of P. aeruginosa, some CF clinical isolates are capable of RhlR activation in the absence of functional LasR. However, the origin of this LasR-independent regulation remains unclear. In this study, we investigate whether the LasR–RhlR hierarchy is conserved in environmental P. aeruginosa. We collected several environmental isolates and genetically engineered them through allelic exchange mutagenesis to disrupt LasR function. We then transformed wild type and LasR-null isolates with reporter constructs that produce GFP (Green Fluorescent Protein) in response to either LasR or RhlR. In these two conditions, we measured protein levels of LasR/RhlR, signal concentrations and the production of several QS-regulated products. In most cases, we observed that RhlR activity was substantially reduced in the absence of functional LasR. The observed decrease in RhlR function is so far consistent with the idea that the QS hierarchy is a conserved feature of P. aeruginosa. However, mechanisms of quorum regulation in environmental strains may differ from laboratory strains and a proper understanding of them can potentially contribute to the development of non-antibiotic therapies.

Currently, only metals are known to be affected by nearby magnets. Ferromagnetic materials, such as iron and nickel, exhibit a strong attraction when exposed to an external magnetic field. Paramagnetic materials, such as magnesium and lithium, also experience an attraction to magnetic fields, but the attraction is weaker than that experienced by ferromagnetic objects. This experiment shows that non-metallic materials can be attracted to magnetic fields when floating in water. When a small piece of paper was buoyed in a closed petri-dish filled with water and exposed to an external magnetic field, the piece of paper was observed to float towards the magnet. Additionally, the speed of the paper continuously increased as the paper got closer to the magnet. This effect was also observed with small pieces of Nafion and Teflon tubes. This means that items that we would consider to be non-magnetic may in fact be magnetic in a very small scale. Future work will be needed to find out what allows these materials to experience positive susceptibility to magnetic fields.

Macrophages are specialized cells of the immune system that direct the pro-inflammatory response and its resolution. Matrix Metalloproteinase 28 (MMP28) is an extracellular matrix proteinase expressed in alveolar macrophages during lung inflammation and is involved in macrophage recruitment and polarization. Our aim was to explore the regulation of Mmp28 expression in macrophages. We hypothesized that LPS (lipopolysaccharide, a component of bacterial cell membranes which simulates bacterial infection) stimulation would increase Mmp28 expression via a TLR4-dependent (Toll-Like Receptor 4) pathway involving Type I Interferon (IFN) signaling. We also hypothesized that Polyinosinic-polycytidylic acid (Poly(I:C), a synthetic form of double-stranded RNA which simulates viral infection) would increase Mmp28 by a TLR3-dependent pathway. To test these hypotheses, we generated bone-marrow derived macrophages (BMDM) from wild-type mice. We exposed these cells to purified recombinant IFNα/β (two forms of IFN involved in antiviral responses), LPS, Poly(I:C), or control media; 4-6 h after treatment, we isolated RNA from the BMDM, synthesized cDNA, and used qPCR to measure mRNA levels of several inflammatory genes. We also included BMDMs from knockout mice for several proteins thought to be part of the Mmp28 regulatory pathway. We confirmed that LPS and Poly(I:C) increased Mmp28 expression in BMDM. By comparing knockout mice for two adaptor molecules shared by TLR4, we found that LPS-stimulated Mmp28 expression was retained in MyD88-/- but decreased in TRIF-/- macrophages. Since both poly(I:C) and the TRIF-dependent response to LPS induce a type I IFN response, we tested if Mmp28 expression was dependent on type I IFN by using IFNAR-/- (IFN Receptor) macrophages. We found that both TLR4 and TLR3 ligands require IFNAR for Mmp28 expression. These data link type I IFN responses to regulation of MMP28 and development of experimental COPD and emphysema models of lung disease, and further our understanding of how MMP28 may regulate macrophage responses.

There currently exists no device capable of accurately quantifying bone loss around dental implants. Epithelial down growth causes resorption of bone that supports the implant which can eventually result in reduced stability of the implant, and if continues unchecked, lead to mobility and ultimately implant failure. It is crucial that clinicians are able to determine implant stability using a method that is sensitive, rapid, accurate and precise. With an increasing number of dental implants being placed throughout the world, dentists need more definitive information to improve the determination of implant health and improved diagnosis. Previous studies have found that an increase in bone loss around an implant leads to decreased natural frequency of the implant in the frequency response of the system. Piezoelectric drivers are ideal to measure natural frequency shifts as they are small, sensitive, and accurate. This project proposes the use of a piezoelectric driver-sensor pair to measure the frequency response of implants to determine implant stability and bone loss. It will be divided into three sections. The first section has shown that piezoelectric driver-sensor pairs can both actuate and sense the frequency response of a model cantilever beam system and are currently being tested together. The second section consists of testing piezoelectric driver-sensor pairs in different implant conditions to determine how bone loss and other factors influence the results seen in the frequency response of the system. The final section consists of designing a prototype handpiece implementing a piezoelectric driver-sensor pair and creating a LabView Virtual Instrument that can help clinicians assess implant stability and bone loss. This project has the potential to help improve the quality of oral implants for many patients worldwide.

Our goal is to quantify the fraction of H-α emitting B stars in M31. A small subset of massive stars emits strongly in H-α, which indicates the presence of hotter gas than is typically associated with such stars. These stars’ line emission may be related to stellar rotation, but it is unclear what causes the rotation or if there are other possible causes for the line emission. In order to shed light on this problem, we’ve begun a project to identify a homogeneous sample of these stars in M31. Our data are comprised of imaging from six fields in M31. Using resolved photometry of over two million stars, we have found 50 candidates. Using a ratio of these candidates to ordinary stars of the same temperature that are not emitting in the H-α, we can put observational constraints on models of the formation of stars in this mass range, as any viable model must be able to reproduce the observational statistics of these rare stars. Therefore, despite their rarity, they provide a key diagnostic of stellar evolution models fundamental to many fields of astrophysics.

Spinal-cord injuries can lead to loss of motor function in affected limbs. Our laboratory has demonstrated enhanced recovery of forelimb motor function after spinal-cord injury using a neuroprosthetic intervention, targeted activity-dependent spinal stimulation (TADSS), to direct plasticity in spared motor pathways. TADSS uses a recurrent neural-computer interface to detect muscle activity in the affected forelimb and deliver (electromyographic) activity-triggered electrical stimulation at a functionally related spinal site below the lesion. To further enhance motor recovery outcomes and make them longer lasting, we are combining TADSS with delivery of plasticity-enhancing neuromodulators, such as brain-derived neurotrophic factor (BDNF) and quipazine, a serotonin-receptor agonist. Rats with cervical contusion injuries have been receiving TADSS in conjunction with delivery of BDNF and quipazine for 5 hours/day through chronically implanted Pt/Ir tubes. Recovery of forelimb movements over a 12-week therapy period is assessed through performance on a trained food pellet reach-grasp retrieval task, administered twice per day during the 5-hour therapy period, which further serves as physical retraining of the injured limb. We expect our combined electrochemical therapeutic strategy to lead to enhanced motor recovery outcomes compared to delivery of TADSS alone. This study may lead to development of novel therapies for targeting spinal-cord injuries.

Suicide is one of the leading causes of death in the United States and in the world. Suicide has a profound economic, psychological, and social impact. Unfortunately, many of those who experience suicidal thoughts and behavior are not in touch with traditional mental health treatment. One method to reduce the structural and attitudinal barriers for suicidal individuals not enrolled in treatment is the use of technology. In this study, we recruit approximately 800 college student age 18 to 24 from University of Washington psychology subject pool. Subjects fill out surveys on demographics, suicidal ideation (Beck Scale of Suicide Ideation; Beck, Kovacs, Weissman, 1979), help-seeking preference (Rickwood, Deane, Wilson & Ciarrochi, 2005), stigma (Internalized Stigma of Mental Illness Inventory; Ritsher, Otilingam, & Grajales, 2003), self-concealment (Self-Concealment Scale; Larson and Chastain, 1990), interpersonal relationship (Inventory of Interpersonal Problems), and cultural values (Culture Orientation Scale; Triandis & Gelfand, 1998). First, we examine whether one’s level of suicide ideation predict one’s treatment preference (face-to-face vs online) using a multinomial logistic regression with three groups (individuals unlikely to seek help either online or face-to-face; individuals likely to seek help online only; and individuals likely to seek help face-to-face). In the second phase of analyses, we investigate potential mediators of the relationship between suicidality and help-seeking preference with mediation using bootstrapping, a method based on repetition and resampling with replacement. The results can be used to direct the development of future interventions to better fit suicidal individuals’ needs according to their preferences of being helped.

Exploring the ocean always requires human divers going undersea. Due to the complicated environment, undersea human activity is normally dangerous. Additionally, there are limits to what human operators can collect in terms of underwater data, often relying on larger, more expensive manned vehicles to acquire the desired data. We want a remotely-operated device that captures the undersea views, with geological data attached. Thus, we are presenting a submersible ROV, tethered to a clean-energy, self-sustained buoy for power and user communication, that carries cameras and positioning sensors to explore the ocean.The ROV can capture video and images that are transmitted to a host computer, which will interface with Booz Allen Hamilton’s Ocean Lens VR platform to create a user-friendly 3D environment. Based on the design requirements stated above, we built a system block diagram of the project, showing the planned system component hierarchy. From that, we assigned design subtasks according to each team member’s expertise. The primary design sections are as follows: power (conversion and distribution of power from buoy), controls (physical operation and movement of ROV), sensors (video, temperature, positioning, etc.), and mechanics (ROV chassis design, construction, and hydrodynamics). The ROV comes with an inertial measurement unit to track vehicle orientation, as well as a GPS, to provide accurate positioning data. The ROV will operate up to 100 meter clear- and salt-water depths and will collect pressure, ambient light, pH, and temperature data while giving the user near-real-time control. The ROV comes in a watertight acrylic chassis with 3 thrusters to control forward, backward, and vertical movement while in motion and idle. Receiving power from the buoy, the ROV will charge up internal batteries that will allow it to operate for 1.5 hours.

The medial geniculate body (MGB) is part of the auditory thalamus and is critical to the processing of sound. It receives input from the inferior colliculus (IC), projects to the auditory cortex (AC), then receives extensive descending projections from the AC, forming a feedback loop. Some evidence suggests that the MGB may remain plastic longer than other parts of the subcortical auditory pathway, and may play a role in learning and experience-dependent plasticity. To further understand the development of the MGB, we measured the volume of the MGB in the Big Brown Bat (Eptesicus fuscus) relative to the rest of the brainstem from prenatal ages through adulthood and found that its relative size did increase postnatally. To look for causes of volume expansion, we measured neuron size and cell density. We found an increase in neuron size and a decrease in cell density. This suggests that not only are neurons becoming larger, there is also a proliferation of connections as evidenced by greater volume of neuropil. These findings are consistent with the idea that new connections are forming as young bats learn. As the next step in our study, we are currently working on collecting the same data for the IC.

Real-time quantification of pollutants in urban and non-urban environments has far-reaching prospective benefits for our society. Such a possibility could provide instantaneous detection of hazardous chemicals, identification of areas that require air quality restoration, or estimation of agricultural effectiveness. However, currently dynamic and efficient platforms to perform quantification do not exist and only a few static sensors are deployed in sparse locations. While several mobile devices have been constructed, they are typically large, expensive and imprecise. In our project, we address this problem by building inexpensive, mobile multi-sensor devices. Each of these devices was designed to support an array of inexpensive sensors that measure the surrounding air quality and send the information in real-time to either a mobile phone or server-side application. We designed the device such that all the parts are of the lowest cost and least energy consuming as possible. To enhance the reading performance of the device, we are implementing a neural network, based on the insect olfactory system, to calibrate, recognize, track and learn signatures of scents. Early experiments show that the neural network is capable to discriminate and track more precise values of contaminants, and we are currently working on integrating sensor data from multiple devices to show the quantification of pollutants on a heat map. Overall, this project showed many possible ways to make air quality data useful and accessible. It is expected that the scale of this project can be increased by increasing the number of deployed devices such that more data can be obtained and used to train the neural network while more communities can be informed about their surrounding air quality.

Developed in the 1990s, the Implicit Association Test (IAT) is one of the most widely used measurements for implicit attitudes. Previous research has found that mental rehearsal of unfamiliar stimuli can increase both implicit liking and association with self. The present study further explored the mental rehearsal procedure and its potential mitigating effects on bias in implicit attitudes and identities. Subjects learned and remembered faces and names of either African American or European American jazz musicians. They then completed two types of IAT, the race attitude and the identification IAT. The race attitude IAT measures one’s implicit preference for one group of people over the other; the identification IAT measures whether one identifies with one group over the other. Three experiments were conducted and varied the difficulty of the rehearsal task. Face picture stimuli in IATs were either those rehearsed in the experiment or those used in the standard race attitude IAT. We hypothesize that the mental rehearsal procedure can reduce implicit bias for both attitude and identification IATs. The IAT and self-report results of this study allow an evaluation of the possible use of the mental rehearsal procedure to reduce implicit biases.

Understanding the role of fluids in active fault zones and the coevolving relationship between fault activity and fluid movements is significant in tectonics research. Calcite (calcium carbonate) veins are a record of past fluids in previously fractured rocks, giving insight into the past environments and conditions for faulting. Studying the characteristics of faulting in the past is key to understanding faulting in the present. I studied carbonate fluid precipitates in carbonate fault rocks from three major fault zones located in the Apennine fold and thrust belt of Italy to better understand this complex coexistence. The faults in this area are active and produce earthquakes. By taking thin sections of rocks from specific domains within the fault zone and using a petrographic microscope, I identified different phases of calcite relating to different stages of fault deformation. Carbon and oxygen isotopes from the CaCO₃ act as tracers for fluid source and evolution. After I identified the characteristics of the minerals that make up the fluid precipitates, I analyzed the minerals for relative abundances of C¹³ and O¹⁸, to better characterize the source of the fluids. These measurements have already been made on rocks from the same area, so I aimed to reproduce the isotopic tracers previously found. This research and characterization of the specific faults and fluids is key in preparing for future clumped isotope measurements.This further work will involve the measurement of relative abundances of a specific isotopologue of CO₂ in our samples with clumped O¹⁸ and C¹³. Temperature and clumping of these isotopes is directly related by the laws of thermodynamics and thus allow us further insight into the origin of fluid originated carbonate veins.

The increase use of composites in manufacturing complex aerospace structure calls for a more robust method of inspection. Conventional metallic strain gauges currently used by the aerospace industry require extensive preparation and can be time consuming. An embedded sensor system could tremendously improve on inspection time, as well as continuously monitoring the health of a structure. The sensors being developed are able to withstand larger tensile strain compare to traditional metal based strain gauges, and can be manufactured inexpensively. The embedding offers additional protection against harsh environment for the sensor; however, the sensor could affect the performance of a composite as a foreign body defect. This study aims to investigate the capacity and effects of an embedded uniaxial fiber strain gauges, arranged in orthogonal grid network configuration, inside a fiberglass fabric-reinforced laminate. These sensor fibers are made by hot extrusion of thermoplastic elastomer, and finished with a piezoresistive chemical coating. Linear behavior of resistance changes by strains was observed when applied loadings. Bending and torsion tests have shown the embedded sensors network to be effective in distinguishing mode of deformation in the test specimens.

Numerous imaging studies have reported brain chemical alterations in children with autism spectrum disorder (ASD). Neurochemical concentrations have been linked to specific cognitive abilities and behaviors in both typical and atypical populations. This preliminary study investigates the relationship between neurochemical profiles in high-functioning children between 8 and 12 years of age with ASD (n = 17) and typically developing (TD) children (n = 21) in relation to IQ as measured by the Wechsler Abbreviated Scale of Intelligence (WASI). All data were collected as part of an ongoing study. Cognitive, behavioral, and diagnostic evaluations were performed by clinical psychologists at the UW Autism Center. Concentrations of N-acetylaspartate, choline, creatine, and glutamate in the left cerebellum and left amygdala were measured using single-voxel proton magnetic resonance spectroscopy (1H-MRS) on a 3T MR system. Significant positive correlations between choline concentration and Performance (p<0.01), Verbal (p<0.05), and Full Scale (p<0.01) IQ were found in the ASD sample in the amygdala, and between choline concentration and Performance IQ (p<0.01) in the cerebellum. No significant correlations were found between brain metabolites and IQ scores in the TD sample, or between other metabolites and WASI IQ scores in the ASD sample. Additionally, there were no significant group differences in metabolite concentrations between the ASD and TD groups. Choline is a precursor to the neurotransmitter acetylcholine, which plays an important role in memory, attention, and cognitive flexibility. These preliminary findings suggest that small differences in brain choline concentrations may have a larger impact on cognition in children with ASD than in TD children.

Contrary to the previous belief that itch (pruritis) is simply a less intense form of pain, recent findings suggest that itch and pain are distinct sensations mediated via independent neuronal circuits. Nevertheless, itch and pain share similar molecular machinery. In mammals, itch sensations are the product of the coupling of pruritic receptors, usually GPCRs, and TRP ion channels such as TRPA1, nociceptors that normally encode noxious, painful stimuli. We identified an itch selective compound, imiquimod (IMQ), which was found to mediate pruritic responses via direct activation of TRPA1. However, other TRPA1 agonists such as allyl isothiocyanate (AITC) specifically evoke nociceptive responses. We found that IMQ is a weak agonist of TRPA1 and specifically activates itch selective neurons. These itch selective neurons are primed to respond to TRPA1 agonists while having no effect on TRPA1-expressing nociceptors. We hypothesized that the differences in sensitivity of itch-selective versus nociceptive sensory neurons to TRPA1 agonists could be caused by differences in the amount of TRPA1 channels they contain, the activation mechanism of TRPA1, or the trafficking of TRPA1 following activation. Thus, we developed a strategy to visualize TRPA1 by creating fusion proteins that tether various indicators to TRPA1. We have created a fusion construct of TRPA1 and green fluorescent protein (GFP) by using overlap polymerase chain reactions (PCR) to amplify segments of DNA and tethering the two pieces together via direct, rigid, and flexible linkers. This experiment will lead to the fusion of TRPA1 to other molecules, such as genetically-encoded calcium indicators, GCaMP and CaMPARI. These fusions will give us the ability to visualize the localization, expression, activation, and trafficking of TRPA1. These studies will help elucidate the molecular mechanisms underpinning itch versus pain sensation and show how a single ion channel can mediate distinct sensations via differential activation.

In recent decades, coral reef ecosystems have experienced mass mortality events related to increased sea surface temperatures and coral bleaching. Although numerous studies have documented the loss of coral diversity and abundance after bleaching, less is known about the potential of these communities to recover over longer time periods. We investigated whether reef communities in Bocas del Toro, Panama, were able to recover from two bleaching events: a moderate event in 2005 and a severe event in 2010. Ten reefs were randomly sampled using underwater photography before bleaching occurred (2004) and four years post-bleaching (2013). We identified marine organisms (corals, sea urchins, sponges, algae, etc.) and substrate type for 3,500 random points across a depth gradient on each reef. Subsequently, we investigated trends in reef community composition across the two different sampling periods. We hypothesize that coral recovery will be influenced by different life history strategies such as reproductive behavior and tolerance to environmental perturbation. Preliminary data suggest that sea urchins have increased in abundance since 2004, while algal cover has declined. We hope that this study will provide more information on the contribution of invertebrate herbivores to reef recovery and resilience.

One of the research goals of the Stoll lab is to characterize how the behavior of tryptophan radicals is influenced by their nano-environment. Tryptophan is an aromatic amino acid that facilitates electron transport during reduction or oxidation reactions within a protein. The ability for a protein to fine-tune tryptophan’s reduction potential by modulating its nano-environment is a crucial aspect of the function of enzymes such as cryptochromes, which are involved in regulation of circadian rhythm, and photolyases, which are involved in DNA repair. In order to study the relationship between protein environment and tryptophan oxidation, we have designed and synthesized model peptides to engineer specific environments. The peptides used in this study fold into small cages surrounding a tryptophan residue. Mutations to the peptide change the local environment around tryptophan from charged to hydrophobic with varying levels of solvent exposure. The reduction potential of the tryptophan in these peptides was then determined using differential pulse voltammetry experiments. Each peptide was tested over a pH range from 1-7.5 in order to determine how the pH dependence of tryptophan's reduction potential responds to changes in its nano-environment. We have identified some initial trends such as the tendency for more cationic environments to increase the potential at which tryptophan oxidation occurs. Further analysis will allow us to develop a model which will be able to directly predict tryptophan oxidation behavior as a function of the pKas of surrounding residues and a weighting metric based on solvent exposure. We hope to extend this model to be able to describe the tryptophan redox behavior in full proteins.

    Optical microscopy is a cornerstone of biological research. It reveals to scientists phenomena that explain many of life’s secrets. Despite its capabilities, traditional optical microscopy techniques are limited to a spatial resolution of ~250 nm due to the diffraction limit of light. Consequently, features smaller than ~250 nm cannot be distinguished. However, the development of super-resolution microscopy techniques such as the Stochastic Optical Reconstruction Microscopy (STORM) now enables resolution up to ~60 nm. An essential criteria of STORM requires that the fluorophores turn on and off (photoswitch) stochastically. Up to date, the fluorophores used in these techniques are commercially available fluorophores with photophysical properties optimized for traditional optical microscopy methods. Since the photoswitching properties of these commercial dyes for use in STORM is not well understood, my project thus involves tuning the electronic properties of the cyanine scaffold by adding various functional groups to the dye in order to improve the photoswitching properties. I am currently working on synthesizing cyanine fluorophores in the red region that have electron-withdrawing groups on the indole molecule and a carboxylate linker attached for bioconjugation. Organic synthetic chemistry techniques will be used in the creation of these dyes. The structure of the dyes will then be characterized by proton NMR and mass spectrometry, while the photophysical properties will be characterized by Ultraviolet-Visual spectroscopy. The dyes will eventually be applied to fixed sample staining and STORM. If successful, these new dyes will improve multi-color STORM and researchers will be able to view cells in greater detail.

The Olympia oyster, Ostrea lurida, benefits the ecosystem for the health of marine environments. Olympia oysters provide natural filtration and restore complex nearshore habitats. Along the estuarine habitats of Washington State, native oyster populations have been declining. Invertebrates reproduce by producing thousands of larvae to ensure survival of a few. Most larvae will not survive due to natural causes such as predation or advection, or human causes such as poor water quality or ocean acidification. The larval stage is the most mobile form of development, and it determines whether they settle away from where they were reproduced or remain in the same area. Larval identification allows for the study of larval behavior and provides data for environmental organizations as to whether restored populations self-sustain or export new offpsring. In this study, we collected samples from Fidalgo Bay, WA from eight stations: five intertidal and three subtidal including three replicates of each sample. Larval samples have been quantified by microscopy and visual identification to describe spatial and temporal distribution of O. lurida larvae. Although the results are pending, data is being collected to find out if larvae will be found in intertidal sites near suitable substrate for adult settlement and adult populations. Results will also indicate if their narrow band as adults is due to their settling everywhere and then die when they get bigger, or if they settle only in their narrow band. If successful, restored populations of Ostrea lurida will attract fish, invertebrates and other marine organisms throughout Fidalgo Bay and therefore prevent these species from going extinct.

Youth at risk of justice involvement often have histories of complex trauma. Complex trauma is exposure to traumatic stress at a critical age and is found to affect emotional self-regulation. Difficulties with self-regulation can lead to higher levels of aggression, which is associated with criminal behavior. Consequently, I am interested in how complex trauma affects overall aggression in youth. I used the results of a survey conducted by the Seattle Youth Violence Prevention Initiative to examine the relationships between items on the trauma inventory and levels of aggression. I hypothesized that multiple trauma exposures, a lack of normal emotion, and increased arousal and vigilance would be positively associated with aggression. I also hypothesized that re-experiencing trauma and avoiding trauma reminders would not be associated with aggression. I found that multiple trauma exposures and re-experiencing trauma were significantly and positively associated with aggression. Avoiding trauma reminders, lacking normal emotion, and increased arousal and vigilance were not significantly associated with aggression. These results support my hypothesis that multiple exposures to trauma is positively associated with aggression in youth, but the results do not support my hypotheses that lacking normal emotion and increased arousal and vigilance are positively associated with aggression in youth as well. The significance of positive associations between multiple exposures to trauma and aggression as well as re-experiencing trauma and aggression suggests that there is a relationship between complex trauma and aggression in youth, which may also be related to criminal behavior. This research suggests the importance of addressing trauma in aggressive youth before their behavior progresses in order to prevent youth from being involved involved in criminal behavior.

Social media plays a significant role in the lives of college students today. Most young adults (82%) use Facebook while 59% use Instagram. Research has begun to examine the effects that social media usage has on mental health issues, but conclusions have been mixed. Anxiety is the most common mental health issue among college students, with 17% of undergraduates having been diagnosed with an anxiety disorder in the past year. Few studies have focused on social media’s effect on anxiety in young adults. The purpose of this study was to produce a stakeholder-driven conceptual model of the relationship between social media and anxiety. This qualitative focus group study examined whether college students perceive a relationship between social media and anxiety and what factors play into the relationship. College students ages 18-22 were recruited through student organizations and class settings. Participants were asked questions about their perceptions of social media and anxiety and to construct a consensus-based conceptual model. During the conceptual modelling activity, participants arranged notecards representing factors derived from the literature, including social comparison and approval, self-esteem, stress, and idealized selves on social media. Participants also had the opportunity to write in additional factors on blank notecards. Then, participants collaborated with the other focus group members to construct a collective model on a whiteboard. Analysis included comparing models that the participants created and synthesizing them into one final model. Additionally, the themes and key points of the focus group discussions were analyzed using qualitative methods. The results of this study will inform future research by creating an innovative conceptual model of the relationship between social media and anxiety. Future studies will be able to form targeted research questions based on the model or more closely examine contributing factors and outcomes.

At low obliquity, planets like Earth will tend to form permanent ice sheets at the poles, if anywhere. However, at high obliquity, the poles receive more sunlight on average than the tropics, and so ice sheets might be expected to form at the equator. We have used a simple energy balance model for investigating the orbital, atmospheric, and obliquity parameters that allow for the formation of permanent ice at the equator. We find there is a narrow range of semi-major axis and obliquity values that allow for such “ice-belts”. First, we experimented with the range of stability of ice-belt formation by increasing the semi-major axis to the edge of the habitable zone, increasing CO2 levels to counteract the decrease in solar flux. This led to a very small range of conditions that allow for stable ice-belts. To further investigate the cause of this, we decreased the luminosity of the host star while increasing CO2 levels (to simulate the control temperatures). This stabilizes the otherwise unstable system, although the range of ice-belt formation is still narrow. Further, we find that this equatorial ice formation often leads to instability due to the ice-albedo feedback. This work suggests that it should not be common for Earth-like worlds at high obliquity to maintain large ice sheets.

The large-scale configuration of the cosmos has long been a mystery to astronomers. My research project aims to shed light on this mystery. The overarching goal for my research project is to further understand the distribution of matter on 10⁵ –10⁷ light-year distances by comparing numerical simulations to actual observations of the universe. This comparison tests the current theoretical parameters of the physics governing the largest scales of the cosmos. I used simulated data from models made by Professor Matthew McQuinn and observations of quasar spectra from the Very Large Telescope (VLT). From the VLT data, I analyzed quasar spectra for absorption lines at a hydrogen electronic transition called the Lyman-alpha line. The IGM absorbs amply at this transition, creating absorption lines throughout the quasars’ spectra. I processed this data by programming Python scripts to compare flux probability distribution functions (PDFs) of the Lyman-alpha absorption in the observed quasar spectra to the simulated quasar spectra. Through this comparison, PDFs of observations fell within one standard deviation of the simulated PDFs. Although at higher redshifts (further back in cosmological time), there was considerably more variance. In order to further constrain the observational flux PDFs, I aim to eliminate systematic and statistical uncertainties that may make the observations spuriously differ from models. The salient errors include fitting precise continua (spectra without absorption) and removing metal (predominantly carbon-4) contamination from the Lyman-alpha forest. I used Principal Component Analysis to predict the continuum spectrum but found that a fit by eye was more effective. Through analysis of absorption probabilities outside the Lyman-alpha forest, I statistically removed carbon-4 from the data inside the Lyman-alpha forest. Ensuring the most precise flux PDF measurements are used is critical to the comparison of observations to simulations and to the constraints of parameters governing the universe.

Within computed tomography (CT), increased image quality comes at the expense of amplified radiation exposure to patients. To reduce radiation exposure, techniques have emerged which use iterative methods to reconstruct images from limited-angle or sparse-view CT data. Superiorization is a recently proposed framework which “superiorizes” an existing reconstruction algorithm with respect to some secondary objective function. As a result, superiorization can reduce artifacts in the reconstructed image caused by sparse-view and/or limited-angle data. Within certain constraints, the superiorization method guarantees a reconstruction which fits the data equally well as the original reconstruction. More importantly, superiorization is expected to produce an image with higher quality. Existing research within superiorization has been confined to reconstructing images given data generated from monoenergetic X-ray beams. Despite the promise of this existing research, polyenergetic X-ray beams are common in clinical settings and existing methods need to be extended to handle polyenergetic data. To this end, our research goal was to extend the superiorization method to reconstruct images from polyenergetic data, using total variation (TV) and anistropic total variation (ATV) as the secondary objectives to be superiorized. In addition to successfully reconstructing images from polyenergetic data, we also demonstrate our method is capable of reconstructing images with “missing” data which is a result of sparse-view and limited-angle scans. This research further extends the potential of utilizing iterative methods in CT and thus reducing radiation exposure to patients.

When cold-season oceanic midlatitude cyclones make landfall over coastal mountain ranges, forecasters commonly assume that locations at higher elevations and closer to the mountainous interior will receive greater orographic precipitation enhancement under all synoptic conditions. In this study, we test this assumption by examining precipitation data obtained at two stations on the windward slopes of the Olympic Mountains in the northwest corner of Washington State. These stations are located 35.2 and 45.4 km from the coast along the Quinault river valley, only 10.2 km apart. It was found that the two stations often recorded drastically different amounts of rain for the same storm system. The U.S. Climate Reference Network (CRN) site is located immediately northeast of Lake Quinault at 86.7 meters elevation, only slightly lower than the second station in Bunch Field at 115.8 meters elevation, which is located slightly closer to the higher terrain of the Olympic Range. This study examines daily rainfall at the CRN and Bunch Field sites over 9 winter seasons (Sept 1 -Mar 31) from October 2006 through December 2015. Dates were identified where one station received 15 millimeters more precipitation than the other in a 24 h period. Composite synoptic maps were made using the NCEP North American Regional Reanalysis data for dates when CRN exhibited higher rainfall totals and for when Bunch exhibited higher rainfall totals. It was found that when CRN had significantly more rainfall than Bunch Field, the flow was predominantly from the SW at all levels and warmer than climatological average. When Bunch Field had significantly more rainfall than CRN, the synoptic flow was more southerly with a large trough and a strongly baroclinic structure. These subtle differences illustrate that orographic enhancement patterns vary strongly with large-scale environmental conditions. The results can be applied to how flooding events can vary on small scales, which in term will inform the management of hydrologic resources.

We live in a world driven by efficiency, optimized by technology. In this project, our team is developing specialized commercial smart home products as we investigate their efficacy. With the industry’s unprecedented growth over the past several years, we believe that smart home technology will quickly become a cornerstone of today’s automation paradigm. To focus our scope, we are working with an industry leader: Wally, who has already launched a successful commercial package. Our job is to ideate, design, and create prototypes for new sensors that will augment and enhance their existing product. We are focusing on utilitarian applications of smart sensors. As such, the first product we are developing is a sensor that detects various gas leaks, ranging from volatile organic compounds to natural gases. The second product we are developing is a water pressure sensor that measures flow rate throughout a house. Both sensors will connect to a cloud infrastructure and data is sent to a mobile application where users will be alerted in real-time and can view analytics. Additionally, both sensors will interface with other automated products such as shutoff valves. If our research and development goes as planned, we hope to add both products to the existing Wally line and release them commercially.

36.7 million people are living with HIV worldwide, with most in low- to middle-income countries. Among many clinical interventions for controlling HIV, Antiretroviral Therapy (ART) has shown success in reducing HIV transmission. However, the effectiveness of these ART is undermined by rising drug-resistant HIV. The lab-based Oligonucleotide Ligation Assay (OLA) can specifically detect HIV drug resistance as an alternative to expensive complete-genome sequencing, but its reliance on moderately expensive lab equipment prevents its universal access. Our lab, in collaboration with the Frenkel Lab (Seattle Children’s Research Institute) and the Lai Lab (UW Bioengineering), is developing a simplified OLA-based HIV drug resistance test. In the OLA, PCR amplification of the HIV Pol region is necessary before subsequent detection of Single Nucleotide Polymorphisms (SNPs) conferring HIV drug resistance. Current amplification relies on a thermal cycler, a >$2.5k instrument not widely available in low resource settings. Isothermal Helicase Dependent Amplification (HDA) eliminates need for a thermal cycler. HDA has been used to amplify the conserved Gag and Env regions of HIV, however it has never been used for amplification of the Pol region, a non-conserved region containing mutations that interfere with DNA annealing in isothermal amplification. We have designed and validated three sets of primers that successfully amplified three regions in the HIV Pol gene covering six common SNPs that confer HIV drug resistance against 1st-line ART. These primer sets each could detect down to 10 copies of HIV Pol DNA per reaction.  We are currently working on a multiplexed HDA assay that simultaneously amplifies three regions in a single tube to reduce cost and consolidate workflow. Implementation of HDA will allow OLA to be further simplified for use in low resource settings by allowing accurate diagnoses of HIV-infected individuals, controlling transmission rates, and reducing the evolution of drug-resistant HIV within entire populations.

Lake Kapowsin in Pierce County, formed approximately 500 years ago when the Electron Mudflow from Mount Rainer dammed a valley creek, was recently designated Washington’s first freshwater aquatic reserve. In light of this new designation, the Washington Department of Natural Resources contracted University of Washington Tacoma staff to collect summertime water quality data to be used in future watershed management decisions. Field sampling was carried out once a month from June to October 2016 at three stations along a lake transect from inlet to outlet. Water samples were collected at surface and near-sediment depths to monitor chlorophyll a, total suspended solids, turbidity, and alkalinity, and in situ vertical profiles of temperature, pH, specific conductivity, and dissolved oxygen were measured. Secchi depth was also recorded and phytoplankton samples were collected by vertical net (20 µm mesh) tows. Elevated chlorophyll concentrations in the water column, coupled with a shallow Secchi depth, give indication of high algal productivity leading to poor light penetration. Consequently, Lake Kapowsin, despite its shallow depth, experiences thermal stratification between June and September due to the inhibited light penetration. This thermal stratification, in concert with the high algal productivity and bacterial aerobic respiration at the bottom of the lake, results in anoxic conditions and reduced pH in the bottom waters. These resultant conditions lead to a release of phosphorus and, to a lesser degree, nitrogen from the sediment. Ultimately, the persistent anoxic bottom waters capped with warmer waters makes the lake more suitable for warm water fish than cold water fish. The elevated productivity and anoxia in Lake Kapowsin is likely due primarily to naturally occurring eutrophication owing to the flooded forest inundated during the creation of the lake.

Iron is the fourth most abundant element in the Earth’s crust. Both the behavior of Fe and isotope fractionations are associated with redox state. Therefore, the study of Fe isotopes is often used in tracing change of oxygen in the atmosphere and hydrosphere. My research will be divided into two parts — chemical separation of iron and mass spectrometry. Purification of iron requires a two-step anion exchange column scheme in HCl medium. With concentrated HCl, matrix and interfering elements will be eluted as waste while Fe will be retained on the resin, because iron forms strong anion chloride complexes. Iron will subsequently be eluted at lower HCl molarity. Iron isotopic analyses will be performed with HR-MC-ICPMS — an instrument that measures isotopic ratios of elements. I’ll have different combinations of the acids tested, and will find out the most efficient scheme which can maximize the elimination of unwanted elements by the day of my Symposium presentation. Once the method is devised, it can be utilized by a broad implication. It can be applied to the analysis of Fe isotopes in rocks, minerals, and extraterrestrial samples. Ultimately, iron isotope measurements will be used to extrapolate geochemical cycling of iron in the ocean and sediments. Measurements can also be used to extrapolate the conditions of core formation and can provide context for Fe fractionation during magmatic processes in extraterrestrial bodies. In the marine realm, Fe isotopes can be used to trace various sources of Fe to the ocean such as atmospheric dust, hydrothermal vents, and oxic seafloor sediments.

The ability to replicate results and data from a research project is important for validating published research and improving the transparency and trustworthiness of the scientific process. Still, reproducibility issues, like failure to reproduce experiments, are not uncommon. Towards this, we set out to replicate of a recent (2017) publication, “Realization of compact tractor beams using acoustic delay-lines”. We selected this publication because the methods were explicitly written to be accessible and reproducible. During the reproduction, we evaluated the level of accessibility of materials required and ease of reproducing the published results. We also further explored the properties of the replicated system by assessing the limitations of the items that can be captured with an acoustic tractor beam. This work helps to bridge the translation of published research to non-experts, as well as evaluate the transparency and reproducibility of science.

Dead reckoning is the process of calculating one's current position by using a previously determined position, or fix, and advancing that position based upon known or estimated speeds over elapsed time.  It is typically used when external systems, such as GPS satellites or acoustic tracking, are unavailable to track a vehicle relative to earth-fixed coordinates. When used on an autonomous underwater vehicle (AUV) it cannot account for the influence of tidal currents changing the vehicle’s displacement without influencing internal speed and heading data. My research experiments with an Ecomapper AUV in Australia in September 2016 illustrate the influence of such tidal currents on dead-reckoning AUV tracks. The experiment was completed at the Morton Bay Research Center of Queensland University and the research data were collected by Ecomapper AUV from designed runs. As a result, the comparison of actual AUV track with the predicted one that was calculated by Matlab with the application of Dead reckoning method showed how much the tidal current influenced the predicted track, and how to reduce this difference by setting the AUV's heading with the tidal current in particular angles. The results are useful for planning new AUV missions in similar tidal currents for reducing position errors generated by this technique.

It has been well documented in the literature that many children with autism spectrum disorder (ASD) display abnormalities in sensory processing, which is often displayed through hyper- or hyposensitivity to sensory stimuli. Auditory processing difficulties in particular are shown to be the most common. However, there is limited research into the association between abnormal auditory processing with other factors like language and cognitive ability. This study aims to further examine auditory processing difficulties in children with ASD by first looking at the auditory items on the Short Sensory Profile (SSP), a 38-item caregiver questionnaire measuring sensory reactivity. While the SSP is a measure often used in studies to quantify sensory behaviors and has commonly shown atypical behaviors within the Auditory Filtering domain in children with ASD, it should be noted that additional auditory items are also present within other SSP domains. This study examines all of the items on the SSP that have to do with auditory processing, and scores will be compared between two groups: children (nine to ten years old) with ASD and children with other developmental disorders within the same age range. This study also investigates the relationship between SSP auditory scores with receptive and expressive language scores from the Peabody Picture Vocabulary Test and the Expressive Vocabulary Test, as well as cognitive scores from the Differential Ability Scales. Data analysis is ongoing, and it is hoped that the results will help to increase conversation around this topic and the need for further research into the functional impact of sensory processing difficulties in children with ASD. 

The embryonic development of vertebrates involves the anterior movement of progenitor cells, which form and proliferate at the posterior end of the embryonic body and then move anteriorly to differentiate into neural and muscle cells. Our interest is in understanding how these progenitor cells migrate, and how their movement is regulated, using the zebrafish as a model system because of its outstanding optical qualities. In my project, I am examining the role of specific proteins that are hypothesized to be important in this migration. To alter the expression of these proteins, I injected messenger RNAs encoding these proteins and also some newly developed transgenic lines where the expression of these proteins is regulated by a short period heat shock. For the mRNA injections I examined the phenotype of the whole embryo, whereas with the transgenic lines I microdissected the most posterior end of the growing embryo (called the “tailbud”) and imaged cell movement using a confocal microscope, with image analysis using software such as Slidebook and ImageJ. My results show that a change in the expression of several interesting proteins produced abnormal embryos and yielded abnormal movement of progenitor cells and impaired formation of the tail. These results provide useful information that helps us understand vertebrate development, including human embryogenesis.

During brain surgery for intractable epilepsy, clinicians utilize electrodes placed on the cerebral cortex to monitor seizure activity and map functional brain areas in patients so that the epileptic brain regions may be removed. This recording method is called electrocorticography (ECoG) and is essential to prevent the accidental removal of brain regions that are crucial for daily activities. To reduce tremors produced by Parkinson’s disease, electrical stimulation is applied to electrodes implanted within the deep brain near the subthalamalic nucleus. In both cases, to optimize treatment, a more detailed study of current flow during electrical stimulation than is currently done, due to surgical limitations of electrode placement, is needed. In addition, the time available to work with patients during brain surgery is extremely limited, necessitating a non-clinical research platform. I developed a saline-enriched gelatin phantom brain that mimics the electrical resistivity of the brain to study brain stimulation outside the surgery room, and I compared measurements taken with this model to analogous measurements of the brains of human patients made by my group. I found that my measurements agree within 10-15% with those made by my group on human brains. Using the phantom brain will allow us to easily and safely study electrical brain stimulation. Also, an electrical phantom that can be used offline to determine the most effective measurements will be extremely valuable to inform future research questions while working with patients. We are utilizing the phantom brain model to study how various electrode geometries and the cerebrospinal fluid layer that covers the brain have an effect on the amount of current that flows into the brain. Findings in our phantom brain will help us more effectively treat neurological disorders like intractable epilepsy and Parkinson’s and perform safer stimulations through optimized stimulation protocols.

Surveillance data has shown extremely high rates of bacterial sexually transmitted diseases (STDs) among men who have sex with men (MSM). STDs increase the risk of acquiring and transmitting HIV. HIV infection is associated with lower education levels; however, it remains unknown whether STDs are associated with lower education levels among. To determine whether education level is associated with diagnosis of bacterial STDs, we plan to conduct a cross-sectional study using baseline data from a larger cohort study on MSM and bacterial STDs. All enrollees were MSM who met inclusion criteria for either >4 sexual partners (female or male) in the past 2 months, ≥10 sexual partners in the past 12 months, or diagnosis of a bacterial STD in the past 12 months. Data consists of an enrollment demographics survey that includes age, race, educational attainment, income, number of sexual partners in the past 2 months, and STD testing results (gonorrhea and chlamydia at the pharynx and rectum, and syphilis). We will evaluate the relationship between educational level (categorized as <college education or ≥college education) and STD test results (aggregated as any positive STD result), as well as by specific STD (gonorrhea, chlamydia, and syphilis). We will also conduct a multivariable regression model to adjust for the confounders of age, race, income and number of sexual partners. Results are anticipated by April 2017. The analysis is expected to show a negative correlation between educational levels and positive STD test results. Research is needed to address the rising rates of STDs in MSM in relation to education levels.

The smart parking group has developed an Application Program Interface, or API, with analytics tools to support analysis, policy development, and innovation by Seattle Department of Transportation, or SDOT, and third party solution providers. An API is a set of routines, protocols, and tools for building software applications. It specifies how software components should interact and is also used when programming graphical user interface components. Python computer code is used to make calculations and place relevant data in a public database, where it can be remotely accessed for applications by the API. To demonstrate our API’s functionality, the team is currently developing a visualization tool that overlays analytics produced by the API on a map. The analytics algorithms we will implement integrate a wide variety of data streams available through our collaboration with SDOT. The API will provide the needed support for apps that deliver a dynamic summary of information that can help citizens determine the best and worst places to find parking in Seattle metro area, at specific times and dates.

The purpose of this research project is to demonstrate the feasibility of incorporating a new photocatalytic Diels-Alder reaction into the curriculum of undergraduate organic chemistry laboratories at University of Washington. Although the traditional Diels-Alder reaction is thermally allowed and does not require UV light catalysis, it fails to succeed when both the dienophile and diene are electron rich due to electrostatic repulsion between the reagents. Recently, an alternative photocatalytic Diels-Alder reaction was reported in the chemical literature involving a ruthenium bipyrazine catalyst, Ru(bpz)₃X, where X denotes the counter anion. This new photocatalytic method is advantageous because it allows the Diel-Alder reaction to work when both the diene and the dienophile are electron rich. Consequently, we believe that the new photocatalytic Diels-Alder reaction would be an attractive addition to our undergraduate laboratory curriculum by increasing the versatility of Diels Alder reagents to accommodate both electron rich substrates. Because the ruthenium catalyst is not commercially available, it must be synthesized from chloropyrazine. Two equivalents of chloropyrazine were coupled together under heated conditions and a Pd(Ph₃)₄ catalyst to yield a solution mixture containing bipyrazine, which was purified via silica flash chromatography. Subsequently, bipyrazine was reacted with RuCl₃xH₂O in ethylene glycol at 170°C to afford a solution mixture containing Ru(bpz)₃Cl₂. This chloride salt was reacted with NaBArF to precipitate the desired catalyst, Ru(bpz)₃X, where X represents the anion BArF-. Finally, the catalyst was purified through alumina flash chromatography. We intend to test the efficacy of this new photocatalytic method using the electron rich substrates trans anethole (dienophile) and isoprene (diene) in the upcoming months. We expect that the two photocatalysts we have synthesized will catalyze the cycloaddition reaction between isoprene and trans anethole with endo stereoselectivity.

An object's color is composed of three dimensions: hue, brightness, and saturation. According to Opponent Process Theory, hue is determined by two opposed chromatic color mechanisms (red versus green, and blue versus yellow). The red-green and blue-yellow balances of an object are determined by the activity of the three retinal cone-photoreceptors: S-, M-, and L-cones which have peak activations at short, medium, and long wavelengths of light. S- and L-cones are responsible for the perception of red and oppose M-cones which signal green, and M- and L-cones the perception of yellow and oppose S-cones which signal blue. Previous research has shown how achieving a red-green balance can be influenced by various brightness effects. This current study demonstrates how these same stimuli are influenced by manipulating saturation. We displayed a variety of colored stimuli that varied in their saturation and red-green balance. Saturation was manipulated by the experimenter by adding blue phosphor to a colored stimulus composed primarily of red and green phosphor on a CRT monitor in order to create 6 different experimental conditions. Participants were asked to set a colored stimulus on the screen to a red-green balance by varying the amount of red and green phosphor with a keyboard, with the blue phosphor levels being held constant. Results show that as more S-cone activating blue phosphor was added to the stimulus participants adjusted the stimulus more toward the direction of M-cone activation (green). This is the predicted direction of Opponent Process Theory since L- and S-cone activity add together to signal redness. We conclude that as you reduce the saturation of a red-green balanced stimulus by adding blue, the stimulus will appear redder. Psychophysical researh like this can help guide ideas to design safety signs, materials for people working in low-light environments, or other real-world physiological investigations.

Lai et al. tested nine interventions to reduce implicit racial biases. Although all interventions immediately reduced biases, the effects produced did not last for long with duration of a couple of hours. Kirby and Greenwald made a discovery, termed "mental ownership", whereby active rehearsal of novel stimuli increases implicit liking for the stimuli and identification of it with self. Inspired by this finding, we conducted a series of experiments in attempt to test if a procedure developed from "mental ownership" could be effective in terms of mitigating implicit biases. We randomly assigned University of Washington undergraduate students to complete mental rehearsal procedures of one out of two complementary categories; they actively rehearsed and memorize exemplars of either insects or flowers. To measure their implicit biases, subjects then completed Implicit Association Tests. D scores, measuring their performances on the tests, were used to assess the effects of the mental rehearsal procedures. We expect for the procedure to produce a significant effect, and we should see an increase in subjects' implicit liking for the category from which exemplars have been rehearsed. If our hypothesis is correct, more studies will be conducted to test the duration of the effect and compare it to that of the traditional nine interventions.

Post-surgical endophthalmitis is a serious vision-threatening inflammation following any ocular procedure. Detection of endophthalmitis with culture has been traditionally used; however, endophthalmitis caused by viruses are overlooked. In a 1995 Endophthalmitis Vitrectomy Study by RK Forster, 33% of patient samples were culture negative. This represents a significant portion of patients who could have had endophthalmitis due to viruses. Using alternative techniques, we can characterize viral pathogens found in post-surgical endophthalmitis with qPCR (quantitative polymerase chain reaction) and polymerase chain reaction (PCR). qPCR quantifies the number of viral DNA strands whereas PCR indicates presence or absence of the viral DNA. This allows doctors to treat more patients which would have been previously thought to not have endophthalmitis. Patients diagnosed with endophthalmitis following intraocular procedures were recruited from Wills Eye Hospital and University of Washington from Sept 1st , 2014 to Dec 31st , 2015. All patients underwent vitreous or aqueous tap. The specimens were sent for microbial culture. Following DNA extraction, qPCR for Torque Teno Virus (TTV), Merkel Cell Polyomavirus (MCV), and actin were performed in all samples. qPCR results were verified via PCR and gel electrophoresis. A total of 29 samples were collected (12 culture-positive [41.38%], 17 culture-negative [58.62%]). Overall, 18 (62.07%) samples were positive for TTV and 15 (51.72%) for MCV. The presence of TTV or MCV was not associated with culture positivity by Fisher’s exact test. Viral DNA sequences are unexpectedly common in endophthalmitis. The role of these viruses in this disease process remains unclear.

Current research in microelectronic devices made of semiconducting silicon wafers focuses on developing high aspect ratio vertical connections between stacked wafers, known as through-silicon vias (TSV), which result in 3D integrated circuits. These 3D integrated circuits are used to increase density of memory chips, enhance performance, and reduce power requirements, resulting in smaller and more efficient electronic devices. After processing silicon wafers with preliminary photolithography, a dense pattern of high-aspect ratio TSV “holes” is achieved with a deep reactive ion etching (DRIE) tool which utilizes the Bosch process. When a hardware change in the electrostatic chuck of the DRIE tool used for this project resulted in poor etch quality and inconsistent sidewall profile, this experiment attempted to develop a new sequence of steps, or “recipe,” for the DRIE tool in order to correct the new issues. Etching parameters were refined in order to achieve the primary goal, to improve sidewall conditions which were not consistently 90 degrees, as well as secondary goals, to obtain the tool’s etch rate and selectivity towards photoresist against silicon. Sidewall profile, etch rate, and selectivity were determined by sample analysis via scanning electron microscopy. The conclusion of this experiment was a process recipe which could reliably achieve the desired high aspect ratio vias and continue downstream processing of TSV development.

Restoration of threatened marine invertebrate species necessitates information on cryptic life history stages, including the distribution and abundance of the crucial larval stage. The larval stage is the most mobile stage of development, and information on their location and migratory tendencies gives insight into whether or not a restored population will be able to persist after initial aid from humans. Traditionally, larval samples have been quantified through visual identification methods that rely on knowledge of shell morphology. More recently there has been a shift to using molecular methods including DNA extraction and quantitative polymerase chain reaction (qPCR) to quantify and identify larvae. QPCR is a time efficient and cost effective way to identify species by specific DNA code, which is copied thousands of times and quantified using fluorescent markers. The results, when placed against known standard samples, can describe the abundance of specific larvae in environmental samples. Limited studies have successfully used qPCR with environmental samples due to potential inhibitors within the sample that can influence amplification. In this study, two methods of DNA extraction were tested on native Olympia oysters for qPCR using environmental samples from Puget Sound. Of the two methods, one was a more expensive DNA extraction and isolation kit while the other was a cheaper, modified DNA extraction method using proteinase-K that did not include DNA isolation. Results indicate issues with inhibition in the proteinase-K modified method that could potentially be due to high sediment collected in the samples. The DNA extraction and isolation kit was proven to work, but at a cost 3 times the amount of the modified proteinase-K method. A fast, accurate method of processing environmental samples such as qPCR is a crucial step for successful restoration efforts of marine invertebrates, but cost must also be taken into account moving forward.

Our goal is to create an inverted pendulum in the form of a cube. An inverted pendulum is an object with a center of mass above its pivot point. Having the center of mass above the pivot point makes for an unstable system. The object will tend to fall over when it is not perfectly balanced, or when no external force is applied. To balance the object and keep it stable, a controlled counter force must be applied. Generating the counter force for our cube will be accomplished by using angular momentum. Controlling the angular momentum will be achieved through counter-torque generated by the three internal motors, each spinning on an axis perpendicular to the others. These motors will spin flywheels in order to store angular momentum. The stored angular momentum can be utilized by controlling the torque and direction of the motors. This will ultimately cause a net force to be applied to the cube in a direction determined by the sensor feedback and control algorithm. This project has its significance in the field of aerospace. For instance, having an object fixed in a certain angular position in the outer space. Moreover, the control system, which takes input of gyroscopes and accelerometers to control flywheels and motors, has various applications in the field of automobiles and construction machinery.  

Mammalian cardiac myocytes (CM) stop proliferating soon after birth, and heart growth afterwards predominately occurs with hypertrophy, increasing in cell size, rather than of hyperplasia, increasing in cell number. However under stress or injury stimuli, adult myocytes are able to re-enter the cell cycle and proliferate at a low rate. The lab of Dr. YiQiang Zhang has found that such processes are associated with histone epigenetic modifications important for gene transcription, and cellular dedifferentiation, resulting in adult CMs becoming more primitive and competent in re-entering the cell cycle. Additionally, many studies have suggested that the CREB-Binding Protein (CBP) and its homologue p300 act as transcriptional co-activators responsible for the acetylation of histone H3 residues lysine 18 and 27 (H3K18ac and H3K27ac), and contribute to other locus-specific histone acetylation events leading to cellular proliferation. CBP/p300 participates in transcriptional control through various mechanisms such as serving as a physical “bridge” between gene-specific transcription factors (GSTFs) and components of the basal transcriptional machinery or exhibiting acetyl-transferase activity. We hypothesize that the acetyl-transferase activity of CBP/p300, rather than the formation of a “bridge”, is responsible for the expression of genes during adult CM dedifferentiation and cell cycle re-entry. To test this hypothesis, we transfected C2C12 mice myoblast cells with a plasmid containing the CBP/p300 gene sequences as well as kinase-dead forms of both. Using quantitative-reverse-transcription-PCR and Western Blot, we can quantify the expression of the constitutive and inactive forms of CBP/p300 and compare it to the expression of essential cell cycle proteins, H3K18ac and H3K27ac. Our experiment is currently still ongoing, but data collected suggest that the enzymatic domain of CBP/p300 may be essential for re-entering the cell cycle. We hope that we can eventually apply our findings to work to adult myocytes which is highly demanded for regeneration in heart failure patients.

Our group built a drone that will unfoul utility lines. In other words, it removes shoes tossed over and around power cables. The current method of removing shoes of powerlines is expensive and dangerous for the workers involved. This is why we see so many shoes polluting our lines. We built a quad-rotor drone aircraft fitted with a special cutting device that is encapsolated in a specially engineered shell. Our main sensor is a camera and the visual inputs are used to identify the shoes on the utility line. The drone also has an extendable arm that cuts the shoelace. At the end of the arm is the boom which is a shelled device with a cutter inside. This project involved robotics, control systems, and computer vision. There are different versions of the drone, each one differing in the level of autonomy. The first version is human pilot operated:; the user controls the drone’s altitude, orientation, and cut. The other version is semi-automatic; the human takes the drone to the shoe's location. The drone's computer vision system which recognize the shoes and laces. It will then cut the laces itself and fly back to the user. 

Hepatocellular Carcinoma is the most common form of liver cancer accounting for a quarter of a million deaths annually across the globe. Microbubbles have been shown to be an effective vector for both gene and drug delivery. In combination with ultrasound, the microbubbles can be locally burst to deliver a drug payload via sonoporation – the temporary disturbance of the cell membrane. Ultrasound and microbubbles drug delivery is a strong candidate for the treatment of HCC because the liver that is easily accessible with ultrasound and the organ itself is highly perfused allowing for easy microbubble access. In microbubble gene and drug delivery, ultrasound parameters that induce cell death when used in combination with microbubbles are not well defined. While the literature agrees on general trends that greater exposure times and greater pressure lead to higher cell death, the specific combination of parameters that cause cell death are still disputed. We will design an enclosure such that cells can be cultured and exposed to ultrasound with microbubbles. We hypothesize that at greater exposure times, higher duty cycle, and greater pressures the cell viability will decrease. We hypothesize when multiple parameters are increased in combination, there will be a compounded decrease on cell viability. The rationale behind these hypotheses is that with greater power and force, greater stress and strain will be placed on the cell causing lowered cell viability. The goal of the project is to conclude specific values at which cell viability is compromised. From the study, it was found that while stronger acoustic parameters caused higher cell death, the variable that made the most difference was bubble concentration.